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Rabbit study of novel delivery vehicles for rhBMP
Abstracts The 7th International
166s
Congress
Bone Vol. 19, No. 3, Supplement September 1996: 129%169s
of Bone Morphometry
149
147 TRABECULAR
BONE
CHARACTERISTICS B. van Rietw, Biomechanics
QUALITY
MEASURED
OF TRABECULAR H. Weinans
FROM
MECHANICAL
ARCHITECTURE
and R. Huiskes
Sectmn, Inst. of Orthopaedics,
Univ. Nljmegen, The Netherlands
It is generally believed that trabecular bone adapts its architecture such that it is optimally
adapted to the external loading. However,
from stereological measurements
alone the quality of the hone architecture with respect to its load carrying capacity can not be determined. In the present study an alternative measure for the mechanical quality of bone is introduced, which is determined directly from the mechanical characteristics of bone architecture. This measure is
based on the notion
that bone architecture is well
adapted to the loading if all trabeculae are equally loaded (uniform stress distribution). To investigate this, the local trabecular stresses are calculated, using Finite Element models of the trabecular architecture, and represented by stress distribution curves. The uniformity
of the element loading was expressed a the width of these curves. A cubic specimen ofbabecular bone was reconstructed in a computer, using
serial sectioning and digitization’.
With this procedure the trabecular architecture was
represented by 521,090 voxels (bricks) which were converted to elements in a finite element (FE) model’. The FE-model was used to calculate the local trabecular stresses and strains in the elements due to external loads. From these results, local trabecular loads for any external deformation could be obtained. The stresses and strains were plotted in distribution curves to evaluate the uniformity of the load transfer. The most uniform distribution was found when the specimen was loaded in its anatomical direction. When loaded in the hvo orthogonal transversal directions, the standard deviations increased by a factor 4.0 and 8.9 respectively, indicating a much lesser efficient load transfer through the structure. These analyses show that under non-physiological loading conditions the structure is not well arranged to withstand the load. It h.w a low mechanical efficiency for these loading conditions. In fact the presently inInduced method provides the inherent mechanical etXciency of a trabecular stmch~re, which can now be classified by means of the uniformity of the load transfer. Less uniform stress distributions signify relatively larger areas of unloaded and overloaded bone. This is not only detrimental for the risk of frachxe, hut it may also contribute to the adaptation process in the hzbecular bone structllre, thereby possibly enhancing fracture risk in the future. References:
‘Odgaard et al., J.Micros.
Acknowledgments: (NCF),
159:691 ‘Van Riethergen et al. JBiomech.
HIGH FREQUENCY OF IRON BONE DEPOSITS IN A MEXICAN POPULATION WITH RENAL OSTEODISTROPHY. Vel&ouez-Forero F. TrImdad P. Altamrrano E, P&a JC Laboratorlo de Patologia y Metabollsmo Mineral Oseo del Hospital Motel. Depatiamento de Nefrologia y Metabollsmo Mmeral del Instltuto Naclonal de la Nutrwjn Salvador Zubrren y Departamento de Nefrologia del Hospital de Espewlldades del CMN Slglo XXI Mexico D F M&+x Renal osteodlstrophy (ROD) is a mulbfactorlal drsease Aluminum deposrts have been lmpllcated m Its phystopathology Iron depwts have been seldom described m this enbty The present report Includes 60 pabents wth advanced renal fallure submrtted to pentoneal dralysls (45 patients 75%) and hemodlalysls (15 pabents 25%) for a period of 32 r 24 months, 31 males and 29 women mean age 46 r 25 years A bone biopsy was obtamed I” each patient after double tetracyclmg labeling Blochemrcal markers of bone remodeling were taken at the time of the bone biopsy Ca 6 6+ 0 9 mgldl, P 6 I+ 1 5 mgldl, Total Alk Phosphatase 197? 