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MicroRNA (miR) is a class of highly conserved non-coding single-stranded RNA widely existing in mammals, which can negatively regulate the expression of targeting genes after transcription. As a key regulator, miR negatively regulates the expression of the targeting genes and disrupts important molecular signaling pathways, leading to the imbalance of multiple pathways such as tissue repair and inflammation involved in the fibrotic process. Among them, miR-15a/16 can participate in regulating and controlling the fibrotic process of various organs, including liver, lung, heart, kidney and other fibrotic diseases by acting on cell proliferation and transformation, extracellular matrix proteins production and degradation, inflammation and other important cell functions. It has potential diagnostic and therapeutic value. Clarifying the biological function of miR-15a/16 and its mechanism for action and therapeutic application prospects in various fibrotic lesions are of great significance for the molecular targeted treatment of fibrotic diseases.
Subject(s)
Humans , Fibrosis/genetics , MicroRNAs/genetics , Signal Transduction , InflammationABSTRACT
OBJECTIVES@#Nephrotic syndrome is a common disease of the urinary system. The aim of this study is to explore the effect of astragalus polysaccharides (APS) on multidrug resistance gene 1 (MDR1) and P-glycoprotein 170 (P-gp170) in adriamycin nephropathy rats and the underlying mechanisms.@*METHODS@#A total of 72 male Wistar rats were divided into a control group, a model group, an APS low-dose group, an APS high-dose group, an APS+micro RNA (miR)-16 antagomir group and an APS+miR-16 antagomir control group, with 12 rats in each group. Urine protein (UP) was detected by urine analyzer, and serum cholesterol (CHOL), albumin (ALB), blood urea nitrogen (BUN), and creatinine (SCr) were detected by automatic biochemical analyzer; serum interleukin-6 (IL-6), IL-1β, tumor necrosis factor α (TNF-α) levels were detected by ELISA kit; the morphological changes of kidney tissues were observed by HE staining; the levels of miR-16 and MDR1 mRNA in kidney tissues were detected by real-time RT-PCR; the expression levels of NF-κB p65, p-NF-κB p65, and P-gp170 protein in kidney tissues were detected by Western blotting; and dual luciferase was used to verify the relationship between miR-16 and NF-κB.@*RESULTS@#The renal tissue structure of rats in the control group was normal without inflammatory cell infiltration. The renal glomeruli of rats in the model group were mildly congested, capillary stenosis or occlusion, and inflammatory cell infiltration was obvious. The rats in the low-dose and high-dose APS groups had no obvious glomerular congestion, the proliferation of mesangial cells was significantly reduced, and the inflammatory cells were reduced. Compared with the high-dose APS group and the APS+miR-16 antagomir control group, there were more severe renal tissue structure damages in the APS + miR-16 antagomir group. Compared with the control group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 in the model group were significantly increased (all P<0.05); the levels of ALB and miR-16 were significantly decreased (both P<0.05). Compared with the model group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of pNF-κB p65 and P-gp170 in the low-dose and high-dose APS groups were significant decreased (all P<0.05); and the levels of ALB and miR-16 were significantly increased (both P<0.05). Compared with APS+miR-16 antagomir control group, the UP, CHOL, BUN, SCr, IL-6, IL-1β, and TNF-α levels, MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 were significantly increased (all P<0.05). The levels of ALB and miR-16 were significantly decreased in the APS+miR-16 antagomir group compared with the APS+miR-16 antagomir control group (both P<0.05).@*CONCLUSIONS@#APS can regulate the miR-16/NF-κB signaling pathway, thereby affecting the levels of MDR1 and P-gp170, and reducing the inflammation in the kidney tissues in the adriamycin nephropathy rats.
Subject(s)
Animals , Male , Rats , ATP Binding Cassette Transporter, Subfamily B, Member 1/genetics , Antagomirs , Doxorubicin/toxicity , Genes, MDR , Interleukin-6/metabolism , Kidney Diseases/genetics , MicroRNAs/metabolism , NF-kappa B/metabolism , Polysaccharides/pharmacology , RNA, Messenger , Rats, Wistar , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Abstract Ulcerative colitis is one of the IBDs. Its etiology and pathogenesis remain undefined with an interaction between environmental, genetic and immunological factors is the most accepted explanation. Several recent studies have examined microRNA expression in the peripheral blood and tissues from IBD patients. The study aims at assessing the expression of serum miR-16 in ulcerative colitis patients and its correlation with disease extent, activity and severity. It included 30 treatment naïve ulcerative colitis patients of different presentations. Serum miR-16 expression was assessed using reverse transcriptase quantitative real time PCR (RT-qPCR), and then correlated with that of a group of 20 healthy subjects to assess its role in diagnosis of ulcerative colitis. Also, it was correlated with disease extent (proctitis, left sided colitis, extensive colitis) and disease activity and severity indices (Truelove and Witts criteria, fecal calprotectin and UCEIS). Thirty ulcerative colitis patients were enrolled, 53% had mild, 37% had moderate, while 10% had severe disease. Concerning endoscopic extent, 8 had proctitis, 14 had left sided colitis and 8 had extensive colitis. Serum expression of miR-16 in the 30 patients were compared to that of the healthy control subjects. The patients' group showed median serum miR-16 expression of 1.91, 1.13 for the control group with a significant difference between both groups. Correlation between serum miR-16 expression with disease extent, activity and severity showed no significant relation. From the current study we can conclude that increased serum expression of miR-16 is associated with ulcerative colitis despite no significant relation to disease activity extent or severity.
Resumo A colite ulcerativa é uma das DII. Sua etiologia e patogênese permanecem indefinidas; a interação entre fatores ambientais, genéticos e imunológicos é a explicação mais aceita. Vários estudos recentes avaliaram a expressão de microRNA no sangue e tecidos periféricos em pacientes com DII. O presente estudo teve como objetivo avaliar a expressão do miR-16 sérico em pacientes com colite ulcerativa e sua correlação com a extensão, atividade e gravidade da doença. Foram incluídos 30 pacientes de colite ulcerativa, com diferentes apresentações, que ainda não haviam sido submetidos a nenhum tipo de tratamento. A expressão sérica de miR-16 foi avaliada usando transcrição reversa seguida de reação em cadeia da polimerase quantitativa (RT-qPCR) e, em seguida, correlacionada com a de um grupo de 20 indivíduos saudáveis para avaliar seu papel no diagnóstico de colite ulcerativa. Além disso, foi feita uma correlação com a extensão da doença (proctite, colite do lado esquerdo, colite extensa) e com os índices de atividade e gravidade da doença (critérios de Truelove e Witts, calprotectina fecal e UCEIS). Trinta pacientes com colite ulcerativa foram incluídos no estudo, classificada como leve em 53%, moderada em 37% e grave em 10%. Quanto à extensão endoscópica, oito apresentavam proctite, 14 apresentavam colite do lado esquerdo e oito apresentavam colite extensa. A expressão sérica de miR-16 nos 30 pacientes foi comparada à dos indivíduos controle saudáveis. No, grupo de pacientes, a expressão sérica de miR-16 foi de 1,91 (grupo controle: 1,13), uma diferença estatisticamente significativa entre os dois grupos. Não foi observada relação significativa entre a expressão sérica de miR-16 e a extensão, atividade e gravidade da doença. A partir do presente estudo, pode-se concluir que o aumento da expressão sérica do miR-16 está associado à colite ulcerativa, apesar de não haver relação significativa com a extensão ou gravidade da atividade da doença.
