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Bone Microarchitecture Analysis Add

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1. 1 Download data set BMA SC micro CT AVW from File C Data BMA Data BMA SC micro CT avw WWW an alyzed i rect CO m d ata Intensities Flip Shift SubRegion Resize Pad Orient Output 2 Open Analyze 12 0 and select File gt Load As igi PR Axis Voxel Voxel Axis i Dim Size Size 3 Inthe Load As module use the File button to navigate to EL us and select the BMA SC micro CT AVW data set v 208 ai z 615 00 4 Inthe Load As window click on the Resize tab if the Memor 9272 Mb 92 72 Mb voxels are not isotropic check the Force Cubic option c o p nterpolation Linear Figure 5 1 5 Now select the Intensities tab FRECUENTES SUE LU 6 Note that Scanco data imports with a float data type To Hen ce decrease the volume size and thus decrease processing Figure 5 1 Load As Module time load the data set as signed 16 bit Select Signed 16 bit from the DataType drop down menu option under Output The Output Maximum and Minimum values will be set to the default for the 16 bit signed 32767 and 32768 Round the Maximum and Minimum values for the Input data and enter these values in the Output Maximum and Minimum fields Figure 5 2 Note that in this example the data set size reduces from 92 72 Mb to 46 36 Mb Input uy os Output E B Input 7 Output nm DataType Float Float iv DataType o Float Signed 16 bit kd Maximum 1229746777 12297467 Maimum 1229746777 12297 CEN LES li Lond Cort E Le en
2. In the Morph Object window returned set the following arameters p Figure 4 23 Hole in the a Object to Trabecular Tissue WARECO CSS b Operation to Fill Holes c Fill Type to 3 pass 2D d Transverse e Connectivity to 4 connected f Defined Object to Trabecular Tissue A Objects VolumeEdit S Next click Morph Figure 4 24 Control by e Object O Attribute a i Add m EE ee ES Object Larger holes and gaps in the Trabecular En Delete Tissue object are now filled and assigned Object Trabecular Tissue RE to the Trabecular Tissue object Note Desks HERES Object that this is also a quick way to reassign Bis Remove incorrectly identified regions of Cortex Omp OB 3pas2D Bui back to the Trabecular Tissue and could Transverse Coronal C Sagittal Hue provide a utility for Cortex to Trabecular Connectivity I Lead Tissue reassignment without having to Le se A ees conduct any manual reassignment as Defined Object Trabecular Tissue us described in the Reassigning Cortex to I Morph Trabecular Tissue steps above CRE Filter T Objects Once editing is complete use the File D Save Object Map option to save the Danse object map as Cortex edited obj and then close the Volume Edit module Figure 4 24 The Morph Object Window In the BMA add on module use the Load Segmented Cortex Object Map to load the Cortex edited obj map saved in step 60 a
3. 1999 2014 AnalyzeDirect Inc 1999 2014 BIR Mayo Clinic Contents 1 Introduction to the Bone Microarchitecture Analysis Add 0n 4 1 1 Bone Microarchitecture Analysis Workflow ss 5 1 1 1 Data Preprocessing OVErVIEW NT 5 1 1 2 Bone Microarchitecture Analysis Overview sn 5 1 1 3 Processing Single Bone SOSCIMENS wissuncisdensdverscitesilreiuadiasiserdsseisirsivesusundewdsidavilan 5 2 The Bone Microarchitecture Analysis Add on Interface Overview RL 6 ZN The Man BMA Program WINGOW ERE TM T PRA E a aes oe cere ESE PA A O E E E E E E T E E A E 8 P o C M 8 KAMERUN 8 LRO GNI E P 9 ERER E a E E PIE Pautas raat cusses av IS hpc ee MUN Pat 10 FANS M Tere erc iE 10 er Sty SCI NU T TN TNT NEMO 11 Nel r iBI e EU UU NU ETUR 11 Bea Rondernng WiNGOW MR UU I TT 12 REM DMO SE IN PAPAS nas a a a ein nn 14 2 6 BMA Workflow TaDS PERENNE T on 14 eo E O ame E NETS 14 220 2 es We Al EP E RR NEC 15 2 5 MON UO ON a E s ncauch n NI UR KO nIS IPM N RECIEN IUE EDDIE RM DUDEN uU tU REDEMIT DRE 17 3 Bone Microarchitecture Analysis Measurements ss 19 3 1 Trabecular Bone Microarchitecture Measurements senes 19 3 2 Cortical Bone Morphology Measurements ss 20 4 Bone Microarchitecture Analysis PerkinElmer QuantumFX microCT Data Processing
4. e Cortical Moments Values per transverse section saved in a file named VolumeName M csv e 3D Trabecular Thickness This produces histograms of trabecular thickness and intertrabecular space over the entire 3D extent of the bone The histogram csv files VolumeName TTH csv for trabecular thickness and VolumeName TSBP csv for intertrabecular space declare the range of thicknesses in mm in each bin in the first line and the values in the following lines are the volume of trabecular bone or intertrabecular space in mm of that thickness The mean and standard deviation of these measurements are stored in VolumeName TrSt csv e 3D Trabecular Structure and Connectivity Two global characterizations of trabecular structure that are also stored in VolumeName_TrSt csv e 2D Characterization Multiple values per transverse section in file VolumeName Ch2D csv Note that detailed descriptions of each measurement generated for each group of bone measurements are provided in the next section 3 Bone Microarchitecture Analysis Measurements Measure Bone Begins the measurement process All the measurement groups selected will be performed When complete the csv file viewer will be displayed Show Measurements If you dismiss the csv file viewer it can be recalled using the Show Measurements button Analyze lirecl 18 3 Bone Microarchitecture Analysis Measurements Bone Mineral Density BMD mg cm is an important
5. 23 DA a N eT ET DT Em 23 dels IDEO SUE ct RE 23 cm AB cuoio d acte B TI o UOTE 23 4 1 3 S bregioning DEA na nbniaasEi nick ragircna dick inta ducta o Unc dal aad ed Ema Ua rd und ce 2D LN M elt qais EB ir NE Rm 21 41 9 Data PRES STE OMR nee es ee nn 28 v AnalyzeDirect AE VU Ce ESS de EET TO TET 30 ORENK E ON ENNIO 30 4 2 2 Manual Refinement of Cortex Object Map using Volume Edit 32 4 2 2 1 Reassigning Cortex to Trabecular TiSSue nn ain san initie 32 4 2 2 2 Filling Holes and Gaps in the Trabecular Tissue 33 4a s Parco apro e ili TIE 34 4 2 4 Manual Refinement of the Final Object Map using Volume Edit 36 4 2 0 Measure m6 1 Erbe Rm ENERE ANT EDS EEEE Drag dixi ed CVM NH AE PEEPU IUE 37 5 Bone Microarchitecture Analysis Scanco microCT Data Processing 38 9 1 Data e eccle NT mE 38 BOR d SP LOS Re dam adul b Er UHR MM FEIN ESTEE D cietansde E TE E NEUE 38 9 12 Da OS eI RET NE o E PT MT ERR 39 De roure oning Dalase ei TL De a 40 SLA Dala alice Bici e TTE E a S QE 42 siu uiae ecco MM ITE 44 5 2 2 Manual Refinement of Cortex Object Map using Volume Edit 46 5 2 2 1 Reassigning Cortex to Trabecular Tissue 46 5 2 2 2 Filling Holes and Gaps in the Trabecular TISSUB iesus is Rd lxs Ca Mese via bus sri ue Cobra Eds 47 VE Ae eR LE sc nioa tartan ce wrechearsantiovainansciaeaiaenatnatamiage toa reakasiensrsbiunalicedeandainecnianecad
6. Amling M P sl M Hahn M amp Delling G The thickness of human vertebral cortical bone and its changes in aging and osteoporosis a histomorphometric analysis of the complete spinal column from thirty seven autopsy specimens J Bone Miner Res 12 89 95 1997 Arlot M E et al Histomorphometric and microCT analysis of bone biopsies from postmenopausal osteoporotic women treated with strontium ranelate J Bone Miner Res 23 215 22 2008 Umemura Y Ishiko T Yamauchi T Kurono M amp Mashiko S Five jumps per day increase bone mass and breaking force in rats J Bone Miner Res 12 1480 5 1997 Wallace J M et al Exercise Induced Changes in the Cortical Bone of Growing Mice Are Bone and Gender Specific Bone 40 1120 1127 2007 Giladi M et al Stress Fractures and Tibial Bone Width J bone Jt Surg 69 B 326 329 1987 Stock J T amp Shaw C N Which measures of diaphyseal robusticity are robust A comparison of external methods of quantifying the strength of long bone diaphyses to cross sectional geometric properties Am J Phys Anthropol 134 412 23 2007 Zanchetta J R et al Effects of teriparatide recombinant human parathyroid hormone 1 34 on cortical bone in postmenopausal women with osteoporosis J Bone Miner Res 18 539 43 2003 Jiang Y et al Recombinant human parathyroid hormone 1 34 teriparatide improves both cortical and cancellous bone structure J Bone Miner
7. intial Object Map OMA PE micro CT reform Bone Object Bone Segment Cortex BMD Scaling Parameters SigmaCT Slope 1 0684 BetaCT Offset 1023 2 Point Ped 7 118 27V VA s Figure 2 19 BMA Segment Cortex Analyze lirecl 14 Cortex Thresholds Provides access to a threshold slider bar to allow for segmentation of the cortex Note that the module will automatically compute the minimum threshold value via histogram analysis however this may be adjusted by the user before segmenting the cortex Threshold Rendering This option turns on and off the interactive 3D rendering display as the threshold slider changes It is useful to switch off this option if you are working with extremely large data sets and the interactive 3D update is slow Use Initial Segmentation If the image volume is a single bone this option is not needed If the image volume contains several bones it must be pre segmented by use of an object map to indicate the bone of interest This pre segmentation will only be used to avoid segmenting and measuring the other bones in the image The cortex and trabecular bone will still be identified by the segmentation routines in the program If the module finds an object map that has the same name as the input volume the module will automatically set this option and the object map will be loaded Initial Object Map Opens a file browser to allow users to navigate to and select the pre segmentation object map
8. and postmenopausal osteoporotic women receiving strontium ranelate experienced a significant increase in Ct Th Periosteal perimeter Ps Pm mm is the perimeter of the periosteum the membrane around the outside of the cortex and is calculated for each transverse section Ps Pm was found to increase as a result of jump training and strontium ranelate treatment in female rats and as a result of treadmill running in male mice Endocortical perimeter Ec Pm mm is the perimeter of the endosteum the membrane that forms the boundary between the cortex and medulla and is calculated for each transverse section Ec Pm was found to increase along with periosteal perimeter Ps Pm above as a result of treadmill running in male mice indicating endocortical resorption along with periosteal formation of bone Moment of inertia about the anteroposterior axis I mm is a measure of how material is distributed about the anatomical anteroposterior front to back axis through the area centroid of each slice Area moments of inertia are also known as second moments of area or SMAs and characterize resistance to bending around a given axis Exercise has been shown to increase ls in the tibia of male mice Moment of inertia about the mediolateral axis l mm is a measure of how material is distributed about the anatomical mediolateral left to right or right to left axis through the area centroid of each slice Area moments of inertia ar
9. 4 11 Spatial Filters Save Option 26 Close the Spatial Filters module a AnalyzeDirect 27 4 1 5 Data Presegmentation This data set contains multiple bones therefore it is necessary to conduct a preliminary segmentation to isolate the bone of interest This will be used as an input in the BMA add on module to guide the cortical segmentation process 27 Select the filtered data BMA PE micro reformat sub Med and open Segment gt Volume Edit 28 Inthe Object control panel click the Add Object button and name the object Bone Figure 4 12 29 Next select the Semi Automatic tab and choose the Region Grow option a Original C Name Bone Color Red 5 Add Objet p Display Locked Figure 4 12 Volume Edit Object Control Panel 30 Click on the bone of interest in any of the 3 orthogonal displays to set a seed Figure 4 13 then use the threshold slider to define a global threshold range that represents bone Now click Extract Object Figure 4 14 All voxels that fall within the specified minimum and maximum threshold range and that are spatially connected to the seed will be assigned to the Bone object For this data set a Min value of 3745 and a Max value of 9463 works well Edge Strength Walls Semi Automatic Manual IC Browse Threshold c Region Grow Connected Components Object Extractor _ Object Separator Oblique Cutting C3 Fill Holes _