258, PTH medium molecule 4 9t 4 05 ng/ml( normal 0 4-O 7 ng/ml), Calcitonlne ll? 6 pglml (normal values 1-26 pglml) The histomorphometnc analysts was done followmg Sherrard et al crlterla with slight modrficabons, bewde the usual stains aluminum, iron and amyloid stammgs were done m all bone specunens RESULTS.- Osterbs flbrosa cysbca (OFC) was found I” 27 pabents (45%). Mild bone disease I” 17 cases (26%). Mlxed bone disease ‘2 patients (3%). AdynamIc bone lesron (ABL) m 12 cases (20%). and Osteomalacra(OM) I” 2 cases (3%) Amylotd depowts were negative m all cases Alummum was detected m 6 patients (6 3%) and I” less than 10% of the splculae surface Iron deposrts were found m the mmeralrzatlon front m 31 patients (55 7%) Percent of iron deposits in the mineralization front Mixed ABL OM OFC Mlld 0 <25X 6 3 1 2 1 > 25% 2 1 11 1 3 Total 13/27 212 9117 5/12 2/2 CONCLUSIONSThe hrstomorphometrlc spectrum of ROD m our populabon IS slmrlar to the results reported m other serves and conhrms the tendency to an mcrease in the Incidence of ABL The study clearly showed a high frequency of rron deposits in the mlneralizatlon front We have no explanation for this flndmg and may be a phenomenon mfluenced by the geographlcal Impact
28:69
The work was sponsored by the Netherlands Computer Facility
by a Cray Research Grant and by the Royal Netherlands Academy of Arts and
Sciences.
148 RABBIT
150 STUDY
H. Vasconez’.
OF NOVEL
J. Rodgem’,
DELIVERY B. Fink’,
VEHICLES
S. Duggiral??,
FOR
rhBMP
P. Deluca2.
M. Faugere’.
H.
Malluche’ ‘Department of Surgery. Unwersity of Kentucky, College of Medrcine, Lexington. KY. ‘College of Pharmacy, Umvers~ty of Kentucky. and ‘Department of Nephrologylaone,M,neral Metabohsm, UnNerslty of Kentucky
The purpose of this study was to evaluate a new carrier matnx for bone morphogenetic protein (BMP) in bone defects. Although BMP is known lo induce bone growth, difficulty in keeping the material at the local site for sufficient periods of time has limited its successful application. Issues of BMP dose and pharmacokinetics must be resolved in order to assure an optimal response on the target tissue. From the basic research standpoint, a controlled delivery vehicle would greatly facilitate studies of the bone healing process. In this experiment, five groups of 8 mature male New Zealand White rabbits each underwent creation of a circular “critical size” defect. Experimental groups consisted of untreated controls, carboxymethylcellulose (CMC) matrix alone, methylcellulose (MC) matrix alone and one group of each matrix containing 90 ug per device of cloned recombinant human BMP-2 (rhBMP). Matrix implants were formulated. encoded and provided to our lab for implantation by our coinvestigators.’ Following implantation the rabbits were sacrificed at 2, 4, or 8 weeks postop. At necropsy. afler gross inspection, a 30 x 30 mm specimen including the defect area was obtained. The samples were processed in methylmethacrylate and stained with modified Masson-Goldner trichrome stains for histologic analysis. Qualitative histology revealed that there were distinct differences between methylcellulose (MC) and carboxymethylcellulose (CMC) implants. In controls, only small amounts of peripheral growth occurred. The general effect of the plain implants was to eliminate contact between the periosteum and the dura. thereby allowing some bone regrowth from the periphery of the defects. CMC implants performed much better in this regard than MC implants, possibly due to the increased hydrophilicity of CMC. In fact, CMC-implanted defects were bridged with new bone at 2 weeks in one of two animals, and at 8 weeks in 3/3 rabbits. MC implants showed only minimal bone healing. When BMP was added, results improved in that abundant new bone was produced over the &week observation especially in MC implants. New bone arose from the edges of the defect, and also from the center of the defect inside and around the implants. Remaining issues to be addressed include the reasons for bone growth in plain CMC implants, establishment of the minimal effective dose healing model.
