Up-regulation of LncRNA UCA1 by TGF-ß promotes doxorubicin resistance in breast cancer cells.

BACKGROUND: Doxorubicin (DOX) resistance remains a major challenge for adriamycin-based treatment of breast cancer (BC). Transforming growth factor ß (TGF-ß) has been reported to contribute to drug resistance. Although the role of long noncoding RNAs (LncRNAs) in cancer progression has been widely studied, its effect on TGF-ß-induced resistance remains limited. This study aimed to investigate the role of LncRNA on the regulation of TGF-ß-induced drug resistance. METHODS: Cell counting kit-8 (CCK-8) and an EdU assay were used to evaluate cell viability and proliferation. The level of LncRNA mRNA expression in BC tissues and cells was examined by quantitative real-time PCR. Changes in epithelial-mesenchymal transition (EMT) and cell apoptosis were quantified by Western blot and immunofluorescence. RESULTS: TGF-ß induced EMT and promoted DOX resistance. LncRNA urothelial carcinoma-associated 1(lncRNA UCA1) associated with TGF-ß was upregulated in BC cells and tissues. LncRNA UCA1 silencing enhanced sensitivity to DOX decreased cellular proliferation and increased apoptosis in BC cells. The effect of TGF-ß on EMT and DOX resistance disappeared following a lncRNA UCA1 knockdown. CONCLUSIONS: These findings suggest that lncRNA-UCA1, a mediator of TGF-ß signaling, could predispose BC patients to EMT and DOX resistance.

Knockdown of miR-182 promotes apoptosis via regulating RIP1 deubiquitination in TNF-α-treated triple-negative breast cancer cells.

Overexpression of microRNA-182 (miR-182) is found in multiple cancers, but the association of miR-182 expression with the sensitivity of triple-negative breast cancer (TNBC) cells to tumor necrosis factor-alpha (TNF-α) remains unknown. In this study, up-regulation of miR-182 was validated in TNBC patients and cell lines. Knockdown of miR-182 was observed to hinder the proliferation of BT-549 cells. More importantly, knockdown of miR-182 significantly promoted the apoptosis induced by TNF-α treatment in BT-549. JC-1 staining and western blot assays revealed that the K63-linked ubiquitin chains on receptor-interacting protein 1 (RIP1) were removed and the outer mitochondrial membrane potential (MMP) and permeability was altered upon combination of TNF-α with anti-miR-182. We then demonstrated that knockdown of miR-182 up-regulated the expression of cylindromatosis (CYLD) deubiquitinase, which promoted the formation of death-inducing signaling complex (DISC) and subsequent caspase-8 activation in TNF-α-treated BT-549 cells. Collectively, the results of the present study improve our understanding of the role of miR-182 in TNBC, knockdown of which facilitates the degradation of ubiquitin chains on RIP1, leading to the caspase-8 activation and apoptosis in TNF-α-treated TNBC cells. This may be valuable for the development of cancer therapy.

Allicin prevents H2O2-induced apoptosis of HUVECs by inhibiting an oxidative stress pathway.

BACKGROUND: Allicin, a primary ingredient of garlic, has been proposed to possess cardioprotective properties, which are commonly mediated by improved endothelial function. METHODS: To investigate the effect and mechanism of allicin on the apoptosis of human umbilical vein endothelial cells (HUVECs), we used Propidium iodide (PI) staining and Annexin V/ PI staining assays to establish a model of oxidative stress apoptosis induced by H2O2. MTT, RT-PCR and western-blot assays were used to detect the effects and mechanism of allicin on the model. RESULTS: PI staining, Annexin V/ PI staining assays and morphological assessment suggest that the cell death induced by 0.5 mM H2O2 is primarily apoptotic. Conversely, allicin reverses the effect of H2O2 on cell death, suggesting a role in protecting HUVECs from apoptosis. We demonstrated that H2O2 activates PARP cleavage, reduces pro-Caspase-3 levels and activates Bax expression; however, allicin inhibits each of these apoptotic signaling indicators. Allicin also reduces the levels of malondialdehyde and increases the levels of superoxide dismutase, nitric oxide release and endothelial nitric oxide synthase mRNA, but has no significant effect on inducible nitric oxide synthase mRNA levels. CONCLUSION: These results demonstrate that allicin has powerful effects in protecting HUVECs from apoptosis and suggest that protection occurs via a mechanism involving the protection from H2O2-mediated oxidative stress.

