Chung-Hwan Chen

Proliferation and differentiation potential of human adipose‐derived mesenchymal stem cells isolated from elderly patients with osteoporotic fractures

Aging has less effect on adipose‐derived mesenchymal stem cells (ADSCs) than on bone marrow‐derived mesenchymal stem cells (BMSCs), but whether the fact holds true in stem cells from elderly patients with osteoporotic fractures is unknown. In this study, ADSCs and BMSCs of the same donor were harvested and divided into two age groups. Group A consisted of 14 young patients (36.4 ± 11.8 years old), and group B consisted of eight elderly patients (71.4 ± 3.6 years old) with osteoporotic fractures. We found that the doubling time of ADSCs from both age groups was maintained below 70 hrs, while that of BMSCs increased significantly with the number of passage. When ADSCs and BMSCs from the same patient were compared, there was a significant increase in the doubling time of BMSCs in each individual from passages 3 to 6. On osteogenic induction, the level of matrix mineralization of ADSCs from group B was comparable to that of ADSCs from group A, whereas BMSCs from group B produced least amount of mineral deposits and had a lower expression level of osteogenic genes. The p21 gene expression and senescence‐associated β‐galactosidase activity were lower in ADSCs compared to BMSCs, which may be partly responsible for the greater proliferation and differentiation potential of ADSCs. It is concluded that the proliferation and osteogenic differentiation of ADSCs were less affected by age and multiple passage than BMSCs, suggesting that ADSCs may become a potentially effective therapeutic option for cell‐based therapy, especially in elderly patients with osteoporosis.

Parathyroid hormone 1-34 inhibits terminal differentiation of human articular chondrocytes and osteoarthritis progression in rats.

OBJECTIVE Parathyroid hormone 1-34 (PTH[1-34]), a parathyroid hormone analog, shares the same receptor, PTH receptor 1, with parathyroid hormone-related peptide (PTHrP). This study was undertaken to address the hypothesis that PTH(1-34) inhibits terminal differentiation of articular chondrocytes and in turn suppresses the progression of osteoarthritis (OA). METHODS We studied the effect of PTH(1-34) on human articular chondrocytes with azacytidine (azaC)-induced terminal differentiation in vitro and on papain-induced OA in the knee joints of rats. In the in vitro study, we measured the levels of messenger RNA for SOX9, aggrecan, type II collagen, type X collagen, alkaline phosphatase (AP), Indian hedgehog (IHH), Bcl-2, and Bax by real-time polymerase chain reaction, levels of glycosaminoglycan (GAG) by dimethylmethylene blue assay, and rate of apoptosis by TUNEL staining. In the in vivo study, we evaluated the histologic changes in GAG, type II collagen, type X collagen, and chondrocyte apoptosis in the articular cartilage of rat knees. RESULTS AzaC induced terminal differentiation of human chondrocytes, including down-regulation of aggrecan, type II collagen, and GAG and up-regulation of type X collagen, alkaline phosphatase, and IHH. Apoptosis was reversed by 3-10 days of treatment with 10 nM PTH(1-34). SOX9 expression was not changed by either azaC or PTH(1-34) treatment. Bcl-2 and Bax were up-regulated on day 10 and day 14, respectively, after azaC induction of terminal differentiation, but PTH(1-34) treatment did not reverse this effect. Furthermore, PTH(1-34) treatment reversed papain-induced OA changes (decreasing GAG and type II collagen, and increasing type X collagen and chondrocyte apoptosis) in the knee joints of rats. CONCLUSION Our findings indicate that PTH(1-34) inhibits the terminal differentiation of human articular chondrocytes in vitro and inhibits progression of OA in rats in vivo, and may be used to treat OA.

The effect of the local delivery of alendronate on human adipose-derived stem cell-based bone regeneration.