I
D”gg,rsla. S
of rhBMP.
and
the
delineation
of the
dura’s
etal, Pharmaceubcal Development and Technaloqy, “olumel
role
in lhts
bone
AGE-DEPENDENT LOSS OF HORIZONTAL AND VERTICAL TRABECULAE IN THE HUMAN SPINE ‘M. Voael, ‘*M. Hahn, +T. Smit, “G. Delling ‘Orthopadische Klinik. “Osteopathologie und +Zentrum Biomechanik der Universittit, Martinistr. 52, D-20246 Hamburg It is of controversial discussion wether age-dependent loss of trabecular bone in the spine results in a preferential loss of horizontal trabeculae or not. This question is of substantial interest in regard to the biomechanical interpretation of histomorphologic data and therefore essential for the understanding of osteoporotic fractures of the spine. A review of the literature shows that most authors are convinced that horizontal trabeculae disappear faster than vertical ones. Nevertheless, quantitative analyses have been performed only in a very few cases. We analyzed the 2nd lumbar vertebral body of 25 autopsy cases without skeletal diseases. In each vertebral body we measured in a 9’9mm large area, 3mm from the endplate, and in the central part of the vertebral body. Surface stained block grindings were used which are comparable to 1Km thick conventional sections. The MIL (mean intercept length vector) in bone was measured in steps of one degree from 0 to 180 degree. In contrast to the common opinion, the MIL vector does not represent an ellipsoid. The MIL vector has two peaks, representing the preferential vertical and horizontal orientation. Age-dependent loss of trabecular bone volume (BV/TV) was significantly faster in the area close to the endplate than in the central part. In the central part of the vertebral body there was an equal agedependent loss of the MIL for horizontal and vertical orientations; in the part close to the endplate there was a significant faster decrease of MIL in the horizontal than in the vertical orientation. Our data demonstrate: the preferential age-dependent loss of horizontal trabeculae depends on the location within the vertebrae.
166s
Congress
Bone Vol. 19, No. 3, Supplement September 1996: 129%169s
of Bone Morphometry
149
147 TRABECULAR
BONE
CHARACTERISTICS B. van Rietw, Biomechanics
QUALITY
MEASURED
OF TRABECULAR H. Weinans
FROM
MECHANICAL
ARCHITECTURE
and R. Huiskes
Sectmn, Inst. of Orthopaedics,
Univ. Nljmegen, The Netherlands
It is generally believed that trabecular bone adapts its architecture such that it is optimally
adapted to the external loading. However,
from stereological measurements
alone the quality of the hone architecture with respect to its load carrying capacity can not be determined. In the present study an alternative measure for the mechanical quality of bone is introduced, which is determined directly from the mechanical characteristics of bone architecture. This measure is
based on the notion
that bone architecture is well
adapted to the loading if all trabeculae are equally loaded (uniform stress distribution). To investigate this, the local trabecular stresses are calculated, using Finite Element models of the trabecular architecture, and represented by stress distribution curves. The uniformity
of the element loading was expressed a the width of these curves. A cubic specimen ofbabecular bone was reconstructed in a computer, using
serial sectioning and digitization’.
With this procedure the trabecular architecture was
represented by 521,090 voxels (bricks) which were converted to elements in a finite element (FE) model’. The FE-model was used to calculate the local trabecular stresses and strains in the elements due to external loads. From these results, local trabecular loads for any external deformation could be obtained. The stresses and strains were plotted in distribution curves to evaluate the uniformity of the load transfer. The most uniform distribution was found when the specimen was loaded in its anatomical direction. When loaded in the hvo orthogonal transversal directions, the standard deviations increased by a factor 4.0 and 8.9 respectively, indicating a much lesser efficient load transfer through the structure. These analyses show that under non-physiological loading conditions the structure is not well arranged to withstand the load. It h.w a low mechanical efficiency for these loading conditions. In fact the presently inInduced method provides the inherent mechanical etXciency of a trabecular stmch~re, which can now be classified by means of the uniformity of the load transfer. Less uniform stress distributions signify relatively larger areas of unloaded and overloaded bone. This is not only detrimental for the risk of frachxe, hut it may also contribute to the adaptation process in the hzbecular bone structllre, thereby possibly enhancing fracture risk in the future. References:
‘Odgaard et al., J.Micros.
