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2.
AIM: To explore the effects of microRNA-129-3p (miR-129-3p) on the viability and migration of NIH3T3 cells during transforming growth factor-β (TGF-β)-induced transformation into myofibroblasts and the underlying molecular mechanisms. METHODS: RT-qPCR was used to examine the relative expression of miR-129-3p in renal cell carcinoma (RCC)-adjacent tissues and fibrotic renal tissue. NIH3T3 cells were stimulated with TGF-β to transform into myofibroblasts, and miR-129-3p expression level was detected. After transfection with miR-129-3p mimics for 48 h in vitro, the cell viability was measured by MTT assay, the protein expression level of Ki-67 was determined by Western blot, and the cell migration was observed by wound healing assay. The direct target of miR-129-3p was predicted by online database TargetScan and confirmed by dual-luciferase reporter assay. The expression level of target protein was further confirmed by Western blot. RESULTS: Compared with the RCC-adjacent tissues, the expression of miR-129-3p was down-regulated in fibrotic renal tissue (P<0.01). In TGF-β-induced NIH3T3 cell transformation into myofibroblasts, the expression of miR-129-3p was also decreased (P<0.01). Transfection with miR-129-3p mimics followed by TGF-β stimulation in the NIH3T3 cells inhibited the viability, Ki-67 expression and migration. TargetScan analysis showed miR-129-3p had binding sites in the 3'-UTR of Smad3, which was confirmed by dual-luciferase reporter assay. The results of Western blot further confirmed that miR-129-3p affected the expression of Smad3. CONCLUSION: miR-129-3p inhibits the viability and migration ability of NIH3T3 cells during TGF-β-induced transformation into myofibroblasts by directly targeting Smad3.  相似文献   

3.
AIM:To observe the expression of long noncoding RNA TTTY15 in osteosarcoma tissues and cell lines and to explore its effect on the viability and invasion ability of osteosarcoma cell lines. METHODS:qPCR was used to detect the expression of TTTY15 in 11 cases of osteosarcoma and its adjacent tissues. The mRNA levels of TTTY15 in osteosarcoma cell lines (143B, Saos2, MG-63, U2OS and HOS) and human osteoblast cell line hFOB1.19 were also tested. TTTY15 was down-regulated after transfected with small interfering RNA in MG-63 cells, the cell line with the highest level of TTTY15. The effect of TTTY15 knockdown on the viability of MG-63 cells was measured by CCK-8 assay. The cell cycle distribution was analyzed by flow cytometry. The effect of TTTY15 knockdown on the cell invasion ability was detected by Transwell assay. The levels of miR-216b-5p and FOXM1 mRNA were detected by qPCR, and the changes of the related proteins were determined by Western blot. RESULTS:Compared with the adjacent tissues, the expression of TTTY15 increased in the osteosarcoma tissues (P<0.01). Compared with the human osteoblast cell line, the expression of TTTY15 increased in the osteosarcoma cell lines (P<0.05), and the level of TTTY15 in the MG-63 cells was the highest (P<0.01). After knockdown of TTTY15 expression in the MG-63 cells, the cell viability was decreased (P<0.05), cell cycle progression was inhibited (P<0.01), and the cell invasion ability was decreased (P<0.01). The expression of miR-216b-5p was increased (P<0.01) and the expression of FOXM1 mRNA was decreased (P<0.01). The protein expression of FOXM1, CDK4, cyclin D1, MMP-2 and N-cadherin was decreased, while the protein expression of E-cadherin was increased (P<0.05). CONCLUSION:The expression of TTTY15 is increased in the osteosarcoma tissues and cell lines. The low expression of TTTY15 inhibits the cell viability and invasion ability of osteosarcoma cells. The possible mechanism is that the knockdown of TTTY15 expression results in the increase in miR-216b-5p expression and the down-regulation of FOXM1 expression.  相似文献   

