Cartilage degeneration is a basic pathological feature of osteoarthritis (OA), and there is growing evidence that it is associated with inflammation. ACY-1215, a selective HDAC6 inhibitor, has been reported to have anti-inflammatory effects. Here, we investigated the anti-inflammatory and chondroprotective effects of ACY-1215 in IL-1 beta-stimulated human primary chondrocytes and C28/I2 cells. The results suggested that ACY-1215 can markedly suppress the expression of inflammatory factors, including IL-1 beta and IL-6 in human primary chondrocytes and C28/I2 cells. Furthermore, ACY-1215 exerts potent chondroprotection through the amelioration of cartilage degradation by inhibiting the expression of matrix-degrading proteases, including MMP-1 and MMP-13 in chondrocytes. These effects may be related to ACY-1215 induced down-regulation of NF-kappa B and STAT3 pathways in OA chondrocytes. Taken together, our results show that ACY-1215 may be a potential and promising therapeutic drug for the management of OA.
Thyroid cancer (TC) is a familiar cancer, which accounts for approximately 1% of the malignant tumors of all cancers worldwide. Recently, icariin (ICA) has been reported to play an anti-tumor role in different cancers. The study aimed to investigate the effect of ICA on TC cells to uncover the regulatory mechanism. The different concentrations of ICA were stimulated SW579 and TPC1 cells, and cell viability, apoptosis, migration, invasion and main factors of these processes were detected by CCK-8, flow cytometry, Transwell and western blot. The expression of miR-625-3p in TC tissues or in ICA-treated cells was examined by qRT-PCR. MiR-625-3p mimic and the negative control were transfected into SW579 and TPC1 cells to investigate the effect of miR-625-3p on TC. Finally, the signaling pathways of PI3K/AKT and MEK/ERK were examined by western blot. ICA significantly suppressed cell viability in a dose-dependent manner and induced apoptosis by regulating Bcl-2, Bax and cleaced-Caspase-3/-9 expression in SW579 and TPC1 cells (p < 0.001). Moreover, ICA inhibited cell migration and invasion by down-regulating MMP-9 and Vimentin in SW579 and TPC1 cells (p < 0.01 or p < 0.001). The expression level of miR-625-3p was decreased by ICA, and miR-625-3p overexpression reversed the anti-tumor effect of ICA on SW579 and TPC1 cells (p < 0.05, p < 0.01 or p < 0.001). Furthermore, ICA inactivated PI3K/AKT and MEK/ERK signaling pathways by mediating miR-625-3p in SW579 and TPC1 cells. ICA exerted antitumor effect by inhibiting cell proliferation, migration and invasion by down-regulating miR-625-3p in TC cells.
Zerumbone has been reported to maintain the anti-cancer effects in many malignant cells. However, the effect and mechanism of Zerumbone on esophageal squamous cell carcinomas (ESCC) is rarely investigated. Here we report the inhibitory effect of Zerumbone (hereinafter referred to as ZER) on ESCC migration and the underlying molecular mechanism. ZER could inhibit the migration of human esophageal squamous cancer KYSE-30 cells and KYSE-150 cells. ZER induced Rac1 protein down-regulation in a dose-and time-dependent manner. The reduction of Rac1 protein was crucial for ZER-induced inhibition of cell migration, as Rac1 knockdown could enhance ZER-induced inhibition of cell migration. We further demonstrated that the decrease of Rac1 after ZER treatment is via proteasome-dependent degradation pathway, and ZER treatment drastically enhanced ubiquitination of Rac1, which finally caused Rac1 degradation. Collectively, our results indicated that ZER inhibits cell migration by suppressing Rac1 expression. This suppresion is achieved through promoting Rac1 ubiquitination and degradation. Thus, the study raises the possibility of ZER as a potential drug for ESCC due to its ability to inhibit cell migration.
