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Non-vesicle-bound free microRNAs could enter cells and affect gene expression

Advances in Bioscience and Biotechnology 2012年第3期3卷 181-185页

作者： Gang Wang Ka-Bik Lai bonnie ching-ha kwa Kai-Ming Chow Philip Kam-Tao Li Cheuk-Chun Szeto Department of Medicine & Therapeutics The Chinese University of Hong Kong Hong Kong China

Background: Cell-free microRNAs (miRNAs) exist in body fluid. Previous studies showed that cell-free mi-RNAs are partly bound in microvesicles, and could transfer between cells via fusion with cell membrane. Methods: We quantified the amount of a panel of mi-RNA targets in and outside microvesicles in human proximal tubular epithelial cell (HK2) medium by microarray and real-time quantitative polymerase chain reaction (RT-QPCR). Intercellular miRNA transfer was explored by medium transfer experiments. Results: We identified a portion of cell-free miRNAs that exists as non-vesicle bound, truly naked form. More importantly, these non-vesicle bound free miRNA could transfer between cells and exert biological effects. By miRNA microarray, we showed that the expression of many miRNA targets in HK-2 cells were altered, either up- or down-regulated, after exposure to extrinsic free miRNAs. The miRNA-200 family was the most affected in our model, with a corresponding alteration in the messenger RNA expression of down-stream targets including ZEB1 and vimentin. Conclusion: Our results suggest that free miRNA may serve as an intercellular messenger, a phenomenon that needs further exploration.

关键词： Cell Signaling Fibrosis Cytokine

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Vesicle-bound and free microRNAs in spent cell culture medium and biological fluids

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Advances in Bioscience and Biotechnology 2012年第8期3卷 1091-1095页

作者： Gang Wang Ka-Bik Lai bonnie ching-ha kwan Kai-Ming Chow Philip Kam-Tao Li Cheuk-Chun Szeto Department of Medicine & Therapeutics The Chinese University of Hong Kong Hong Kong China

Background: In addition to the control of gene transla- tion intra-cellularly, microRNAs (miRNAs) have been found to exist extracellularly. However, extracellular miRNA identified in previous studies were largely confined to microvesicles. It remains uncertain whether free extracellular miRNA exists. Methods: We quantify a panel of miRNAs (miRNA 200 family, miR-205, and miR-192) in spent culture medium of renal tubular epithelial cells, as well as serum and urine from patients with systemic lupus erythematosus and healthy controls. Microvesicle bound and free miRNA were separated by ultracentrifugation. Results: In spent cell culture medium, we found substantial amount of miR-200a, miR-200b, miR-200c, miR-141, miR-429, miR-205, and miR-192 in microvesicles, as well as the presence of miR-200a, miR-200b, miR-429, miR-205 and miR-192, but not miR-200c or miR-141 as non-vesicle bound free form. In healthy individuals, we found substantial amount of miR-200a, miR-200b, miR-200c, miR-141 and miR-192 in microvesicles from serum and urine, while only miR-141 exists as free form in both serum and urine. There was no significant difference in serum or urinary free miR-141 levels between SLE patients and healthy controls. In SLE patients, urinary free miR-141 level was signifiantly higher than serum. Neither serum nor urinary free miR-141 levels correlated with lupus disease activity. Conclusion: We found miRNA is present extracellularly, both within microvesicles and as free form, in spent cell culture medium, serum and urine. The biological role of extracellular miRNA requires further study.

关键词： Cell Signaling Fibrosis Cytokine

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The gene expression of NGAL and TLR9 in glomerulus and tubulo-interstitium of patients with lupus nephritis

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Advances in Bioscience and Biotechnology 2011年第1期2卷 33-39页

作者： Jianxin Lu bonnie ching-ha kwan Fernand Mac-Moune Lai Paul Cheung-Lung Choi Lai-Shan Tam Edmund Kwok-Ming Li Kai-Ming Chow Gang Wang Philip Kam-Tao Li Cheuk-Chun Szeto 不详

Background The role of neutrophil gelatinase associated lipocalin (NGAL) and Toll-like receptor 9 (TLR9) in the pathogenesis of lupus nephritis remain elusive. Methods We quantified the glomerular and tubulointerstitial mRNA expression of NGAL and TLR9 in 42 patients with lupus nephritis (LN group) and 10 controls. Results As compared to controls, LN group had higher glomerular expression of TLR9, and higher tubulointerstitial expression of NGAL and TLR9. Tubulointerstitial NGAL expression significantly correlated with proteinuria (r = 0.492;p = 0.003), renal function (r = -0.386;p = 0.022) and histological chronicity index (r = 0.540;p = 0.004). Proteinuria had significant correlation with glomerular (r = 0.554;p = 0.001) and tubulointerstitial (r = 0.379;p = 0.043) TLR9 expression. Furthermore, there was a significant difference in tubulointerstitial expression of NGAL between treatment response groups. Conclusion There is an increase in intra-renal mRNA expression of NGAL and TLR9 in LN. Although tubulointerstitial expression of NGAL does not correlate with systemic disease activity, it correlates with proteinuria, renal function, and therapeutic response. The role of NGAL in the pathogensis in LN, as well as its application as biomarker for lupus nephritis, requires further study.

关键词： Lupus Nephritis Immunology Cytokine

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