10. 8 Next select Apply then Save and then Done Save Restore Select All Figure 5 8 The Image Calculator Button Configuration Tool a AnalyzeDirect 40 18 19 20 21 22 23 From the main Image Calculator window click the Region Pad button Region the Subregion window will open Figure 5 9 Pad SubRegion Low High X 80 292 Y Fd Figure 5 9 The Subregion Tool In the Subregion window click the Interactive button In the Interactive window Figure 5 10 set the yellow box around the bone of interest Use the orientation buttons to navigate between views Once the crop is set in all three orientations click Apply When prompted select Save a Copy of the Loaded Data The cropped data will be saved to the Analyze 12 0 workspace Close the Image Calculator module Select the subregioned data set from the Analyze 12 0 workspace and then right click and select Rename from the menu Change the name of the data set from BMA SC micro reformatO to BMA SC micro reformat sub j m D U a AnalyzeDirect Figure 5 10 The Interactive Tool 41 5 1 4 Data Presegmentation This data set contains multiple bones therefore it is necessary to conduct a preliminary segmentation to isolate the bone of interest This will be used as an input in the BMA add on module to guide the cortical segmentation process 24 Select the
11. AnalyzeDirect 33 DMD ating Parameters Sigmalt Hope 1 0 4 2 3 Segment Trabeculae The second step of the BMA process provides for segmentation of the Trabecular region into Trabecular Bone and Intra Trabecular Space This segmentation is achieved via threshold based segmentation 62 To segment the Trabecular region into the Trabecular Bone and Intra Trabecular Space objects set a threshold range using the Trabeculae Thresholds slider Figure 4 25 W e View CHher Piein Bee 2e Sgen ceama Moi Certes Trabeculse Bone Save Secgmemted Corker Lead Segmented Cortes Object Map Cyect Map Tisbeculsr Threchalk Min HERS EF I 4 ET ist Thrachold Rendering zular Tissue trabecular Dore Betac tse 0 0 128 Clap Eiche Ea Figure 4 25 Segment Trabeculae Tab with Thresholds Set to Segment Trabecular Bone e AnalyzeDirect 34 63 Once the threshold has been specified click the Segment Trabeculae button The voxels specified by the threshold range will be assigned to the Trabecular Bone object The module will automatically proceed to the Measure Bone tab Figure 4 26 File View Other Help Bee Om f Segment Segment Measure Cortex Trabeculae Bone Save Final Load Final Otyect Map Object Map Measurement Dets Directory CuData Data tor AnehyzeDvect Data Measure Optsonn V Bone Mineral Densrty v Volurmes Surface Areas and Ratios Cortical Porosity Cortical
12. Off All Slices C Slices Specified in Slice Menu Sequence Display On _ Off Stat Type STEREO Et Reset Sum Sample Intensity _ 2D Shape _ 3D Shape _ Fractal Boundary Coordinates Region Pixels Decimal Places 2 Log Stats On Off Configure Log Stats Figure 6 5 The ROI Sample Options Window 8 AnalyzeDirect 53 6 2 Calculation of SigmaCT and BetaCT 19 14 19 16 Import the measurements into MS Excel as a space delimited file y 1 06843x 1023 2 2000 R 0 989 Plot the grayscale value versus mineral density of the phantom xs Create a linear regression of measured grayscale value versus BMD Figure 6 6 a a m a m Le S m L i Record the slope SigmaCT and offset BetaCT values In the example shown Figure 6 6 SigmaCT E 400 ux soni 1250 1 0684 and BetaCT 1023 2 Eq BMD mg HA cm 3 Figure 6 6 Plot of the Grayscale Values vs Mineral Density of Phantom 6 3 Conversion of CT numbers to Bone Mineral Density 17 Select the bone image data and open the BMA add on module from the Apps menu 18 To calibrate the image data to BMD enter the slope SigmaCT and offset BetaCT values in the BMD Scaling Parameters in the lower left hand corner of the BMA module interface Figure 6 7 Note that these values can be entered at any time in the BMA process prior to calculation of bone measurements For da
13. Res 18 1932 41 2003 Nishiyama K K Macdonald H M Buie H R Hanley D A amp Boyd S K Postmenopausal women with osteopenia have higher cortical porosity and thinner cortices at the distal radius and tibia than women with normal aBMD an in vivo HR pQCT study J Bone Miner Res 25 882 90 2010 Kazakia G J et al Age and Gender Related Differences in Cortical Geometry and Microstructure Improved Sensitivity by Regional Analysis Bone 52 623 631 2013 Sietsema W Animal Models of Cortical Porosity Bone 17 2978 3058 1995 Patsch J M et al Increased cortical porosity in type 2 diabetic postmenopausal women with fragility fractures J Bone Miner Res 28 313 24 2013 Maurel D B Boisseau N Benhamou C L amp Jaffr C Cortical bone is more sensitive to alcohol dose effects than trabecular bone in the rat Joint Bone Spine 79 492 9 2012 Maurel D B et al Low bone accrual is associated with osteocyte apoptosis in alcohol induced osteopenia Bone 49 543 52 201 1 Li X et al Sclerostin antibody treatment increases bone formation bone mass and bone strength in a rat model of postmenopausal osteoporosis J Bone Miner Res 24 578 88 2009 Analyze lirecl 56
14. a median filter to reduce noise Note when using QuantumFX data a For the 30mm fast scan a median filter with an 11x11x11 kernel works well b For the 24mm fast scan a median filter with a 5x5x5 kernel works well 4 Select the QRM BMD phantom data and open Measure Region of Interest 5 Use the slice slider to navigate into the data set Use the Oval tool to define a circular ROI over one of the inserts Figure 6 1 Note there may not be contrast insert for 0 in this case define the ROI anywhere in the phantom casing 6 Select the Copy ROI button and copy the ROI to the 4 remaining inserts Figure 6 2 Inberuec Ber became part of Dbi Te Define A Lichred 7 Exrbna Hee The T uni Inberuechorn bone part of i 4D Mese d 3D Mise A0 Mess Dep ng Tpi m HH Mays Dae AUS yc 3283 FE Valac EUR Figure 6 1 Defining Inserts in a BMD Phantom with ROI Figure 6 2 Copy Regions a AnalyzeDirect 52 9 10 11 12 Go to View gt Objects and rename objects with associated mg CC values Figure 6 3 Determine last slice where objects should be defined then open Generate gt Slice a Set Slice to the current slice first slice with objects b Set Increment to number of slices between first slice and last desired slice Next click the Copy Regions Forward button opy Open Tools Propagate Regions Check all of the objects except Original Then click the Pro
15. aging does Total pore volume Po V mm is the total volume occupied by cortical pores in the volume of interest Average pore volume AvgPo V mm is calculated as the total pore volume Po V divided by the pore number Po N AvgPo V is higher in postmenopausal than in premenopausal women and higher in women than in men s Pore size has also been shown to be larger in diabetic postmenopausal women exhibiting fragility fractures As mentioned above the increase of cortical porosity with aging is understood to be due to an increase in average pore volume rather than an increase in pore number Standard deviation of pore volume Po V SD mm is a measure of the heterogeneity in size of cortical pores in the volume of interest Diabetic postmenopausal women exhibiting fragility fractures had a higher variability in pore size Pore density Po Dn 1 mm is calculated as the pore number Po N divided by the total cortical compartment volume Ct V giving the number of pores per unit volume in the cortex Analyze lireci 22 4 Bone Microarchitecture Analysis PerkinElmer QuantumFX microCT Data Processing 4 1 Data Preprocessing 4 1 1 Data Loading The BMA add on module requires that input bone specimens have isotropic voxels To ensure your image data is loaded with isotropic voxels load the data using the Load As module and resample anisotropic data using the Resize option 1 Download data set BMA PE micro CT AVW from
16. are listed below Tables 1 and 2 For further information about each bone measurement calculated please refer to section 3 Bone Microarchitecture Analysis Measurements Table 1 Trabecular Bone Morphometric Indices Table 2 Cortical Bone Morphometric Indices Abbreviation Description Unit Abbreviation Description Unit TV Total Volume mm Tt Ar Total Cross Sectional Area mm BV Bone Volume mm Ct Ar Cortical Bone Area mm BS Bone Surface mm Ma Ar Medullary Area mm BV TV Bone Volume Fraction Ct Ar Tt Ar Cortical Area Fraction BS TV Bone Surface Density mm mm Ct Th Average Cortical Thickness mm BS BV Specific Bone Surface mm mm Ps Pm Periosteal perimeter mm Conn D Connectivity Density 1 mm Ec Pm Endocortical Perimeter mm SMI Structure Model Index a Moment of Inertia mm Tb N Trabecular Number 1 mm Anteroposterior Axis Tb Th Trabecular Thickness mm b Moment of Inertia mm Mediolateral Axis HERD Uaec aee pde MOD d I Maximum Moment of Inertia mm Tb Th SD Standard Deviation of mm Trabecular Thickness are Minimum Moment of Inertia mm Tb Sp SD Standard Deviation of mm J Polar Moment of Inertia mm Trabecular Separation Ct Po Cortical Porosity DA Degree of Anisotropy Po N Pore Number n MIL Mean Intercept Length Po V Total Pore Volume mm AvgPo V Average Pore Volume mm Po V SD Standard Deviation of Pore mm Volume Po Dn Pore Density 1 mm Analyze Il 1 1 Bone Microarchitecture Analysis Workfl
17. dominant trabecular structure Perfect plates are represented by 0 perfect rods by 3 and perfect spheres by 4 Most real trabecular structures will lie somewhere between the ideal plate and ideal rod structure At very high or low BV TV values the SMI may be outside of the defined range The SMI impacts the mechanical properties of trabecular tissue In the femoral neck and vertebrae the SMI increases significantly with age moving Analyze lireci 19 from a plate like structure to a weaker rod like structure as trabeculae undergo perforation and or thinning mechanisms SMI has been shown to exhibit a strong negative correlation with BV TV As bone volume fraction decreases trabeculae tend to become more rod like Different bones tend to have different trabecular structure the lumbar spine tends to be more rod like and the femoral head tends to be more plate like for example Trabecular number Tb N 1 mm is the average number of trabeculae per unit length Tb N is calculated by finding the mean distance between mid axes of the trabecular structures then taking the inverse This mean distance is determined using a 3D sphere fitting method Low Tb N is significantly associated with vertebral fracture and the decrease in BV TV with age Tb N tends to be larger in the femoral head and smaller in the lumbar spine Trabecular thickness Tb Th mm is the mean thickness of trabeculae as determined by a 3D sphere fitting met
18. for specimens containing multiple bones Bone Object If the object map contains multiple objects the Bone Object drop down menu allows users to select objects corresponding to the bone of interest Segment Cortex The Segment Cortex button begins the cortical segmentation process Depending on the input data and the resources of the system this process could take several minutes Segmentation progresses by transverse slice and varies in speed by the characteristics of the section Mid shaft sections are generally fastest sections containing significant trabecular tissue and or articulating surfaces are slower and sections with cortical defects or traumatic damage can be slower to resolve 2 6 2 Segment Irabeculae The Segment Trabeculae tab is displayed at the end of the Segment Cortex procedure Figure 2 20 The cortical segmentation algorithm has been designed to perform well on intact long bones but it may make minor errors on unusually shaped bones or on complicated sections near the metaphysis or on sections containing cortical defects On this tab the user may save the cortical segmentation as an object map and use any of the manual segmentation tools in Analyze to correct regions believed to be inappropriately assigned Likewise this tab may be selected when starting the program and a corrected object map loaded to proceed At this point the trabecular bone and intertrabecular space are combined as a single object as this is easier to