Background: Acute respiratory distress syndrome is associated with a mortality of 45%. The authors investigated the possible mechanisms and effect of vascular endothelial growth factor on alveolar epithelial barrier permeability in acute respiratory distress syndrome mice model. Methods: Eighty Male BALB/c mice were randomly assigned to four group: PBS group, LPS group, sFlt group, or LPS+ sFlt group. The levels of vascular endothelial growth factor and total protein in bronchoalveolar lavage fluid were compared, together with lung injury score and the histopathology of alveolar epithelial barrier. The expressions of vascular endothelial growth factor and tight junction proteins mRNA in lung tissue were also studied. Results: Lipopolysaccharide (LPS) inhaling was accompanied with increasing lung vascular endothelial growth factor (VEGF) expression. Anti-VEGF with soluble fms-like tyrosine kinase-1 (sFlt-1) attenuated the lung injury effectively. Conclusions: Our data indicate that anti-vascular endothelial growth factor with soluble fms-like tyrosine kinase1 could maintain the normal structure and function of respiratory membrane in acute respiratory distress syndrome mice model and might be a suitable therapeutic tool for the treatment of acute respiratory distress syndrome.
Macrophages' function play a vital role in the progression of atherosclerosis (AS), and miRNAs can modulate inflammatory cytokine secretion, lipid uptake and apoptosis of macrophages. miR-152 is down-regulated in the serum samples of AS patients and inhibits the migration of human umbilical vein endothelial cell, suggesting that miR-152 exerts a role in the atherogenesis. Nevertheless, the function of miR-152 in the inflammatory reaction of macrophages remains unexplored. Besides, bioinformatics shows that KLF5 is a direct target of miR-152. As a result, the objective of this study is to investigate the effects and mechanism of miR-152/KLF5 in the inflammatory reaction of macrophages. ApoE knockdown mouse (ApoE(-/-)) fed with high fat diet (HFD) was used as animal AS models. Ox-LDL treated RAW264.7 cell was used as cell model. Results showed that miR-152 expression was reduced, while KLF5 expression was elevated in the aortic tissues of AS mice, as compared with that of the control mice. Up-regulation of miR-152 significantly reduced the elevated expression of IL-1, IL-6 and TNF-alpha mediated by ox-LDL in the cultural supernatant of RAW264.7 cells and reduced beta-catenin expression, whereas these effects were all neutralized when KLF5 was up-regulated in the base of miR-152 up-regulation. In conclusion, this study illustrates that miR-152 alleviates the pathogenesis of AS through inhibiting inflammatory responses by targeting KLF5, in which beta-catenin might involves in. Our study provides a possibility of consideration of miR-152/KLF5 as a target for AS treatment.
Objective: Compound sophorae decoction, a Chinese medicinal formulae composed of six Chinese herbs, is effective for the clinical treatment of ulcerative colitis (UC). Some of its effective monomers had been proven to have suppressive effect on UC models. The aim of this study is to further explore the mechanism whether compound sophorae decoction ameliorates dextran sodium sulfate (DSS)-induced mice colitis by regulating the balance between T helper (Th) 17 and regulatory T (Treg) cells. Methods: Experimental model of UC, established by drinking water with DSS, was treated with compound sophorae decoction and mesalazine. The stool, activity, body weight of the mice, colon length and colon histopathology were observed to evaluate severity of colitis. The concentration of cytokines in colonic tissues were detected by ELISA. The expression of phosphorylated nuclear factor-kappaB (NF-kappa B) p65, STAT3 and phosphorylated STAT3 in colonic tissues were determined by western blotting and immunohistochemistry. The percentage of Th17 and Treg cells in spleen and mesenteric lymph nodes (MLNs) were detected by flow cytometry. The levels of transcription factor ROR-gamma t and FOXP3 in colon tissues were detected by qRT-PCR and immunohistochemistry. Results: The aqueous extract of compound sophorae decoction was able to improve the symptoms and pathological damage of mice. The body weight of mice were increased and DAI were significantly decreased; ulcers were slighter than DSS group. The administration of compound sophorae decoction reduced the level of inflammatory factors interleukin (IL)-1 beta, tumor necrosis factor (TNF)-alpha and phospho-NF-kappa B p65, and also decreased the proportions of Th17 cells in spleen and MLNs and the expression of ROR-gamma t, IL-17A, STAT3, IL-6 in colonic tissues; while the percentage of Treg cells in spleen and MLNs and the expression of FOXP3, transforming growth factor (TGF)-beta 1, IL-10 in colonic tissues were upregulated. Conclusion: Overall, this study suggested that compound sophorae decoction significantly improves the symptoms and the pathological damage of mice with colitis and influences the immune function by regulating Th17/Treg cell balance in DSS-induced colitis in mice.
