The Biomechanical and Stereological Effects of Pathological Proximal Femoral Lesions and Bisphosphonate Treatment in a Murine Model BA Rogers,

L Buchan, MK Akens, CM Whyne

Introduction
metastatic pathological fractures Up to 27% of all cancers Variable histology Osteolytic & osteoblastic mix Affected by bisphosphonate Rx (not uniform) Unknown biomechanics Variable stereology
Bone mineral density Bone volume fraction

Aim
Quantify the comparative structural impact of..
Osteolytic lesions Mixed osteolysis/osteoblastic lesions Bisphosphonate treatment

..on metastatic femoral lesions

Assess the reliability of clinical fracture risk assessment.

Methods
Murine model Outcomes
CT-based stereological analysis

Mechanical testing
Clinical fracture risk assessment
Mirels size & type criteria

Animal Care Committee (University Health Network) Approved all procedure

Controls

Healthy N=6

N=32

MT-1 human breast cancer genes

Osteolytic lesions N=8

5-6 week old Nude athymic rats

MT-1 human breast cancer genes

+ systemic bisphosphonates Osteolytic lesions N=9 Mixed osteolytic/osteoblastic lesions, N=9

Canine Ace-1 prostate cancer genes

Stereology
CT imaging Region of interest
600 slices distal physis

proximal

CT slices per 0.01 mm Each scan reconstructed

continuous tone (.ct) format greyscale bitmap (.bmp)

Amira Dev 5.2.2 imaging software

Parameter

Symbol (units)

Bone volume, BV (mm3) Tissue Mineral Density, TMD, gHA/cm3

Average cross-sectional surface area Total volume Bone volume Tissue mineral density Bone volume fraction

A (mm2) TV (mm3) BV (mm3) TMD (gHA/cm3) BV/TV (unitless)

Average cross-sectional area, A (mm2) Total volume, TV (mm3)

Bone 0.25gHA/cm3

Mechanical Testing
Potted with PMMA
To LT proximal 3mm distal
Potting zone

Strain gauge
20mm of distal femur

Region of interest (ROI)

8mm of distal femur

8mm ROI

20mm gauge length

Axial Loading
6mm/min strain rate Load-displacement at 0.01s intervals
3mm potting depression

Clinical Fracture Risk Assessment

2 independent observers Coronal CT images of ROI (all femora) Repeated +2 weeks Assessed
Type Size

Inter- & intra-observer agreement (kappa coefficient)

Clinical Feature Site Pain Type of metastasis Size (cortical destruction)

Score 1 Upper Limb Mild Osteoblastic

Score 2 Lower limb Moderate Mixed

Score 3 Peritrochanteric Functional Osteolytic

< 1/3

1/3 - 2/3

> 2/3

CORR 1989; 249: 256 - 264

Cortical destruction

>2/3

1/3-2/3

<1/3

Results
Tissue Mineral Density
0.85 0.8 0.75
TMD gHA/cm3

0.7 0.65 0.6 0.55 0.5

Osteolytic BP-Osteolytic Mixed Control

Results
Bone Volume Fraction, BV/TV
0.75
0.7 0.65 0.6

BV/TV

0.55 0.5 0.45 0.4 Osteolytic BP-Osteolytic Mixed Control

Results
Mechanical Testing
Ultimate Compressive Stress, UC (Mpa)
200 180 160 140 120 uc (MPa) 100 80 60 40 20 0 0 2 1 EA
(kN)

Axial Rigidity, EA, kN

7 6 5 4 3 E
(GPa)

Youngs Modulus, E, GPa

4.5 4 3.5 3 2.5 2 1.5 1

0.5
0

Results
UC stress vs Tissue Mineral Density
250
200 150 uc
(MPa)

R2=0.72

100 50

0
0.35 0.55 0.75 TMD (gHA/cm3) 0.95

Cortical involvement
by lesion type
8
Number of Occurences 7 6 5 4 3 2 1 0 8 7 6 5 4 3 2 1 0

8
7 6 5 4 3 2 1 0 1 2 3

Osteolytic

BP- Osteolytic

Mixed

Quantitative correlation with bone volume fraction (BV/TV)

Mirels Size Agreement.

A1 B1 0.625 A2 0.940 B2 0.824

B1 A2

0.700

0.700 0.880

Mirels 'Type' Agreement.

A1 B1 A2

B1
0.244

A2
0.183 0.291

B2
0.077 0.677 0.282

substantial = Blue ; excellent = Yellow significant agreement Landis & Koch guidelines

Discussion
Histology - variable biomechanical affect Superior
UC stress Youngs Mod Axial rigidity osteolytic + BP treatment mixed osteolytic/osteoblastic group

Discussion
Bisphosphonate treatment increases
BV/TV by 19% (p<0.05) stiffness by 71% (p<0.05) Compressive strength (p=0.1) Lower Mirels score
cf untreated osteolytic lesions

Discussion
Mirels size correlates
with bone volume fraction Not with tissue mineral density

Ultimate compressive stress correlates

Tissue mineral density Not with bone volume

Mirels size does not correlate ultimate compressive stress

Discussion
CT improves
definition of cortical involvement enhances reliability of Mirels score

Previous published inter-observer correlation

Size..0.48 0.56 Type.0.35 0.61

Strengths & Weaknesses

Few studies with controlled histological pathology
Biomechanical In assessing clinical fracture risk

Limitations
Compressive not bending stress Histological variation within groups No 3D Ct analysis

Thank You