Mechanical Stretch Promotes the Osteogenic Differentiation of Bone Mesenchymal Stem Cells Induced by Erythropoietin.

INTRODUCTION: The effects of erythropoietin (EPO) on the behaviors of bone marrow mesenchymal stem cells (BMSCs) subjected to mechanical stretch remain unclear. This study was therefore aimed at establishing the dose-response effect of EPO stimulation on rat BMSCs and investigating the effects of mechanical stretch combined with EPO on the proliferation and osteogenic differentiation of BMSCs. MATERIAL AND METHODS: The proliferation and osteogenic differentiation of rat BMSCs were examined and compared using EPO with different concentrations. Thereafter, BMSCs were subjected to 10% elongation using a Flexcell strain unit, combined with 20 IU/ml EPO. The proliferation of BMSCs was detected by Cell Counting Kit-8, colony formation assay, and cell cycle assay; meanwhile, the mRNA expression levels of Ets-1, C-myc, Ccnd1, and C-fos were detected by reverse transcription and real-time quantitative PCR (qPCR). The osteogenic differentiation of BMSCs was detected by alkaline phosphatase (ALP) staining, and the mRNA expression levels of ALP, OCN, COL, and Runx2 were detected by qPCR. The role of the extracellular signal-regulated kinases 1/2 (ERK1/2) in the osteogenesis of BMSCs stimulated by mechanical stretch combined with 20 IU/ml EPO was examined by Western blot. RESULTS: Our results showed that effects of EPO on BMSCs included a dose-response relationship, with the 20 IU/ml EPO yielding the largest. Mechanical stretch combined with 20 IU/ml EPO promoted proliferation and osteogenic differentiation of BMSCs. The increase in ALP, mineral deposition, and osteoblastic genes induced by the mechanical stretch-EPO combination was inhibited by U0126, an ERK1/2 inhibitor. CONCLUSION: EPO was able to promote the proliferation and osteogenic differentiation of BMSCs, and these effects were enhanced when combined with mechanical stretch. The underlying mechanism may be related to the activation of the ERK1/2 signaling pathway.

Intensity-dependent effect of treadmill running on rat Achilles tendon.

It is understood that mechanical loading may affect tendon properties. However, how different mechanical loading conditions may affect tendons remains unknown. The present study aimed to investigate the effect of treadmill running at various intensities on rat Achilles tendon. A total of 18 male Wistar rats were randomly assigned to one of three groups: Control (CON), medium-intensity running (MIR), and high-intensity running (HIR). Following 8 weeks of treadmill running protocols, all Achilles tendons were harvested for histological observation and gene expression analysis. Significant morphological changes were observed with regular and large diameter collagen fibrils in the MIR group, whereas irregular and small diameter collagen fibrils were observed in the HIR group. Collagen type I was significantly upregulated in the MIR group compared with the CON group, and downregulated in the HIR group compared with the CON or MIR groups (P<0.05). However, collagen type III was significantly upregulated in the HIR group in comparison with the CON or MIR groups (P<0.05). Furthermore, the expression of matrix metallopeptidase-13 was significantly increased in the MIR and HIR groups compared with the CON group (P<0.05). The expression of tissue inhibitor of metalloproteinases-1 was increased in the MIR group compared with the CON group, but decreased in the HIR group compared with the CON and MIR groups (P<0.05). Additionally, decorin expression was significantly higher in the MIR group compared with the CON group, and significantly decreased in the HIR group compared with the CON or MIR groups (P<0.05). A converse pattern of changes in biglycan expression was identified among the three groups. Aggrecan expression was significantly higher in the HIR group compared with the CON or MIR groups (P<0.05). These findings indicated that moderate exercise may induce increased collagen synthesis and organize regular and large collagen fibers, thus benefiting the Achilles tendon. However, overuse during exercise may result in collagen degradation and disturbance, which predisposes individuals to injury.

Response of decorin to different intensity treadmill running.

Decorin is widely understood to affect collagen fibrillogenesis. However, little is understood about its response to various mechanical loading conditions. In the present study, 36 Wistar rats were randomly divided into control (CON), moderate treadmill running (MTR) and strenuous treadmill running (STR) groups. Animals in the MTR and STR groups were subjected to a 4­ or 8­week treadmill running protocol. Subsequently, all Achilles tendons were harvested to perform histological and biochemical analyses. Decorin expression was markedly increased in the MTR group compared with the CON group at 4 and 8 weeks. Conversely, decorin expression was markedly decreased in the STR group compared with the CON and MTR group at 4 and 8 weeks. Furthermore, between the two time points, decorin expression levels were significantly increased in the MTR group, whereas they were markedly decreased in the STR group. These results suggested that MTR exercise may induce increased decorin expression via a balance of MMP­2 and TIMP­2, improving tendon structure and function. However, STR exercise may result in degradation of decorin due to an imbalance of MMP­2 and TIMP­2, with a bias to MMP­2, resulting in a predisposition to tendinopathy.