19. for the selected orientation is From Size Menu this means the display size of the selected orientation is dictated to that From Size Menu Size F set globally via the Size menu or the increase decrease size buttons Quarter s on the toolbar The other options will apply the selected size action to Third the selected slice orientation only BgEEHEREEEEEE E Halve 7 Single Double Triple Quadruple 2 4 2 Rendering Window The upper right quadrant is the rendering quadrant which displays a 3D rendering of the image volume Figure 2 13 Right Click Size Menu Linked Cursor A left click on this rendered surface Figure 2 14 will move the linked cursor to that surface voxel and select all three sectional images Figure 2 14 The Linked Cursor Tool Rotating Holding the middle mouse button down in the rendering quadrant will enable interactive rotation of the rendering viewpoint For those users without a middle mouse button the Ctrl key left mouse button will enable interactive rotation of the rendering viewpoint a Analyze 12 Right click menu Right clicking in the rendering quadrant will open the right click menu Figure 2 15 The right click menu provides access to the following options e Reset Rotation Provides the ability to reset the rendering display to one of the 6 orthogonal viewpoints Front Back Left Right Top and Bottom Reset Rotation e Size When selected provides acc
20. set is another optional step Image filtering can help improve the segmentation of the cortex and is often a necessary step for segmentation of data acquired on certain microCT systems 21 Select the subregioned data set BMA_PE_micro_reformat_sub from the Analyze 12 0 workspace and open Process gt Spatial Filters 22 To visually review any filter applied to the image data prior She TESI to filtering open Generate Preview Options in the Preview SSS hy Options window returned Figure 4 9 set the following sha os BE L a Loaded and Preview Volumes Number _ 1 b Slice to 233 EE c Number to 1 Done 23 Next open Generate Filters In the Filters window set a Filter Type to Median b Set the Kernel size to 5 by 5 by 5 Figure 4 9 Preview Options Window 24 Click the Preview button to review the original slice 233 and a copy of slice 233 with the selected filter and kernel size applied Figure 4 10 Figure 4 10 Side by Side Display of Unfiltered and Median Filtered Image 25 Next click Filter to apply the filter to the entire data set When prompted select Change a Copy of the Loaded Volume Figure 4 11 A filtered copy of the data set named BMA PE micro reformat sub Med will be saved to the current Analyze 12 0 workspace This action Filtering modifies the loaded volume Change the Change a Copy of Cancel Loaded Volume the Loaded Volume Don t Change Anything Figure
21. surface density and specific bone surface as defined below Bone volume fraction BV TV 95 is the ratio of the segmented bone volume to the total volume of the region of interest Trabecular BV TV has been found to be lower in patients who have sustained vertebral fracture BV TV decreases with age in both women and men contributing to osteoporosis Bone surface density BS TV mm mm is the ratio of the surface area of mineralized bone to the total volume as defined above BS TV decreases with aging as bone volume decreases so does bone surface area Specific bone surface BS BV mm mm is the ratio of the bone surface area to the volume of mineralized bone BS BV remains constant with aging suggesting that the shape of trabecular microarchitecture rod like plate like or a combination does not change much with age Connectivity density Conn D 1 mm is a measure of the degree of connectivity of trabeculae normalized to the total volume Conn D contributes to bone strength and decreases significantly with age in the femoral neck and vertebra In women the lower levels of estrogen associated with menopause lead to an increase in osteoclastic resorption which causes the perforation of trabeculae and thus a decrease in Conn D Trabecular disconnectivity due to various causes can also lead to vertebral fractures in men Structure model index SMI is a dimensionless number ranging from O to 4 that describes the
22. www analyzedirect com data 2 Open Analyze 12 0 and select File Load As 3 Inthe Load As module use the File button to File C Oata BMA Data BMA PE micro CT AVW avw Intensities Flip Shift SubRegion Resize Pad Orient Output Input Output Axis Voxel Voxel Dim Size Size A 31 2 D Y 512 0 02 pi 31 2 uL Memory 256 00 Mb 296 00 Mb Interpolation Linear ID navigate to and select the BMA_PE_micro_ CT AVW data set and then click Open 4 Inthe Load As window click on the Resize Auto Exit After Load Size 256 00 Mb tab and then if the voxels are not isotropic check the Force Cubic option Figure 4 1 Load Preview Cancel 5 Select the Load button to load the data set Figure 4 1 Load As Module into the Analyze workspace 4 1 2 Data Reorientation The BMA add on also requires that the input bone specimens are orientated into the correct anatomical orientation Figure 4 2 The Oblique Sections module provides the tools necessary to quickly reorientate the data 6 Select the BMA_PE_micro_CT data set from the Analyze workspace and open Display gt Oblique Sections 7 Select Tools gt Orthogonal Figure 4 2 Correct Bone Orientation for BMA a AnalyzeDi 23 8 Use the Interactive Perp Axis tool to draw a line down the center of the bone to be analyzed To do this click on the image and drag the cursor to a new position in this example this is done in the coronal orientation Figur
23. Auto Exit After Load Size 02 72 Mb Auto Exit After Load ize 46 36 Mb Load Preview admo Cancel Load Preview do Cancel Figure 5 2 Converting Float Data to Signed 16 bit Data 7 Select the Load button to load the data set into the Analyze workspace 38 a AnalyzeDirect 5 1 2 Data Reorientation The BMA add on requires that the input bone specimens are orientated into the correct anatomical orientation Figure 5 3 The Oblique Sections module provides the tools necessary to quickly reorientate the data 8 Select the BMA_SC_micro_CT data set from the Analyze workspace and open Display gt Oblique Sections 9 Select Tools gt Orthogonal 10 Use the Interactive Perp Axis tool to draw a line down the center of the bone to be analyzed To do this click on the image and drag the cursor to a new position in this example this is done on the sagittal orientation Figure 5 4 An oblique plane Shown in yellow will be defined perpendicular to the red line Figure 5 3 Correct Bone Orientation for BMA Figure 5 4 The Orthogonal Tool 11 Note that the end of the bone we are interested in analyzing is at the bottom of this data set To orientate this to the top of the data set open Tools gt Fly from the main Oblique Sections window Set the Fly Value to 180 and then click the Pitch Up button once Figure 5 5 Pitch dh Figure 5 5 The Fly Tool Pitching 180 Degrees a AnalyzeDirect 39 12 Now set
24. Moments v 30 Trabecular Thickness V 30 Trabeculae Structure and Connectivity v 7D Charactenmabon 200 12 j Figure 4 26 The Measure Bone Tab Showing the Segmented Trabecular Bone in Orange c v 64 Atthis point in the workflow the user will again need to visually review the segmentation result to determine if the trabecular bone segmentation is acceptable or not If the result is acceptable then processing can continue and the user can proceed to the Measure Bone step step 76 If the segmentation result requires manual refinement the user has the option to output and modify the object map using the Volume Edit module available within Analyze 12 0 The steps neccasary for manual refinement are given in section 4 2 4 65 Use the linked cursor tool to inspect the 3 orthogonal slice displays and the 3D rendering of the bone to review the trabecular bone region segmentation result 66 To save the object map select the Save Final Object Map button Name the object map Final obj 67 Minimize the BMA add on module do not close the module Analyze lirecl 35 4 2 4 Manual Refinement of the Final Object Map using Volume Edit Here we will review manual segmentation tools in the Volume Edit module that will allow for the correction of small regions of trabecular bone identified as cortex Object map refinement will improve the accuracy of the measurements generated by the BMA add on module 68 With the BMA_PE_micro_reformat_sub d
25. Propagate Objects Reassign Object Min Threshold 37544 e M Threshold Rendering Extract Object Ret Seed Reset Limits Redo j Undo Figure 4 13 Region Grow Seed Point Figure 4 14 Volume Edit Semi Automatic Tab 31 In the Object control panel add a new object and name it Extra v AnalyzeDirect 28 32 From the Semi Automatic tab select the Object Separator tool Click on a bone that is not the bone of interest this will set the first seed point location Next click on the bone of interest this will set the second seed point Figure 4 15 33 Click Separate to disconnect the two bone objects The area of bone selected first should be disconnected from the main bone object and assigned to the Extra object Figure 4 16 Figure 4 15 Object Separator Seed Points Figure 4 16 Rendering of the Separated Bone Objects 34 Continue to disconnect any superfluous bones surrounding the bone of interest 35 Once all bones surrounding the bone of interest have been reassigned to the Extra object delete the object by clicking on the trash can button Figure 4 17 Le Pde CWamw Other Help Bee 15659 Color Era Add Object A Display D Load D Edge Strecsih Walls Serm oberste Manual ke Edge Sirena Figure 4 17 Volume Edit Segmented Bone of Interest 36 Select File Save Object Map Name the object map initial obj Close the Volume Edit module AnalyzeDirec 4 2 BMA P
26. a Transverse Rendering Coronal i Sagittal Figure 2 11 The BMA Display Screen 2 4 1 Slice Display The upper left quadrant displays the selected transverse image from the volume the lower left the selected coronal image and the lower right the selected sagittal image Linked Cursor A linked cursor identifies a single voxel in all three sectional images Figure 2 12 Aleft click in one of the sectional Eom Dore A images moves the cursor to that point and selects the other two sectional images displayed The left button may be held down in one of the sectional images to continuously scroll the cursor and the other two images Figure 2 12 The Linked Cursor Tool Arrow Keys The arrow keys can also be used to move the linked cursor The keys will move the linked cursor in the corresponding direction up down left and right for the active orientation Page Up and Down Keys The page up and page down keys can be used to move through the slices in the active orientation a AnalyzeDirect 11 Sliders The sliders below each slice display quadrant can be used to navigate through the slices in that orientation The middle mouse scroll button can also be used to move through the slices this will provide the ability to scroll through slices in the active orientation Right Click Menu Right clicking on any of the slice display quadrants provides access to a size menu Figure 2 13 The default Size option
