LINC01485 contributes to colorectal cancer progression by targeting miR-383-5p/KRT80 axis.

Long non-coding RNAs (lncRNAs) are important in tumorigenesis and the development of multiple malignant human tumors, including colorectal cancer (CRC). We aimed to determine the regulatory mechanism of LINC01485 and its biological function in CRC. We estimated the expression of miR-383-5p, KRT80, and LINC01485 in CRC cells and tissues using quantitative reverse transcription polymerase chain reaction (qRT-PCR) and western blotting. The results were confirmed using RNA immunoprecipitation (RIP) and dual-luciferase assays. Binding relationships among miR-383-5p, LINC01485, and KRT80 were assessed. We explored the molecular mechanisms and functions of the LINC01485/miR-383-5p/KRT80 axis using CCK-8 and colony formation assays. Expression of the apoptotic markers Bcl-2 and Bax was quantified by western blotting, and the effects of LINC01485 on tumor development in vivo were investigated using xenograft tumors. Both LINC01485 and KRT80 were upregulated, whereas miR-383-5p was downregulated in CRC cells and tissues. Knockdown of LINC01485 attenuated CRC cell growth and xenograft tumor formation in vivo, whereas LINC01485 enhanced the proliferative capacity of CRC cells but inhibited apoptosis by sponging miR-383-5p to increase KRT80 expression in CRC cells. The regulatory molecular mechanism of the LINC01485/miR-383-5p/KRT80 axis plays a crucial role in CRC progression. Our findings highlight novel pathways and promising biomarkers for diagnostic and therapeutic application to patients with CRC.

Circular RNA SMARCA5 inhibits cholangiocarcinoma via microRNA-95-3p/tumor necrosis factor receptor associated factor 3 axis.

Enhancing research indicatedthat circular RNA (circRNA) acted a critical part in cholangiocarcinoma (CHOL) development. This research aims to discover the role of circRNA SWI/SNF-related, matrix-associated, actin-dependent regulator of chromatin, subfamily a, member 5 (SMARCA5) in CHOL bio-progression, which has been proved to be downregulated in CHOL tissues. In this study, quantitative reverse transcription polymerase chain reaction was used to reveal the level and linkage of circRNA SMARCA5, miRNA-95-3p and TNF receptor-associated factor 3 gene (TRAF3) in CHOL tissues and cancer cells. The target sites of circRNA SMARCA5 and miRNA-95-3p were forecast by Starbase, and Targetscan was conducted to forecast the potential linkage points of TRAF3 and miRNA-95-3p, and which is affirmed by double luciferase reporter assay. CCK-8 and flow cytometry assay was carried to indicate cell viability. And apoptosis-related protein was counted by caspase3 activity and Western blot assay. CircRNA SMARCA5 was downregulated in CHOL cell lines and cancer samples. Besides, over-expression of SMARCA5 inhibited cell growth and promoted apoptotic rate. Dual-luciferase reporter assays presented that miRNA-95-3p could link with circRNA SMARCA5. Moreover, miRNA-95-3p was discovered highly expressed in CHOL. Interference of miRNA-95-3p repressed cell proliferation and raised the apoptosis. Importantly, TRAF3 was validated to be a downstream of miRNA-95-3p. Strengthen of miRNA-95-3p reversed the inhibitory impact of circRNA SMARCA5-plasmid transfection, and the results of miRNA-95-3p inhibitor were reversed by si-TRAF3. CircRNA SMARCA5 is involved in CHOL development by interosculating miRNA-95-3p/TRAF3 axis and may become a novel approach for treating CHOL.

miR-542-3p inhibits the growth and invasion of colorectal cancer cells through targeted regulation of cortactin.