Triptolide (TP) has an anti-proteinuric effect and is used for the treatment of podocytopathies. TP has also been shown to act directly on immortalized podocytes in culture to protect them from injury. In the present study, we examined the effect of TP on healthy podocytes both in vitro and in vivo to better understand the action of TP on podocytes. We found that treatment of TP at 10 ng/ml, a concentration that is routinely used for podocyte protection, was sufficient to activate pro-apoptotic signaling of MAPK p38, p53 and BAX and induced apoptosis in cultured podocytes; and higher concentrations of TP exacerbated the p38, p53 and BAX activations and apoptosis. Moreover, TP severely downregulated the genes that are essential for podocyte structure and function. Interestingly, in contrast with other agents TP-induced podocyte injury was not prevented by glucocorticoids. In vivo, high-dose TP treatment for prolonged time did not cause podocyte injury, essential genes downregulation, and proteinuria in mice. TP was also not toxic to the podocytes with isolated glomeruli ex vivo. In summary, TP is toxic to immortalized podocytes in culture but not to the podocytes in animals or isolated glomeruli ex vivo. Our study suggests that immortalized podocytes might have genetically evolved to become sensitive to TP toxicity and thus caution should be taken in interpreting data from immortalized podocytes. Nevertheless, in vivo TP could be as safe as glucocorticoids in treating podocytopathies. Finally, TP may be used as a unique in vitro model for studying steroid-resistant podocytopathies.
Background: The anti-tumor properties of Alpinia oxyphylla Miguel (A. oxyphylla) extracts and their petroleum ether (PE) fractions have long attracted scientific attention. These extracts' anti-tumor activity and mechanisms in vivo are still unclear. This study was designed to investigate the anti-tumor activity and the underlying mechanism of PE's effect on hepatocellular carcinoma (HCC) in vitro and in vivo. Materials and method: The anti-tumor activity of PE was evaluated by MTT assay and xenograft study. Mechanistic studies of PE were analyzed by Hoechst 33342 staining, Annexin V-FITC/PI double-staining assay, immunohistochemical staining and western blot assay. The toxicity of the PE treatment was verified by the levels of liver and kidney function in nude mice and the H&E staining of their liver and kidney tissues. Result: PE significantly inhibited the growth of HepG2, BEL-7402, SMMC-7721 and Hep3B cells in a concentration- and time-dependent manner. Specifically, PE inhibited the growth of Hep3B cells by inducing apoptosis. PE treatment at the doses of 0.25, 0.5 and 1 g/kg for 21 days caused a respective 35.7 percent, 49.3 percent and 58.8 percent inhibition of the tumor volume, and a 14.8 percent, 40.2 percent and 55.6 percent decrease in the tumor weight, respectively, as compared with the vehicle group in tumor-loaded mice in vivo. PE promoted the release of cytochrome c from mitochondria to cytosol in a concentration-dependent manner. The expression levels of BAX (p < 0.01), cleaved caspase-9 (p < 0.01) and cleaved caspase-3 (p < 0.05) were increased significantly in the PE-treated group at the dose of 1 g/kg; the expression level of BAX (p < 0.05) was increased significantly in the PE-treated group at the dose of 0.5 g/kg, and the expression level of Bcl-2 (p < 0.01) was decreased significantly in the PE-treated group in a concentration-dependent manner. Apoptosis was induced by PE through up-regulating the expression of PTEN, down-regulating the expression of PI3K and inhibiting the phosphorylation of Akt. The liver and kidney function of the plasma and the morphology of the liver and kidney were normal in each group. Conclusion: These findings suggested that PE exhibited anti-cancer efficacy on Hep3B cell in vitro and in vivo. The induction of apoptosis might be one mechanism that underlies PE's ability to combat cancer by inhibiting the PI3K/Akt pathway. PE has no obvious toxicity in vivo when it exerts anti-tumor effects and has the potential to develop into an alternative anti-cancer drug for HCC treatment.