Acknowledgments: (NCF),
159:691 ‘Van Riethergen et al. JBiomech.
HIGH FREQUENCY OF IRON BONE DEPOSITS IN A MEXICAN POPULATION WITH RENAL OSTEODISTROPHY. Vel&ouez-Forero F. TrImdad P. Altamrrano E, P&a JC Laboratorlo de Patologia y Metabollsmo Mineral Oseo del Hospital Motel. Depatiamento de Nefrologia y Metabollsmo Mmeral del Instltuto Naclonal de la Nutrwjn Salvador Zubrren y Departamento de Nefrologia del Hospital de Espewlldades del CMN Slglo XXI Mexico D F M&+x Renal osteodlstrophy (ROD) is a mulbfactorlal drsease Aluminum deposrts have been lmpllcated m Its phystopathology Iron depwts have been seldom described m this enbty The present report Includes 60 pabents wth advanced renal fallure submrtted to pentoneal dralysls (45 patients 75%) and hemodlalysls (15 pabents 25%) for a period of 32 r 24 months, 31 males and 29 women mean age 46 r 25 years A bone biopsy was obtamed I” each patient after double tetracyclmg labeling Blochemrcal markers of bone remodeling were taken at the time of the bone biopsy Ca 6 6+ 0 9 mgldl, P 6 I+ 1 5 mgldl, Total Alk Phosphatase 197? 258, PTH medium molecule 4 9t 4 05 ng/ml( normal 0 4-O 7 ng/ml), Calcitonlne ll? 6 pglml (normal values 1-26 pglml) The histomorphometnc analysts was done followmg Sherrard et al crlterla with slight modrficabons, bewde the usual stains aluminum, iron and amyloid stammgs were done m all bone specunens RESULTS.- Osterbs flbrosa cysbca (OFC) was found I” 27 pabents (45%). Mild bone disease I” 17 cases (26%). Mlxed bone disease ‘2 patients (3%). AdynamIc bone lesron (ABL) m 12 cases (20%). and Osteomalacra(OM) I” 2 cases (3%) Amylotd depowts were negative m all cases Alummum was detected m 6 patients (6 3%) and I” less than 10% of the splculae surface Iron deposrts were found m the mmeralrzatlon front m 31 patients (55 7%) Percent of iron deposits in the mineralization front Mixed ABL OM OFC Mlld 0 <25X 6 3 1 2 1 > 25% 2 1 11 1 3 Total 13/27 212 9117 5/12 2/2 CONCLUSIONSThe hrstomorphometrlc spectrum of ROD m our populabon IS slmrlar to the results reported m other serves and conhrms the tendency to an mcrease in the Incidence of ABL The study clearly showed a high frequency of rron deposits in the mlneralizatlon front We have no explanation for this flndmg and may be a phenomenon mfluenced by the geographlcal Impact
28:69
The work was sponsored by the Netherlands Computer Facility
by a Cray Research Grant and by the Royal Netherlands Academy of Arts and
Sciences.
148 RABBIT
150 STUDY
H. Vasconez’.
OF NOVEL
J. Rodgem’,
DELIVERY B. Fink’,
VEHICLES
S. Duggiral??,
FOR
rhBMP
P. Deluca2.
M. Faugere’.
H.
