Factors regulating bone remodeling processes in aseptic implant loosening

This study was undertaken to screen periprosthetic tissues (PPTs) under specified conditions for a series of molecular components and describe them in bone remodeling processes within aseptic loosening. PPT samples were obtained from patients undergoing revision surgery of endoprostheses (n = 24) and synovial tissues from patients with OA (control) (n = 18), patients with any form of inflammatory arthritides were excluded. Tissue samples were examined via microbiology, histology (H&E, TRAP), immunohistochemistry (CD68/anti‐S100a4), quantitative real‐time PCR (ALP, COL1A1, cathepsin K, M‐CSF, MMP13, OPG, RANK, RANKL, TNF‐α, and TRAP) and an endotoxin‐assay. PPT samples contained a variety of cellular components and stained positive for TRAP (56%), CD68 (100%), and S100a4 (100%). Wear debris were found in cells staining positive for CD68 and S100a4. In PPTs significantly higher ALP, COL1A1, MMP‐13, RANK, RANKL, and TRAP expression were found along with a significantly higher RANKL/OPG ratio and a significantly lower OPG expression. No significant difference was observed for M‐CSF, TNF‐α, cathepsin K, and endotoxin levels. In conclusion we found osteogenic proteins (ALP, COL1A1), a proteolytic enzyme (MMP‐13), markers for osteoclast differentiation (RANK, RANKL), and osteoclast activity (TRAP) to be increased in PPT, whereas OPG expression decreased significantly in comparison to control. We present data about a large series of molecular components in PPT and describe novel and key findings about their expression levels in regards to aseptic implant loosening. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:248–257, 2017.

     

Low‐magnitude high‐frequency vibration enhances gene expression related to callus formation,mineralization and remodeling during osteoporotic fracture healing in rats

Low magnitude high frequency vibration (LMHFV) has been shown to improve anabolic and osteogenic responses in osteoporotic intact bones and during osteoporotic fracture healing; however, the molecular response of LMHFV during osteoporotic fracture healing has not been investigated. It was hypothesized that LMHFV could enhance osteoporotic fracture healing by regulating the expression of genes related to chondrogenesis (Col‐2), osteogenesis (Col‐1) and remodeling (receptor activator for nuclear factor‐ κ B ligand (RANKL) and osteoproteger (OPG)). In this study, the effects of LMHFV on both osteoporotic and normal bone fracture healing were assessed by endpoint gene expressions, weekly radiographs, and histomorphometry at weeks 2, 4 and 8 post‐treatment. LMHFV enhanced osteoporotic fracture healing by up‐regulating the expression of chondrogenesis‐, osteogenesis‐ and remodeling‐related genes (Col‐2 at week 4 (p = 0.008), Col‐1 at week 2 and 8 (p < 0.001and p = 0.008) and RANKL/OPG at week 8 (p = 0.045)). Osteoporotic bone had a higher response to LMHFV than normal bone and showed significantly better results as reflected by increased expression of Col‐2 and Col‐1 at week 2 (p < 0.001 for all), larger callus width at week 2 (p = 0.001), callus area at week 1 and 5(p < 0.05 for all) and greater relative area of osseous tissue (p = 0.002) at week 8. This study helps to understand how LMHFV regulates gene expression of callus formation, mineralization and remodeling during osteoporotic fracture healing. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:1572–1579, 2014.     