Colorectal cancer is one of the most common malignancies. Previous studies have reported that cortactin (CTTN) is often overexpressed in tumors and is associated with metastasis and poor prognosis of patients. The abnormal expression of microRNAs (miRNAs or miRs) is closely related to the development and progression of various types of cancer, including colorectal cancer. However, little is known about the miRNAs targeting cortactin. In the present study, prediction using biological software revealed that cortactin has binding sites for miR-542-3p. Transfection with miR-542-3p mimic demonstrated that miR­542-3p reduced the expression of cortactin in colorectal cancer cells. Dual luciferase reporter assays further demonstrated that miR-542-3p regulated cortactin in a targeted manner and that miR-542-3p expression was significantly downregulated in colorectal cancer cells. A cell proliferation assay and Transwell migration assay were undertaken: we noted that miR­542-3p inhibited the proliferation and invasion of colorectal cancer cells while promoting their apoptosis. By contrast, cortactin acted antagonistically. When co-transfected with miR-542-3p mimic and CTTN overexpression vector, the inhibitory effect of miR-542-3p was blocked. This indicates that miR-542-3p regulates CTTN in a targeted manner to modulate the growth and invasion of colorectal cancer cells. The present study thus provides new targets for the prevention and treatment of colorectal cancer.

Anti-tumor effect of LTA combined with 5-FU on H22 tumor bearing mice.

OBJECTIVE: To study the effect of lipoteichoic acid (LTA) and 5-FU on the expression of caspase-3, EGFR, TGF-α proteins of tumor tissue of H22 cancer bearing mice and its anti-tumor mechanism. METHODS: A total of 40 SPF grade Kunming mice were selected to establish H22 liver cancer model, and then the mice were divided into 4 groups at random with ten mice in each group. Group A was given saline lavage treatment, Group B was treated with 5-FU by intraperitoneal injection, Group C was treated with LTA by lump body injection; Group D was treated with LTA by lump body injection and 5-FU by intraperitoneal injection. Two weeks after the treatment, the mice in each group were executed and the tumor tissue was stripping and weighted, and the tumor growth inhibition ratio was calculated. Then the tumor tissue was processed for conventional embedding, sectioned to observe the expression of caspase-3, EGFR, TGF-α by immunohistochemical staining method. RESULTS: The tumor inhibitory rate o f Group D was significantly higher than Groups B and C (P < 0.05); B, the tumor inhibitory rate o f Group B had no statistical difference compared with Group C (P > 0.05). The IDO values of TGF-α, EGFR proteins in Groups B, C, D mice tumor tissue were significantly lower than that in group A (P < 0.05); while IDO value of caspase-3 in Groups B, C, D group mice tumor tissue was significantly higher than that in Group A (P < 0.05). The IDO value of TGF-α, EGFR in Group D mice tumor tissue were significantly lower than that in Groups B and C; While IDO value of aspase-3 in Group D was significantly higher than that in Groups B and C (P < 0.05). CONCLUSIONS: LTA combined with 5-FU can effectively inhibit the tumorigenesis of H22 tumor bearing mice, increase the caspase-3 protein expression, inhibit TGF-α and EGFR protein expression, further promote tumor cell apoptosis and play a synergistic antitumor effect.

Effects of Local Pancreatic Renin-Angiotensin System on the Microcirculation of Rat with Severe Acute Pancreatitis.

Severe acute pancreatitis (SAP) is normally related to multiorgan dysfunction and local complications. Studies have found that local pancreatic renin-angiotensin system (RAS) was significantly upregulated in drug-induced SAP. The present study aimed to investigate the effects of angiotensin II receptors inhibitor valsartan on dual role of RAS in SAP in a rat model and to elucidate the underlying mechanisms. 3.8% sodium taurocholate (1 ml/kg) was injected to the pancreatic capsule in order for pancreatitis induction. Rats in the sham group were injected with normal saline in identical locations. We also investigated the regulation of experimentally induced SAP on local RAS expression in the pancreas through determination of the activities of serum amylase, lipase and myeloperoxidase, histological and biochemical analysis, radioimmunoassay, fluorescence quantitative PCR and Western blot analysis. The results indicated that valsartan could effectively suppress the local RAS to protect against experimental acute pancreatitis through inhibition of microcirculation disturbances and inflammation. The results suggest that pancreatic RAS plays a critical role in the regulation of pancreatic functions and demonstrates application potential as AT1 receptor antagonists. Moreover, other RAS inhibitors could be a new therapeutic target in acute pancreatitis.

