Ze-Qi-Tang Formula Induces Granulocytic Myeloid-Derived Suppressor Cell Apoptosis via STAT3/S100A9/Bcl-2/Caspase-3 Signaling to Prolong the Survival of Mice with Orthotopic Lung Cancer.

Non-small-cell lung cancer (NSCLC) remains the most common malignancy with the highest morbidity and mortality worldwide. In our previous study, we found that a classic traditional Chinese medicine (TCM) formula Ze-Qi-Tang (ZQT), which has been used in the treatment of respiratory diseases for thousands of years, could directly inhibit the growth of human NSCLC cells via the p53 signaling pathway. In this study, we explored the immunomodulatory functions of ZQT. We found that ZQT significantly prolonged the survival of orthotopic lung cancer model mice by modulating the tumor microenvironment (TME). ZQT remarkably reduced the number of MDSCs (especially G-MDSCs) and inhibited their immunosuppressive activity by inducing apoptosis in these cells via the STAT3/S100A9/Bcl-2/caspase-3 signaling pathway. When G-MDSCs were depleted, the survival promotion effect of ZQT and its inhibitory effect on lung luminescence signal disappeared in tumor-bearing mice. This is the first study to illustrate the immunomodulatory effect of ZQT in NSCLC and the underlying molecular mechanism.

microRNA-1236 promotes chondrocyte apoptosis in osteoarthritis via direct suppression of PIK3R3.

AIMS: Chondrocyte degeneration is the main cause of osteoarthritis (OA) and increased evidence suggests that miRNAs could have vital roles in the pathology of various cartilage illnesses. miR-1236 has been found to contribute to inflammation in diseases such as pneumonia. However, the exact role of miR-1236 in OA is poorly understood. MATERIALS AND METHODS: H&E staining and saffron fixation experiments were employed to determine OA tissues. qRT-PCR and immunohistochemistry were used to detect the expression levels of miR-1236 and PIK3R3. Western blot was performed to detect the expression levels of proteins. Luciferase reporter assays were utilized to investigate the interaction between miR-1236 and PIK3R3. Cell counting assays and AO/EB were used to quantify cell growth and apoptosis. KEY FINDINGS: miR-1236 was up-regulated in OA knee cartilage compared to normal cartilage. Up-regulated expression of miR-1236 suppressed cell proliferation as well as induced apoptosis in chondrocytes. Bioinformatics identified PIK3R3 as a target of miR-1236. Co-transfection with miR-1236 and PIK3R3 could reverse cell apoptosis induced by the miR-1236 mimic. SIGNIFICANCE: These data enhance our understanding on the role of miR-1236 in OA and identifies miR-1236 as a potential biomarker or possible treatment target within OA.

The molecular mechanism of platelet lysate promotes transformation of non-union cells into osteoblasts.

BACKGROUND: Platelet lysate (PL) had a remarkable therapeutic effect on bone repair related diseases, such as delayed fracture healing, femoral head necrosis and meniscal tear. In this study, we investigated the effect of PL on patients with nonunion, cartilage repair and osteonecrosis, and to evaluate the effect of PL on nonunion cells proliferation and the effect of PL on OPG/RANKL signaling pathway in nonunion cell of male rats. To reveal the molecular mechanism of PL for bone healing. METHODS: We used different concentrations of PL to treat nonunion cells, then detected cell proliferation and protein expression levels of osteoprotegerin (OPG), RANKL, osteopontin (OPN), osteocalcin (OCN) and alkaline phosphatase (ALP). RESULTS: The proliferation rate of nonunion cells treated by 5% PL, was significantly higher than that of the control group (P<0.05). Surprisingly, there were no significant difference among the proliferation rates of nonunion cells treated by 8% PL, 10% FBS and the control group (P>0.05). the results of western blot analysis and immunofluorescence analysis showed that PL improved the expression of OPG, OPN, OCN and ALP proteins in nonunion cells, but PL had no effect on the expression of nuclear factor-κB ligand (RANKL) protein. CONCLUSIONS: We found that PL had a remarkable therapeutic effect on bone repair related diseases; 5% PL significantly improved the proliferation rate of the nonunion cells; 10% PL had a significantly positive effect on improving the expression levels of osteogenic related genes.