Changes in the expression of matrix extracellular genes and TGFB family members in rotator cuff tears

Lack of synthesis of extracellular matrix compounds may contribute to degeneration of the tendons. Thus, we aimed to evaluate the expression of extracellular matrix and TGFB family members in ruptured and non‐ruptured tendons of the rotator cuff, as well as the effect of clinical factors on gene expression in tendon samples, and the relationship between histological findings and altered gene expression. Injured and non‐injured supraspinatus tendon samples and subscapular non‐injured tendon samples were collected from 38 patients with rotator cuff tears. Non‐injured supraspinatus tendons were obtained from eight controls. Specimens were used for histological evaluation, quantification of collagen fibers, and mRNA and protein expression analyses. Increased COL1A1, COL1A2, COL3A1, COL5A1, FN1, TNC, and TGFBR1 mRNA expression was observed in the tear samples (p < 0.05). Duration of symptoms was correlated with the levels of collagen type I/III fibers (p = 0.032; ρ = 0.0447) and FN1 immunostaining (p = 0.031; ρ = 0.417). Smoking was associated with increased frequency of microcysts, myxoid degeneration, and COL5A1, FN1, TNC, and TGFB1 mRNA expression (p < 0.05). FN1 immunostaining was correlated with the number of years of smoking (p = 0.048; ρ = 0.384). Lower levels of collagen type I/III fibers were detected in samples with fissures (0 = 0.046). High frequency of microcysts was associated with increased COL5A1, FN1, and TNC expression (p < 0.05, for all comparisons). Neovascularization was associated with reduced FN1 (p = 0.035) and TGFBR1 expression (p = 0.034). Our findings show differential expression of matrix extracellular genes and TGFB family members in the degeneration process involved in rotator cuff tears. These molecular alterations are influenced by clinical factors. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:2542–2553, 2018.

     

硫酸钙对人骨髓基质干细胞向成骨细胞转化时基因表达的影响

目的 观察硫酸钙(CS)在人骨髓基质干细胞(BMSCs)向成骨细胞转化过程中对成骨基因表达的影响,探讨硫酸钙修复骨缺损可能的生物学机制.方法 制备硫酸钙浸提的成骨诱导液(实验组)与常规的成骨诱导液(对照组),分别加入人BMSCs培养瓶(各3例),使其向成骨细胞的诱导分化,注意观察细胞的分化和生长.培养至第7天时进行RNA的抽提、纯化和质量检测,并合成cDNA,采用成骨功能基因芯片分别检测实验组和对照组中各种成骨基因的变化.结果 实验组和对照组细胞均生长良好,缓慢增殖,但实验组向成骨细胞分化的趋势要明显较对照组好.成骨基因芯片共检测到89种基因,其中有23种基因表达改变显著(Fold change＞2,P＜0.05).表达上调超过2倍的基因包括:AMELY、BMP2、COL4A3、COMP、EGF、FLT1、IGF1、ITGA2、MMP10、MMP2、TGFB2、TGFBR1、VDR和VEGFA.表达下调超过2倍的有:COL2A1、COL15A1、COL1A1、COL1A2、COL5A1、CSF2、FGF1、ITGA3和MMP8.结论 硫酸钙促进了人BMSCs向成骨细胞转化的过程,这种作用与硫酸钙促进成骨基因表达上调、合成活性因子增加相关,说明硫酸钙具有潜在的骨诱导活性,可以作为良好的骨修复替代材料,促进细胞的骨修复能力.     

Polymorphisms in BMP4 and FGFR1 genes are associated with fracture non‐union

Fracture healing is a complex process influenced by a multitude of factors and expression of several thousand genes. Polymorphisms in these genes can lead to an extended healing process and explain why certain patients are more susceptible to develop non‐union. A total of 16 SNPs within five genes involved in bone repair pathogenesis (FAM5C, BMP4, FGF3, FGF10, and FGFR1) were investigated in 167 patients with long bone fractures, 101 with uneventful healing, and 66 presenting aseptic non‐unions. Exclusion criteria were patients presenting pathological fractures, osteoporosis, hypertrophic and infected non‐unions, pregnancy, and children. All genetic markers were genotyped using TaqMan real‐time PCR. Chi‐square test was used to compare genotypes, allele frequencies, and haplotype differences between groups. Binary logistic regression analyzed the significance of many covariates and the incidence of non‐union. Statistical analysis revealed open fracture to be a risk factor for non‐union development (p < 0.001, OR 3.6 [1.70–7.67]). A significant association of haplotype GTAA in BMP4 (p = 0.01) and FGFR1 rs13317 (p = 0.005) with NU could be observed. Also, uneventful healing showed association with FAM5C rs1342913 (p = 0.04). Our work supported the role of BMP4 and FGFR1 in NU fracture independently of the presence of previously described risk factors. © 2013 Orthopaedic Research Society Published by Wiley Periodicals, Inc. J Orthop Res 31:1971–1979, 2013     

