| Co-Investigator(Kenkyū-buntansha) |
ASO Yoshinori 東京医科歯科大学, 大学院・医歯学総合研究科, 寄付講座教員 (50345279)
四宮 謙一 東京医科歯科大学, 大学院・医歯学総合研究科, 教授 (20111594)
竹田 秀 慶應義塾大学, 医学部, 准教授 (30376727)
立川 哲彦 昭和大学, 歯学部, 教授 (10085772)
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| Budget Amount *help |
¥4,550,000 (Direct Cost: ¥3,500,000、Indirect Cost: ¥1,050,000)
Fiscal Year 2011: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
Fiscal Year 2010: ¥1,430,000 (Direct Cost: ¥1,100,000、Indirect Cost: ¥330,000)
Fiscal Year 2009: ¥1,560,000 (Direct Cost: ¥1,200,000、Indirect Cost: ¥360,000)
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| Research Abstract |
Purpose
Proinflammatory cytokines, such as IL-1 and tumor necrosis factorα(TNFα), are suspected of causing damage to osteoarthritis(OA) cartilage. TNFαcontent is elevated in the synovial fluid of OA joints. There are 2 cell surface receptors for TNFαincluding p55(TNFRI) and p75(TNFRII). TNFαand TNFRs I and II are up-regulated in OA cartilage. For bone metabolism, mice null for TNFR1 have significantly increased peak bone mass, resulting from elevated bone formation. In vitro, TNFαinhibited mineralization of osteoblasts through regulation of NF-kB activity. Thus, whereas TNFR1 plays roles in osteoblast differentiation, its functions in chondrocyte metabolism is unknown. Endochondral ossification is an essential process for development of OA, which is characterized by cartilage degradation and osteophyte formation. The aim of this study is to investigate the role of TNFR1 in the maintenance of articular tissues.
Materials and methods
Histological evaluation of aged TNFR1-/-mice
Knee joints o
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f TNFR1-/-and their wild-type(WT) littermates(52-60 weeks of age) were evaluated immunohistologicaly. Whole knee joints were removed by dissection, fixed in 4% paraformaldehyde, and decalcified in EDTA. After dehydration and paraffin embedding, serial 5-□m sagittal sections were made from the whole medial compartment of the knee joint. Three sections were picked up at 100-□m intervals from the weight-bearing region of each medial plateau of tibia, and were stained with Safranin O-fast green and HE. Osteophyte area and articular cartilage thickness were quantified in each slide using Image-Pro Plus 4. 1 software.
Surgical induction of OA in TNFR1-/-mice.
TNFR1-/-and their wild-type(WT) littermates were surgically induced to develop OA by medial collateral ligament transection and medial meniscectomy. Four weeks after surgery, the mice were euthanized.
Results
At 52-60 weeks of age, whereas osteophyte formation was detected at the medio-anterior edge of tibial plateau regardless of genotype, osteophyte area was significantly increased in TNFR1-/-mice. Moreover, articular cartilage thickness was reduced at medial end region of tibia plateau in TNFR1-/-mice, suggesting ossification of articular chondrocyte was advanced by TNFR1 deficiency. We next compared osteoarthritis development between adult littermates of wild-type and TNFR1-/-mice by creating a surgical osteoarthritis model through induction of instability to the knee joints. Histological evaluation confirmed that the TNFR1 deficiency caused significant acceleration for osteophyte formation. Degree of cartilage destruction was almost comparable between TNFR1-/-and WT. Real time PCR analysis for epiphyseal chondrocyte from new born mice revealed mRNA expression of type X collagen, a marker for chondrocyte hypertrophy, was significantly elevated in TNFR1-/-mice compared to WT littermate. Currently, we are picking up a number of candidate genes for targets of TNFR1 in the course of chondrocyte hypertrophy using cDNA microarray analysis.
Conclusion
Endogenous TNFR1 had a protective effect against osteophyte formation through inhibition of type X collagen synthesis. Less
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