Exercise affects biological characteristics of mesenchymal stromal cells derived from bone marrow and adipose tissue.

Both bone marrow mesenchymal stromal cells (BMSCs) and adipose-derived mesenchymal stromal cells (ADSCs) are good sources for tissue engineering. To maximize therapeutic efficacy of MSCs, an appropriate source of MSCs should be selected according to their own inherent characteristics for future clinical application. Hence, this study was conducted to compare proliferative, differential and antiapoptosis abilities of both MSCs derived from exercised and sedentary rats under normal and hypoxia/serum deprivation conditions (H/SD). Our results showed that exercise may enhance proliferative ability and decrease adipogenic ability of BMSCs and ADSCs. However, positive effect of exercise on osteogenesis was only observed for BMSCs in either environment. Little effect was observed on the antiapoptotic ability of both MSC types. It was also suggested that biological characteristics of both types were partly changed. It is therefore believed that BMSCs derived from exercised rat on early passage may be a good cell source for bone tissue engineering.

MiR-451 suppresses proliferation, migration and promotes apoptosis of the human osteosarcoma by targeting macrophage migration inhibitory factor.

Previous studies have shown that MiR-451 plays an important role in human osteosarcoma carcinogenesis, but the underlying mechanism by which MiR-451 affects the osteosarcoma has not been fully understood. This study intends to uncover the mechanism by which MiR-451 functions as a tumor suppressor. The expression of MiR-451 in osteosarcoma tissues and osteosarcoma cell lines was monitored by real-time PCR. The proliferation ability was examined by MTT and cell cycle assay. The migration and apoptosis of cells were monitored by migration assay and flow cytometry, respectively. Moreover, the angiogenesis of HUVEC cells transfected with MiR-451 mimics was examined by tube formation assay. The effect of MiR-451 on MIF was determined by luciferase assays and Western blot assay. The results showed that MiR-451 expression level was significantly reduced in the osteosarcoma compared with normal bone tissues. Overexpression of MiR-451 significantly attenuated the proliferation and migration, and induced the apoptosis of osteosarcoma cells. Furthermore, the angiogenesis of HUVEC cells transfected with MiR-451 mimics was assayed and the decreased angiogenic ability was detected compared to the controls. Finally, we demonstrated that MiR-451 overexpression inhibited the malignant behavior of osteosarcoma by downregulating MIF. These findings suggest that MiR-451 may act as a tumor suppressor in osteosarcoma. MiR-451 inhibited cell proliferation, migration and angiogenesis and promoted apoptosis of human osteosarcoma cells, at least partially, by inhibiting the expression of MIF. MiR-451/MIF may be a novel therapeutic target in treatment of osteosarcoma.

miR-451 inhibits cell growth, migration and angiogenesis in human osteosarcoma via down-regulating IL 6R.

Osteosarcoma (OS) has become one of the most common primary malignant tumors in the children and adolescents with a poor prognosis owing to its high malignant and metastatic potential. Although increasing evidence indicates that miR-451 could inhibit the growth and metastasis of OS, its effect on angiogenesis in OS is still very poor. What is more, the mechanism by which miR-451 affects the OS has not been fully elucidated. In the present study, miR-451 was reduced in human osteosarcoma tissues compared with the adjacent bone tissues, and the introduction of miR-451 dramatically inhibited the growth, migration and angiogenesis in OS. Additionally, it was suggested that IL 6R is a direct target gene of miR-451. Silencing of IL 6R suppressed the growth, migration and angiogenesis of OS, which was consistent with the effect of overexpression of miR-451. In conclusion, our data demonstrate that miR-451 may function as a potential suppressor of tumor growth, migration and angiogenesis in OS via down-regulating IL 6R, suggesting a promising therapeutic avenue for managing OS.

Spatial and temporal changes of subchondral bone proceed to articular cartilage degeneration in rats subjected to knee immobilization.

This study was aimed to investigate the spatial and temporal changes of subchondral bone and its overlying articular cartilage in rats following knee immobilization. A total of 36 male Wistar rats (11-13 months old) were assigned randomly and evenly into 3 groups. For each group, knee joints in 6 rats were immobilized unilaterally for 1, 4, or 8 weeks, respectively, while the remaining rats were allowed free activity and served as external control groups. For each animal, femurs at both sides were dissected after sacrificed. The distal part of femur was examined by micro-CT. Subsequently, femoral condyles were collected for further histological observation and analysis. For articular cartilage, significant changes were observed only at 4 and 8 weeks of immobilization. The thickness of articular cartilage and chondrocytes numbers decreased with time. However, significant changes in subchondral bone were defined by micro-CT following immobilization in a time-dependent manner. Immobilization led to a thinner and more porous subchondral bone plate, as well as a reduction in trabecular thickness and separation with a more rod-like architecture. Changes in subchondral bone occurred earlier than in articular cartilage. More importantly, immobilization-induced changes in subchondral bone may contribute, at least partially, to changes in its overlying articular cartilage.