Sclerostin Expression in Bile Ducts of Patients With Chronic Cholestasis May Influence the Bone Disease in Primary Biliary Cirrhosis

Sclerostin is involved in the regulation of osteoblastogenesis and little is known about its role in the development of bone disease in primary biliary cirrhosis (PBC), characterized by low bone formation. Therefore, we have assessed the circulating levels and the liver expression of sclerostin in this cholestatic disease. Serum sclerostin levels were measured in 79 women with PBC (mean age 60.6 ± 1.2 years) and in 80 control women. Lumbar and femoral bone mineral density (BMD), as well as parameters of mineral metabolism and bone remodeling, were measured. Moreover, sclerostin gene (SOST) expression in the liver was assessed by real‐time PCR in samples of liver tissue taken by biopsy in 11 PBC patients and in 5 normal liver specimens. Presence and distribution of sclerostin was evaluated in liver slices from 11 patients by immunohistochemistry. The severity of histologic lesions was assessed semiquantitatively in the same liver samples. PBC patients had higher sclerostin levels than controls (75.6 ± 3.9 versus 31.7 ± 1.6 pmol/L, p < 0.001). Serum sclerostin correlated inversely with markers of bone formation and resorption. Sclerostin mRNA in the liver was overexpressed compared with control samples (2.7‐fold versus healthy liver). Sclerostin was detected by immunohistochemistry in 7 of the 11 liver samples, mainly located in the bile ducts. Liver sclerostin was associated with the severity of cholangitis (p = 0.02) and indirectly with the degree of lobular inflammation (p = 0.03). Sclerostin mRNA expression was higher in samples that tested positive by immunohistochemistry and particularly in those with lobular granuloma (p = 0.02). The increased expression of sclerostin in the liver and the association with histologic cholangitis may explain the high serum levels of this protein in patients with PBC, thus suggesting that sclerostin may influence the decreased bone formation in this cholestatic disease. © 2016 American Society for Bone and Mineral Research.     

Bone Morphogenetic Protein 7 (BMP7) Gene Polymorphisms Are Associated With Inverse Relationships Between Vascular Calcification and BMD: The Diabetes Heart Study

Inverse relationships have been observed between BMD and vascular calcification (VC), suggesting an underlying metabolic pathway linking these processes. Bone morphogenetic proteins (BMPs) are potential candidate genes that may mediate this relationship. Four single nucleotide polymorphisms (SNPs) in the BMP2 gene, 2 SNPs in BMP4, and 16 SNPs in BMP7 were tested for association with measures of VC using CT (coronary and carotid arteries, abdominal aorta), and BMD was measured using DXA (lumbar spine, hip, and distal radius) and quantitative CT (QCT; thoracic and lumbar spine) in 920 European Americans from 374 Diabetes Heart Study families: 762 with type 2 diabetes. Variance components quantitative trait locus association analysis was computed using SOLAR software, and a bivariate principal component analysis (PCA) assessed for genetic relationships between BMD and VC. Association was observed between several measures of BMD and BMP7 rs17404303 (thoracic spine QCT p = 0.03; lumbar spine QCT p = 0.02; hip DXA p = 0.06, dominant models). In addition, 6 of 16 BMP7 SNPs showed significant and opposing effects on the bivariate PCA for VC and BMD (two‐sided exact test, p = 0.0143). Polymorphisms in BMP7 are associated with inverse relationships between bone mineralization and VC in the coronary, carotid, and abdominal aorta in a diabetes‐enriched cohort of European Americans.     

