Stromal cell-derived factor-1α regulates chondrogenic differentiation via activation of the Wnt/ß-catenin pathway in mesenchymal stem cells.

BACKGROUND: Mesenchymal stem cells (MSCs) have been applied to treat degenerative articular diseases, and stromal cell-derived factor-1α (SDF-1α) may enhance their therapeutic efficacy. However, the regulatory effects of SDF-1α on cartilage differentiation remain largely unknown. Identifying the specific regulatory effects of SDF-1α on MSCs will provide a useful target for the treatment of degenerative articular diseases. AIM: To explore the role and mechanism of SDF-1α in cartilage differentiation of MSCs and primary chondrocytes. METHODS: The expression level of C-X-C chemokine receptor 4 (CXCR4) in MSCs was assessed by immunofluorescence. MSCs treated with SDF-1α were stained for alkaline phosphatase (ALP) and with Alcian blue to observe differentiation. Western blot analysis was used to examine the expression of SRY-box transcription factor 9, aggrecan, collagen II, runt-related transcription factor 2, collagen X, and matrix metalloproteinase (MMP)13 in untreated MSCs, of aggrecan, collagen II, collagen X, and MMP13 in SDF-1α-treated primary chondrocytes, of glycogen synthase kinase 3ß (GSK3ß) p-GSK3ß and ß-catenin expression in SDF-1α-treated MSCs, and of aggrecan, collagen X, and MMP13 in SDF-1α-treated MSCs in the presence or absence of ICG-001 (SDF-1α inhibitor). RESULTS: Immunofluorescence showed CXCR4 expression in the membranes of MSCs. ALP stain was intensified in MSCs treated with SDF-1α for 14 d. The SDF-1α treatment promoted expression of collagen X and MMP13 during cartilage differentiation, whereas it had no effect on the expression of collagen II or aggrecan nor on the formation of cartilage matrix in MSCs. Further, those SDF-1α-mediated effects on MSCs were validated in primary chondrocytes. SDF-1α promoted the expression of p-GSK3ß and ß-catenin in MSCs. And, finally, inhibition of this pathway by ICG-001 (5 µmol/L) neutralized the SDF-1α-mediated up-regulation of collagen X and MMP13 expression in MSCs. CONCLUSION: SDF-1α may promote hypertrophic cartilage differentiation in MSCs by activating the Wnt/ß-catenin pathway. These findings provide further evidence for the use of MSCs and SDF-1α in the treatment of cartilage degeneration and osteoarthritis.

Successful management of lower limb nonunion using locking plates and bone graft with retention of intramedullary nail.

This study aims to investigate the clinical effect of the combined an additional locking plate with bone graft based on retaining the original intramedullary nail for the treatment of lower limb nonunion.From June 2008 to December 2012, 39 patients were admitted and treated, who developed non-infectious bone nonunion after intramedullary nail fixation for long bone fracture in the lower limb. Additional locking plate and autogenous iliac bone grafting were performed for these patients, in which the original intramedullary nail was retained. Follow-ups were performed once at postoperative months 1, 2, 3, 6, and 12, and every year onwards. During these follow-ups, imaging and clinical function examinations were performed, in order to observe callus growth and the fractured limb functions.All patients have been followed-up, in which the duration of these follow-ups ranged between 8 and 24 months. All patients gained bony union within 6 to 11 months, and the healing rate was 100%. Radiographic healing time ranged between 8 and 15 months. Full weight-bearing time ranged between 2 and 10 months. According to Harris hip scores and Hospital for Special Surgery (HSS) Knee joint scores, 17 cases were excellent, 2 cases were good, and 1 case was acceptable; with an excellent and good rate of 95.00%. According to HHS score for the knee, 15 cases were excellent, 3 cases were good, and 1 case was acceptable; with an excellent and good rate of 94.74%.The combined treatment of the additional blocking plate with bone grafting based on retaining the original intramedullary nail for bone nonunion could effectively eliminate lateral and rotatory instability of the fractured ends. This surgical method had a short operation time, high healing rate and other advantages.

Necrostatin-1 Attenuates Trauma-Induced Mouse Osteoarthritis and IL-1ß Induced Apoptosis via HMGB1/TLR4/SDF-1 in Primary Mouse Chondrocytes.

Necrostatin-1 (Nec-1) is a specific small molecule inhibitor of receptor-interacting protein kinase 1 (RIPK1) that specifically inhibits phosphorylation of RIPK1. RIPK1 regulates inflammation and cell death by interacting with receptor-interacting serine/threonine protein kinases 3(RIPK3). We hypothesized that Nec-1 may have anti-inflammatory efficacy in patients with osteoarthritis (OA), as the pathophysiology of OA involves the activation of inflammation-related signaling pathways and apoptosis. In this study, we explored the effects of Nec-1 on interleukin (IL)-1ß-induced inflammation in mouse chondrocytes and the destabilised medial meniscus (DMM) mouse model. Inhibiting RIPK1 with Nec-1 dramatically suppressed catabolism both in vivo and in vitro, but did not inhibit changes in subchondral bone. Nec-1 abolished the in vitro increases in matrix metalloproteinase (MMP) and ADAM metallopeptidase with thrombospondin type 1 motif 5 (ADAMTs5) expression induced by IL-1ß. However, adding high-mobility group box 1 (HMGB1) partially abrogated this effect, indicating the essential role of HMGB1 and Nec-1 in the protection of primary chondrocytes. Furthermore, Nec-1 decreased the expression of Toll-like receptor 4 (TLR4) and stromal cell-derived factor-1 (SDF-1), and attenuated the interaction between TLR4 and HMGB1. Western blot results suggested that Nec-1 significantly suppressed IL-1ß-induced NF-κB transcriptional activity, but not MAPK pathway. Micro-computed tomography, immunohistochemical staining, and Safranin O/Fast Green staining were used in vivo to assess the degree of destruction of OA cartilage. The results show that NEC-1 can significantly reduce the degree of destruction of OA cartilage. Therefore, Nec-1 may be a novel therapeutic candidate to treat OA.

