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Mutations in Pycr1 cause progeroid changes in skin and bone due to reduced mitochondrial stress resistance
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Abstracts / Bone 47 (2010) S72–S241
spine BMD. There was significant interaction between Cdx-2 genotype and metPA on BMD at both hips and at spine (p < = 0.004). No interaction was observed between metPA and any other SNP genotype/haplotype with respect to BMD at any site, nor between mechPA and any SNP. These data provide evidence in support of a role for the VDR Cdx-2 polymorphism in modulating the effect of PA on BMD, and may provide some explanation for the conflicting roles reported for VDR SNPs in BMD. Disclosure of Interest: None declared Keywords: bone mineral density, habitual physical activity, vitamin D receptor gene doi:10.1016/j.bone.2010.04.354
PP219 Polymorphisms in the osteoprotegerin gene are associated with BMD and osteoporotic fractures L.B. Husted1,⁎, T. Harsløf1, K. Brixen2, P. Eiken3, J.-E.B. Jensen4, L. Rejnmark1, L. Mosekilde1, B.L. Langdahl1 1 Department of Endocrinology and Metabolism C, Aarhus University Hospital, Aarhus C, Denmark 2 Department of Endocrinology M, Odense University Hospital, Odense C, Denmark 3 Department of Cardiology and Endocrinology, Hillerød Hospital, Hillerød, Denmark 4 Department of Endocrinology, Hvidovre Hospital, Hvidovre, Denmark Osteoprotegerin (OPG) plays a key role in bone remodeling together with receptor activator of nuclear factor-κB (RANK) and RANK ligand (RANKL). Binding of OPG to RANKL prevents RANKL from binding to RANK and thereby osteoclast differentiation, activity, and survival are inhibited. We have previously demonstrated that SNPs in the promoter and exon1 of the OPG gene are associated with BMD and risk of vertebral fractures. Recently, a Spanish study has found an effect of SNPs in the promoter and intron 1 on BMD and fractures risk, and genome-wide association studies have identified SNPs associated with BMD and fracture risk upstream and downstream of the gene. The aim of this study was to investigate the effect of these SNPs on perimenopausal BMD, postmenopausal bone loss, and the risk of incident osteoporotic fractures, and to examine whether there is an interaction between these SNPs and response to hormone therapy (HT). The study population comprised 1713 perimenopausal women participating in the Danish Osteoporosis Prevention Study (DOPS). Genotyping was performed using the Sequenom MassARRAY Genotyping system, TaqMan assays or RFLP and BMD was measured by DXA at baseline and after 5 and 10 years. We found no effect of the SNPs identified in genome-wide association studies on perimenopausal BMD, and there was no consistent effect on postmenopausal bone loss or response to HT. However, for two of these SNPs, rs6993813 and rs6469804, individuals homozygous for the variant allele had a reduced risk of incident non-vertebral osteoporotic fractures compared with carriers of the normal allele (HR: 0.69 (CI: 0.49-0.97), p = 0.03 and HR: 0.66 (CI: 0.46-0.94), p = 0.02, respectively). The promoter SNPs, rs3102735 and rs3134069, were associated with decreased perimenopausal BMD at the lumbar spine (dominant model: p = 0.01 and p = 0.03, respectively), and the intron 1 SNP rs1032129 was associated with decreased perimenopausal BMD at the femoral neck and total hip (recessive model: p = 0.03 for both sites). Rs7844539 in intron 4 was not associated with perimenopausal BMD, but this SNP was associated with increased bone loss at the femoral neck (p = 0.047) and total hip (p = 0.009) after 5 years and with increased risk of incident non-vertebral osteoporotic fractures (dominant model: p = 0.03).