The masquelet induced membrane technique with BMP and a synthetic scaffold can heal a rat femoral critical size defect

Long bone defects can be managed by the induced membrane technique together with autologous bone graft. However, graft harvest is associated with donor site morbidity. This study investigates if a tricalcium phosphate hydroxyapatite scaffold can be used alone or in combination with bone active drugs to improve healing. Sprague Dawley rats (n = 40) were randomized into four groups. (A) scaffold, (B) BMP‐7, (C) BMP‐7 + scaffold, and (D) BMP‐7 + scaffold + systemic bisphosphonate at 2 weeks. Locked femoral nailing was followed by 6 mm segment removal and implantation of an epoxy spacer. At 4 weeks, the spacers were removed and the defects grafted. Eleven weeks later, the bones were explanted for evaluation with radiography, manual assessment, micro‐CT, histology, and Fourier Transform Infrared spectroscopy (FTIR). Isolated scaffolds (A) did not heal any defects, whereas the other treatments led to healing in 7/10 (B), 10/10 (C), and 9/10 (D) rats. Group D had greater volume of highly mineralized bone (p < 0.01) and higher bone volume fraction (p < 0.01) compared to all other groups. A synthetic scaffold + BMP‐7 combined with a bisphosphonate improved the callus properties in a rat femoral critical size defect, compared to both BMP‐7 and scaffold alone or the two combined. © 2015 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 33:488–495, 2015.     

Methylation of Bone SOST,Its mRNA,and Serum Sclerostin Levels Correlate Strongly With Fracture Risk in Postmenopausal Women

Inhibition of sclerostin, a glycoprotein secreted by osteocytes, offers a new therapeutic paradigm for treatment of osteoporosis (OP) through its critical role as Wnt/catenin signaling regulator. This study describes the epigenetic regulation of SOST expression in bone biopsies of postmenopausal women. We correlated serum sclerostin to bone mineral density (BMD), fractures, and bone remodeling parameters, and related these findings to epigenetic and genetic disease mechanisms. Serum sclerostin and bone remodeling biomarkers were measured in two postmenopausal groups: healthy (BMD T‐score > –1) and established OP (BMD T‐score < –2.5, with at least one low‐energy fracture). Bone specimens were used to analyze SOST mRNAs, single nucleotide polymorphisms (SNPs), and DNA methylation changes. The SOST gene promoter region showed increased CpG methylation in OP patients (n = 4) compared to age and body mass index (BMI) balanced controls (n = 4) (80.5% versus 63.2%, p = 0.0001) with replication in independent cohorts (n = 27 and n = 36, respectively). Serum sclerostin and bone SOST mRNA expression correlated positively with age‐adjusted and BMI‐adjusted total hip BMD (r = 0.47 and r = 0.43, respectively; both p < 0.0005), and inversely to serum bone turnover markers. Five SNPs, one of which replicates in an independent population‐based genomewide association study (GWAS), showed association with serum sclerostin or SOST mRNA levels under an additive model (p = 0.0016 to 0.0079). Genetic and epigenetic changes in SOST influence its bone mRNA expression and serum sclerostin levels in postmenopausal women. The observations suggest that increased SOST promoter methylation seen in OP is a compensatory counteracting mechanism, which lowers serum sclerostin concentrations and reduces inhibition of Wnt signaling in an attempt to promote bone formation. © 2014 American Society for Bone and Mineral Research.     