27. a spreadsheets will be saved Figure 5 25 Directory C Data BMA Data Statistics 71 By default all of the Measure Options Figure 5 26 Figure 5 25 Directory Selector for will be checked Uncheck any option you are not Specification of Output Directory interested in Measure Options Measure Bor Bone Mineral Densit 72 Click the Measure Bone ri ty button to start the measurement process Note Volumes Surface Areas and Ratios it could take several minutes to calculate all of the bone morphometric indices Once the Cortical Porosity measurements have been generated they will be displayed in a new tabbed window Figure 5 27 Each of the tabs provides access to a different set of measurements Cortical Moments 3D Trabecular Thickness ee 3D Trabecular Structure and Connectivity 73 Right clicking on a row or column will allow users to plot the row or column when supported TP handra bon 74 When the process is complete several csv files will Figure 5 26 Measurement Options be saved to the location specified in step 70 The csv files can be opened in MS Excel or any other stats analysis package All Measures BMD CP Ch2D M TSP TTH TrSt VSR am i File slice angle measurement BMA SC micro reformat sub all all CortexMean BMD BMA SC micro reformat sub all all CortexStd BMO BMA 5C micro reformat sub all all IntraTrabecularMean BMD BMA SC micro re
28. an object select the color square next to the object name Segment Trabeculae When pressed the Segment Trabeculae button commits the threshold decision to the object map populating the Trabecular Bone object Analyze ire 16 2 6 3 Measure Bone The Measure Bone tab Figure 2 21 is displayed at the end of the Segment Trabeculae procedure The final segmentation may be saved as an object map if corrections are desired or the tab can be selected when data is loaded and a corrected final object map loaded This tab allows the selection of the specific bone measurements desired all measurements are selected by default and initiates the measurement process File View Other Help Be om Segment Segment Measure Cortex Trabeculae Bone Save Final Load Final Object Map Object Map Measurement Data i C Data Measure Options V Bone Mineral Density Volumes Surface Areas and Ratios v Cortical Porosity v Cortical Moments 3D Trabecular Thickness V 3D Trabecular Structure and Connectivity v 2D Characterization Measure Bone BMD Scaling Parameters SigmaCT Slope 1 0684 BetaCT Offset 1023 2 Point Picked 82 118 239 Value 940 Object Original Figure 2 21 BMA Measure Bone Each measurement group is saved to a comma separated values csv file named using the image volume name and an additional extension described below The first line o
29. ata set selected open Segment gt Volume Edit 69 Select File gt Load Object Map and load the Final object map saved in step 66 0 In the Coronal orientation use the slice slider to navigate through the slice display Note that there are a number of cortical regions shown in red that need to be assigned to the trabecular bone shown in orange 71 Lock the Intra Trabecular Space Tissue yellow object Figure 4 27 then select the Trabecular Bone object and then click on the Manual tab and select the fed Trace tool S lorgina EB cortex E cortical Pores CT S Trabecular Bone 72 Draw around the regions of cortex to reassign them to the trabecular bone Figure 4 28 Figure 4 27 Lock Intra Trabecular Space 73 Continue working through the slice data to correct regions in this manner When completed select File gt Save Object Map and save the object map as Final edited obj Figure 4 28 Steps to Reassign Incorrectly Assigned Regions to the Trabecular Bone 74 Close the Volume Edit module 75 In the BMA add on module use the Load Final Object Map to load the Final edited obj map saved in step 73 a AnalyzeDirect 36 4 2 5 Measure Bone The third and final step of the BMA processing workflow is the generation of the bone morphometric indices 76 Specify the directory where all of the measurement Measurement Data data spreadsheets will be saved Figure 4 29 Di ssmnmumamuunuuuuunuunuau
30. ataniues 48 5 2 4 Manual Refinement of the Final Object Map using Volume Edit 50 D WS US DONG ne de UN ee de ee a LR RUD ei Scd DUDEN 51 em ci Egi AU PASSER SR EP NUESTRAS 52 SDS SAS gE NS METTE I E 52 6 2 Calculation f SigmaC T and BOE PE rrai i 54 6 3 Conversion of CT numbers to Bone Mineral Density ccccsecceeeceeeeseecaeeeceeeceeeeeaeeesaeenaees 54 ES SN LE PR TII LN UN TE 55 v AnalyzeDirect 1 Introduction to the Bone Microarchitecture Analysis Add on The Bone Microarchitecture Analysis BMA module is a new add on available for Analyze 12 0 The BMA add on is a powerful pre clinical research application designed for the evaluation of 3D CT and microCT image data The module provides researchers the ability to process single whole bone specimens and isolated 3D samples of trabecular tissue It generates an extensive set of bone morphometric indices as described by Bouxsein et al The BMA add on provides automatic and user guided isolation of bone from non bone tissue by means of a novel segmentation process This process also provides for the separation of the cortical and trabecular bone in whole bone specimens BMA uses the segmented regions to automatically drive the calculation of common bone morphometric indices that provide researchers with a quantitative description of bone microarchitecture The microarchitectural characteristics of the trabecular and cortical bone
31. correct than the detailed object map Analyze lirecl 15 75 File View Other Help Eee Segment Segment Measure Cortex Trabeculae Bone Load Segmented Cortex Object Map Save Segmented Cortex Trabeculae Thresholds Min Max 10364 441501 v Threshold Rendering Segment Trabeculae Ei Origina mE cortex cortical Pores trabecular Tissue trabecular Bone BMD Scaling Parameters SigmaCT Slope 1 0684 BetaCT Offset 1023 2 Point Picked 82 118 239 Value 940 Object Original Figure 2 20 BMA Segment Trabeculae Save Segmented Cortex Object Map This option opens a save window allowing the user to save the segmented cortex object map Load Segmented Cortex Object Map This open opens a load window allowing the user to load a corrected cortex object map Trabeculae Thresholds The minimum threshold value is set from the Cortical Thresholds slider on the Segment Cortex tab This is usually the best threshold to use but the user may adjust this slider if a different segmentation is desired within the trabecular compartment As the slider is adjusted the sectional images show the resulting trabecular bone Object Map List The list of objects shows the color associated with each object and whether the object is displayed in the rendering or not To turn off the display of an object select the monitor icon next to the object name To change the color of
32. crinol 2013 213234 2013 Stauber M amp M ller R Age related changes in trabecular bone microstructures global and local morphometry Osteoporos Int 17 616 26 2006 Parkinson amp Fazzalari N Characterisation of trabecular bone structure Stud Mechanobiol Tissue Eng Biomater 5 31 51 2013 Hildebrand T Laib A M ller R Dequeker J amp R egsegger P Direct Three Dimensional Morphometric Analysis of Human Cancellous Bone Microstructural Data from Spine Femur lliac Crest and Calcaneus J Bone Miner Res 14 1999 Burghardt A J Link T M amp Majumdar S High resolution computed tomography for clinical imaging of bone microarchitecture Clin Orthop Relat Res 469 2179 93 2011 Macdonald H M Nishiyama K K Kang J Hanley D a amp Boyd S K Age related patterns of trabecular and cortical bone loss differ between sexes and skeletal sites a population based HR pQCT study J Bone Miner Res 26 50 62 2011 Nishiyama K K et al Cortical porosity is higher in boys compared with girls at the distal radius and distal tibia during pubertal growth an HR pQCT study J Bone Miner Res 27 273 82 2012 Lieberman D E Polk J D amp Demes B Predicting long bone loading from cross sectional geometry Am J Phys Anthropol 123 156 71 2004 Burghardt A J Kazakia G J Ramachandran S Link T M amp Majumdar S Age and gender related differences i
33. ct map initial obj a AnalyzeDirecl 43 5 2 BMA Processing Once data preprocessing is complete the image data is ready for processing with the Bone Microarchitecture Analysis add on module 30 Select the BMA_SC_micro_reformat_sub data set from the Analyze workspace and then open the BMA add on module from the Apps menu 31 As mentioned previously this data set contains multiple bones to allow the module to distinguish between the bone it should process and all other bones it is necessary to load the initial segmentation object map created using the Volume Edit module To do this check the Use Initial Segmentation option Figure 5 15 and then load the object map initial obj saved in step 29 Use Initial Segmentation Bone Object Bone w Figure 5 15 Initial Segmentation Option 32 Although an object map has been loaded it is necessary to specify a threshold range that identifies the bone in order for the cortex segmentation algorithm to function Use the Cortex Thresholds slider to specify a min max threshold range to segment the bone or leave the values calculated automatically by the module Figure 5 16 For this data set a Minimum value of 2803 and a Maximum value of 9449 work well i P f de Wew Other biaip He Sm GMD Sc airig Far iare BetacT Offset OU ii drca td 8 374 FIRE tape Soe Figure 5 16 BMA Module with Initial Segmentation Object Map Loaded 33 Next select the Segment Cortex butt
34. d for the user to enter a specific slope SigmaCT and offset BetaCT value to convert CT numbers Hounsfield units to bone mineral density BMD in units of mg CC Figure 2 17 BMD Scaling Parameters SigmaCT Slope 10 BetaCT Offset 0 0 Figure 2 17 BMD Scaling Parameters Generic values are provided but the user may modify these values as required Depending upon the vendor and scanning protocol these values may be available in the header information of the data e g Scanco For other data such as that acquired from the PerkinElmer QuantumFX or the Rigaku RH mCT2 microCT scanner these values need to be calculated For information on how to determine the SigmaCT and BetaCT values for data acquired from these systems please refer to Section 6 BMD Calibration 2 6 BMA Workflow labs The BMA workflow tabs Figure 2 18 provide access to the three Segment Segment Measure processing steps in the BMA workflow Segment Cortex Segment Cortex Trabeculae Bone Trabeculae and Measure Bone Figure 2 18 BMA Workflow Tabs 2 6 1 Segment Cortex The first step of the BMA workflow process is to segment the cortex from the trabecular bone and intertrabecular space Figure 2 19 If the image volume is of sufficient resolution to reveal cortical pores they will be identified at this stage Tr File View Other Help M Cortex Thresholds M M an ax 10364 411501 v Threshold Rendering Vi Use Initial Segmentation
35. e 4 3 An oblique plane shown in yellow will be defined perpendicular to the red line defined Figure 4 3 Definition of Oblique Plane in Coronal View 9 Next select Tools gt Fly from the main Oblique Sections window Use the Fly tool to roll the oblique plane to the correct position in the corresponding orientation Note that in this example the axis shown in yellow on the sagittal image on right should be rotated slightly in a clockwise direction Figure 4 4 this is achieved using the Roll left button Orthogonais Oblique Sections Figure 4 4 The Orthogonals Tool Showing Fly Adjustment a Analyze 24 10 To output the reformatted data open the File gt Output option Figure 4 5 Destination File WorkSpace Transformed volume size should be O Maintained Change to best fit data Reformat Images Object Map Generate Slices Figure 4 5 Oblique Sections Output Window 11 In the Output window set a he output Name to BMA PE micro CT reformat b Method to Reformat Entire Volume and select Change to best fit data c Finally select Generate Slices The reformatted data set will be saved to the Analyze workspace 12 Close the Oblique Sections module 4 1 3 Subregioning Data The subregioning or cropping of the input image data set is an optional step however cropping the data to the bone of interest will reduce the total size of the volume and thus decrease proc