Purpose: We intended to evaluate expression and mechanisms of human microRNA 1270 (hsa-miR-1270) in papillary thyroid cancer (PTC). Methods: In PTC cell lines and human PTC tumors, hsa-miR-1270 expressions were evaluated by qRT-PCR. HsamiR-1270 was downregulated in TPC1 and K1 cells via lentiviral transduction. Its effects on PTC cancer cell proliferation, migration and in vivo transplantation were assessed, respectively. Potential targeting of hsa-miR-1270 on downstream gene, Suppressor Of Cancer Cell Invasion (SCAI), was assessed. In hsa-miR-1270-downregulated PTC cells, SCAI was further downregulated to examine its associating role in hsa-miR-1270-mediated regulation on cancer cell proliferation and migration. Results: Hsa-miR-1270 was aberrantly upregulated in PTC cell lines and human PTC tumors. In TPC1 and K1 cells, lentivirus-mediated hsa-miR-1270 downregulation suppressed cancer cell proliferation, migration and in vivo transplantation. Hsa-miR-1270 binds SCAI and inversely regulated SCAI gene expression in PTC cells. SCAI downregulation removed the suppressing effects of hsa-miR-1270 downregulation in human PTC cells. Conclusion: Hsa-miR-1270 downregulation may suppress human PTC cell development both in vitro and in vivo. The regulatory mechanism of hsa-miR-1270 in PTC may be primarily associated with SCAI.
Background: Higenamine (HG) is an active compound derived from Aconiti root with a cardiotonic effect. It has been approved by the Chinese SFDA for clinical trials due to its effect as a potent inotropic and chronotropic agent in the heart. However, the direct mode of action of HG on cardiac electrophysiology is unclear. Methods: The experiments were performed at both cell levels and the isolated organ. The major cardiac ion currents and the action potential duration (APD) were measured using patch-clamps in single guinea-pig left ventricular myocytes. ECG was recorded in isolated guinea pig hearts. Results: In the left ventricular myocytes, HG increased ICa-L and I-Ks in concentration- and voltage-dependent manners in the left ventricular myocytes. It potentiated the ICa-L and I-Ks, simultaneously for synchronization. The EC50 values were 0.27 mu M and 0.64 mu M for the ICa-L and I(Ks )respectively. HG (0.1 mu M, 0.5 mu M and 1 mu M) had no effect on the I-Kr and I-Na . HG slightly prolonged APD at lower concentrations, and shortened the APD at higher concentrations. HG can induce the delayed after depolarization (DAD), which showed some pro-arrhythmic effect. In the isolated perfused heart, HG increased the heart rate via an action on the sinoatrial node cells, but did not induce cardiac arrhythmias, even at high concentrations. The EC(50 )value for the sinoatrial node that controls the heart rate was 0.13 mu M. The sinoatrial node cells appeared to be more sensitive than ventricular myocytes to HG. The effects of HG on ventricular cells and sinoatrial node cells were both mediated through stimulation of beta 1-AR. Conclusion: We show for the first time that HG produced a predominant action on the sinoatrial node. HG appears to control the cardiac electrophysiology through its predominant effect on the sinoarial node cells, without induction of the ectopic activity that causes cardiac arrhythmias. Thus, HG might be useful for the treatment of bradycardia.