In conclusion our results support that SNPs in the OPG gene play a role in the development of osteoporosis, but no interaction between these SNPs and HT could be demonstrated. Disclosure of Interest: None declared Keywords: BMD, osteoprotegerin, polymorphisms doi:10.1016/j.bone.2010.04.355
PP220 Mutations in Pycr1 cause progeroid changes in skin and bone due to reduced mitochondrial stress resistance B. Reversade1, N. Escande-Beillard1, A. Dimopoulou2, B. Fischer2, Y. Li3, H. Kayserili4, L. Al-Gazali5, F. Brancati6, H. Lee7, B. O'Connor7, U. Kornak2,⁎,8, B. O'Connor9, M. Schmidt-von Kegler8, B. Merriman7, S.F. Nelson7, A. Masri10, D. Guerra11, A. nanda12, A. Rajab13, A. Grix14, A. Sommer15, A.R. Janecke16, E. Steichen16, P. Nürnberg17, P. Seemann8, G. Zambruno18, B. Dallapiccola6, M. Schuelke19, S. Robertson20, H. Hamamy21, B. Wollnik3, L. Van Maldergem22, S. Mundlos2 1 Institute of Medical Biology, A⁎STAR, Singapore, Singapore 2 Institute of Medical Genetics, Charité Universitaetsmedizin Berlin, Berlin, Germany 3 Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany 4 Medical Genetics Department, Istanbul Medical Faculty, University of Istanbul, Istanbul, Turkey 5 Department of Paediatrics, Faculty of Medicine and Health Sciences, UAE University, Al Ain, United Arab Emirates 6 CSS-Mendel Institute, Rome, Italy 7 Department of Human Genetics, David Geffen School of Medicine, Los Angeles, United States 8 Max Planck Institute for Molecular Genetics, Berlin, Germany 9 Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, United States 10 Departments of Pediatrics, Obstetrics and Gynecology, The University of Jordan, Amman, Jordan 11 Department of Biomedical Sciences, University of Modena and Reggio Emilia, Modena, Italy 12 As'ad Al-Hamad Dermatology Center, Al-Sabah Hospital, Kuwait, Kuwait 13 Genetic Unit, DGHA, Ministry of Health, Muscat, Oman 14 The Permanente Medical Group, Sacramento, United States 15 The Ohio State University College of Medicine and Nationwide Children's Hospital, Columbus, United States 16 Innsbruck Medical University, Innsbruck, Austria 17 Cologne Center for Genomics (CCG), Universität zu Köln, Cologne, Germany 18 Laboratory of Molecular and Cell Biology, IDI-IRCCS, Rome, Italy 19 Department of Neuropediatrics and NeuroCure Clinical Research Center, Charité Universitaetsmedizin Berlin, Berlin, Germany 20 Department of Paediatrics and Child Health, University of Otago, Dunedin, New Zealand 21 National Center for Diabetes, Endocrinology and GeneticsNational Center for Diabetes, Endocrinology and Genetics, Amman, Jordan 22 Centre de génétique humaine, CHU du Sart-Tilman, Liège, Belgium Autosomal recessive cutis laxa (ARCL) describes a group of syndromal disorders that are often associated with a progeroid appearance, lax and wrinkled skin, osteopenia, and joint hypermobility. Homozygosity mapping in several kindreds with ARCL identified a candidate region on chromosome 17q25. By highthrouput sequencing of the entire candidate region we detected disease-causing mutations in the gene PYCR1 encoding pyrroline-5carboxylate reductase, an enzyme involved in proline biosynthesis. However, serum proline levels were not significantly reduced in patients. Expression analysis revealed extremely high PYCR1 mRNA levels in osteoblasts and skin fibroblasts. All mutations entailed a
Abstracts / Bone 47 (2010) S72–S241
reduction of protein expression to a variable degree. We found that the gene product localizes to mitochondria. Altered mitochondrial morphology, membrane potential and increased apoptosis rates upon oxidative stress were evident in patient fibroblasts. Knockdown of the orthologous gene in zebrafish and frog embryos led to epidermal abnormalities that were accompanied by a massive increase of apoptosis. Thus, local production of proline in mitochondria seems to be important for the resistance of osteoblasts and other cells to oxidative stress. As increasing oxidative stress is discussed as a general aging mechanism our results might have relevance for common age-related osteoporosis. Disclosure of Interest: None declared Keywords: mitochondria, osteoporosis, oxidative stress resistance doi:10.1016/j.bone.2010.04.356