36. e Load Volume Choosing the Load option from the File menu within the BMA add on module opens the Load module This allows users to load data sets from disk that may not already be loaded into Analyze e Exit Selecting the Exit option closes the Bone Microarchitecture Analysis add on module 2 2 2 View The View menu provides access to the Intensities and Size tools e Intensities When the Intensities option is selected the Intensity window Figure 2 5 for the module opens The Intensity window allows users to change the intensity display of the image data Users also have the ability to choose the intensity processing type Max Min or Level Width Maximum 1367 15353 Range Volume _ DataType Figure 2 5 The Intensity Window v AnalyzeDirect 8 e Size When the Size option is selected the Size window Figure 2 6 for the module opens The window allows users to specify the display size of the 2D slices and 3D rendering Table 2 1 Size Option Action Quarter Sets slice and rendering display to 1 4 of i O Third Halve HOMME SE e Single C Double C Triple Third Sets slice and rendering display to 1 3 of i actual size A Quadruple _ Specified Factor Halve Sets slice and rendering display to 1 2 of x10 actual size Interpolate te Single Sets slice and rendering display to actual Y 0 size Double Sets slice and rendering display to twice D the actual size v Triple Sets slice and re
37. e also known as second moments of area or SMAs and characterize resistance to bending around a given axis Low _ has been determined to be a risk factor for stress fracture in the tibia Maximum moment of inertia l_ mm is a measure of how material is distributed about the major centroidal axis which is orthogonal to the minor centroidal axis Maximum moment of inertia correlates to the maximum bending strength of a bone Area moments of inertia are also known as second moments of area or SMAs and characterize resistance to bending around a given axis Analyze lirecl 21 Minimum moment of inertia l mm is a measure of how material is distributed about the minor centroidal axis which is orthogonal to the major centroidal axis Minimum moment of inertia correlates to the minimum bending strength of a bone Area moments of inertia are also known as second moments of area or SMAs and characterize resistance to bending around a given axis Polar moment of inertia J mm is the sum of any two perpendicular area moments of inertia I or lan and characterizes resistance to torsion Polar moment of inertia has been found to increase with parathyroid hormone treatment in postmenopausal women with osteoporosis A reduction in weight bearing in female mice led to a reduction in polar moment of inertia Cortical porosity Ct Po 95 is calculated as the pore volume Po V divided by the total cortical compartme
38. e the slice slider to navigate through the slice display Note that there are a number of cortical regions shown in red that need to be assigned to the trabecular bone shown in orange Lock the Intra Trabecular Space yellow and Cortical Pores green objects then select the Trabecular Bone object and then click on the Manual tab and select the E Trace tool Draw around the regions of cortex to reassign them to the trabecular bone Figure 5 23 Figure 5 23 Steps to Reassign Incorrectly Assigned Regions to the Trabecular Bone Note that locking the Cortical Pores and Intra Trabecular Space objects Figure 5 24 will allow you to only reassign cortex M original BB cortex em Cortical Pores a mm Intra Trabecular Space E E NN Figure 5 24 Locking Objects Continue working through the slice data to correct regions in this manner When completed select File gt Save Object Map and save the object map as Final edited obj Close the Volume Edit module In the BMA add on module use the Load Final Object Map to load the Final edited obj map saved in step 67 a AnalyzeDirect 50 0 2 9 Measure Bone The third and final step of the BMA processing workflow is the generation of the bone morphometric indices 70 Specify the directory where all of the measurement Measurement Data dat
39. ess to a Size menu for t 2m the Rendering quadrant The default Size option is From oize Menu this means the display size of the rendering is dictated to that set globally via the Size menu or the increase decrease size buttons on the toolbar The other options will apply the selected size action to the rendering only Transparency Background Color Print Figure 2 15 The Rendering Quadrant e Interactive Rotate Allows users to select between Right Click Menu Visible Surface Only useful for interactive rotation of extremely large image volumes and Full Render e Transparency The transparency option turns transparent rendering on or off The transparency rendering mode is useful for visualizing objects contained within objects or objects with views obscured by other objects e Background Color Allows users to set the color of the rendering background The option opens a Color selection window enabling selection of desired color e Background Opacity The Background Opacity option is only available when Transparency rendering is enabled The option allows users to specify the background opacity in the transparency rendering Figure 2 16 Figure 2 16 Background Color and Opacity Options e Print Opens the Print command window facilitating the selection of a printer for the printing of the current rendering quadrant a Analyze 13 2 0 BMD Scaling Parameters A set of input fields has been provide
40. essing time 13 Select the reformatted data BMA PE micro CT reformat and open Process Image Calculator I To Place 14 Right click on the Calculator interface and select Buttons In the buttons window select the Region Pad button and drag it from the Palette to the Key Pad Figure 4 6 Next select Apply then Save and then Done Apply Reset Done Save Restore Select All Figure 4 6 The Image Calculator Button Configuration Tool a AnalyzeDirect 25 15 From the main Image Calculator window click the Region Pad Region button the Subregion window will open Figure 4 7 Pad 16 In the Subregion window click the Interactive button 17 In the Interactive window Figure 4 8 set the yellow box around the bone of interest Use the orientation buttons to navigate between views SubRegion Low High X 214 437 183 440 Figure 4 7 Subregion Tool Figure 4 8 Interactive Subregion Tool 18 Once the crop is set in all three orientations click Apply When prompted select Save a Copy of the Loaded Data The cropped data will be saved to the Analyze 12 0 workspace 19 Close the Image Calculator module 20 Select the subregioned data set from the Analyze 12 0 workspace and then right click and select Rename from the menu Change the name of the data set from BME_PE_micro_reformat0 to BMA_PE_micro_reformat_sub Analyze ire 26 4 1 4 Filtering Data The filtering of the image data
41. f each csv file contains the variable names of the measurements in the following row s An additional csv file named VolumeName csv is also saved containing all of the measures calculated in a format including explicit units for all measurements Save Final Object Map This option opens a save window allowing the user to save the final object map Load Final Object Map This open opens a load window allowing the user to load a corrected final object map Measurement Data Directory This is initially set to the OriginalDir value in the image header Directory This option allows the user to set the measurement data directory the location where the csv files will be saved to Analyze ire 17 Measure Options The Measure Options allow the user to specify groups of bone measurements to sample Unchecking the checkbox to the left of the group name before a measurement excludes the measurement For detailed information on each of the measurements generated please refer to section 3 Bone Microarchitecture Analysis Measurements e Bone Mineral Density Means and standard deviations measured in the grayscale image volume under the various objects and converted by use of the SigmaCT and BetaCT to mg cc units These are saved in a single file named VolumeName BMD csv e Volumes Surface Areas and Ratios Global values saved in a file named VolumeName VSR csv e Cortical Porosity Global values saved in a file named VolumeName_CP csv
42. format sub all all IntraTrabecularStd BMD BMA SC micro reformat sub all all TrabeculaeMean BMD BMA SC micro reformat sub all all TrabeculaeStd BMD BMA SC micro reformat sub all all TrabecularTissueMean BMD BMA SC micro reformat sub all all TrabecularTissueStd BMD EMA SC micro reformat sub all TV BMA 5C micro reformat sub all BV BMA SC micro reformat sub all BS BMA SC micro reformat sub all BMA SC micro reformat sub all BS TV BMA SC micro reformat sub all EMA SC micro reformat sub all BMA SC micro reformat sub all Figure 5 27 Measures Window 8 AnalyzeDirect 51 6 BMD Calibration For certain vendor data it is necessary to calculate BMD scaling parameters for proper conversion of CT or grayscale numbers to BMD units mg CC This requires that a QRM BMD phantom be scanned with the image data using the same acquisition protocol This calibration exercise has been developed for microCT data acquired on the PerkinElmer QuantumFX and the RH mCT2 microCT scanner however the exercise methodology can be applied to microCT image data acquired using other microCT systems 6 1 Data Analysis 1 Scan your images and QRM BMD phantom with appropriate acquisition protocol 2 Load the image data for the QRM phantom into Analyze Ensure it was acquired using the same protocol used for the bone images 3 Select the image and open Process Spatial Filters Apply
43. hod A decrease in strength of trabecular bone with aging in men is usually associated with a decrease in Tb Th In women Tb Th does not generally decrease after menopause Trabecular separation Tb Sp mm is the mean distance between trabeculae as determined by a 3D sphere fitting method Tb Sp is sometimes called trabecular spacing Higher Tb Sp is associated with vertebral fracture Tb Sp generally increases with age exhibiting a linear dependence on age in the lumbar spine Standard deviation of trabecular thickness Tb Th SD mm is a measure of the heterogeneity of trabecular thickness Tb Th above Standard deviation of trabecular separation Tb Sp SD mm is a measure of the heterogeneity of trabecular separation Tb Sp above Degree of anisotropy DA is a dimensionless variable defined as the length of the longest mean intercept length vector divided by the length of the shortest mean intercept length vector For an isotropic perfectly oriented system DA is equal to 1 As the system becomes more anisotropic less well oriented the DA increases to some value greater than 1 DA is related to the directional dependence of the mechanical properties of trabecular bone and does not appear to vary with age Mean intercept length MIL is a measure that is part of the algorithm used to determine the degree of anisotropy DA above 3 2 Cortical Bone Morphology Measurements Total cross sectional area inside the pe
