Knockdown of lncRNA MALAT1 Alleviates LPS-Induced Acute Lung Injury via Inhibiting Apoptosis Through the miR-194-5p/FOXP2 Axis.

PURPOSE: We aimed to identify and verify the key genes and lncRNAs associated with acute lung injury (ALI) and explore the pathogenesis of ALI. Research showed that lower expression of the lncRNA metastasis-associated lung carcinoma transcript 1 (MALAT1) alleviates lung injury induced by lipopolysaccharide (LPS). Nevertheless, the mechanisms of MALAT1 on cellular apoptosis remain unclear in LPS-stimulated ALI. We investigated the mechanism of MALAT1 in modulating the apoptosis of LPS-induced human pulmonary alveolar epithelial cells (HPAEpiC). METHODS: Differentially expressed lncRNAs between the ALI samples and normal controls were identified using gene expression profiles. ALI-related genes were determined by the overlap of differentially expressed genes (DEGs), genes correlated with lung, genes correlated with key lncRNAs, and genes sharing significantly high proportions of microRNA targets with MALAT1. Quantitative real-time PCR (qPCR) was applied to detect the expression of MALAT1, microRNA (miR)-194-5p, and forkhead box P2 (FOXP2) mRNA in 1 µg/ml LPS-treated HPAEpiC. MALAT1 knockdown vectors, miR-194-5p inhibitors, and ov-FOXP2 were constructed and used to transfect HPAEpiC. The influence of MALAT1 knockdown on LPS-induced HPAEpiC proliferation and apoptosis via the miR-194-5p/FOXP2 axis was determined using Cell counting kit-8 (CCK-8) assay, flow cytometry, and Western blotting analysis, respectively. The interactions between MALAT1, miR-194-5p, and FOXP2 were verified using dual-luciferase reporter gene assay. RESULTS: We identified a key lncRNA (MALAT1) and three key genes (EYA1, WNT5A, and FOXP2) that are closely correlated with the pathogenesis of ALI. LPS stimulation promoted MALAT1 expression and apoptosis and also inhibited HPAEpiC viability. MALAT1 knockdown significantly improved viability and suppressed the apoptosis of LPS-stimulated HPAEpiC. Moreover, MALAT1 directly targeted miR-194-5p, a downregulated miRNA in LPS-stimulated HPAEpiC, when FOXP2 was overexpressed. MALAT1 knockdown led to the overexpression of miR-194-5p and restrained FOXP2 expression. Furthermore, inhibition of miR-194-5p exerted a rescue effect on MALAT1 knockdown of FOXP2, whereas the overexpression of FOXP2 reversed the effect of MALAT1 knockdown on viability and apoptosis of LPS-stimulated HPAEpiC. CONCLUSION: Our results demonstrated that MALAT1 knockdown alleviated HPAEpiC apoptosis by competitively binding to miR-194-5p and then elevating the inhibitory effect on its target FOXP2. These data provide a novel insight into the role of MALAT1 in the progression of ALI and potential diagnostic and therapeutic strategies for ALI patients.

microRNA-30a inhibits the liver cell proliferation and promotes cell apoptosis through the JAK/STAT signaling pathway by targeting SOCS-1 in rats with sepsis.

Sepsis is a systemic inflammatory response that may be induced by trauma, infection, surgery, and burns. With the aim of discovering novel treatment targets for sepsis, this current study was conducted to investigate the effect and potential mechanism by which microRNA-30a (miR-30a) controls sepsis-induced liver cell proliferation and apoptosis. Rat models of sepsis were established by applying the cecal ligation and puncture (CLP) method to simulate sepsis models. The binding site between miR-30a and suppressor of cytokine signaling protein 1 (SOCS-1) was determined by dual luciferase reporter gene assay. The gain-of-and-loss-of-function experiments were applied to analyze the effects of miR-30a and SOCS-1 on liver cell proliferation and apoptosis of the established sepsis rat models. The expression of miR-30a, SOCS-1, Janus kinase 2 (JAK2), signal transducer and activator of transcription 3 (STAT3), Bcl-2 associated X protein (Bax), B cell lymphoma-2 (Bcl-2), toll-like receptor 4 (TLR4), and high-mobility group box 1 (HMGB1), and the extent of JAK2 and STAT3 phosphorylation were all determined. Sepsis led to an elevation of miR-30a and also a decline of SOCS-1 in the liver cells. SOCS-1 was negatively regulated by miR-30a. Upregulated miR-30a and downregulated SOCS-1 increased the expression of JAK2, STAT3, Bax, TLR4, and HMGB1 as well as the extent of JAK2 and STAT3 phosphorylation whereas impeding the expression of SOCS-1 and Bcl-2. More important, either miR-30a elevation or SOCS-1 silencing suppressed liver cell proliferation and also promoted apoptosis. On the contrary, the inhibition of miR-30a exhibited the opposite effects. Altogether, we come to the conclusion that miR-30a inhibited the liver cell proliferation and promoted cell apoptosis by targeting and negatively regulating SOCS-1 via the JAK/STAT signaling pathway in rats with sepsis.