Hsa-miR-132 inhibits proliferation of hepatic carcinoma cells by targeting YAP.

MicroRNAs and Yes-associated protein (YAP) play an important role in the occurrence and development of hepatic carcinomas. However, the interaction between microRNAs and YAP was seldom elucidated. In this study, we showed that miR-132 could target YAP gene by using dual-luciferase reporter system. Further quantitative polymerase chain reaction analysis and western blotting showed that miR-132 could significantly reduce the expression of YAP at mRNA and protein levels. Results of annexin V-fluorescein isothiocyanate, 5-ethynyl-2'-deoxyuridine staining and transwell assays showed that miR-132 significantly promoted the cell apoptosis and effectively inhibited the proliferation and invasion of hepatoma cells. These results indicated that miR-132 could inhibit the growth of hepatoma cells by targeting YAP gene and reducing its expression level. Taken together, results from this study would help us to understand the mechanisms for occurrence and development of hepatic carcinoma and provide new targets for diagnosis and treatment of liver cancer.

Lentivirus vectors construction of SiRNA targeting interference GPC3 gene and its biological effects on liver cancer cell lines Huh-7.

OBJECTIVES: To build GPC3 gene short hairpin interference RNA (shRNA) slow virus vector, observe expression of Huh-7 GPC3 gene in human liver cell line proliferation apoptosis and the effect of GPC3 gene influencing on liver cancer cell growth, and provide theoretical basis for gene therapy of liver cancer. METHODS: Hepatocellular carcinoma cell line Huh-7 was transfected by a RNA interference technique. GPC3 gene expression in a variety of liver cancer cell lines was detected by fluorescence quantitative PCR. Targeted GPC3 gene sequences of small interfering RNA (siRNA) PGC-shRNA-GPC3 were restructured. Stable expression cell lines of siRNA were screened and established with the help of liposomes (lipofectamine(TM2000)) as carrier transfection of human liver cell lines. In order to validate siRNA interference efficiency, GPC3 siRNA mRNA expression was detected after transfection by using RT-PCR and Western blot. The absorbance value of the cells of blank group, untransfection group and transfection group, the cell cycle and cell apoptosis were calculated, and effects of GPC3 gene on Huh-7 cell proliferation and apoptosis were observed. RESULTS: In the liver cancer cell lines Huh-7, GPC3 gene showed high expression. PGC-shRNA-GPC3 recombinant plasmid was constructed successfully via sequencing validation. Stable recombinant plasmid transfected into liver cancer cell lines Huh-7 can obviously inhibit GPC3 mRNA expression level. CONCLUSIONS: The targeted GPC3 siRNA can effectively inhibit the expression of GPC3.

Overexpression of GPC3 inhibits hepatocellular carcinoma cell proliferation and invasion through induction of apoptosis.

Glypican­3 (GPC3) is a membrane heparan sulfate proteoglycan involved in cell proliferation, differentiation, adhesion, migration and the development of the majority of mesodermal tissues and organs. GPC3 has been found to be important for the occurrence and development of hepatocellular carcinoma (HCC). Therefore, it may be suitable for use as a novel molecular marker for the diagnosis of primary liver cancer. In the present study, the role of GPC3 in the occurrence and development of HCC was determined. GPC3 recombinant vector was transfected into two HCC cell lines, Huh7 and SK­HEP­1, to upregulate the expression of GPC3 and examine changes in the biological behavior of the cells. Results indicate that overexpression of GPC3 in Huh7 and SK­HEP­1 cells effectively inhibited cell proliferation and cell invasion through induction of apoptosis. However, cotreatment of the cells with insulin­like growth factor 2 (IGF2) and fibroblast growth factor 2 (FGF2) was found by Annexin V­PI flow cytometric analysis to significantly inhibit the apoptotic cell death induced by GPC3 overexpression. These observations indicate that GPC3 may act as a negative regulator of IGF2 and FGF2 pathways. Taken together, these results demonstrate that overexpression of GPC3 inhibits the occurrence and development of HCC.