PP221 Association of CER1 polymorphisms with osteoporosis in Greek women; correlations with biochemical markers and bone density T. Koromila1,⁎, Z. Dailiana2, S. Samara3, C. Chassanidis3, P. Georgoulias4, V. Aleporou-Marinou1, P. Kollia1 1 Laboratory of Human Genetics, Department of Biology, University of Athens, Athens, Greece 2 Department of Orthopaedic Surgery, University of Thessalia, Larissa, Greece 3 Laboratory of Medical Genetics and Cytogenetics, University of Thessalia, Larissa, Greece 4 Department of Nuclear Medicine, School of Medicine, University of Thessalia, Larissa, Greece Osteoporosis is a common skeletal disease where the rate of bone turnover can be assessed by measuring biochemical markers. A number of studies suggest an important genetic component in the determination of peak bone mass and susceptibility to later fractures–. In this study we evaluated the contribution of singlenucleotide polymorphisms (SNPs) of CER1 gene in osteoporotic and control groups. 350 osteoporotic and 100 healthy Greek women (4585 age) included in the study. BMD measured by dual energy x-ray absorptiometry(DXA). Bone markers [osteocalcin/calcitonin/insulin growth factor I(IGF-I)/parathormon(PTH)/leptin] were measured in the serum. CER1 SNPs were studied by PCR and sequencing of the whole gene. IGF-I and osteocalcin values were the only statistically different among the osteoporotic and control group; however, IGF-I differentiated women with low BMD at greater level than the other studied bone markers(P < 0.0001). A weak and none correlation between leptin/BMD was observed in patients(P = 0.045) and controls(P = 0.575), respectively, while no association was detected between calcitonin/PTH and BMD values at any group. CER1 gene analysis revealed 5 SNPs at the positions 239C > G(rs3747532), 1058G > T(rs1494360), 2160A > G, 2355A > G(rs17289263), and 2749T > C. In patients, the SNPs frequencies were: 78.6% heterozygotes and 3.6% homozygotes for 239C > G SNP, 66.7% and 4.3% heterozygotes and homozygotes, respectively, with T allele at the position 1058, 52.4% heterozygotes and 9.5% homozygotes for the polymorphic site A > G nt.2160, 51.2% heterozygotes and 2.4% homozygotes for the G allele at nt.2355, whereas only heterozygotes (38.9%) for the polymorphism 2749T > C were determined (P < 0.001). However, in controls the SNPs detected only in heterozygosity and the overall distributions of the SNPs 239C > G, 1058G > T, 2160A > G, 2355A > G, and 2749T > C, were 38.9%, 31.3%, 15.6%, 9.4%, 6.9% (P < 0.001), respectively. Our results showed that among the studied markers only IGF-1 and osteocalcin differentiated the osteoporotic from the normal women. Furthermore, all the above SNPs, except rs3747532, were firstly correlated with osteoporotics. Allele frequencies of the controls were significantly lower than those of patients for any of the 5 polymorphisms. These data provide the first evidence of
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an association between the aforementioned genotypes in CER1 gene and osteoporosis while a correlation of the above genotypes with bone markers could be useful for the prediction of the disease. Disclosure of Interest: None declared Keywords: biochemical markers, CER1 gene polymorphisms, osteoporosis doi:10.1016/j.bone.2010.04.357
PP222 Genetic differences in the osteogenic differentiation potency according to the classification of the ossification of the posterior longitudinal ligament of the cervical spine H. Kudo1,⁎, A. Ono1, T. Numasawa1, S. Tanaka1, S. Toh1, S. Motomura2, K.-I. Furukawa2 1 Orthopaedic Surgery, Hirosaki University Graduate School of Medicine, Hirosaki, Japan 2 Pharmacology, Hirosaki University Graduate School of Medicine, Hirosaki, Japan Introduction: Ossification of the posterior longitudinal ligament (OPLL) of the spine is characterized by ectopic bone formation in the spinal ligaments. OPLL causes compression of the spinal cord and leads to various degree of myelopathy. Ossification types have been classified into four types by X-rays (Figure 1): continuous, segmental, mixed, and circumscribed type. It was reported that patients with mixed or continuous types of OPLL had the greatest risk for progression of OPLL clinically. Thus, we investigated genetic differences in osteogenic differentiation potency according to the classification of OPLL. Methods: The diagnosis of OPLL or non-OPLL was confirmed using X-rays and CT image. We categorized into three groups; non-OPLL group, OPLL segmental group (segmental or circumscribed type) or OPLL continuous group (continuous