4.
AIM:To compare the cDNA sequences of ASBT (apical Na+/bile acid cotransporter) gene cloned from human kidney and intestine tissues, and to determine subcellular localization of ASBT in renal tubular epithelial cells and expression site of ASBT in human kidney tissue.METHODS:The total RNA was extracted from human kidney and intestine tissues. The full-length cDNA gene of ASBT was amplified by RT-PCR technique using primers with 8-peptide FLAG tag, sequenced and inserted into eukaryotic expression vector to construct ASBT protein expression vector, which was then transfected into swine renal tubular epithelial cell line LLC-PK1. The subcellular localization of ASBT protein was determined by immunofluorescence staining and laser confocal microscope. Immunohistochemical analysis was used to observe expression position of ASBT in kidney tissue.RESULTS:The results revealed that the sequence of kidney ASBT cDNA gene was identical to that of human intestine ASBT gene. Western blotting analysis indicated that ASBT protein was correctly expressed in LLC-PK1 cells. Confocal scanning analysis showed that ASBT protein was mainly localized at the cellular plasma membrane, consistent with the predicting result obtained by bioinformatics. The results of immunohistochemistry revealed that ASBT was expressed at brush border membrane of proximal renal tubular cells, but not expressed in distal tubule and renal interstitium.CONCLUSION:The gene sequence of kidney ASBT is the same as that of intestine ASBT and ASBT protein is expressed at the lumen (apical) membrane of proximal renal tubular epithelial cells.  相似文献   

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6.
AIM: To investigate the dysfunction of renal cell and tissue in Npc1 mutant mice, in order to provide support for the treatment of Niemann-Pick disease type C1 (NPC1) patients.METHODS: The kidneys of wild-type (Npc1+/+) and Npc1 mutant (Npc1-/-) mice on postnatal day 60 were isolated. HE staining was performed to examine the morphological changes of the renal tissues. Oil red O staining was used to examine the lipid deposition in the renal tissues. The apoptosis of the renal cells was detected by TUNEL staining. The expression of apoptosis-related proteins in the renal tissue was determined by Western blot, and immunofluorescence was performed to examine the expression of α-smooth muscle actin (α-SMA) and vimentin in the renal tissues. RESULTS: Compared with Npc1+/+ mice, the morphological observation showed obvious vacuoles and no lipid deposition in the renal tissue of Npc1-/- mice. Subsequently, TUNEL staining showed significant increase in the apoptotic cells in the renal tissue of Npc1-/- mice (P<0.01), and the expression levels of Bax and Bad were up-regulated in the renal tissues of Npc1-/- mice (P<0.01), but Bcl-2 was down-regulated (P<0.05). Furthermore, the expression of α-SMA and vimentin was significantly up-regulated in the renal tissues of Npc1-/- mice (P<0.01). CONCLUSION: Npc1 gene mutation causes abnormal lipid metabolism in the renal cells, which induces the apoptosis of renal cells and promotes the fibrosis of renal tissue.  相似文献   

7.
AIM: To investigate the expression of microRNA-625-3p (miR-625-3p) in colorectal carcinoma (CRC) and its underlying mechanism. METHODS: Quantitative real-time PCR was employed to detect the levels of miR-625-3p expression in different CRC cell lines, CRC tissues and pair-matched adjacent normal tissues. The relationships between the expression levels of miR-625-3p and the patients' clinicopathological parameters were estimated. The effects of miR-625-3p on the apoptosis and the cell mitotic cycle of CRC cells were analyzed with propidium iodide staining and flow cytometry. The effect of miR-625-3p on the apoptosis-related proteins was analyzed by Western blot. RESULTS: The expression level of miR-625-3p in the CRC tissues was higher than that in the pair-matched adjacent normal tissues (P<0.05). The expression of miR-625-3p in the CRC tumor tissues was significantly correlated with the tumor infiltrative depth, TNM stage and distant metastasis (P<0.05). The expression levels of miR-625-3p in CRC SW620 cells were higher than that in SW480 cells. The CRC cell mitotic cycle was significantly inhibited and cell apoptosis was significantly promoted when the expression of miR-625-3p was inhibited (P<0.05). The expression of Bax protein didn't change and the expression of Bcl-2 protein increased after miR-625-3p mimics were transfected into CRC SW620 cells(P<0.05). CONCLUSION: miR-625-3p may be a promising approach for the treatment of CRC by promoting cell proliferation and inhibiting apoptosis.  相似文献   