44. in Orange 57 At this point in the workflow the user will need to visually review the segmentation result to determine if the trabecular bone segmentation is acceptable or not If the result is acceptable then processing can continue and the user can proceed to the Measure Bone step step If the segmentation result requires manual refinement then the user has the ability to output and refine the object map using the Volume Edit module available within Analyze 12 0 The steps neccasary for manual refinement are given in section 5 2 4 58 Use the linked cursor tool to inspect the 3 orthogonal slice displays and the 3D rendering of the bone to review the trabecular bone region segmentation result 59 To save the object map select the Save Final Object Map button Name the object map Final obj 60 Minimize the BMA add on module do not close the module O AnalyzeDi 49 5 2 4 Manual Refinement of the Final Object Map using Volume Edit Here we will review manual segmentation tools in the Volume Edit module that will allow for the correction of small regions of trabecular bone identified as cortex Object map refinement will improve the accuracy of the measurements generated by the module 61 62 63 64 65 66 67 68 69 With the BMA_SC_micro_reformat_sub data set selected open Segment gt Volume Edit Select File gt Load Object Map and load the Final object map saved in step 59 In the Coronal orientation us
45. ling of holes and gaps in the trabecular tissue region After object refinement is complete the modified object map can be reloaded into the BMA add on module at this point in the workflow using the Load Segmented Cortex Object Map option Use the linked cursor tool to inspect the 3 orthogonal slice displays and the 3D rendering review both the cortex and trabecular region segmentation results To save the object map select the Save Segmented Cortex Object Map button Name the object map Cortex obj Minimize the BMA add on module do not close the module Analyze i 31 4 2 2 Manual Refinement of Cortex Object Map using Volume Edit The Volume Edit module provides all of the necessary semi automatic and manual segmentation tools required to modify the cortex object map Here we will review some common tools that can be applied to the manual adjustment of the object map to correct small regions of trabecular tissue identified as cortical bone and to fill holes in the trabecular region Editing the object map will allow for greater accuracy in the measurements generated by the BMA add on module 47 With the BMA PE micro reformat sub data set selected open Segment Volume Edit 48 Select File Load Object Map and load the Cortex object map saved in step 45 4 2 2 1 Reassigning Cortex to Trabecular Tissue 49 n the coronal orientation use the slice slider to navigate through the slice display Note that there are a number
46. measure in the evaluation of osteoporosis and other bone related conditions The X ray attenuation coefficients produced in uCT are converted to an equivalent BMD value This conversion is accomplished by using phantoms liquid K HPO or solid resin embedded hydroxyapatite of known densities to generate a calibration line Low bone mineral density is a risk factor for fragility fracture although many other factors also contribute to fracture risk and BMD alone should not be used to diagnose osteoporosis The World Health Organization has developed an algorithm called FRAX that takes multiple factors into account to evaluate fracture risk BMD has been shown to increase with parathyroid hormone treatment 3 1 Trabecular Bone Microarchitecture Measurements Total volume TV mm is the volume of the region of interest In the BMA module this volume of interest includes the cortex cortical pores trabecular tissue and trabecular bone objects TV is used to calculate the bone volume fraction and bone surface density indices below that allow comparison between bones of different patients or animals Bone volume BV mm is the volume of the region segmented as mineralized bone including cortical and trabecular bone BV is used in the calculation of bone volume fraction as defined below Bone surface BS mm is the surface area of the region segmented as mineralized bone including trabecular and cortical bone BS is used to calculate the bone
47. n the geometric properties and biomechanical significance of intracortical porosity in the distal radius and tibia J Bone Miner Res 25 983 93 2010 Analyze rec 55 22 23 24 29 20 2 2D 29 30 31 32 33 34 35 36 37 38 39 40 41 42 Ellman R et al Partial reductions in mechanical loading yield proportional changes in bone density bone architecture and muscle mass J Bone Miner Res 28 875 85 2013 Ammann P et al Strontium ranelate improves bone resistance by increasing bone mass and improving architecture in intact female rats J Bone Miner Res 19 2012 20 2004 Stein M S Feik S a Thomas C D Clement J G amp Wark J D An automated analysis of intracortical porosity in human femoral bone across age J Bone Miner Res 14 624 32 1999 Sugiyama T et al Bones adaptive response to mechanical loading is essentially linear between the low strains associated with disuse and the high strains associated with the lamellar woven bone transition J Bone Miner Res 27 1784 93 2012 Li X et al Inhibition of sclerostin by monoclonal antibody increases bone formation bone mass and bone strength in aged male rats J Bone Miner Res 25 2647 56 2010 Eddleston A et al A short treatment with an antibody to sclerostin can inhibit bone loss in an ongoing model of colitis J Bone Miner Res 24 1662 71 2009 Ritzel H
48. ndering display to three times the actual size Figure 2 6 The Size Window Quadruple Sets slice and rendering display to four times the actual size Table 2 1 Explanation of Size Options e Specified Factor The Specified Factor option allows users to set the display size of the 2D slices and 3D Rendering Selecting the Specified Factor radio button enables the X and Y fields below Enter the desired value into these fields by double clicking on the current value and entering in the new value X specifies the display width of the 2D slices and 3D Rendering Y specifies the display length of the 2D slices and height of the 3D volume e interpolate When selected specifies that the pixels displayed in the 2D slices will be interpolated rather than replicated Figure 2 7 when scaling the slices for display Figure 2 7 Interpolated vs Replicated Image Display 2 2 3 Other e Options When selected opens the Options window Figure 2 8 This window allows users to specify window attributes for the module Auto Scroll Automatic sub window close gi Threads e Threads The Threads option allows users to specify the number of processing cores the multithreaded code can utilize By default the threads option is set to the maximum number of processors on the system e PowerBar Editor Opens the PowerBar Editor window This window allows users to add or remove buttons from the Toolbar in the BMA Figure 2 8 The O
49. nt volume Ct V Ct Po is negatively correlated with bone strength and is thought to increase during puberty and is higher in boys than girls during puberty because of increased calcium demands during growth spurts In adults Ct Po increases with age 9 exponentially so in women after menopause The increase of porosity with age is understood to be caused by an increase in pore size and not by a change in pore number Ct Po can be increased by diseases and treatments such as kidney disease hyperparathyroidism or parathyroid hormone treatment thyroid hormone treatment or prostaglandins Ct Po was higher in diabetic women exhibiting fragility fractures than in diabetic women not exhibiting fractures in the femoral neck of patients exhibiting a hip fracture in men with idiopathic osteoporosis exhibiting vertebral fractures and in osteopenic and osteoporotic postmenopausal women Ct Po has been found to increase with chronic alcohol consumption in rats and decreased as a result of sclerostin inhibitor treatment in female rats Pore number Po N n is a count of the number of cortical pores in a given volume Po N does not appear to increase with age rather the size of each pore increases leading to an increase in overall porosity In rats chronically dosed with alcohol pore number was increased rather than pore size indicating that chronic alcohol consumption increases porosity through a different mechanism than
50. odule with Initial Segmentation Object Map Loaded a AnalyzeDirect 30 40 41 42 At this point in the workflow the user will need to visually review the segmentation result to determine if the result is acceptable or not If the result is acceptable then processing can continue and the user 43 44 45 46 Next select the Segment Cortex button Note that the cortex segmentation algorithm is a computationally intense process and could take several minutes to complete Once processing is complete the module will automatically move to the Segment Trabeculae tab Figure 4 20 The three orthogonal slices displayed will update to show the Cortex Cortical Pores and Trabecular Tissue objects Fie Vos Ohe Help mT F E iemet tegmen Leurs 2 Cene Tinb ivler Boer G r Segmented Caries load Sesrnenped Cortes Oe Klap Diyet Map Tease uli Th eibsckd on ban ie D 1 4047 v Tiras Rerdenag p imr Trabsculas e I Hi Figure 4 20 BMA Segment Trabeculae Tab can proceed to the second step in the BMA processing workflow Segment Trabeculae step 62 If the segmentation result requires manual refinement then the user has the ability to output and refine the object map using Volume Edit available within Analyze 12 0 The steps neccasary for manual refinement are given in section 4 2 2 Examples of manual refinement that may be needed include the reassignment of incorrectly labeled bone structures and the fil
51. of cortical regions shown in red that should be assigned to the trabecular object Figured ee eee shown in yellow Figure 4 21 Tabecular Tissue 50 Select the Trabecular Tissue object and then click on the Manual tab and choose the Draw tool Figure 4 22 Steps to Reassign Incorrectly Labeled Regions Using the Draw Tool 51 Next click on the image and draw over the Cortex regions to be reassigned Figure 4 22 52 Continue moving through the slice data and correcting regions using this method a AnalyzeDirect 32 4 2 2 2 Filling Holes and Gaps in the Trabecular Tissue 53 o4 55 56 D 98 59 60 61 The Fill Holes option in the Volume Edit module can be used to correct for smaller holes in the trabecular tissue Select the Semi Automatic tab and then select the Fill Holes tool then click on the Trabecular Tissue object to initiate the fill holes routine Note that small holes and gaps will be resolved To quickly correct larger holes in the Trabecular Tissue object Figure 4 23 it may be necessary to utilize the Fill Holes morphological operation from the Objects menu Select View gt Objects to open the Objects window In the Objects window select the Morph object option from the button options to the right
52. on Note that the cortex segmentation algorithm is a computationally intense process and could take several minutes to complete v AnalyzeDi 44 34 39 36 37 38 39 Once processing is complete the module will automatically move to the Segment Trabeculae tab Figure 5 17 The three orthogonal slices displayed will update to show the Cortex Cortical Pores and Trabecular Tissue objects P File View Om Help H9 Segment Segment Mesure Cortex Trabecuse Bone wee Segenented Cones load Segrented Comme Object Mag Object Map Trabeculee Thresholds Mn Mes vug ghs v Threihoid Rendennq Segment Teabecul e s Ld aD Figure 5 17 BMA Segment Trabeculae Tab At this point in the workflow the user will need to visually review the segmentation result to determine if the result is acceptable or not If the result is acceptable then processing can continue and the user can proceed to the second step in the BMA processing workflow Segment Trabeculae step 55 If the segmentation result requires manual refinement then the user has the ability to output and refine the object map using the Volume Edit module available within Analyze 12 0 The steps neccasary for manual refinement are given in section 5 2 2 Examples of manual refinement that may be needed include the reassignment of incorrectly labeled bone structures and the filling of holes and gaps in the trabecular tissue region After object refinement is comple