or mixed type). Paraspinal ligaments were harvested aseptically from patients during surgery, and cultured by the explant method. Cells were cultured in osteogenic medium (OS: DMEM + 1% FBS + 0.1 μM dexamethasone) or control medium. The mRNA expressions of bone-related markers; BMP-2, Osterix, ALP were quantified by real time RT-PCR. To evaluate ALP activity staining, the stained area was measured, and the ratio of stained cells to total cells was determined. Results: In OPLL continuous group, mRNA expressions of BMP-2, ALP and Osterix were increased respectively after exposing to OS (Figure 2). In ALP activity staining, OPLL continuous group had higher ALP activity than OPLL segmental group. OPLL continuous group showed high osteogenic potency in comparison to OPLL segmental group. We propose to distinguish OPLL continuous group from OPLL segmental group. Image/Graph:
Abstracts / Bone 47 (2010) S72–S241
spine BMD. There was significant interaction between Cdx-2 genotype and metPA on BMD at both hips and at spine (p < = 0.004). No interaction was observed between metPA and any other SNP genotype/haplotype with respect to BMD at any site, nor between mechPA and any SNP. These data provide evidence in support of a role for the VDR Cdx-2 polymorphism in modulating the effect of PA on BMD, and may provide some explanation for the conflicting roles reported for VDR SNPs in BMD. Disclosure of Interest: None declared Keywords: bone mineral density, habitual physical activity, vitamin D receptor gene doi:10.1016/j.bone.2010.04.354
PP219 Polymorphisms in the osteoprotegerin gene are associated with BMD and osteoporotic fractures L.B. Husted1,⁎, T. Harsløf1, K. Brixen2, P. Eiken3, J.-E.B. Jensen4, L. Rejnmark1, L. Mosekilde1, B.L. Langdahl1 1 Department of Endocrinology and Metabolism C, Aarhus University Hospital, Aarhus C, Denmark 2 Department of Endocrinology M, Odense University Hospital, Odense C, Denmark 3 Department of Cardiology and Endocrinology, Hillerød Hospital, Hillerød, Denmark 4 Department of Endocrinology, Hvidovre Hospital, Hvidovre, Denmark Osteoprotegerin (OPG) plays a key role in bone remodeling together with receptor activator of nuclear factor-κB (RANK) and RANK ligand (RANKL). Binding of OPG to RANKL prevents RANKL from binding to RANK and thereby osteoclast differentiation, activity, and survival are inhibited. We have previously demonstrated that SNPs in the promoter and exon1 of the OPG gene are associated with BMD and risk of vertebral fractures. Recently, a Spanish study has found an effect of SNPs in the promoter and intron 1 on BMD and fractures risk, and genome-wide association studies have identified SNPs associated with BMD and fracture risk upstream and downstream of the gene. The aim of this study was to investigate the effect of these SNPs on perimenopausal BMD, postmenopausal bone loss, and the risk of incident osteoporotic fractures, and to examine whether there is an interaction between these SNPs and response to hormone therapy (HT). The study population comprised 1713 perimenopausal women participating in the Danish Osteoporosis Prevention Study (DOPS). Genotyping was performed using the Sequenom MassARRAY Genotyping system, TaqMan assays or RFLP and BMD was measured by DXA at baseline and after 5 and 10 years. We found no effect of the SNPs identified in genome-wide association studies on perimenopausal BMD, and there was no consistent effect on postmenopausal bone loss or response to HT. However, for two of these SNPs, rs6993813 and rs6469804, individuals homozygous for the variant allele had a reduced risk of incident non-vertebral osteoporotic fractures compared with carriers of the normal allele (HR: 0.69 (CI: 0.49-0.97), p = 0.03 and HR: 0.66 (CI: 0.46-0.94), p = 0.02, respectively). The promoter SNPs, rs3102735 and rs3134069, were associated with decreased perimenopausal BMD at the lumbar spine (dominant model: p = 0.01 and p = 0.03, respectively), and the intron 1 SNP rs1032129 was associated with decreased perimenopausal BMD at the femoral neck and total hip (recessive model: p = 0.03 for both sites). Rs7844539 in intron 4 was not associated with perimenopausal BMD, but this SNP was associated with increased bone loss at the femoral neck (p = 0.047) and total hip (p = 0.009) after 5 years and with increased risk of incident non-vertebral osteoporotic fractures (dominant model: p = 0.03).