8.
AIM:To investigate the significance and changes of p14ARF gene in gastric cancer.METHODS:The tumors and gastric tissues neighboring carcinoma from 48 patients with gastric cancer were studied. The homozygous deletions, mutations, methylation of the CpG islands, and mRNA expression of p14ARF gene were assessed by PCR, PCR-SSCP, PCR based methylation assay, and RT-PCR.RESULTS:①The homozygous deletion rate of p14ARF was 31.3% (15/48), and no homozygous deletions were examined in all the gastric tissues neighboring tumor. ②There were no point mutations of p14ARF in 33 gastric cancers without homozygous deletion and in the matched gastric tissues adjacent to tumor. ③Methylation rate of the CpG islands of p14ARF was significantly higher in gastric cancers(47.9%, 23/48) than that in gastric tissues neighboring cancer (4.2%, 2/48)(P＜0.01).④ No expression of p14ARF mRNA was detected in 45.8%(22/48) of gastric cancers. Moreover, the negative rate (100%, 3/3) of p14ARF mRNA of gastric cancers with the combined methylation of exons 1β and 2 was significantly higher than that (15%, 3/20) of the sole methylation of exon2(P＜0.05). CONCLUSION:p14ARF gene is frequently inactivated by homozygous deletion and methylation of the 5' CpG islands in gastric cancer, which may play an important role in the development of gastric cancer.  相似文献   

9.
CAI Zi-wei  ZHENG Xue-zhi  HU Jing 《园艺学报》2007,23(11):2191-2194
AIM: To study the expression of nucleostemin (NS) gene in human breast tumor tissues and the relations of NS gene expression level with histological grades,histological types and TNM stages of the tumor.METHODS: Total RNA was isolated from human breast tumor tissue.The methods of electrophoresis and RT-PCR were used in measuring NS gene expression level,and the relations of NS gene expression level with histological grades,histological types and TNM stages of the tumor were analyzed.RESULTS: The results indicated that there was no NS gene expression detected in normal breast tissues,and NS gene expression in malignant breast tumor tissues (P<0.01) was higher than that in the benign breast tumor tissues.The higher histological grades of the breast cancer showed the stronger NS gene expression (P<0.01),the higher TNM stages of the breast cancer showed the stronger NS gene expression (P<0.01),and the level of NS gene expression had not correlation with the histological types (P>0.05).CONCLUSION: It is suggested that there is no relation of NS gene expression level with histological types of the breast cancer,but there is a marked correlation of NS gene expression level with the histological grades and TNM stages.  相似文献   

10.
AIM: To investigate the expression of calprotectin(CALP) in the rats with renal ischemia-reperfusion injury(IRI). METHODS: Male Sprague-Dawley rats were randomly divided into sham operation and IRI group(n=25 in each group). Blood samples and the kidneys were obtained at 6 h, 12 h, 24 h, 48 h and 72 h after reperfusion. The pathological changes of the kidneys were observed. The serum concentrations of blood urea nitrogen(BUN) and serum creatinine(SCr) were measured. The serum levels of CALP, tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) were detected by ELISA, and the expression of CALP, Toll-like receptor 4(TLR4) and NF-κB p65 in the renal tissues were determined by the methods of immunohistochemistry and Western blot. RESULTS: Different serial ischemia changes were observed in the renal tissues, mainly in the renal tubular epithelial cells and the mesenchyma, with the infiltration of inflammatory cells. The serum levels of BUN, SCr, CALP, TNF-α and IL-6 in IRI group were markedly increased as compared with sham group(P<0.05). The protein expression of CALP, TLR4 and NF-κB p65 in the renal tubular epithelial cells in IRI group was greatly enhanced in comparison with that in sham group(P<0.05). CONCLUSION: The serum concentrations of CALP, TNF-α and IL-6, and the protein expression levels of CALP, TLR4 and NF-κB p65 in the renal tissue are significantly increased in the rats with IRI, suggesting that calprotectin plays an important role in the inflammation in rats with IRI.  相似文献   

