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Knockdown of KLK12 inhibits viability and induces apoptosis in human colorectal cancer HT-29 cell line.

ABSTRACT: Kallikrein?related peptidase 12 (KLK12) is overexpressed in cancer tissues including gastric, breast and prostate cancer. However, the role of KLK12 in colorectal cancer is not fully understood. In the present study, the level of KLK12 was determined by performing reverse transcription?polymerase chain reaction (RT?qPCR) in colorectal cancer tissues and cell lines. Lipofectamine® 2000 was used to transfect HT?29 cells to overexpress and knockdown KLK12. Cell viability, migration, invasion and apoptosis were detected by MTT, wound healing, Transwell and flow cytometry assays, respectively. The mRNA and protein expression levels of EMT?associated proteins, apoptosis?associated proteins, phosphorylated adenosine monophosphate?activated protein kinase (p?AMPK) and phosphorylated mammalian target of rapamycin (p?mTOR) were determined by RT?qPCR and western blot analysis. It was identified that the KLK12 mRNA levels were increased significantly in colorectal cancer tissues and cell lines. KLK12 small interfering RNA inhibited cell viability, migration and invasion. Furthermore, epithelial?mesenchymal transition (EMT)?associated proteins were altered by siKLK12. Cell apoptosis was induced by KLK12 downregulation, which was demonstrated by the changes in apoptosis?associated proteins; however, KLK12 overexpression produced the opposite effect. SiKLK12 enhanced the expression of p?AMPK and suppressed the expression of p?mTOR, while KLK12 overexpression had the opposite effect. Promotion of KLK12 overexpression?induced cell viability was reversed by 5?aminoimidazole?4?carboxamide ribonucleotide, an activator of the AMPK signaling pathway, and rapamycin, a specific inhibitor of the mTOR signaling pathway. Taken together, the results of the present study indicated that KLK12 was overexpressed in colorectal cancer and may regulate cell behavior, potentially via the AMPK and mTOR pathways.

SUBMITTER: Li Q

PROVIDER: S-EPMC6777684 | biostudies-literature | 2019 Nov

REPOSITORIES: biostudies-literature

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Knockdown of KLK12 inhibits viability and induces apoptosis in human colorectal cancer HT-29 cell line.

Li Qianyuan Q Zhou Xiukou X Fang Zhengyu Z Zhou Huamiao H

International journal of molecular medicine 20190830 5

Kallikrein‑related peptidase 12 (KLK12) is overexpressed in cancer tissues including gastric, breast and prostate cancer. However, the role of KLK12 in colorectal cancer is not fully understood. In the present study, the level of KLK12 was determined by performing reverse transcription‑polymerase chain reaction (RT‑qPCR) in colorectal cancer tissues and cell lines. Lipofectamine® 2000 was used to transfect HT‑29 cells to overexpress and knockdown KLK12. Cell viability, migration, invasion and ap ...[more]

PMID: 31485623

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Project description:The transcription factor STAT6 is strongly expressed in various tumours and is most highly expressed in human malignant lymphomas and pancreatic, colorectal, prostate and breast cancers. STAT6 is associated with cancer cell proliferation, an increased malignancy and poor prognosis. Thus, techniques aimed at reducing or blocking STAT6 expression may be useful in treating STAT6high cancers. Among these cancers, colorectal and breast cancers represent two of the most common worldwide and their incidence is increasing every year. In 2018, colorectal and breast cancers represented 10.2% and 11.6% of all new cases of cancer diagnosed, respectively. In this study, four proprietary STAT6 specific small interfering RNA (siRNA) sequences were tested in vitro using the human colon adenocarcinoma cell line, HT-29, and the breast/duct carcinoma cell line, ZR-75-1. Decreases in STAT6 mRNA and protein levels were analysed to confirm the transfection was successful and STAT6 knockdown effects were measured by analysing cell proliferation and apoptosis. Results showed that 100nM siRNA concentration was the most effective and, although all individual sequences were capable of significantly inhibiting cell proliferation, STAT6 siRNA sequences 1 and 4 had the largest effects. STAT6 silencing also significantly induced apoptotic events. In conclusion, these results demonstrate that STAT6 siRNA sequences are capable of inhibiting proliferation of and inducing apoptosis of HT-29 colorectal cancer cells and ZR-75-1 breast cancer cells, halving the number of cancer cells in a short period of time. These experiments will be repeated in other STAT6high cancers in vitro, and animal studies in immunocompromised mice have been planned using xenografts of STAT6-expressing human colorectal and breast cancer cells. The STAT6 siRNA sequences therefore represent a potential treatment for STAT6high colorectal and breast cancers and a wide variety of other STAT6-expressing cancers.

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Knockdown of KLK12 inhibits viability and induces apoptosis in human colorectal cancer HT-29 cell line.

Publications

Knockdown of KLK12 inhibits viability and induces apoptosis in human colorectal cancer HT-29 cell line.

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