miR-182-5p Promotes Growth in Oral Squamous Cell Carcinoma by Inhibiting CAMK2N1.

BACKGROUND/AIMS: Emerging evidence suggests that the propagation of oral squamous cell carcinoma (OSCC) is influenced by the abnormal expression of microRNAs (miRNAs). This study aimed to characterize the involvement of miR-182-5p in OSCC by targeting the calcium/ calmodulin-dependent protein kinase II inhibitor CAMK2N1. METHODS: miR-182-5p expression was quantified in OSCC tissues and cell lines with reverse transcription polymerase chain reaction (RT-PCR). Cell colony formation, Cell Counting Kit-8 (CCK-8), Ki-67, and nude mouse xenograft assays were used to characterize the role of miR-182-5p in the proliferation of OSCC. A miR-182-5p target gene was identified with western blotting, RT-PCR, and luciferase activity assays. OSCC patient survival based on CAMK2N1 expression was also analyzed. RESULTS: miR-182-5p was up-regulated in in vitro cell lines and in vivo clinical OSCC samples. CCK-8, colony formation, and Ki-67 assays revealed that miR-182-5p promoted the growth and proliferation of OSCC cells. miR-182-5p directly targeted CAMK2N1, as evidenced by luciferase assays and target prediction algorithms. CAMK2N1 operated as a tumor suppressor gene in patients with OSCC. Down-regulating miR-182-5p expression in the CAL-27 cell line restored CAMK2N1-mediated OSCC cell proliferation. miR-182-5p expression inhibited the activation of AKT, ERK1/2, and NF-κB. Mice injected with CAL-27 cells transfected with miR-182-5p-inhibitor demonstrated a significant increase in tumor size and weight and increased CAMK2N1 mRNA and protein expression compared with the miR-negative control group. CONCLUSION: The miR-182-5p-CAMK2N1 pathway can be potentially targeted to regulate the proliferation of OSCC cells.

Hemin inhibits NLRP3 inflammasome activation in sepsis-induced acute lung injury, involving heme oxygenase-1.

NLRP3 inflammasome activation contributes to acute lung injury (ALI), accelerating caspase-1 maturation, and resulting in IL-1ß and IL-18 over-production. Heme oxygenase-1 (HO-1) plays a protective role in ALI. This study investigated the effect of hemin (a potent HO-1 inducer) on NLRP3 inflammasome in sepsis-induced ALI. The sepsis model of cecal ligation and puncture (CLP) was used in C57BL6 mice. In vivo induction and suppression of HO-1 were performed by pretreatment with hemin and zinc protoporphyrin IX (ZnPP, a HO-1 competitive inhibitor) respectively. CLP triggered significant pulmonary damage, neutrophil infiltration, increased levels of IL-1ß and IL-18, and edema formation in the lung. Hemin pretreatment exerted inhibitory effect on lung injury and attenuated IL-1ß and IL-18 secretion in serum and lung tissue. In lung tissues, hemin down-regulated mRNA and protein levels of NLRP3, ASC and caspase-1. Moreover, hemin reduced malondialdehyde and reactive oxygen species production, and inhibited NF-κB and NLRP3 inflammasome activity. Meanwhile, hemin significantly increased HO-1 mRNA and protein expression and HO-1 enzymatic activity. In contrast, no significant differences were observed between the CLP and ZnPP groups. Our study suggests that hemin-inhibited NLRP3 inflammasome activation involved HO-1, reducing IL-1ß and IL-18 secretion and limiting the inflammatory response.