11.
AIM: To investigate the expression of renal peroxisome proliferator-activated receptor gamma (PPARγ) in rats with adrimycine nephrosis (ADR), and the effect of rosiglitazone on the activation of NF-κB p65 in renal tissue rats with ADR. METHODS: The rats were randomly assigned to following groups: control (CTR) group, adrimycine nephrosis (ADR) group, and ADR treated with rosiglitazone (5 mg·kg-1·d-1) group（RGL）. The levels of urinary protein, albumin, total cholesterol, triglyceride and renal function change in rats were measured after 12 weeks. The nuclear-translocation of cortical NF-κB p65 was detected by immunohistochemistry. The activity of cortical NF-κB p65 was measured by sandwich ELISA. The mRNA levels of cortical PPARγ and TGF-β1 were detected by RT-PCR. The protein expressions of PPARγ and TGF-β1 in the rat kidney tissues were detected by Western blotting. RESULTS: As compared to ADR group, the urinary protein excretion in RGL treatment group was decreased and the serum albumin levels were increased, but the serum total cholesterol and triglyceride were decreased and the renal pathological lesion was ameliorated. The activity of NF-κB p65 and the expressions of TGF-β1 mRNA and protein were significantly decreased in rosiglitazone group, while the expression of PPARγ mRNA and protein was increased in RGL group (P<0.01). The correlation analysis was manifested: in ADR and RGL group, a negative correlation between the activity of NF-κB p65 and the expression of PPARγ in renal tissue (r=-0.8305, P<0.01) was observed. There was a negative correlation between the expression of TGF-β1 and PPARγ in renal tissues (r=－0.7938, P<0.01). CONCLUSION: The expression of renal cortical PPARγ is up-regulated in rats with adrimycine nephrosis by rosiglitazone. Rosiglitazone inhibits the activation of renal cortical NF-κB p65 in part, so it inhibits the gene expression of renal TGF-β1 and relieves the renal pathological lesion.  相似文献   

12.
AIM:To preliminarily explore the relationship between microRNA-7-5p (miR-7-5p) and Itch gene and their relationship with insulin resistance by establishing insulin resistance model of HepG2 cells in vitro for detecting differential expression of miR-7-5p and its predicted target gene Itch in the state of insulin resistance. METHODS:The insulin resistance model of HepG2 cells was induced by suitable concentration of plamitic acid. The possible target genes and the associated signaling pathways of miR-7-5p were predicted based on bioinformatic analysis. The expression levels of miR-7-5p and Itch were detected by RT-qPCR and Western blot in the HepG2 cells with insulin resistance. RESULTS:The HepG2 cell model of insulin resistance was successfully induced by treatment with 0.25 mmol/L palmitic acid for 24 h. Compared with negative control group, the expression level of miR-7-5p detected by RT-qPCR in insulin resistance group was significantly decreased (P<0.01). Bioinformatic analysis showed that a considerable number of target genes of miR-7-5p were enriched in the ubiquitin proteasome system. Among them, E3 ubiquitin ligase Itch gene was the most relevant target gene to insulin resistance. The results of Western blot showed that the protein expression of Itch was up-regulated in the HepG2 cells under insulin resistance (P<0.01). CONCLUSION:miR-7-5p may be involved in the pathophysiological process of insulin resistance, which may directly or indirectly affect the normal transduction of insulin signaling pathway by targeting Itch gene.  相似文献   