Atractylenolide II combined with Interferon-γ synergistically ameliorates colorectal cancer progression in vivo and in vitro by blocking the NF-kB p65/PD-L1 pathway.

Purpose: Atractylodes macrocephala Koidz is a widely used classical traditional Chinese herbal medicine, that has shown remarkable efficacy in cancers. Colorectal cancer (CRC) is the most common malignant tumor globally. Interferon (IFN)-γ, a prominent cytokine involved in anti-tumor immunity that has cytostatic, pro-apoptotic, and immune-stimulatory properties for the detection and removal of transformed cells. Atractylenolides-II (AT-II) belongs to the lactone compound that is derived from Atractylodes macrocephala Koidz with anti-cancer activity. However, whether AT-II combined with IFN-γ modulates CRC progression and the underlying mechanisms remain unclear. The present study aimed to elucidate the efficacy and pharmaceutical mechanism of action of AT-II combined with IFN-γ synergistically against CRC by regulating the NF-kB p65/PD-L1 signaling pathway. Methods: HT29 and HCT15 cells were treated with AT-II and IFN-γ alone or in combination and cell viability, migration, and invasion were then analyzed using Cell Counting Kit-8 (CCK-8) and Transwell assays, respectively. Furthermore, the underlying mechanism was investigated through western blot assay. The role of AT-II combined with IFN-γ on tumor growth and lung metastases was estimated in vivo. Finally, the population of lymphocytes in tumor tissues of lung metastatic C57BL/6 mice and the plasma cytokine levels were confirmed by flow cytometry and enzyme-linked immunosorbent assay (ELISA). Results: AT-II or the combination IFN-γ significantly inhibited the growth and migration abilities of CRC cells in vitro and in vivo. The biological mechanisms behind the beneficial effects of AT-II combined with IFN-γ were also measured and inhibition of p38 MAPK, FAK, Wnt/ß-catenin, Smad, and NF-kB p65/PD-L1 pathways was observed. Moreover, AT-II combined with IFN-γ significantly inhibited HCT15 xenograft tumor growth and lung metastases in C57BL/6 mice, which was accompanied by lymphocyte infiltration into the tumor tissues and inflammatory response inactivation. Conclusions: The results showed that the AT-II in combination with IFN-γ could be used as a potential strategy for tumor immunotherapy in CRC. More importantly, the mechanism by which AT-II suppressed CRC progressions was by inhibiting the NF-kB p65/PD-L1 signal pathway.

[Effects of noncoding RNA NRON gene regulation on human umbilical vein endothelial cells functions].

OBJECTIVE: To determine the effects of noncoding repressor of NFAT (NRON) overexpression or silencing on human umbilical vein endothelial cells (HUVECs) functions. METHODS: Stable HUVECs cell lines with NRON overexpression and short hairpin RNA (shRNA) interference were obtained. HUVECs, the empty vector pBABE-cell line and the empty vector pSuper-cell line served as controls. Cell proliferations of these cell lines were tested using MTS method, tube formation capacity and migration function were also examined. RESULTS: MTS experiments evidenced dose-dependent cells proliferations in all cell lines after 48 h culture with fetal bovine serum (HUVECs, r = 0.91;pBABE empty vectors cell-line, r = 0.88;NRON overexpression cell-line, r = 0.89;pSuper empty vectors cell-line, r = 0.95;shRNA infererence cell-line, r = 0.97). Proliferation capacity was lower in NRON overexpressed HUVECs and was higher in NRON silencing HUVECs compared with pBABE empty vectors treated and normal HUVECs (all P < 0.05). Tube formation and migration functions were also reduced in NRON overexpressed HUVECs [(8.33 ± 0.12) roots, (1857 ± 65) cells] and increased in shRNA infererence of NRON treated HUVECs [(36.00 ± 0.51) roots, (6987 ± 50) cells] compared with pBABE empty vectors treated HUVECs [(19.67 ± 1.42) roots, (4411 ± 117) cells], pSuper empty vectors treated HUVECs [(17.33 ± 2.93) roots, (3883 ± 109) cells] and normal HUVECs [(23.33 ± 3.01) roots, (5145 ± 72) cells, all P < 0.05]. CONCLUSION: NRON overexpression could reduce and NRON silencing could increase proliferation, tube formation and migration capacities of HUVECs.

