TUFM-knockdown inhibits the migration and proliferation of gastrointestinal stromal tumor cells.

Gastrointestinal stromal tumors (GISTs) are the most common pathologic type of mesenchymal tumor in the digestive tract. Patients with GIST face the risk of metastasis, postoperative recurrence and imatinib mesylate (IM) resistance. Mitochondrial Tu translation elongation factor (TUFM) is highly expressed in GISTs, and is associated with oncogenesis, progression and prognosis. There is evidence that TUFM is involved in tumor invasion and metastasis. However, the effect of TUFM on GIST-T1 cells and the IM-resistant GIST-IR cell line remains unclear. The present study aimed to evaluate the effects of TUFM on the proliferation, migration and apoptosis of GIST cells in vitro. TUFM short hairpin (sh)RNA expression plasmids were transfected into GIST-T1 and GIST-IR cells by electroporation. The expression levels of enhanced green fluorescent protein were observed by fluorescence microscopy to evaluate the electroporation efficiency. The expression levels of TUFM were detected by western blot analysis and reverse transcription-quantitative PCR. Cell proliferation was assessed by counting cells and using a Cell Counting Kit-8 assay. Cell migration was analyzed using wound healing and Transwell migration assays. Cell cycle distribution and late apoptosis were assessed by flow cytometry. TUFM shRNA expression plasmids were successfully transfected into the GIST cell line by electroporation. The transfection efficiency was >75%, and the TUFM gene silencing efficiency was 73.2±1.4%. TUFM-knockdown decreased the proliferation and migration capacity of GIST-T1 and GIST-IR cells. The proportion of cells in the pre-G1 stage was increased without change in the proportions of cells in the G1, S and G2/M stages after TUFM silencing in GIST-T1 and GIST-IR cells. TUFM may be related to GIST infiltration and metastatic recurrence, suggesting that TUFM may be an effective target for preventing the progression and metastasis of GISTs.

Establishment of a GIST-T1 gastrointestinal stromal tumour cell line resistant to imatinib mesylate.

In the present study, imatinib mesylate (IM) was used to induce resistance in the gastrointestinal stromal tumour (GIST) cell line, GIST-T1, to establish a stable resistant cell line. The growth characteristics and expression profile of the established cell line were compared with those of the parental cell line. Additionally, the resistance mechanism of the gastrointestinal stromal tumours was preliminarily investigated. The GIST-T1 cells were cultured in vitro, and the drug was administered in the logarithmic phase of cell growth using intermittent dosing with increasing concentrations to obtain a drug-resistant cell line by repeated induction. Differences in the biological behaviours of the parental cells and drug-resistant cells were examined, and changes in the expression profiles were compared in the two cell lines. The results showed that the IM-resistant GIST-T1 cell line (GIST-T1 IR) was successfully established. Analysis of the biological behaviours of the two cell lines revealed that the average doubling times of the parental cells and drug-resistant cells were 26.59 and 33.63 h, respectively. The results of a scratch migration assay revealed that the migration ability was enhanced in the GIST-T1 IR cells. The results of CCK-8 detection indicated that the half maximal inhibitory concentration values of the two types of cells were 10.5 and 42.0 µM, respectively, which represented an increase of ~4-fold in the GIST-T1 IR cells. Flow cytometric cell cycle analysis indicated that the numbers of cells in the G0/G1, S and G2 phases increased following the induction treatment. Taken together, an IM-resistant GIST T1 cell line was successfully established, which opens novel avenues for individualized tumour chemotherapy.

MicroRNA hsa-let-7e-5p as a potential prognosis marker for rectal carcinoma with liver metastases.

MicroRNAs (miRNAs) are post-transcriptional regulators of gene expression that target mRNAs for translational repression or cleavage. The present study was conducted to identify differentially expressed miRNAs in primary tumor tissues of rectal carcinoma (RC) that may be associated with heterochrony hepatic metastasis (HHM). Samples were collected exclusively from patients with RC but not colon cancer (CC); Next-generation high-throughput sequencing technology and bioinformatics tools were used to profile and analyze small RNAs and their corresponding targets in primary tumor tissues with HHM (n=2) or without metastases (non-metastatic, NM; n=2). A total of 24 known miRNAs were identified to be differentially expressed (P<0.01; absolute value of log2-fold change ≥1). Hsa-let-7e-5p exhibited the most significant elevation in tissues with HHM (log2-fold change=2.62). By combining online informatics resources and previous mRNA sequencing data, it was identified that 54 validated target genes of let-7e were downregulated in primary tumor tissues with HHM. A number of these target genes have been demonstrated to be directly involved in tumor metastasis (including MYC proto-oncogene, bHLH transcription factor, high-mobility group AT-Hook 2, peptidase inhibitor 3, KIT proto-oncogene receptor tyrosine kinase, Jun proto-oncogene, AP-1 transcription factor subunit and ribonuclease T2), or have physiological associations to immunity (including C-C motif chemokine receptor 4 and cluster of differentiation 40 ligand) and cellular metabolism (including peroxisome proliferator-activated receptor γ, coactivator 1 α). Next, 14 target genes were selected for reverse transcription-quantitative polymerase chain reaction analysis in non-sequenced samples, and the downregulation of 10 target genes in RC samples with HHM was confirmed. In addition, it was demonstrated that hsa-let-7e-5p stimulated colorectal cancer cell migration in vitro. The miRNA hsa-let-7e-5p may serve as a potential biomarker for rectal carcinoma-associated HHM, facilitating the identification of patients with RC who are at risk of developing HHM.