BMPR1A antagonist differentially affects cartilage and bone formation during fracture healing

A soluble form of BMP receptor type 1A (mBMPR1A‐mFC) acts as an antagonist to endogenous BMPR1A and has been shown to increase bone mass in mice. The goal of this study was to examine the effects of mBMPR1A‐mFC on secondary fracture healing. Treatment consisted of 10 mg/kg intraperitoneal injections of mBMPR1A‐mFC twice weekly in male C57BL/6 mice. Treatment beginning at 1, 14, and 21 days post‐fracture assessed receptor function during endochondral bone formation, at the onset of secondary bone formation, and during coupled remodeling, respectively. Control animals received saline injections. mBMPR1A‐mFC treatment initiated on day 1 delayed cartilage maturation in the callus and resulted in large regions of fibrous tissue. Treatment initiated on day 1 also increased the amount of mineralized tissue and up‐regulated many bone‐associated genes (p = 0.002) but retarded periosteal bony bridging and impaired strength and toughness at day 35 (p < 0.035). Delaying the onset of treatment to day 14 or 21 partially mitigated these effects and produced evidence of accelerated coupled remodeling. These results indicate that inhibition of the BMPR1A‐mediated signaling has negative effects on secondary fracture healing that are differentially manifested at different stages of healing and within different cell populations. These effects are most pronounced during the endochondral period and appear to be mediated by selective inhibition of BMPRIA signaling within the periosteum. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 34:2096–2105, 2016.     

Novel Genetic Variants Associated With Increased Vertebral Volumetric BMD,Reduced Vertebral Fracture Risk,and Increased Expression of SLC1A3 and EPHB2

Genome‐wide association studies (GWASs) have revealed numerous loci for areal bone mineral density (aBMD). We completed the first GWAS meta‐analysis (n = 15,275) of lumbar spine volumetric BMD (vBMD) measured by quantitative computed tomography (QCT), allowing for examination of the trabecular bone compartment. SNPs that were significantly associated with vBMD were also examined in two GWAS meta‐analyses to determine associations with morphometric vertebral fracture (n = 21,701) and clinical vertebral fracture (n = 5893). Expression quantitative trait locus (eQTL) analyses of iliac crest biopsies were performed in 84 postmenopausal women, and murine osteoblast expression of genes implicated by eQTL or by proximity to vBMD‐associated SNPs was examined. We identified significant vBMD associations with five loci, including: 1p36.12, containing WNT4 and ZBTB40; 8q24, containing TNFRSF11B; and 13q14, containing AKAP11 and TNFSF11. Two loci (5p13 and 1p36.12) also contained associations with radiographic and clinical vertebral fracture, respectively. In 5p13, rs2468531 (minor allele frequency [MAF] = 3%) was associated with higher vBMD (β = 0.22, p = 1.9 × 10^–8) and decreased risk of radiographic vertebral fracture (odds ratio [OR] = 0.75; false discovery rate [FDR] p = 0.01). In 1p36.12, rs12742784 (MAF = 21%) was associated with higher vBMD (β = 0.09, p = 1.2 × 10^–10) and decreased risk of clinical vertebral fracture (OR = 0.82; FDR p = 7.4 × 10^–4). Both SNPs are noncoding and were associated with increased mRNA expression levels in human bone biopsies: rs2468531 with SLC1A3 (β = 0.28, FDR p = 0.01, involved in glutamate signaling and osteogenic response to mechanical loading) and rs12742784 with EPHB2 (β = 0.12, FDR p = 1.7 × 10^–3, functions in bone‐related ephrin signaling). Both genes are expressed in murine osteoblasts. This is the first study to link SLC1A3 and EPHB2 to clinically relevant vertebral osteoporosis phenotypes. These results may help elucidate vertebral bone biology and novel approaches to reducing vertebral fracture incidence. © 2016 American Society for Bone and Mineral Research.     

Gene Expression for Extracellular Matrix Proteins in Shockwave-Induced Osteogenesis in Rats

To clarify the mechanisms underlying shockwave-induced osteogenesis, we applied shockwave to rat femoral shafts from the ventral side. We assessed bone mineral content (BMC) and bone mineral density (BMD), and analyzed the spatial and temporal gene expression for pro-1 (I) collagen (COL1A1), pro-1 (II) collagen (COL2A1), pro-1 (X) collagen (COL10A1), osteocalcin (OC) and osteopontin (OPN) using in situ hybridization. On the 21st day post-exposure, BMC and BMD in the exposed femur were elevated by 8.46% and 5.80%, respectively, relative to the unexposed femur. Immediately following exposure, there was evidence of scraping of the cortex and periosteal separation with hemorrhage. On day 4, new periosteal bone formation could be seen on the ventral and dorsal side of the femur. In the newly formed bone, COL1A1, OC and OPN were expressed in osteoblastic cells underlying the periosteum. On day 7, there was progression of periosteal bone and trabeculae formation. COL1A1 and OC were expressed in mature osteoblasts lining the trabeculae, whereas OPN was expressed in immature osteoblastic cells, osteocytes and osteoclasts. On day 14, bone remodeling commenced in the periosteal bone. COL1A1, OC and OPN were still expressed at this stage, however, signals were much weaker. Between 4–7 days, chondrocyte clusters were distributed multi-focally near the exposed site, and there was expression of COL2A1 but not of COL10A1. The results demonstrate that gene expression patterns of shockwave-induced osteogenesis are similar to those of periosteal hard callus formation during fracture healing. Shockwaves can yield dramatic activation of cells in normal long bones, and drive the cells to express genes for osteogenesis.     

Palovarotene inhibits connective tissue progenitor cell proliferation in a rat model of combat‐related heterotopic ossification

Heterotopic ossification (HO) develops in the extremities of wounded service members and is common in the setting of high‐energy penetrating injuries and blast‐related amputations. No safe and effective prophylaxis modality has been identified for this patient population. Palovarotene has been shown to reduce bone formation in traumatic and genetic models of HO. The purpose of this study was to determine the effects of Palovarotene on inflammation, progenitor cell proliferation, and gene expression following a blast‐related amputation in a rodent model (n = 72 animals), as well as the ability of Raman spectroscopy to detect early HO before radiographic changes are present. Treatment with Palovarotene was found to dampen the systemic inflammatory response including the cytokines IL‐6 (p = 0.01), TNF‐α (p = 0.001), and IFN‐γ (p = 0.03) as well as the local inflammatory response via a 76% reduction in the cellular infiltration at post‐operative day (POD)‐7 (p = 0.03). Palovarotene decreased osteogenic connective tissue progenitor (CTP‐O) colonies by as much as 98% both in vitro (p = 0.04) and in vivo (p = 0.01). Palovarotene treated animals exhibited significantly decreased expression of osteo‐ and chondrogenic genes by POD‐7, including BMP4 (p = 0.02). Finally, Raman spectroscopy was able to detect differences between the two groups by POD‐1 (p < 0.001). These results indicate that Palovarotene inhibits traumatic HO formation through multiple inter‐related mechanisms including anti‐inflammatory, anti‐proliferative, and gene expression modulation. Further, that Raman spectroscopy is able to detect markers of early HO formation before it becomes radiographically evident, which could facilitate earlier diagnosis and treatment. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:1135–1144, 2018.

     

Infrared spectroscopy indicates altered bone turnover and remodeling activity in renal osteodystrophy

Renal osteodystrophy alters metabolic activity and remodeling rate of bone and also may lead to different bone composition. The objective of this study was to characterize the composition of bone in high‐turnover renal osteodystrophy patients by means of Fourier transform infrared spectroscopic imaging (FTIRI). Iliac crest biopsies from healthy bone (n = 11) and patients with renal osteodystrophy (ROD, n = 11) were used in this study. The ROD samples were from patients with hyperparathyroid disease. By using FTIRI, phosphate‐to‐amide I ratio (mineral‐to‐matrix ratio), carbonate‐to‐phosphate ratio, and carbonate‐to‐amide I ratio (turnover rate/remodeling activity), as well as the collagen cross‐link ratio (collagen maturity), were quantified. Histomorphometric analyses were conducted for comparison. The ROD samples showed significantly lower carbonate‐to‐phosphate (p < .01) and carbonate‐to‐amide I (p < .001) ratios. The spatial variation across the trabeculae highlighted a significantly lower degree of mineralization (p < .05) at the edges of the trabeculae in the ROD samples than in normal bone. Statistically significant linear correlations were found between histomorphometric parameters related to bone‐remodeling activity and number of bone cells and FTIRI‐calculated parameters based on carbonate‐to‐phosphate and carbonate‐to‐amide I ratios. Hence the results suggested that FTIRI parameters related to carbonate may be indicative of turnover and remodeling rate of bone. © 2010 American Society for Bone and Mineral Research     

Mechanical stimulation alters tissue differentiation and molecular expression during bone healing

Further understanding of how mechanical cues modulate skeletal tissue differentiation can identify potential means of enhancing repair following injury or disease. Prior studies examined the effects of mechanical loading on osteogenesis, chondrogenesis, and fibrogenesis in an effort to enhance bony union. However, exploring how mechanical stimuli can divert the bone healing process towards formation of other mesenchymal tissues, as an endpoint, may elucidate new avenues for repair and regeneration of tissues such as cartilage and fibrous tissue. This study investigated the use of mechanical stimulation to promote cartilage rather than bone formation within an osteotomy. Our overall goal was to define skeletal tissue distribution and molecular expression patterns induced by the stimulation. Retired breeder Sprague‐Dawley rats (n = 85) underwent production of a mid‐diaphyseal, transverse femoral osteotomy followed by external fixation. Beginning on postoperative day 10 and continuing for 1, 2, or 4 weeks, a cyclic bending motion (+35°/?25° at 1 Hz) was applied in the sagittal plane for 15 min/day for 5 consecutive days/week. Control animals experienced continuous rigid fixation. Histological and molecular analyses indicated that stimulation substantially altered normal bone healing. Stimulated specimens exhibited an increase in cartilage volume over time, while control specimens demonstrated bony bridging. Stimulation induced upregulation of cartilage‐related genes (COL2A1 and COL10A1) and downregulation of bone morphogenetic proteins (BMPs) ‐4, ‐6 and ‐7. However, BMP‐3 was upregulated with stimulation. These findings illustrate that mechanical cues can selectively modulate osteogenesis and chondrogenesis in vivo, and suggest a potential basis for treatment regimens for injured or diseased cartilaginous tissues. © 2009 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res     

Accumulation of advanced‐glycation end products (AGEs) accelerates arthrogenic joint contracture in immobilized rat knee

Joint mobility decreases in the elderly and in diabetics, this process is thought to be caused by accumulation of advanced‐glycation end products (AGEs). Here, we aimed to elucidate the role of AGEs in joint contracture formation in rat knees. Rats were injected with ribose or saline into the knees twice weekly for 8 weeks. Pentosidine (AGE) levels were measured in the knee‐joint tissues. After serial injections, rats were subjected to unilateral knee‐joint immobilization in a flexion position for various periods. At day 21, the passive knee ranges of motions (ROMs) were measured. Knee joint histopathology were assessed, and the expression of fibrotic genes in the posterior joint capsules was examined using real‐time PCR. Ribose injection induced a 7.0‐fold increase in pentosidine levels relative to saline injection. Joint immobilization resulted in equal myogenic ROM restriction in both groups. Arthrogenic ROM restriction was greater with ribose injection in the immobilized joints (p < 0.05), but was not affected in nonimmobilized joints. Type‐I (COL1A1) and type‐III (COL3A1) collagen gene expression increased significantly in immobilized joints relative to nonimmobilized joints in the ribose group, but was not affected in the saline group. Ribose injection increased COL1A1 expression slightly and COL3A1 expression significantly in immobilized joints. Histologically, inflammatory changes appeared at day 3 of immobilization and peaked at day 7. These responses trended to be more severe and prolonged in the ribose group than in the saline group. Our data provide evidence for a causal relationship between AGEs and joint contracture formation following immobilization. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:854–863, 2018.