[Effects of warm and tonify kidney-yang herbs on liver mitochondria proteome of kidney-yang deficiency rats].

OBJECTIVE: To study the effects of warm and tonify kidney-yang herbs on the liver mitochondria proteome of the thyroidectomized kidney-yang deficiency rats. METHOD: Twelve rats were divided into normal group, model group and treated group; each group had four rats. The rats of model group and treated group were excised the two side thyroid gland, and the rats of normal group were done the homologous operation, but didn't excise the thyroid gland. After seven days, the rats of model group and treated group appeared the symptoms of kidney-yang deficiency. From the eighth day, the rats of treated group were fed warm and tonify kidney-yang herbs 6.7 g x kg(-1) once daily, and the rats of other two groups were fed the equal normal. All rats of three groups were killed by decollation after six days of treatment, and liver mitochondria proteins were separated. Each liver mitochondria protein sample was labeled with Cy3 or Cy5 randomly, and one Cy3-labeled sample and one Cy5-labeled sample were mixed on the same 2-D gel along with a Cy2-labeled mixture of all samples as an internal standard and run on the same gel. The gels were scanned under different wavelength light after electrophoresis. All images were analyzed by DeCyder 6. 5 software, and the different proteins were identified by mass spectrum. RESULT: The expression of HSP60, catalase, glutathione peroxidase, carbamoylphosphate synthetase I, ATP synthase, lactotransferrin, H(+)-transporting two-sector ATPase, alpha-ETF and calpain 12 were increased in the thyroidectomized kidney-yang deficiency rats, while the expression of oxoisovalerate dehydrogenase, Acyl-CoA dehydrogenase, ornithine aminotransferase, and GTP-binding regulatory protein were decreased. After the kidney-yang deficiency rats were treated with warm and tonify kidney-yang herbs, the expression of HSP60, catalase, glutathione peroxidase, oxoisovalerate dehydrogenase, Acyl-CoA dehydrogenase, ATP synthase, lacto-transferrin, H(+)-transporting two-sector ATPase, alpha-ETF and GTP-binding regulatory protein were increased, and the expression of carbamoyl-phosphate synthetase I and calpain 12 were decreased. CONCLUSION: The warm and tonify kidney-yang herbs perform its therapeutical effect by regulating the metabolism, protecting the stability of mitochondrial membrance and maintaining the signal conduction in the cells.

Danshen protects against early-stage alcoholic liver disease in mice via inducing PPARα activation and subsequent 4-HNE degradation.

Alcoholic liver disease (ALD) is a type of chronic liver disease caused by long-term heavy ethanol consumption. Danshen is one of the most commonly used substances in traditional Chinese medicine and has been widely used for the treatment of various diseases, and most frequently, the ALD. The current study aims to determine the potential beneficial effect of Danshen administration on ALD and to clarify the underlying molecular mechanisms. Danshen administration improved liver pathologies of ALD, attenuated alcohol-induced increment of hepatic 4-Hydroxynonenal (4-HNE) formation, and prevented hepatic Peroxisome proliferators activated receptor alpha (PPARα) suppression in response to chronic alcohol consumption. Cell culture studies revealed that both hepatoprotective effect and increased intracellular 4-HNE clearance instigated by Danshen supplementation are PPARα-dependent. In conclusion, Danshen administration can protect against ALD via inducing PPARα activation and subsequent 4-HNE degradation.

