appendix to the report
Statement
I have worked in collaboration with several other members of the Bone Metabolism Laboratory at Novartis Pharma AG in the completion of this study. However, I carried out most of the experiments in this project on my own. All the written work and the planning the experimental designs were done together with my industrial supervisor, Dr Jürg A. Gasser.
Author: Jianying Yam yammmy@yahoo.com Date: 30th June 2002
Acknowledgments
I would like to thank the following people for their help and support to make this project a success as well as for creating a delightful atmosphere at Novartis to work in.
Peter Ingold: for his help in all the technical work involved, in designing the holder for the scanner, contribution of many good ideas and giving me his unprecedented support throughout the year.
Adriano Montefusco, Anna Studer and Andrea Rebmann, for teaching and helping me with the histology methods.
Reto Cortesi and Margot Bruderlin for helping out during the ovariectomy procedures.
Dr. Micheala Kneissel for providing me with literature materials.
Dr. Markus Glatt for teaching me how to use the mCT 20 scanner program and allowing me to make measurements on his mCT 20 scanner.
SCANCO Medical for providing the vivaCT 20 scanner and assisting me with the technical problems I had encountered.
And finally
I would like to thank Dr. Jürg Gasser for giving me the opportunity to do this project in his lab as well as giving me all the assistance, guidance, knowledge whenever I needed it.
This year abroad at Novartis would not have been possible without the Pharmacology Department, University of Bristol, England.
I would also like to thank my host family; webhosting pakery@datacomm.ch and Valentin Keller and my friends who have helped me settle in and make my stay in Basel an unforgettable one.
Publications based on this project:
1. J.A. Gasser, J.R. Green, J. Yam, Long-term protective effect of a single IV injection of zoledronic acid on cancellous bone structure and cortical bone in ovariectomized rats. WHAT IS NEW IN BISPHOSPHONATES? 6th Workshop on Bisphosphonates – From the Laboratory to the Patient, March 20th to 22nd, 2002, Davos, Switzerland. Oral communication.
2. J.A. Gasser, J. Yam, J.R. Green, Long-term protective effect of a single intravenous administration of zoledronic acid on cancellous bone structure and cortical bone in ovariectomized rats. IXth Congress of the International Society of Bone Morphometry, Sheraton Hotel, April 7th-10th 2002, Edinburgh, UK. Oral communication. J. Bone Miner. Res., 17(5): 946, (OC18).
3. J.A. Gasser, J. Yam, Non-invasive monitoring of changes in structural cancellous bone parameters over 20 weeks in rats with a novel prototype micro-CT. 3rd International Workshop on Musculoskeletal and Neuronal Interactions, Marbella Hotel, May 30th –June 3rd 2002, Corfu, Greece. Oral communication. J. Musculosk. Neuron. Interact., 2(4): 369, (OR11).
4. J.A. Gasser and J.Yam, In vivo Non-Invasive Monitoring of Changes in Structural Cancellous Bone Parameters with a Novel Prototype MicroCT in Rats. 24th Annual Meeting of the American Society for Bone and Mineral Research, September 20-24th, 2002, San Antonio, USA.. Poster Presentation.
Abbreviations
2D Two Dimensional
3D Three Dimensional
B.Ar Bone Area (mm2)
BMD Bone Mineral Density
BMU Bone Multi-cellular Unit
BV Bone Volume
BV/TV Bone Volume Fraction (%)
Conn.D Connective Density
DXA Dual energy X-ray Absorptiometry
g Grams
hPTH Human Parathyroid Hormone
IL Interleukin
LDL Low Density Lipoprotein
MDx Micro damage
OVX Ovariectomy
pQCT Peripheral Quantitative Computed Tomography
QCT
ROI Quantitative Computed Tomography
Region Of Interest
SERMs Selective Estrogen Receptor Modulators
SMI Structure Model Index
T.Ar Trabecular Area
Tb.N Trabecular Number
Tb.Sp Trabecular Separation
Tb.Th Trabecular Thickness
TBPf Trabecular Bone Pattern Factor
TNF-a Tumour Necrosis Factor-alpha
TV Trabecular Volume
mm Micrometer
mCT
VOI Micro Computed Tomography
Volume Of Interest
Abstract
Osteoporotic fractures are of major socio-economic importance. Degradation of trabecular bone structures often result in significant weakening of bones and osteoporosis. Changes in subchondral bone structures are also related etiologically to rheumatoid arthritis.
The ultimate aim of in vivo bone research is to understand and measure bone structures, and to maintain or regenerate the mechanical properties of bone.
The anti-fracture resistance of a bone depends on 3 major components: bone mass, architecture and material properties. Bone mass can be measured non-invasively by simple X-ray based methods such as DXA (Dual energy X-ray Absorptiometry) and pQCT (peripheral Quantitative Computed Tomography), the latter being also able to determine cortical architectural features.
Recent developments in micro-computed tomography (µCT) techniques have enabled us to measure 3D trabecular architecture of rat bones non-invasively in-vivo. This is of immense interest since the trabecular structures are closely associated with the mechanical properties of the bone. The major advantages are that architectural changes can now be followed in life animals over time in contrast to the invasive histomorphometric-techniques. Also, the µCT measurements do not require any model assumptions in contrast to histomorphometry, which tries to calculate 3D structures from 2D histological sections. With these techniques, the structure and mechanical properties of the trabecular bone can be studied and understood at a higher level of magnification than was previously possible.
The micro-computed tomography scanner (vivaCT 20) is a novel prototype scanner, designed by SCANCO Medical, Bassersdorf, Switzerland. It is the first scanner with the ability to conduct in-vivo measurements at a high resolution of 26mm, requiring inhalation anaesthesia for only 9.3 minutes to map the entire metaphysis of a rat’s tibia.
The aim of this project was to test the scanner stringently by comparing it to other established methods in order to ensure that the results are of good, reliable quality before making the machine available to users worldwide.
Firstly, the threshold value for the cancellous bone detection algorithm was derived by cross-calibrating the vivaCT 20 with an existing, properly calibrated ex vivo mCT 20 scanner. Both these scanners use similar technology except for the long scanning time required by the mCT 20 scanner of several hours, which precludes its use in living animals. The cross-calibration procedure suggested a threshold value of 150, which was then adopted for all further studies.
The various tests carried out includes:
&Mac183; Comparison of the vivaCT 20 with the ex vivo mCT 20.
&Mac183; Determination of its machine- and handling precision (repetitive measurement of the same bone with or without repositioning of the animal).
&Mac183; Definition and selection of the ‘ideal’ region of interest for structural analysis.
&Mac183; Determine its ability to monitor changes in trabecular structures in response to various interventions over a period of time under several experimental conditions.
&Mac183; Determine its ability to discriminate dose-response relationships.
From the results we have obtained, we can confidently conclude that the scanner has passed all the tests and is qualified to take on more daunting research.
Key Words: osteoporosis; rat; trabecular architecture; mechanical properties;
CT scanner; cross-calibration; histomorphometry; densitometry
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