Recent studies have shown that alendronate (Aln) enhances the osteogenesis of osteoblasts and bone marrow mesenchymal stem cells. In this study, we hypothesize that Aln may act as an osteo-inductive factor to stimulate the osteogenic differentiation of human adipose-derived stem cells (hADSCs) for bone regeneration. The in vitro effect of Aln (1-10 μM) on the osteogenic ability of hADSCs was evaluated by examining mineralization and alkaline phosphatase (ALP) activity. Bone morphogenetic protein 2 (BMP2) expression was measured using a real-time polymerase chain reaction and western blot analysis. Our results indicated that 5 μM Aln was sufficient to enhance BMP2 expression, ALP activity and mineralization in hADSCs. The in vivo effect of locally administered Aln on bone repair was examined in a rat critical-sized (7-mm) calvarial defect that was implanted with a hADSC-seeded poly(lactic-co-glycolic acid) (PLGA) scaffold. Aln (5 μM/100 μl/day) was injected locally into the defect site for one week. New bone formation was evaluated by radiographic and histological analyses at 8 and 12 weeks post-implantation. The expression levels of human BMP2 (hBMP2) and hADSC localization in defect sites were examined using immunohistochemistry analysis and fluorescent in situ hybridization, respectively. Results showed that local treatment of Aln on hADSC-seeded PLGA scaffolds at week 12 had a maximal effect on bone regeneration, enhancing mineralization and bone matrix formation. In addition, hADSCs and hBMP2 were also detected at the defect sites. These results demonstrated that local delivery of Aln, a potent osteo-inductive factor, enhances hADSC osteogenesis and bone regeneration.

Controlled-release of rhBMP-2 carriers in the regeneration of osteonecrotic bone.

Untreated osteonecrosis of the hip causes collapse of the femoral head and eventually leads to the development of premature degenerative arthritis. In order to reverse this late complication after the core decompression procedure, we studied three different types of carriers used to entrap recombinant human bone morphogenetic protein-2 (rhBMP-2) in terms of their performance in osteonecrosis regeneration and creeping substitution in Balb/C mice. The rhBMP-2 was loaded into PLGA-HAp microsphere in three different ways. We first verified the therapeutic dose in vitro using D1 and C2C12 cells. Then the individual performance of the three carrier preparations in vivo was examined by soft X-ray observation, histological analysis and immunostaining of bone tissue. In addition, the BMP-2 protein concentration activity in the serum was monitored. The results revealed that the bioactivity of rhBMP-2 released from a carrier with an ideal therapeutic dose was well maintained; this eventually helped to improve the healing and substitution of necrotic bone in vivo. These observations demonstrate that the in vivo performance of these newly developed rhBMP-2 delivery carriers correlates well with their in vitro release profiles. We concluded that sustained controlled-release of rhBMP-2 above a therapeutic dose could not only induce early callus wrapping of the necrotic bone but also produce neovascularization and substitution inside of the dead bone.

Dietary polyphenols and mechanisms of osteoarthritis

Osteoarthritis is a condition caused in part by injury, loss of cartilage structure and function, and an imbalance in inflammatory and anti-inflammatory pathways. It primarily affects the articular cartilage and subchondral bone of synovial joints and results in joint failure, leading to pain upon weight bearing including walking and standing. There is no cure for osteoarthritis, as it is very difficult to restore the cartilage once it is destroyed. The goals of treatment are to relieve pain, maintain or improve joint mobility, increase the strength of the joints and minimize the disabling effects of the disease. Recent studies have shown an association between dietary polyphenols and the prevention of osteoarthritis-related musculoskeletal inflammation. This review discusses the effects of commonly consumed polyphenols, including curcumin, epigallocatechin gallate and green tea extract, resveratrol, nobiletin and citrus fruits, pomegranate, as well as genistein and soy protein, on osteoarthritis with an emphasis on molecular antiosteoarthritic mechanisms.

Alendronate in the prevention of collapse of the femoral head in nontraumatic osteonecrosis: a two-year multicenter, prospective, randomized, double-blind, placebo-controlled study.

OBJECTIVE Osteonecrosis is one of the major debilitating skeletal disorders. Most patients with osteonecrosis of the femoral head eventually need surgery, usually total hip arthroplasty (THA), within a few years of onset. Previous studies showed that alendronate has a pharmacologic effect in reducing osteoclast activity and that it significantly reduced the incidence of collapse of the femoral head in the osteonecrotic hip. The purpose of this study was to determine the cumulative incidence of THA in patients with osteonecrosis of the femoral head and the time-to-event after treatment with alendronate versus placebo during the study period. METHODS A 2-year multicenter, prospective, randomized, double-blind study was performed. From June 2005 to December 2006, 64 patients were enrolled and randomly assigned to the alendronate or placebo group. In patients with bilateral hip osteonecrosis who met the inclusion criteria, both hips were counted in the analyses. Five patients were excluded from the analysis because they did not comply with any of the study regimens. Seven patients were ineligible because they were not diagnosed as having stage IIC or stage IIIC disease according to the University of Pennsylvania system. Thus, a total of 52 patients (65 hips) were assessed in this study. Disease progression was evaluated by radiography and magnetic resonance imaging (MRI). The Harris Hip Score and the Short Form 36 health survey were used to rate hip function and quality of life, respectively. RESULTS There was no significant difference in radiographic and MRI data between the 2 study groups. Four of 32 hips in the alendronate treatment group underwent THA, while 5 of 33 hips in the placebo group had THA (P = 0.837). No differences were noted in disease progression, Harris Hip Scores, or Short Form 36 scores between the 2 groups. CONCLUSION Alendronate has no obvious effect on preventing the necessity for THA, reducing disease progression, or improving life quality.