[Role and Significance of T Help Cells 17 in Pathogenesis of Idiopathic Thrombocytopenic Purpura].

OBJECTIVE: To investigate the role and significance of T help cells 17(Th17) in pathogenesis of idiopathic thrombocytopenic purpura (ITP). METHODS: Peripheral blood samples from ITP patients and normal controls were examined for Th17 cell proportion by flow cytometry (FCM). Expression of IL-17, IL-23, IL-6 and TGF-ß1 in hematoplasma was detected by ELISA. The mRNA expression level of IL-17 and RORγt in peripheral blood mononuclear cells (PBMNC) from patients with ITP and normal controls were measured by RT-PCR technique, and expression levels of pSTAT3 and RORγt proteins were analyzed by Western-blot. RESULTS: Th17 cells in peripheral blood from patients with ITP was greatly increased when compared with normal control group (P<0.05). Expressions of IL-17, IL-23, IL-6 and TGF-ß1 in hematoplasma of ITP patients were all significantly higher than those in normal control group (all P<0.01). mRNA expression levels of IL-17 and RORγt in PBMNC from patients with ITP were much higher than those in normal controls (P<0.05). Protein expressions of pSTAT3 and RORγt in PBMNC of ITP patients were greatly increased as compared with those in control (P<0.05). CONCLUSION: Th17 cell subgroup may play a role in incidence and development of ITP, which may participate in the pathogenesis of ITP by increasing Th17 cell proportion and altering the expression level of Th17-related cytokines as well as regulatory and transcriptional factors.

miR-145 suppresses breast cancer cell migration by targeting FSCN-1 and inhibiting epithelial-mesenchymal transition.

MicroRNAs (miRNAs), small non-coding RNAs, regulate fundamental cellular and developmental processes such as cell growth, apoptosis, migration, and invasion. In our present study, we investigated the inhibitory role of miR-145 on breast cancer cell migration as well as its underlying mechanism. Wound healing assay and transwell migration assay showed that ectopic expression of miR-145 significantly inhibited breast cancer cell migration. Bioinformatics analysis revealed that FSCN-1 was a putative target of miR-145. The expression of FSCN-1 varied among four different breast cancer cells, and inversely correlated with miR-145 levels. Moreover, miR-145 mimic transfection enhanced the expression of FSCN-1 in Bcap-37 and HCC-1937 cells. We also found that siRNA- mediated down-regulation of FSCN-1 inhibited cell motility in breast cancer cells. In addition, we found that up-regulation of miR-145 blocked EMT and decreased the expression of MMP-2/9 in breast cancer cells. These results reveal a new link between miR-145, FSCN-1 and EMT in the regulation of breast cancer migration.

Curcumin up-regulates LDL receptor expression via the sterol regulatory element pathway in HepG2 cells.

Plasma low-density lipoprotein-cholesterol (LDL-C) is mainly taken up and cleared by the hepatocellular LDL receptor (LDL-R). LDL-R gene expression is regulated by the sterol regulatory element binding proteins (SREBPs). Previous studies have shown that curcumin reduces plasma LDL-C and has hypolipidemic and anti-atherosclerotic effects. Herein, we investigated the effect of curcumin on LDL-R expression and its molecular mechanism in HepG2 cells. Curcumin increased LDL-R expression (mRNA and protein) and the resultant uptake of DiI-LDL in a dose- and time-dependent manner. Using a GFP reporter system in a transfected HepG2/SRE-GFP cell line, we found that curcumin activated the sterol regulatory element of the LDL-R promoter. In HepG2/Insig2 cells, curcumin reversed the inhibition of LDL-R expression induced by Insig2 overexpression. These data demonstrate that curcumin increases LDL-R protein expression and uptake activity via the SREBPs pathway. These findings contribute to our further understanding of the cholesterol-lowering and anti-atherosclerotic effects of curcumin.