In conclusion our results support that SNPs in the OPG gene play a role in the development of osteoporosis, but no interaction between these SNPs and HT could be demonstrated. Disclosure of Interest: None declared Keywords: BMD, osteoprotegerin, polymorphisms doi:10.1016/j.bone.2010.04.355
PP220 Mutations in Pycr1 cause progeroid changes in skin and bone due to reduced mitochondrial stress resistance B. Reversade1, N. Escande-Beillard1, A. Dimopoulou2, B. Fischer2, Y. Li3, H. Kayserili4, L. Al-Gazali5, F. Brancati6, H. Lee7, B. O'Connor7, U. Kornak2,⁎,8, B. O'Connor9, M. Schmidt-von Kegler8, B. Merriman7, S.F. Nelson7, A. Masri10, D. Guerra11, A. nanda12, A. Rajab13, A. Grix14, A. Sommer15, A.R. Janecke16, E. Steichen16, P. Nürnberg17, P. Seemann8, G. Zambruno18, B. Dallapiccola6, M. Schuelke19, S. Robertson20, H. Hamamy21, B. Wollnik3, L. Van Maldergem22, S. Mundlos2 1 Institute of Medical Biology, A⁎STAR, Singapore, Singapore 2 Institute of Medical Genetics, Charité Universitaetsmedizin Berlin, Berlin, Germany 3 Center for Molecular Medicine Cologne, University of Cologne, Cologne, Germany 4 Medical Genetics Department, Istanbul Medical Faculty, University of Istanbul, Istanbul, Turkey 5 Department of Paediatrics, Faculty of Medicine and Health Sciences, UAE University, Al Ain, United Arab Emirates 6 CSS-Mendel Institute, Rome, Italy 7 Department of Human Genetics, David Geffen School of Medicine, Los Angeles, United States 8 Max Planck Institute for Molecular Genetics, Berlin, Germany 9 Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, United States 10 Departments of Pediatrics, Obstetrics and Gynecology, The University of Jordan, Amman, Jordan 11 Department of Biomedical Sciences, University of Modena and Reggio Emilia, Modena, Italy 12 As'ad Al-Hamad Dermatology Center, Al-Sabah Hospital, Kuwait, Kuwait 13 Genetic Unit, DGHA, Ministry of Health, Muscat, Oman 14 The Permanente Medical Group, Sacramento, United States 15 The Ohio State University College of Medicine and Nationwide Children's Hospital, Columbus, United States 16 Innsbruck Medical University, Innsbruck, Austria 17 Cologne Center for Genomics (CCG), Universität zu Köln, Cologne, Germany 18 Laboratory of Molecular and Cell Biology, IDI-IRCCS, Rome, Italy 19 Department of Neuropediatrics and NeuroCure Clinical Research Center, Charité Universitaetsmedizin Berlin, Berlin, Germany 20 Department of Paediatrics and Child Health, University of Otago, Dunedin, New Zealand 21 National Center for Diabetes, Endocrinology and GeneticsNational Center for Diabetes, Endocrinology and Genetics, Amman, Jordan 22 Centre de génétique humaine, CHU du Sart-Tilman, Liège, Belgium Autosomal recessive cutis laxa (ARCL) describes a group of syndromal disorders that are often associated with a progeroid appearance, lax and wrinkled skin, osteopenia, and joint hypermobility. Homozygosity mapping in several kindreds with ARCL identified a candidate region on chromosome 17q25. By highthrouput sequencing of the entire candidate region we detected disease-causing mutations in the gene PYCR1 encoding pyrroline-5carboxylate reductase, an enzyme involved in proline biosynthesis. However, serum proline levels were not significantly reduced in patients. Expression analysis revealed extremely high PYCR1 mRNA levels in osteoblasts and skin fibroblasts. All mutations entailed a
Abstracts / Bone 47 (2010) S72–S241
reduction of protein expression to a variable degree. We found that the gene product localizes to mitochondria. Altered mitochondrial morphology, membrane potential and increased apoptosis rates upon oxidative stress were evident in patient fibroblasts. Knockdown of the orthologous gene in zebrafish and frog embryos led to epidermal abnormalities that were accompanied by a massive increase of apoptosis. Thus, local production of proline in mitochondria seems to be important for the resistance of osteoblasts and other cells to oxidative stress. As increasing oxidative stress is discussed as a general aging mechanism our results might have relevance for common age-related osteoporosis. Disclosure of Interest: None declared Keywords: mitochondria, osteoporosis, oxidative stress resistance doi:10.1016/j.bone.2010.04.356