Simvastatin increases osteoblasts and osteogenic proteins in ovariectomized rats

Background  Previous reports have indicated that statins could prevent bone loss in ovariectomized (OVX) rats and increase the expressions of osteogenic genes in cultured osteoblasts. In this study, we hypothesized that simvastatin might increase osteoblast number and protein expressions of osteogenic markers localized in bones in concomitance with the prevention of bone loss in OVX rats.

(–)–Epigallocatechin Gallate Inhibition of Osteoclastic Differentiation via NF-Κb

People who regularly drink tea have been found to have a higher bone mineral density (BMD) and to be at less risk of hip fractures than those who do not drink it. Green tea catechins such as (-)-epigallocatechin gallate (EGCG) have been reported to increase osteogenic functioning in mesenchymal stem cells. However, its effect on osteoclastogenesis remains unclear. In this study, we investigated the effect of EGCG on RANKL-activation osteoclastogenesis and NF-kappaB in RAW 264.7, a murine preosteoclast cell line. EGCG (10-100 microM) significantly suppressed the RANKL-induced differentiation of osteoclasts and the formation of pits in murine RAW 264.7 cells and bone marrow macrophages (BMMs). EGCG appeared to target osteoclastic differentiation at an early stage but had no cytotoxic effect on osteoclast precursors. In addition, it significantly inhibited RANKL-induced NF-kappaB transcriptional activity and nuclear translocation. We conclude that EGCG inhibits osteoclastogenesis through its activation of NF-kappaB.

Fruits and dietary phytochemicals in bone protection

Osteoporosis is a disease of bone characterized by loss of bone matrix and deterioration of bone microstructure that leads to an increased risk of fracture. Cross-sectional studies have shown a positive association between higher fruit intake and higher bone mineral density. In this review, we evaluated animal and cellular studies of dried plum and citrus and berry fruits and bioactive compounds including lycopene, phenolics, favonoids, resveratrol, phloridzin, and pectin derived from tomato, grapes, apples, and citrus fruits. In addition, human studies of dried plum and lycopene were reviewed. Animal studies strongly suggest that commonly consumed antioxidant-rich fruits have a pronounced effect on bone, as shown by higher bone mass, trabecular bone volume, number, and thickness, and lower trabecular separation through enhancing bone formation and suppressing bone resorption, resulting in greater bone strength. Such osteoprotective effects seem to be mediated via antioxidant or anti-inflammatory pathways and their downstream signaling mechanisms, leading to osteoblast mineralization and osteoclast inactivation. In future studies, randomized controlled trials are warranted to extend the bone-protective activity of fruits and their bioactive compounds. Mechanistic studies are needed to differentiate the roles of phytochemicals and other constitutes in bone protection offered by the fruits. Advanced imaging technology will determine the effective doses of phytochemicals and their metabolites in improving bone mass, microarchitecture integrity, and bone strength, which is a critical step in translating the benefits of fruit consumption on osteoporosis into clinical data.

Isolated talonavicular arthrodesis for talonavicular arthritis.

We have reviewed a single surgeons experience with isolated talonavicular fusion in 16 patients with talonavicular arthritis. Fixation was either by staples or screws. Fifteen solid unions were achieved in the 16 patients who were followed (mean: 51 months). The average Ankle-Hindfoot Scale improved from 77.2 preoperatively to 92.9 postoperatively (p < 0.001). Subjectively, 15 patients were satisfied and one patient dissatisfied with the results. Further osteoarthritis in the adjacent joints was noted in five patients. We concluded that isolated talonavicular fusion is an effective method of treatment of talonavicular arthritis regarding pain relief and functional improvement. Though osteoarthritis was found in some adjacent joints postoperatively, the results were still satisfactory.