MicroRNA (miRNA) is known to be involved in regulating the proliferation, migration and apoptosis of cancer cells in osteosarcoma. In this study, We aim to explore the expression of hsa-let-7 g and its role in pathogenesis of osteosarcoma. By analyzing clinical data. We found high expression of hsa-let-7 g in patients with osteosarcoma. The patients with higher expression of hsa-let-7 g showed poorer prognosis and lower survival rate. After downregulation of hsa-let-7 g in cell model and animal model, we found that with downregulation of hsa-let-7 g, the proliferation of osteosarcoma cells was significantly reduced, the level of migration and invasion was downregulated, the cell cycle was inhibited, and cell apoptosis was increased. Through Dual Luciferase Reporter, immunohistochemistry, western blot and other experiments, it was found that hsa-let-7 g down-regulated HOXB1 gene and activated NF-kB pathway to promote the development of osteosarcoma. In conclusion, hsa-let7 g is highly expressed in osteosarcoma tissues, and high expression of hsa-let-7 g can promote the occurrence of osteosarcoma by down-regulating HOXB1 and activating NF-kB pathway.
Objective: Chemoresistance has been a major problem in cancer chemotherapy. The present study aimed to investigate the effect of Rosmarinic acid (RA) on chemoresistance to 5-Fu and its molecular mechanism in gastric carcinoma. Methods: CCK8 cell proliferation and apoptosis assay were used to evaluate the effect of RA on chemoresistance to 5-Fu in GC cells. RNA microarray was used to identify miRNA involved. Expression level of miRNA in GC cells was determined by RT-PCR. Down- or up-regulating of miRNA in the GC cells was performed by transfection of RNA interference or expression vectors in the GC cells. Double luciferase reporter assay was used to verify miRNA target genes. Expression of P-glycoprotein and Bax was analyzed with Western blot. Results: RA treated SGC7901/5-Fu cells showed significant increased chemosensitivity to 5-Fu. The IC50 of 5-Fu was significantly reduced in RA treated SGC7901/5-Fu cells (70.43 +/- 1.06 mu g/mL) compared to untreated SGC7901/5-Fu cells (208.6 +/- 1.09 mu g/mL) (P < 0.05). Apoptosis rate was significantly increased in RA + 5-Fu treated SGC7901/5-Fu cells compared to 5-FU treatment alone (P < 0.01). Two miRNAs, namely miR-642a-3p and miR-6785-5p, were identified to be involved in the chemo-sensitizing effect of RA in the SGC7901/5-Fu cells. RA treated SGC7901/5-Fu cells showed reduced expression levels of miR-642a-3p and miR-6785-5p compared to untreated SGC7901/5-Fu cells (P < 0.05). Down- or up-regulation of miR-6785-5p increased or reduced chemosensitivity of gastric carcinoma cells to 5-Fu, respectively. RA treated SGC7901/5-Fu and the SGC7901/5-Fu-Si cells showed significantly increased FOXO4 expression (P < 0.01). Double luciferase reporter assay confirmed miR-6785-5p directly targets FOXO4 to regulate its expression. RA significantly reduced P-gp expression and increased Bax expression in SGC7901/5-Fu and the SGC7901/5-Fu-Si cells (P < 0.05). Conclusion: RA enhances chemosensitivity of resistant gastric carcinoma SGC7901 cells to 5-Fu by down-regulating miR-6785-5p and miR-642a-3p and increasing FOXO4 expression. These study suggest the potential for RA as a multidrug resistance-reversing agent in GC.
Cisplatin, as an effective chemotherapeutic agent, is widely used to treat verious types of cancers. Nephrotoxicity induced by cisplatin seriously limits its clinical application. Icariin, a major and remarkable flavonoid isolated from Epimedium koreanum, has been reported to exert anti-oxidative stress and anti-inflammation actions. The purpose of this study is to explore the protective effect and possible mechanism of icariin on cisplatin-induced nephrotoxicity on HEK-293 cells. In this study, icariin pretreatment for 24 h significantly ameliorated cisplatin-induced oxidative stress by reducing levels of malondialdehyde (MDA) and reactive oxygen species (ROS), while increasing level of glutathione (GSH) in HEK-293 cells. Furthermore, icariin pretreatment reduced NF-kappa B phosphorylation and nuclear translocation in HEK-293 cells followed by decreased secretion of IL-1 beta, TNF-alpha, and iNOS, suggesting a suppression of inflammatory response. Moreover, icariin pretreatment significantly reduced cellular apoptosis via reduced levels of Bax, cleaved caspase-3/9, and increased anti-apoptotic protein Bcl-2 in the cells. Importantly, LY294002, a specific PI3K inhibitor, abrogated the anti-apoptosis effect of icariin, implicating the involvement of PI3K/Akt pathway. In summary, icariin prevents cisplatin-induced HEK-293 cell injury by inhibiting oxidative stress, inflammatory response, and cellular apoptosis partly via regulating NF-kappa B and PI3K/Akt signaling pathways. Icariin may serve as a potential therapeutic target against cisplatin-induced nephrotoxicity.