PP221 Association of CER1 polymorphisms with osteoporosis in Greek women; correlations with biochemical markers and bone density T. Koromila1,⁎, Z. Dailiana2, S. Samara3, C. Chassanidis3, P. Georgoulias4, V. Aleporou-Marinou1, P. Kollia1 1 Laboratory of Human Genetics, Department of Biology, University of Athens, Athens, Greece 2 Department of Orthopaedic Surgery, University of Thessalia, Larissa, Greece 3 Laboratory of Medical Genetics and Cytogenetics, University of Thessalia, Larissa, Greece 4 Department of Nuclear Medicine, School of Medicine, University of Thessalia, Larissa, Greece Osteoporosis is a common skeletal disease where the rate of bone turnover can be assessed by measuring biochemical markers. A number of studies suggest an important genetic component in the determination of peak bone mass and susceptibility to later fractures–. In this study we evaluated the contribution of singlenucleotide polymorphisms (SNPs) of CER1 gene in osteoporotic and control groups. 350 osteoporotic and 100 healthy Greek women (4585 age) included in the study. BMD measured by dual energy x-ray absorptiometry(DXA). Bone markers [osteocalcin/calcitonin/insulin growth factor I(IGF-I)/parathormon(PTH)/leptin] were measured in the serum. CER1 SNPs were studied by PCR and sequencing of the whole gene. IGF-I and osteocalcin values were the only statistically different among the osteoporotic and control group; however, IGF-I differentiated women with low BMD at greater level than the other studied bone markers(P < 0.0001). A weak and none correlation between leptin/BMD was observed in patients(P = 0.045) and controls(P = 0.575), respectively, while no association was detected between calcitonin/PTH and BMD values at any group. CER1 gene analysis revealed 5 SNPs at the positions 239C > G(rs3747532), 1058G > T(rs1494360), 2160A > G, 2355A > G(rs17289263), and 2749T > C. In patients, the SNPs frequencies were: 78.6% heterozygotes and 3.6% homozygotes for 239C > G SNP, 66.7% and 4.3% heterozygotes and homozygotes, respectively, with T allele at the position 1058, 52.4% heterozygotes and 9.5% homozygotes for the polymorphic site A > G nt.2160, 51.2% heterozygotes and 2.4% homozygotes for the G allele at nt.2355, whereas only heterozygotes (38.9%) for the polymorphism 2749T > C were determined (P < 0.001). However, in controls the SNPs detected only in heterozygosity and the overall distributions of the SNPs 239C > G, 1058G > T, 2160A > G, 2355A > G, and 2749T > C, were 38.9%, 31.3%, 15.6%, 9.4%, 6.9% (P < 0.001), respectively. Our results showed that among the studied markers only IGF-1 and osteocalcin differentiated the osteoporotic from the normal women. Furthermore, all the above SNPs, except rs3747532, were firstly correlated with osteoporotics. Allele frequencies of the controls were significantly lower than those of patients for any of the 5 polymorphisms. These data provide the first evidence of
S155
an association between the aforementioned genotypes in CER1 gene and osteoporosis while a correlation of the above genotypes with bone markers could be useful for the prediction of the disease. Disclosure of Interest: None declared Keywords: biochemical markers, CER1 gene polymorphisms, osteoporosis doi:10.1016/j.bone.2010.04.357
PP222 Genetic differences in the osteogenic differentiation potency according to the classification of the ossification of the posterior longitudinal ligament of the cervical spine H. Kudo1,⁎, A. Ono1, T. Numasawa1, S. Tanaka1, S. Toh1, S. Motomura2, K.-I. Furukawa2 1 Orthopaedic Surgery, Hirosaki University Graduate School of Medicine, Hirosaki, Japan 2 Pharmacology, Hirosaki University Graduate School of Medicine, Hirosaki, Japan Introduction: Ossification of the posterior longitudinal ligament (OPLL) of the spine is characterized by ectopic bone formation in the spinal ligaments. OPLL causes compression of the spinal cord and leads to various degree of myelopathy. Ossification types have been classified into four types by X-rays (Figure 1): continuous, segmental, mixed, and circumscribed type. It was reported that patients with mixed or continuous types of OPLL had the greatest risk for progression of OPLL clinically. Thus, we investigated genetic differences in osteogenic differentiation potency according to the classification of OPLL. Methods: The diagnosis of OPLL or non-OPLL was confirmed using X-rays and CT image. We categorized into three groups; non-OPLL group, OPLL segmental group (segmental or circumscribed type) or OPLL continuous group (continuous or mixed type). Paraspinal ligaments were harvested aseptically from patients during surgery, and cultured by the explant method. Cells were cultured in osteogenic medium (OS: DMEM + 1% FBS + 0.1 μM dexamethasone) or control medium. The mRNA expressions of bone-related markers; BMP-2, Osterix, ALP were quantified by real time RT-PCR. To evaluate ALP activity staining, the stained area was measured, and the ratio of stained cells to total cells was determined. Results: In OPLL continuous group, mRNA expressions of BMP-2, ALP and Osterix were increased respectively after exposing to OS (Figure 2). In ALP activity staining, OPLL continuous group had higher ALP activity than OPLL segmental group. OPLL continuous group showed high osteogenic potency in comparison to OPLL segmental group. We propose to distinguish OPLL continuous group from OPLL segmental group. Image/Graph: