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Jin T.,Chongqing Medical University | Zhu Y.L.,Kunming General Hospital Chengdu Military Command | Li J.,Kunming General Hospital Chengdu Military Command | Shi J.,Kunming General Hospital Chengdu Military Command | And 3 more authors.
Cellular Physiology and Biochemistry | Year: 2013

Background/Aims: Osteomyelitis is a debilitating infectious disease of the bone which is predominantly caused by Staphylococcus aureus (S. aureus). The purpose of this study was to investigate the role of the S. aureus virulence factors, i.e. protein A (SpA), Panton-Valentine leukocidin (PVL) and coagulase (Coa) on osteomyelitis. Methods: The effect of SpA, PVL and Coa on osteoblasts was studied through the following aspects including osteoblast proliferation, apoptosis, bone formation, bone mineralization and RANK-L expression. S. aureus overexpressing PVL, SpA or Coa was constructed and used to study the role of PVL, SpA and Coa, respectively. S. aureus silencing PVL, SpA or Coa was also constructed and used for reversing verification. Osteoblast proliferation was detected by MTT tetrazolium dye reduction assay. Apoptosis was determined by Annexin V-FITC staining. The levels of pro-caspase 3, cleaved-caspase 3, pro-caspase 9 and cleaved-caspase 9 were detected by western blot. Bone formation markers including collagen I, osteopontin and osteocalcin were detected by real time RT-PCR. Alkaline phosphatase activity was measured by adding p-nitrophenyl phosphate as a phosphatase substrate. Von kossa stain and alizarin red stain were applied for determining phosphate and calcium deposition, respectively. The RANK-L expression was tested by ELISA. Results: PVL, SpA and Coa inhibited osteoblast proliferation, induced osteoblast apotosis, prohibited bone formation and mineralization and upregulated RANK-L expression. Conclusions: PVL, SpA and Coa play a critical role on bone loss and bone destruction of osteomyelitis. Copyright © 2013 S. Karger AG, Basel. Source


Jin T.,Kunming General Hospital Chengdu Military Command | Lu Y.,Shanghai JiaoTong University | He Q.X.,Kunming General Hospital Chengdu Military Command | Wang H.,Kunming General Hospital Chengdu Military Command | And 3 more authors.
Journal of Cellular Biochemistry | Year: 2015

Osteomyelitis is a debilitating infectious disease of the bone which is predominantly caused by Staphylococcus aureus (S. aureus). MicroRNAs (miRNAs) have been shown to play a regulatory role in osteogenesis. In the present study, the expression levels of miRNAs proposed to potentially play a regulatory role in bone formation or differentiation (miR-24, miR-29b, miR-200a, miR-208, miR-322) were analyzed in the whole blood of patients with bacterial osteomyelitis or healthy controls, and in MC3T3-E1 cells infected with S. aureus by qRT-PCR. The expression of miR-24 was significantly down-regulated in osteomyelitis patients and S. aureus-infected MC3T3-E1 cells compared with the healthy controls or untreated control cells. Moreover, our results showed that S. aureus inhibited MC3T3-E1 cell proliferation, induced osteoblast apoptosis and prohibited bone formation and mineralization. We found that overexpression of miR-24 could reduce the effects of S. aureus, while inhibition of miR-24 intensified the effects. We also demonstrated that miR-24 suppressed the expression of chitinase 3-like 1 (CHI3L1) mRNA, thought to mediate multiple signaling pathways, by directly binding to the 3′-untranslated region. J. Cell. Biochem. 116: 2804-2813, 2015. © 2015 Wiley Periodicals, Inc. © 2015 Wiley Periodicals, Inc. Source

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