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Inhibition of cell migration and invasion by miR‑29a‑3p in a colorectal cancer cell line through suppression of CDC42BPA mRNA expression

He PY, Yip WK, Chai BL, Chai BY, Jabar MF, Dusa N, et al.

Oncol Rep, 2017 Dec;38(6):3554-3566.
PMID: 29039592 DOI: 10.3892/or.2017.6037

The objective of this study was to determine the effect of miR‑29a‑3p inhibitor on the migration and invasion of colorectal cancer cell lines (CRC) and the underlying molecular mechanisms. miR‑29a‑3p was detected using reverse transcription-quantitative polymerase chain reaction (RT‑qPCR) in the CRC cell lines HCT11, CaCo2, HT29, SW480 and SW620. An invasive subpopulation designated SW480‑7 was derived from the parental cell line, detected by Transwell and Transwell Matrigel assays. Cytoskeleton Regulators RT2 profiler PCR array and western blot analysis were utilized to identify the alterations in expression of downstream mRNAs. siRNA against CDC42BPA was transfected into SW480‑7 and effects on cell migration and invasion were investigated. Data obtained showed that miR‑29a‑3p was detected in these five CRC cell lines. miR‑29a‑3p inhibitor had no effect on viability but stimulated cell migration and invasion of SW480‑7 cells. In contrast, miR‑29a‑3p mimic suppressed cell migration and invasion. TargetScan miRBD and DIANA were employed to identify the potential direct target genes of miR‑29a‑3p in the Cytoskeleton Regulators RT2-Profiler PCR array. Cytoskeleton Regulators RT2-Profiler PCR array data showed that 3 out of the 5 predicted targets genes, CDC42BPA (2.33-fold), BAIAP2 (1.79-fold) and TIAM1 (1.77-fold), in the array were upregulated by miR‑29a‑3p. A significant increase in expression IQGAP2, PHLDB2, SSH1 mRNAs and downregulation of PAK1 mRNA was also detected with miR‑29a‑3p inhibition. Increase in CDC42BPA, SSH1 and IQGAP2 mRNA expression correlated with increased protein level in miR‑29a‑3p transfected SW-480-7 cells. Silencing of CDC42BPA (an enhancer of cell motility) partially abolished miR‑29a‑3p inhibitor-induced stimulation of cell migration and invasion. miR‑29a‑3p expression in stage II and III CRC is relatively lower than that of stage I CRC. However, the data need to be interpreted with caution due to the small sample size. In conclusion, inhibition of miR‑29a‑3p stimulates SW480‑7 cell migration and invasion and downstream expression IQGAP2, PHLDB2, SSH1 mRNAs are upregulated whilst PAK1 mRNA is downregulated. Silencing of CDC42BPA expression partially reduces miR29a‑3p inhibitor-induced migration and invasion of SW480‑7 cells.
Apoptosis induced by para-phenylenediamine involves formation of ROS and activation of p38 and JNK in chang liver cells

Chye SM, Tiong YL, Yip WK, Koh RY, Len YW, Seow HF, et al.

Environ Toxicol, 2014 Sep;29(9):981-90.
PMID: 23172806 DOI: 10.1002/tox.21828

para-Phenylenediamine (p-PD) is a suspected carcinogen, but it has been widely used as a component in permanent hair dyes. In this study, the mechanism of p-PD-induced cell death in normal Chang liver cells was investigated. The results demonstrated that p-PD decreased cell viability in a dose-dependent manner. Cell death via apoptosis was confirmed by enhanced DNA damage and increased cell number in the sub-G1 phase of the cell cycle, using Hoechst 33258 dye staining and flow cytometry analysis. Apoptosis via reactive oxygen species generation was detected by the dichlorofluorescin diacetate staining method. Mitogen-activated protein kinase (MAPK) activation was assessed by western blot analysis and revealed that p-PD activated not only stress-activated protein kinase (SAPK)/c-Jun N-terminal kinases (JNK) and p38 MAPK but also extracellular signal-regulated kinase (ERK). Cytotoxicity and apoptosis induced by p-PD were markedly enhanced by ERK activation and selectively inhibited by ERK inhibitor PD98059, thus indicating a negative role of ERK. In contrast, inhibition of p38 MAPK activity with the p38-specific inhibitor SB203580 moderately inhibited cytotoxicity and apoptosis induction by p-PD. Similarly, SP600125, an inhibitor of SAPK/JNK, moderately inhibited cytotoxicity and apoptosis induced by p-PD, thus implying that p38 MAPK and SAPK/JNK had a partial role in p-PD-induced apoptosis. Western blot analysis revealed that p-PD significantly increased phosphorylation of p38 and SAPK/JNK and decreased phosphorylation of ERK. In conclusion, the results demonstrated that SAPK/JNK and p38 cooperatively participate in apoptosis induced by p-PD and that a decreased ERK signal contributes to growth inhibition or apoptosis.
Fulltext Circulating miRNAs in Type 2 Diabetic Patients with and without Albuminuria in Malaysia

Zahari Sham SY, Ng CT, Azwar S, Yip WK, Abdullah M, Thevandran K, et al.

Kidney Blood Press Res, 2022;47(2):81-93.
PMID: 35158353 DOI: 10.1159/000518866

INTRODUCTION: Diabetic kidney disease (DKD) remains the leading cause of chronic kidney disease. Dysregulation of circulating miRNAs has been reported, suggesting their pathological roles in DKD. This study aimed to investigate differentially expressed miRNAs in the sera of type 2 diabetes mellitus (T2DM) patients with and without albuminuria in a selected Malaysian population.
METHOD: Forty-one T2DM patients on follow-up at a community clinic were divided into normo-(NA), micro-(MIC), and macroalbuminuria (MAC) groups. Differential levels of miRNAs in 12 samples were determined using the pathway-focused (human fibrosis) miScript miRNA qPCR array and was validated in 33 samples, using the miScript custom qPCR array (CMIHS02742) (Qiagen GmbH, Hilden, Germany).
RESULTS: Trends of upregulation of 3 miRNAs in the serum, namely, miR-874-3p, miR-101-3p, and miR-145-5p of T2DM patients with MAC compared to those with NA. Statistically significant upregulation of miR-874-3p (p = 0.04) and miR-101-3p (p = 0.01) was seen in validation cohort. Significant negative correlations between the estimated glomerular filtration rate (eGFR) and miR-874-3p (p = 0.05), miR-101-3p (p = 0.03), and miR-145-5p (p = 0.05) as well as positive correlation between miR-874-3p and age (p = 0.03) were shown by Pearson's correlation coefficient analysis.
CONCLUSION: Upregulation of previously known miRNA, namely, miR-145-5p, and possibly novel ones, namely, miR-874-3p and miR-101-3p in the serum of T2DM patients, was found in this study. There was a significant correlation between the eGFR and these miRNAs. The findings of this study have provided encouraging evidence to further investigate the putative roles of these differentially expressed miRNAs in DKD.

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