EndMT plays an important role in the relationship between endothelial dysfunction and atherosclerosis. This work will elucidate the biofunction induced by miR-449a and lipid rafts in EndMT and development of atherosclerosis. The differential miRNA expression between atherosclerotic plaques and normal arteries were analyzed. The luciferase activities of AdipoR2 3' UTR treated with miR-449a were determined. ECs were dealt with miR-449a mimics or inhibitors, then cell proliferation and migration were assessed. Moreover, the expression of AdipoR2 and mesenchymal cell markers were analyzed. The influences of lipid rafts related to reciprocity between E-cadherin and AdipoR2 on TNF-a-induced damage in ECs were investigated. ApoE KO diabetic mice were used to explore the potential roles of miR-449a on atherosclerosis. Our results indicated that compared with normal arteries, 17 miRNAs were upregulated and 3 miRNAs were down-regulated in atherosclerotic plaques. The relative expression of miR-449a in plaques was significantly higher than that in normal arteries. MiR-449a suppressed AdipoR2 expression, additionally its interaction protein E-cadherin in ECs. MiR-449a enhanced expression of mesenchymal cell markers, induced cell proliferation and migration of ECs, regulated the interaction between E-cadherin and AdipoR2 interceded by lipid rafts. The miR-449a antagomir could protect against the development process of atherosclerosis in ApoE KO diabetic mice. In conclusion, miR-449a targeted to AdipoR2, and was a crucial mediator of EndMT and atherosclerosis in ECs through regulating E-cadherin bindability with AdipoR2 in lipid rafts. These results suggested that aim to lipid rafts and miR-449a in chronic EC inflammation response, was a feasible therapy strategy for atherosclerosis.
Background: The activation of microglia plays a crucial role in neuroinflammation. Previous studies have shown that cerebral dopamine neurotrophic factor (CDNF) has a protective effect on neuroinflammation, but the mechanisms involved have not been fully studied. AKT is a serine-threonine protein kinase widely expressed in mammals through which the downstream pathway FoxO1/mTOR is closely related to cell inflammation, apoptosis, metabolism, etc. Therefore, we examined whether CDNF regulates neuroinflammation through this pathway. Methods: After pretreatment with CDNF and LPS, microglial cells were detected by laser confocal microscopy, coimmunoprecipitation and immunofluorescence to observe whether CDNF was colocalized with AKT. The expression of AKT and its downstream FoxO1/mTOR were determined by Western blot. The effect of CDNF on inflammatory cytokines was detected by ELISA, and the mRNA levels of AKT and FoxO1/mTOR were detected by qRT-PCR. Results: Laser confocal and coimmunoprecipitation experiments significantly reveal the occurrence of interactions between AKT and CDNF in microglia. Western blot results show that CDNF incubation suppressed the activation of AKT/FoxO1/mTOR signaling. Moreover, CDNF clearly decreased the expression of inflammatory cytokines. In qRT-PCR, the expression of mRNA in AKT and its downstream FoxO1/mTOR gradually decreased due to CDNF intervention. Conclusions: CDNF combined with AKT and regulated the downstream pathway FoxO1/mTOR in microglia, eventually suppressing the secretion of inflammatory factors. Therefore, CDNF might play a protective role in the neuroinflammation of microglia via AKT/FoxO1/mTOR signaling.