Subject(s)
Humans , Male , Female , Colitis, Ulcerative/genetics , Colitis, Ulcerative/pathology , MicroRNAs , Inflammatory Bowel Diseases , Reverse Transcriptase Polymerase Chain Reaction , Reverse Transcription , Real-Time Polymerase Chain ReactionABSTRACT
Objective To study the regulatory role of microRNA-16 (miR-16) on human pulmonary surfactant associated protein (SP).Method Human alveolar epithelial A549 cells were transfected by miR-16 analogue,analogue negative control,inhibitor and inhibitor negative control.Blank control group was also set up at the same time.The proliferative abilities of the cells in each group were measured using cell counting kit-8 (CCK8) test.The expressions of miR-16,SP-A,SP-B and SP-C mRNA were examined using reverse transcription polymerase chain reaction (RT-PCR).The protein levels of SP-A,SP-B and SP-C were examined using western blotting method.Result RT-PCR showed that the expression of miR-16 after transfected with miR-16 analogue (34.11± 1.79) was higher than the negative control group (1.65 ± 1.07) and the blank control group (1.07 ±0.50).The expression of miR-16 after transfected with miR-16 inhibitor (0.36±0.05) was lower than the negative control group (0.96±0.13) and the blank control group (1.05±0.20).The differences were significant (all P＜0.05),and indicated that miR-16 over-expression and suppression were successfully achieved.Compared with the blank control group,cell proliferation at different time points in the analogue negative control group and the inhibitor negative control group showed no significant differences (all P＞0.05).Compared with the blank control group (1.02±0.19,1.01±0.09,1.01± 0.12) and the analogue negative control group (1.08±0.24,1.00±0.14,1.00±0.05),miR-16 down-regulated the mRNA expressions of SP-A,SP-B and SP-C (0.58±0.16,0.67±0.05,0.61±0.12).On the other hand,compared with the blank control group (1.02±0.19,1.01±0.09,1.01±0.12) and the inhibitor negative control group (1.05±0.22,0.99±0.13,0.98±0.10),miR-16 up-regulated the mRNA expressions of SP-A,SP-B and SP-C (1.66±0.33,1.29±0.11,1.23±0.12)(all P＜0.05).The trends of protein level of SP-A,SP-B and SP-C were related to their mRNA expression.Conclusion This study indicates that miR-16 inhibits pulmonary surfactant associated protein in A549 cells.
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Objective To investigate the expression of miRNA-16 (miR-16) in breast cancer patients and its effect on the proliferation and migration of breast cancer cells. Methods Polymerase chain reaction (PCR) was used to detect the expression of miR-16 in 30 breast cancer patients. miR-16 mimics was transfected to MDA-MB-231 and MCF-7 cell, and CCK-8 as well as Transwell assay was applied to detect the effect of miR-16 on cell proliferation and migration of MDA-MB-231 and MCF-7 cells. Results Among 30 breast cancer patients, the expression of miR-16 was down-regulated in 23 cases. Cell proliferation and migration of MDA-MB-231 and MCF-7 cells were inhibited significantly after transfection of miR-16 mimics. Conclusion miR-16 is down-regulated in breast cancer patients. miR-16 inhibits significantly the cell proliferation and migration of breast cancer cells in vitro.
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AIM:To study the effect of estradiol (E2) on the viability of mesenchymal stem cells (MSCs) de-rived from the decidua of the placenta by regulating the expression of microRNA-16 (miR-16).METHODS:The concen-tration of E2 in the peripheral blood of normal pregnant women and the patients with severe preeclampsia ( PE) was meas-ured.The effects of E2 at different concentrations on the viability of MSCs were analyzed .The effect of E2 at different con-centrations on the expression of miR-16 in the MSCs was detected , and which estrogen receptor ( ER) mediated the regula-tory effect of E2 on miR-16 expression was determined .RESULTS:The concentration of E2 in peripheral blood of the pa-tients with severe PE was significantly decreased (P<0.01).After treatment with E2 at 5, 10 and 100 nmol/L for 48 h, the viability of MSCs was increased (P<0.05).The expression level of miR-16 was down-regulated in the MSCs treated with E2 at 5, 10 and 100 nmol/L for 12 h.After treatment with E2 at 10 nmol/L for different time (0 h, 3 h, 6 h, 12 h and 24 h) , the expression level of miR-16 in the MSCs showed a clear time-dependent downward trend .E2 significantly promoted the viability of MSCs , and the cell viability was significantly reversed after miR-16 pretreatment.Pretreatment with estrogen receptor antagonists ICI 182780 and tamoxifen for 6 h attenuated the inhibitory effect of E 2 on miR-16 expres-sion.Only ERαagonist propyl pyrazole triol significantly inhibited the expression of miR-16 in MSCs but ERβagonist dia-rylpropionitrile did not .CONCLUSION:E2 promotes the growth of decidua-derived MSCs by inhibiting miR-16 via ERα.