Inhibitory effects of high glucose/insulin environment on osteoclast formation and resorption in vitro.

Patients with type 2 diabetes mellitus (T2DM) exhibit hyperglycemia and hyperinsulinemia and increased risk of fracture at early stage, but they were found to have normal or even enhanced bone mineral density (BMD). This study was aimed to examine the molecular mechanisms governing changes in bone structure and integrity under both hyperglycemic and hyperinsulinemic conditions. Monocytes were isolated from the bone marrow of the C57BL/6 mice, induced to differentiate into osteoclasts by receptor activator of nuclear factor kappa-B ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) and exposed to high glucose (33.6 mmol/L), high insulin (1 µmol/L), or a combination of high glucose/high insulin (33.6 mmol/L glucose and 1 µmol/L insulin). Cells cultured in α-MEM alone served as control. After four days of incubation, the cells were harvested and stained for tartrate resistant acid phosphatase (TRAP). Osteoclast-related genes including RANK, cathepsin K and TRAP were determined by using real-time PCR. The resorptive activity of osteoclasts was measured by using a pit formation assay. Osteoclasts that were derived from monocytes were of multinucleated nature and positive for TRAP, a characteristic marker of osteoclasts. Cell counting showed that the number of osteoclasts was much less in high glucose and high glucose/high insulin groups than in normal glucose and high insulin groups. The expression levels of RANK and cathepsin K were significantly decreased in high glucose, high insulin and high glucose/high insulin groups as compared with normal glucose group, and the TRAP activity was substantially inhibited in high glucose environment. The pit formation assay revealed that the resorptive activity of osteoclasts was obviously decreased in high glucose group and high glucose/high insulin group as compared with normal group. It was concluded that osteoclastogenesis is suppressed under hyperglycemic and hyperinsulinemic conditions, suggesting a disruption of the bone metabolism in diabetic patients.

[Study on the variations of hepatitis B virus status among people having received hepatitis B vaccine].

OBJECTIVE: To study the epidemiological pattern and trends of hepatitis B virus (HBV) in the area where people had been immunized by HBV vaccine for long time. METHODS: Through cluster sampling and cross-sectional study, relative information and blood samples from people in Long-an county by families were collected. Signals of HBV infection were tested by solid-phase reverse immunosorbent test. RESULTS: (1) The average HBsAg positive rate was 7.5% with anti-HBs as 44.5 %, and anti-HBc as 47.8%. The positive rates of HBsAg and anti-HBc among 0-19 year-olds were lower than those of > or = 20 year-olds. (2) The positive rates of HBsAg, anti-HBc and HBV infection among HBV vaccine immunized group were 2.8%, 12.0% and 12.5% respectively, comparing with which among the un-immunized group as 10.2%, 69.8% and 71.2% respectively. (3) The HBsAg positive rate of male was higher than the female's but with no significant difference of anti-HBs and anti-HBc between different sexes. (4) The average HBsAg positive rate of 0-19 years old group was only 2.4%, while that of 20-30 years old group was 13.6%-17.7% and dropped from 60 years old group and on. The anti-HBs positive rate of 0-19 years old people started to drop significantly by age. The anti-HBs and anti-HBc positive rates of > or = 20 years people were showing a rising trend by ages. CONCLUSION: It seemed obviously that the HBV epidemiological patterns had changed after HBV vaccine had been universally used for long time in newborns. The age peak of infection had been pushed backward for nearly 20 years. It had been proved that the HBV vaccine immunization program had obtained excellent efficacy.

[Variations of hepatitis B virus infection epidemic pattern after long-term HBV vaccine immunization].

OBJECTIVE: To study the epidemic pattern and trend of HBV infection in the area where the people had been immunized by HBV vaccine for 20 years. METHODS: The whole sampling method was applied in combination with cross-sectional investigation. Blood samples were taken from every member of families. Markers of HBV infection were determined by using solid-phase radioimmunoassay (SPRIA). RESULTS: (1) The average HBsAg positive rate was 7.5%. The positive rate of markers for HBV infection of 0-19 years old subjects were lower than those of > or = 20 years old subjects. (2) The positive rate of HBsAg of 0-19 years old subjects in 1985 was higher than that in 2005. The anti-HBs positive rate in 1985 stemmed to be higher with age. It was 12.4% in 1- age group to 53.8% in >60 years age group. While the result of 2005 showed that the anti-HBs positive rate of 0-19 years old subjects dropped with age. The anti-HBc positive rate in 1985 also tended to be higher with age. But the result of 2005 showed that the rate of 0-19 years old subjects was just 1.4% to 16.8%. CONCLUSION: The epidemic patterns of HBV infection have had significant variations in the target population. HBV vaccine immunization has obtained excellent efficacy.