53. ow The BMA workflow is divided into two sections Data Preprocessing and Bone Microarchitecture Analysis Data preprocessing is a prerequisite for analysis of data via the BMA add on An overview of the entire workflow is provided in Figure 1 1 Pre processing 1 1 1 Data Preprocessing Overview The BMA add on module requires that input bone specimens have isotropic voxels If the image data contains multiple bones the bone of interest must be isolated and orientated into the correct anatomical orientation Data Preprocessing also includes optional data subregioning cropping and filtering steps to reduce data size and enhance initial object segmentation Optional processing cm Bii Object refinement Ea Object refinement PEE Trabeculae Measurements Figure 1 1 Diagram of BMA Workflow 1 1 2 Bone Microarchitecture Analysis Overview The second part of the Bone Microarchitecture Analysis workflow is conducted within the BMA add on module and is separated into three steps 1 Cortical Bone Segmentation Once the image data is loaded the BMA add on will conduct a preliminary segmentation which will allow the module to identify the cortical shell cortical pores and trabecular tissue regions 2 Trabecular Bone Segmentation Upon user approval the add on will complete the segmentation process by identifying the trabecular bone in the trabecular region Two new regions the trabecular bone and int
54. pagate Regions button Figure 6 4 The objects will be propagated to all slices between the first and last slice the objects are defined on Next open Generate Sample Options and set the following Figure 6 5 e Sample Type Object s e Summing On e Auto Reset On e Sample All Slices e Sequence Display Off e Stat Type Intensity e Decimal Places 4 e Log Stats On Click the Sample Images button and then save the generated stats log file to disk Flt Geanate Teol View Other Hiig mad fud ardd nd EES 5 ut EI E Control by Object Attribute Add Attribute Name Name Field Width 12 Cnet T Reassign 1 Original Object s 2 0 Remove 3 50 Unused 4 200 I teas 5 800 Lect 6 1200 Objects i Done Figure 6 3 Renaming Regions CY 2D Mode X 30 Mode C1 4D Mode Coperigns 82094 BIR Mae Corio IC Original 50 800 Select All Invert Selections Orientation Transverse Coronal Sagittal Figure 6 4 The ROI Propagate Regions Tool Sample Type L Original Selected Region D amp Object s 50 200 800 1200 Select All Invert Selections Combine Objects No Yes C2 Individual Voxels Minimum sample Max Min idea Oh 2592 Range Volume DataType Volume Primary Related Summing On Off Auto Reset 8 On
55. peration to Fill Holes Fill Type to 3 pass 2D Transverse Connectivity to 4 connected 00 0 O Q9 Defined Object to Trabecular Tissue Next click Morph Figure 5 20 Larger holes and gaps in the Trabecular Tissue object are now filled and assigned to the Trabecular Tissue object Note that this is also a quick way to reassign incorrectly identified regions of Cortex back to the Trabecular Tissue and could provide a utility for Cortex to Trabecular Tissue reassignment without having to conduct any manual reassignment as described in the Reassigning Cortex to Trabecular Tissue steps above Once editing is complete use the File Save Object Map option to save the object map as Cortex edited obj and then close the Volume Edit module Objects VolumeEdit 5 r1 Control by 8 Object Attribute Object Trabecular Tissue Operation FillHoles C 3D O 3D Transverse Coronal Sagittal 8 3 pass 2D pU 0 5 Connectivity 8 4 connected 3 B connected Defined Object Trabecular Tissue Add Object Delete Object Reassign Object s Remove rad Object Smoothed Load Object s Unused Load Binary Save Binary Morph Object I Filter Objects Done Figure 5 20 The Morph Object Window In the BMA add on module use the Load Segmen
56. ptions Window module a AnalyzeDirect 9 2 2 4 Help e About The About option opens the Analyze 12 0 Bone Microarchitecture Analysis About window The About window provides detailed information about the system and the Analyze 12 0 software Selecting Done will close the About window e User s Guide The User s Guide option opens the software help documentation 2 3 Toolbar The BMA Toolbar provides quick access to several functions Toggle Region Display Double Display Size Exit Module Be tom NS Intensities Halve Display Size e Toggle Region Display The Toggle Region Display option allows users to quickly switch between the display of the overlaying regions on the slice displays Figure 2 10 The display options are Filled Regions Region Edges and Regions Off Note the keyboard shortcut to toggle region displays is the t key Figure 2 10 Slice Display Showing the 3 Region Display Options Filled Regions Region Edges and Regions Off e Intensities Shortcut to open the View gt Intensities window Figure 2 5 e Double Display Size Doubles the current slice and rendering display size e Halve Display Size Halves the current slice and rendering display size e Exit Closes the BMA add on module a AnalyzeDirect 10 2 4 Display Screen The display screen Figure 2 11 is divided into quadrants displaying the 2D slices in the transverse coronal and sagittal orientations and the 3D rendering of the dat
57. ra trabecular space will be derived 3 Calculation of Bone Morphometric Indices Measurements are automatically calculated for the trabecular and cortical bone and output in several comma separated values csv files Files are saved to a user defined directory Pre processing 1 1 3 Processing Single Bone Specimens Image volumes that contain only a single bone do not require a pre segmentation step to isolate the bone of interest The workflow is summarized in Figure 1 2 Optional processing A Segment Cortex LE Object refinement Object refinement P Trabeculae Measurements Figure 1 2 Diagram of BMA Workflow 2 The Bone Microarchitecture Analysis Add on Interface Overview The BMA add on is started from the Analyze 12 0 Apps Menu Apps gt BMA Figure SISCOM 2 1 To load a data set select the data from the Analyze workspace and open the Mayo 3D Brain Atlas BMA add on Figure 2 2 Alternatively data sets can be dragged from the Analyze E3 workspace and dropped onto the add on module or loaded from disk directly into the add on via the File gt Load option a copy of the data set will also be loaded to the current Analyze workspace DT T2 Projection Volume Metrics Figure 2 1 Starting the BMA Add On Register Measure Apps Other Help SISCOM Mayo 3D Brain Atlas org DTI 4 T2 Project D Volume Metrics a T d b sf af Cop
58. ractures BMJ 312 1254 9 1996 Siris E S et al Identification and Fracture Outcomes of Undiagnosed Low Bone Mineral Density in Postmenopausal Women JAMA 286 2815 2822 2001 Sandhu S K et al Prognosis of fracture evaluation of predictive accuracy of the FRAX algorithm and Garvan nomogram Osteoporos Int 21 863 71 2009 Kanis J A et al Interpretation and use of FRAX in clinical practice Osteoporos Int 22 2395 411 2011 Kanis J A Johnell O Oden A Johansson H amp McCloskey E FRAX and the assessment of fracture probability in men and women from the UK Osteoporos Int 19 385 97 2008 Kanis J A et al Development and use of FRAX in osteoporosis Osteoporos Int 21 Suppl 2 8407 13 2010 Neer H M et al Effect of parathyroid hormone 1 34 on fractures and bone mineral density in postmenopausal women with osteoporosis N Engl J Med 344 1434 41 2001 Qiu S Rao D S Palnitkar S amp Parfitt A M Independent and combined contributions of cancellous and cortical bone deficits to vertebral fracture risk in postmenopausal women J Bone Miner Res 21 1791 6 2006 Ostertag A et al Vertebral fractures are associated with increased cortical porosity in iliac crest bone biopsy of men with idiopathic osteoporosis Bone 44 413 7 2009 Chen H Zhou X Fujita H Onozuka M amp Kubo K Y Age related changes in trabecular and cortical bone microstructure Int J Endo
59. riosteal envelope Tt Ar mm is measured for each transverse section Tt Ar has been found to increase with age with continued periosteal apposition in both men and women at the distal radius and distal tibia however Tt Ar increased more in men than in women at the distal radius with age Boys after the onset of puberty and young men have a larger Tt Ar than girls and women of the same maturity Cross sectional area measurements characterize resistance to axial compression and tension Cortical bone area Ct Ar mm is calculated as the cortical volume Ct V divided by the product of the number of slices times the thickness of each slice The Ct Ar is a separate measurement for each transverse section Men have been found to have a greater Ct Ar than women and Ct Ar has been found to have a weak negative correlation with age The Ct Ar of the femoral midshaft was found to have a linear relationship with Analyze lirecl 20 weight bearing in mice Ct Ar increased in rats receiving strontium ranelate Boys during and after puberty have been found to have a higher Ct Ar than girls of the same maturity Further Ct Ar is greater in men than women and has a positive correlation with height and weight and a negative correlation with age Medullary or marrow area Ma Ar mm is the cross sectional area of the medullary part of the bone on any given transverse section Treatments that tend to increase total cross sectional area T
60. rocessing Once data preprocessing is complete the image data is ready for processing with the Bone Microarchitecture Analysis add on module 4 2 1 Segment Cortex The first step in the BMA add on module is the segmentation of the cortex 37 Select the BMA_PE_micro_reformat_sub data set from the Analyze 12 0 workspace and then open the BMA add on module from the Apps menu 38 As mentioned previously this data set contains multiple bones to allow the module to distinguish between the bone it should process and all other bones it is necessary to load the initial segmentation object map created using the Volume Edit module To do this check the Use Initial Segmentation option Figure 4 18 and then load the object map initial obj saved in step 36 39 Although an object map has been loaded it is necessary to specify a threshold range that identifies Bone Object Bone m Figure 4 18 Initial Segmentation Option the bone in order for the cortex segmentation algorithm to function Use the Cortex Thresholds slider to specify a min max threshold range to segment the bone or leave the values calculated automatically by the module Figure 4 19 F e Vos Cher Help Bee om Segment hemmt Marone Vene Tisbecalec Boca i Lose Teegiheleh Ld hlar mma paes Eaa Rendersg x Lee lahi Sogeti reiil Git Mig it Dory Dot Bona w pe kiakia Cortes e 2 Pla m iris f uk g ifi E LI Figure 4 19 BMA M
61. sub EMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub EMA PE micro CT reformat sub all all all all all all all all all all all all all all all all all all all all all all all all all all all all all all all all CortexMean BMD CortexStd BMD Intra TrabecularMean BMD IntraTrabecularStd BMO TrabeculaeMean BMO TrabeculaeStd BMD TrabecularTissueMean BMD TrabecularTissueStd BMD TV BV B5 BS TV 11 30 36 2688 77 4915 05 481 81 3133 20 1140 99 14 87 5 74 152 53 45 00 10 19 22 64 1 98 4901 Figure 4 31 Measures Window 79 Right clicking on a row or column will allow users to plot the row or column when supported 80 When the process is complete several csv files will be saved to the location specified in step 76 The csv files can be opened in MS Excel or any other stats analysis package a AnalyzeDirect 37 5 Bone Microarchitecture Analysis Scanco microCT Data Processing 5 1 Data Preprocessing 5 1 1 Data Load The BMA add on module requires that input bone specimens have isotropic voxels To ensure your image data is loaded with isotropic voxels load the data using the Load As module and resample anisotropic data using the Resize option