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AIM: To observe the effect of microRNA-16 (miR-16) on the megakaryocytic differentiation of K562 cells, and to explore the potential mechanism.METHODS:miR-16 was over-expressed or silenced by transfection with miR-16 mimics or inhibitor in K562 cells.The level of miR-16 was detected by real-time PCR.The expression of CD41, CD42b and CD61, as megakaryocytic differentiation markers, was detected by flow cytometry.The effect of miR-16 on the expression of myeloblastosis oncogene ( MYB) was measured by Western blotting, and flow cytometry was performed to confirm whether the effect of miR-16 on expression of CD41, CD42b and CD61 was mediated by MYB.RESULTS:Transfection with miR-16 mimics dramatically elevated the level of miR-16 and the expression of CD41, CD42b and CD61 in the K562 cells.Transfection with miR-16 inhibitor decreased the level of miR-16 and the expression of CD41, CD42b and CD61 in the K562 cells (P<0.05).The expression of MYB was regulated by miR-16, and MYB silencing reversed the regulation of CD41, CD42b and CD61 induced by miR-16.CONCLUSION:miR-16 regulates the megakaryocytic dif-ferentiation of K562 cells by targeting MYB.
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AIM:To investigate the effect of microRNA-16 ( miR-16) on the proliferation, invasion and cyto-kine secretion of rheumatoid arthritis ( RA) synovial fibroblasts ( RASFs) from the RA patients.METHODS: miR-16 mimic and miR-16 inhibitor were synthesized, and then Transfected into RASFs isolated from RA patients with lipo-fectamine.MTT assay, Transwell chamber and flow cytometry were used to determine the effect of miR-16 on proliferation, invasion and apoptosis of RASFs.The expression of matrix metalloproteinase 3/13 ( MMP3/13) and interleukin 1β( IL-1β) was measured by RT-PCR and Western blotting.RESULTS: The proliferation and invasion of RASFs were signifi-cantly inhibited by miR-16 mimic.The result of flow cytometry demonstrated that miR-16 had no effect on apoptosis of RASFs.Furthermore, miR-16 down-regulated the expression of MMP3/13 and IL-1β.CONCLUSION:miR-16 plays an important role in the development of RA and may inhibit the proliferation and invasion of RASFs through down-regulating the expression of MMP3/13 and IL-1β.
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Objective To study the expression levels of microRNA (miR)-16 and miR-146a in rat lungs of decompres-sion sickness (DCS) caused by fast buoyancy ascent escape or diving .Methods At 0.5 h after fast buoyancy ascent es-cape or diving, the pathological changes in rat lungs and expression levels of miR-16,and miR-146a were detected by re-verse transcription-quantitive polymerase chain reaction and compared with normal control group .Results The pathological characteristics of lungs in two DCS groups were tissue damage .At 0.5 h after DCS caused by fast buoyancy ascent escape , the lung tissue expression levels of miR-16 and miR-146a did not significantly change compared with normal control and diving DCS groups ,but the rat lung tissue expression level of miR-146 a in diving DCS group was obviously increased , com-pared with normal control group .Conclusion miR-146a may play a role in post-transcriptional regulation in the process of diving DCS .