Identification and characterization of c-raf from orange-spotted grouper (Epinephelus coioides).

C-Raf proto-oncogene serine/threonine kinase is a mitogen-activated protein kinase (MAP) kinase kinase, which can initiate a mitogen-activated protein kinase (MAPK) cascade by phosphorylating the dual-specific MAP kinase kinases (MEK1/2), and in turn activate the extracellular signal-regulated kinases (ERK1/2). To study the function of c-Raf in teleost fish, a c-Raf cDNA sequence from orange-spotted grouper (Epinephelus coioides) was cloned. Ecc-Raf shared 81%-99% amino acid identity with other vertebrate c-Raf molecules, and shared the highest amino acid identity (99%) with Lates calcarifer c-Raf. Genomic structure analysis revealed that grouper c-Raf shared a conserved exon structure with other vertebrates. Tissue distribution showed that Ecc-Raf was mainly transcribed in systemic immune organs. Ecc-Raf was distributed throughout the cytoplasm of transfected GS cells and the overexpression of Ecc-Raf only slightly enhanced the activation of Activator protein 1. The phosphorylation levels of Ecc-Raf can be induced by PMA and H2O2 treatment, in contrast to DMSO or untreated HKLs. Moreover, the phosphorylation level of the Raf-MEK-ERK axis was downregulated after 24 h of SGIV infection. On the other hand, the total level and phosphorylation level of c-Raf significantly increased post C. irritans infection and showed an enhanced level post immunization. The results of this study suggested that the Raf-MEK-ERK cascade was involved in the response to viral or parasitic infections.

miR-590-3p is a novel microRNA which suppresses osteosarcoma progression by targeting SOX9.

Osteosarcoma is the most common primary bone malignancy and arises primarily in the metaphyseal ends of long bones in children and adolescents. m iR-590 has been found to have anti-tumor effects in many other cancers. However, the role of miR-590-3p in osteosarcoma is poorly understood. In this study, we show that miR-590-3p was significantly decreased both in osteosarcoma tissues and cell lines, suggesting a potential role of miR-590-3p in osteosarcoma. Over-expression of miR-590-3p inhibited U2OS cell viability as shown by the CCK-8 assay and clonogenic assay. Ki-67 immunofluorescence staining and cell cycle analysis revealed that up-regulation of miR-590-3p inhibited U2OS cell proliferation. Transfection with miR-590-3p mimics suppressed PCNA, Cyclin D1 and CDK4 expression and increased p53 and p21 expression. In addition, U2OS cells transfected with miR-590-3p mimics exhibited reduced cell invasion and migration, characterized by the wound healing assay and transwell assay. Furthermore, bioinformatics analysis demonstrated that SOX9 was a potential target of miR-590-3p. SOX9 was up-regulated in osteosarcoma tissues. Transfection with miR-590-3p mimics markedly suppressed SOX9 expression both at the mRNA level and protein level. Dual luciferase assay validated the direct binding site of miR-590-3p on SOX9. Exogenous SOX9 expression in U2OS cells at least partially reversed the effects of miR-590-3p in U2OS cells. Enforced SOX9 expression restored cell viability in osteosarcoma cells transfected with miR-590-3p mimics. In addition, over-expression of SOX9 restored decreased cell metastasis properties caused by transfection with miR-590-3p mimics in osteosarcoma cells. In summary, these results indicated that miR-590-3p is an anti-cancer miRNA that can inhibit proliferation and metastasis in osteosarcoma cells. Our findings provide a novel insight into the biological function of miR-590-3p in osteosarcoma and SOX9 may be a potential therapeutic target for osteosarcoma.