Malluche’ ‘Department of Surgery. Unwersity of Kentucky, College of Medrcine, Lexington. KY. ‘College of Pharmacy, Umvers~ty of Kentucky. and ‘Department of Nephrologylaone,M,neral Metabohsm, UnNerslty of Kentucky
The purpose of this study was to evaluate a new carrier matnx for bone morphogenetic protein (BMP) in bone defects. Although BMP is known lo induce bone growth, difficulty in keeping the material at the local site for sufficient periods of time has limited its successful application. Issues of BMP dose and pharmacokinetics must be resolved in order to assure an optimal response on the target tissue. From the basic research standpoint, a controlled delivery vehicle would greatly facilitate studies of the bone healing process. In this experiment, five groups of 8 mature male New Zealand White rabbits each underwent creation of a circular “critical size” defect. Experimental groups consisted of untreated controls, carboxymethylcellulose (CMC) matrix alone, methylcellulose (MC) matrix alone and one group of each matrix containing 90 ug per device of cloned recombinant human BMP-2 (rhBMP). Matrix implants were formulated. encoded and provided to our lab for implantation by our coinvestigators.’ Following implantation the rabbits were sacrificed at 2, 4, or 8 weeks postop. At necropsy. afler gross inspection, a 30 x 30 mm specimen including the defect area was obtained. The samples were processed in methylmethacrylate and stained with modified Masson-Goldner trichrome stains for histologic analysis. Qualitative histology revealed that there were distinct differences between methylcellulose (MC) and carboxymethylcellulose (CMC) implants. In controls, only small amounts of peripheral growth occurred. The general effect of the plain implants was to eliminate contact between the periosteum and the dura. thereby allowing some bone regrowth from the periphery of the defects. CMC implants performed much better in this regard than MC implants, possibly due to the increased hydrophilicity of CMC. In fact, CMC-implanted defects were bridged with new bone at 2 weeks in one of two animals, and at 8 weeks in 3/3 rabbits. MC implants showed only minimal bone healing. When BMP was added, results improved in that abundant new bone was produced over the &week observation especially in MC implants. New bone arose from the edges of the defect, and also from the center of the defect inside and around the implants. Remaining issues to be addressed include the reasons for bone growth in plain CMC implants, establishment of the minimal effective dose healing model.
I
D”gg,rsla. S
of rhBMP.
and
the
delineation
of the
dura’s
etal, Pharmaceubcal Development and Technaloqy, “olumel
role
in lhts
bone
AGE-DEPENDENT LOSS OF HORIZONTAL AND VERTICAL TRABECULAE IN THE HUMAN SPINE ‘M. Voael, ‘*M. Hahn, +T. Smit, “G. Delling ‘Orthopadische Klinik. “Osteopathologie und +Zentrum Biomechanik der Universittit, Martinistr. 52, D-20246 Hamburg It is of controversial discussion wether age-dependent loss of trabecular bone in the spine results in a preferential loss of horizontal trabeculae or not. This question is of substantial interest in regard to the biomechanical interpretation of histomorphologic data and therefore essential for the understanding of osteoporotic fractures of the spine. A review of the literature shows that most authors are convinced that horizontal trabeculae disappear faster than vertical ones. Nevertheless, quantitative analyses have been performed only in a very few cases. We analyzed the 2nd lumbar vertebral body of 25 autopsy cases without skeletal diseases. In each vertebral body we measured in a 9’9mm large area, 3mm from the endplate, and in the central part of the vertebral body. Surface stained block grindings were used which are comparable to 1Km thick conventional sections. The MIL (mean intercept length vector) in bone was measured in steps of one degree from 0 to 180 degree. In contrast to the common opinion, the MIL vector does not represent an ellipsoid. The MIL vector has two peaks, representing the preferential vertical and horizontal orientation. Age-dependent loss of trabecular bone volume (BV/TV) was significantly faster in the area close to the endplate than in the central part. In the central part of the vertebral body there was an equal agedependent loss of the MIL for horizontal and vertical orientations; in the part close to the endplate there was a significant faster decrease of MIL in the horizontal than in the vertical orientation. Our data demonstrate: the preferential age-dependent loss of horizontal trabeculae depends on the location within the vertebrae.