62. subregioned data BMA_SC_micro_reformat_sub and open Segment gt Volume Edit 25 Inthe Object control panel click the Add Object button and name the object Bone Figure 5 11 a E a Original ag Bone Name Bone Color Red Add Object Display Locked Ld Figure 5 11 Volume Edit Object Control Panel 26 Next select the Semi Automatic tab and choose the Region Grow option 27 Click on the bone of interest in any of the 3 orthogonal displays to set a seed Figure 5 12 then use the threshold slider to define a global threshold range that represents bone Now click Extract Object Figure 5 13 All voxels that fall within the specified minimum and maximum threshold range and that are spatially connected to the seed will be assigned to the Bone object For this data set a Min value of 3030 and a Max value of 9385 work well Figure 5 12 Region Grow Seed Point Figure 5 13 Volume Edit Semi Automatic Region Grow Tool a AnalyzeDi 42 28 If necessary use the Object Separator tool to separate any bones connected to the bone of interest to a new object and then delete For this data set this step is not required Figure 5 14 File View Other H Hep Bo Soha E Gergana m Marne Dons Colur Red Add Object x Deiplang Locked Ldge Strength Watts Semi Automatic Menu Ute Dope Strength 150 Figure 5 14 Segmented Bone of Interest 29 Select File gt Save Object Map Name the obje
63. t Ar and cortical area Ct Ar lead to a decrease in Ma Ar as an increase in bone area also increases the ratio of cortical to medullary bone Two examples of such treatments include an increase in mechanical loading and a sclerostin inhibitor 9 Sclerostin is a bone regulatory protein produced by osteocytes that inhibits bone formation Cortical area fraction Ct Ar Tt Ar is the ratio of cortical cross sectional area to total cross sectional area within a given transverse section Ct Ar Tt Ar has been found to significantly increase with a 25 mg kg dose of sclerostin inhibitor in rats Average cortical thickness Ct Th mm is the average thickness of the cortex in each transverse section There have been conflicting results about the correlation of Ct Th with the incidence of vertebral fracture Ct Th has been found by Ostertag et al to be the same in patients with and without vertebral fracture while in a study by Qiu et al Ct Th was found to be an effective measure to discriminate between patients with and without vertebral fracture Vertebral Ct Th has been demonstrated to vary along the length of the spine with higher values in the cervical and lumbar spine and lower values in the thoracic spine Vertebral Ct Th decreases with age in the lower thoracic and lumbar spine and femoral neck but does not vary with gender and is lower in patients with osteoporosis than in a control group Mice receiving a sclerostin inhibitor
64. ta sets previously sampled and not calibrated to BMD it is possible to resample the data if BMD calibration parameters were derived after initial sampling by going to the Measure Bone tab and loading the final object map and setting the SigmaCT and BetaCT values before taking the measurements aa ai File View Other Mele Mee 9 Om Segrnent Segment Measure e Trabeculae Bone te Mas 1046 4 Jats V Tiweshols Kandenny ve inti egenentamon segment Cortex Figure 6 7 BMD Calibration Parameters BMD Scaling Parameters SigmaCT Slope 1 0684 BetaCT Offset 1023 2 AnalyzeDireci Heferences 1 11 12 13 14 15 16 17 18 19 20 21 Bouxsein M L et al Guidelines for assessment of bone microstructure in rodents using micro computed tomography J Bone Miner Res 25 1468 86 2010 Nazarian A Snyder B D Zurakowski D amp M ller R Quantitative micro computed tomography a non invasive method to assess equivalent bone mineral density Bone 43 302 11 2008 Deuerling J M Rudy D J Niebur G L amp Roeder R K Improved accuracy of cortical bone mineralization measured by polychromatic microcomputed tomography using a novel high mineral density composite calibration phantom Med Phys 37 5138 2010 Marshall D Johnell O amp Wedel H Meta analysis of how well measures of bone mineral density predict occurrence of osteoporotic f
65. te the modified object map can be reloaded into the BMA add on module at this point in the workflow using the Load Segmented Cortex Object Map option Use the linked cursor tool to inspect the 3 orthogonal slice displays and the 3D rendering review both the cortex and trabecular region segmentation results Using the parameters specified in this exercise it will be necessary to refine the object map to correct for several regions in the object map at this point To save the object map select the Save Segmented Cortex Object Map button Name the object map Cortex obj Minimize the BMA add on module do not close the module a AnalyzeDi 45 5 2 2 Manual Refinement of Cortex Object Map using Volume Edit The Volume Edit module provides all of the necessary semi automatic and manual segmentation tools required to modify the cortex object map Here we will review some common tools that can be applied to the manual adjustment of the object map to correct small regions of trabecular tissue identified as cortical bone and to fill holes in the trabecular region Editing the object map will allow for greater accuracy in the measurements generated by the BMA add on module 40 With the BMA_SC_micro_reformat_sub data set selected open Segment gt Volume Edit 41 Select File gt Load Object Map and load the Cortex object map saved in step 38 5 2 2 1 Reassigning Cortex to Trabecular Tissue 42 In the coronal orientation use the slice slider
66. ted Cortex Object Map button to load the Cortex edited obj map saved in step 53 a Analyze Fr U m m JY ga D Ss e J 47 5 2 3 Segment Trabeculae The second step of the BMA process provides for segmentation of the Trabecular region into Trabecular Bone and Intra Trabecular Space This segmentation is achieved via threshold based segmentation 55 To segment the Trabecular region into the Trabecular Bone and Intra Trabecular Space objects set a threshold range using the Trabeculae Thresholds slider Figure 5 21 m n View Osher Help Bo Om Segment Segment Measure Cortex Trabeculse Bo Save Segmented Cortex Load Segmented Cortes Obyect Map Otyrct Mag Trabeculse Thresholds Mo x 2067 9 as 35 v Theeshold Rendeneg 207 P sg rab T BD necu Lo BMD Scabreg Poremeten SaqrnaCT Shope 1 0 BetaCT Offset 0 0 117 J Display Mode Edges Figure 5 21 Segment Trabeculae Tab with Thresholds Set to Segment Trabecular Bone a AnalyzeDi 48 56 Once the threshold has been specified click the Segment Trabeculae button The voxels specified by the threshold range will be assigned to the Trabecular Bone object The module will automatically proceed to the Measure Bone tab Figure 5 22 E File View Other Help Pal se gm Segment Segment Messe Cortex Trabeculae Bone an Figure 5 22 The Measure Bone Tab Showing the Segmented Trabecular Bone
67. the Fly value back to 10 Use the Fly tool to roll the oblique plane to the correct position in the corresponding orientation Note that in this example the axis shown in yellow on the coronal middle image should be rotated clockwise this is achieved using the Roll left button Figure 5 6 Rall 13 To output the reformatted data open the File gt Output option Figure 5 7 Destination File WorkSpace n SC micro CT reformat Transformed volume size should be C Maintained Change to best fit data Display Reformat Images Object Map Figure 5 7 Oblique Sections Output Window 14 In the Output window set Figure 5 6 The Fly Tool Rolling Left 10 Degrees a he output Name to BMA SC micro CT reformat b Method to Reformat Entire Volume and select Change to best fit data c Finally select Generate Slices The reformatted data set will be saved to the Analyze workspace 15 Close the Oblique Sections module 5 1 9 Subregioning Data The subregioning or cropping of the input image data set is an optional step however cropping the data to the bone of interest Will reduce the total size of the volume and thus decrease processing time 16 Select the reformatted data BMA SC micro CT reformat and open Process Image Calculator 17 Right click on the Calculator interface and select Buttons In the buttons window select the Region Pad button and drag it from the Palette to the Key Pad Figure 5
68. to navigate through the slice display Note that there are a number of cortical regions shown in red that should be assigned to the trabecular object shown in yellow Figure 5 18 43 Select the Trabecular Tissue object and then click on the Manual tab and choose the Draw tool Lad Figure 5 18 Incorrectly Labeled Trabecular Tissue Figure 5 19 Cortex Reassignment with the Volume Edit Draw Tool 44 Next click on the image and draw over the Cortex object to be reassigned Figure 5 19 45 Continue moving through the slice data and correcting regions using this method a AnalyzeDirect 46 5 2 2 2 Filling Holes and Gaps in the Trabecular Tissue A6 47 48 49 90 51 52 53 54 The Fill Holes option in the Volume Edit module can be used to correct for smaller holes in the trabecular tissue Select the Semi Automatic tab and then select the Fill Holes tool then click on the Trabecular Tissue object to initiate the fill holes routine Note that small holes and gaps will be resolved To quickly correct larger holes in the Trabecular Tissue object it may be necessary to utilize the Fill Holes morphological operation from the Objects menu Select View gt Objects to open the Objects window In the Objects window select the Morph Object option from the button options to the right In the Morph Object window returned set the following parameters Object to Trabecular Tissue O
69. uuuuuuunuauumun 77 By default all of the Measure Options will be checked Uncheck any option you do not wish to generate statistics for Note that data from the QuantumFX microCT scanner does not have sufficient resolution to derive cortical porosity measurements therefore the Cortical Porosity option should be unchecked Figure 4 30 Figure 4 29 Directory Selector for Specification of Output Directory Measure Options Bone Mineral Density Volumes Surface reas and Ratios Measure 8 Click the Measure Bone button 1 LIE to start the measurement process Note that it could take several minutes to calculate all of the bone morphometric indices Once the measurements have been generated they will be displayed in a new tabbed window Figure 4 31 Each of the tabs provides access to a different set of measurements Cortical Moments 30 Trabecular Thickness 3D Trabecular Structure and Connectivity 2D Characterization Figure 4 30 Measurement Options E All Measures BMD cp Ch2D MITSP TTH TrSt VSR as gt neam 5 measurement value EMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub EMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat sub BMA PE micro CT reformat
70. yright 1888 2014 BIR Mayo Clinic Portions of this software are covered under U S Patent 5 588 384 Figure 2 2 Loading Data into the BMA Add On Module 8 AnalyzeDirect 6 2 1 The Main BMA Program Window The main BMA program window Figure 2 3 provides access to all functions and tools within the module Display Screen Rendering Window fe Menu File View Other Help Heo Toolbar a Segment Segment Measure Cortex Trabeculae Bone Cog Thresholds Min Max 10364 4 1501 v Threshold Rendering Segment Workflow zs Tabs BMD Scaling Parameters BMD Scaling Parameters gmaCT Slope 1 0684 BetaCT Offset 1023 2 Point Picked 82 118 239 Value 940 Object Originsl Slice Display Figure 2 3 The BMA Program Window D a rm a AnalyzeDirect i 2 2 Menu The BMA menu provides access to the File View Other and Help menus Zo rile The File menu provides access to the Input Output Ports Load and Exit functions e Input Output Ports Selecting the File gt Input Output Ports option displays an area at the bottom of the Bone Microarchitecture Analysis window which facilitates the loading of image data via drag and drop from the Analyze workspace Figure 2 4 Image data can also be dragged and dropped onto the display screen of the module without opening the Input Output Ports option S hl Figure 2 4 Loading Data Via Input Output Ports

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