13.
ATM: To explore whether the C-reactive protein (CRP) level in microinflammation state induces the apoptosis of renal tubular epithelial cells. METHODS: HK-2 cells were stimulated with recombinant human CRP. Annexin-FITC-PI staining and flow cytometry were used to detect the percentage of apoptotic cells. Morphology observation of apoptosis was assessed by Hoechst 33258 staining. Caspase-3 activity was measured by a colorimetric assay. The expression of apoptotic gene bax and anti-apoptotic gene bcl-2 at mRNA levels was determined by real-time PCR. RESULTS: CRP induced apoptosis of HK-2 cells in a time- and dose-dependent manner. The maximal apoptotic effect of CRP concentration was 10 mg/L CRP at concentration of 20 mg/L. CRP treatment was associated with the characteristic morphological features of apoptosis such as condensation, fragmentation or margination of nuclear chromatin. CRP exposure increased caspase-3 activity, up-regulated the mRNA expression of Bax and down-regulated the mRNA expression of Bcl-2. CONCLUSION: Slightly increased CRP level has the potential to induce apoptosis of renal tubular cells.  相似文献   

14.
AIM:To explore the effect and the underlying mechanisms of microRNA-10b (miR-10b) on high glucose-stimulated epithelial-mesenchymal transition (EMT) of renal tubular epithelial cells. METHODS:The expression level of miR-10b was examined by RT-qPCR in the kidney tissues of the type 2 diabetes patients with kidney fibrosis. The EMT model of HK-2 cells was induced by high glucose stimulation and the miR-10b expression in the process was detected by RT-qPCR. The morphological changes of the HK-2 cells were observed using a microscope. EMT markers, such as fibronectin and N-cadherin, were examined by Western blot. The online database predicted that the 3'-UTR of KLF10 bound to miR-10b and their direct interaction was confirmed by dual luciferase report assay. RESULTS:Compared with the para-carcinoma normal tissues, the expression level of miR-10b was up-regulated in the tissues of type 2 diabetes patients with kidney fibrosis (P<0.01). In high glucose-stimulated HK-2 cells, the expression level of miR-10b was increased in a time-dependent manner (P<0.01). miR-10b inhibitor reversed the morphological changes and the increases expression of the EMT markers including fibronectin, SLUG, N-cadherin and SNAI1 induced by high glucose stimulation. Online database showed miR-10b was able to bind with the 3'-UTR in the promoter region of KLF10, thus negatively regulating its expression. Meanwhile, over-expression of KLF10 inhibited the EMT induced by high glucose. Inhibition of TGF-β/Smad3 activation was observed during the process of KLF10-repressed EMT. CONCLUSION:miR-10b promotes high glucose-stimulated epithelial-mesenchymal transition of renal tubular epithelial cells may through repressing KLF10 expression.  相似文献   

15.
AIM: To observe the influence of high expression of miR-15a-5p on the proliferation and migration of human hepatocellular carcinoma SMMC-7721 cells.METHODS: The miR-15a-5p oligonucleotide, which was reconstructed with additional restriction sites of EcoR Ⅰ and Hind Ⅲ, was chemically synthesized and confirmed by sequencing. The miR-15a-5p eukaryotic expression system was constructed by pcDNA6.2-GW/Em-GFP-pre-miR-15a-5p plasmid. The miR-15a-5p was transfected into the SMMC-7721 cells transiently by plasmid, and quantified by quantitative real-time PCR at the mRNA level. The cell viability was measured by CCK-8 assay, and the living cell counting was performed by the method of Trypan blue exclusion. The migration ability of the SMMC-7721 cells with high expression of miR-15a-5p was detected by wound healing test.RESULTS: The sequence of miR-15a-5p oligonucleotide 100% matched the designed sequence. Compared with control group, the miR-15a-5p expression was increased significantly (P<0.05). The viability, the living cell number and the migration ability of the SMMC-7721 cells were decreased in high expression of miR-15a-5p group with statistically significant difference (P<0.05).CONCLUSION: The abilities of proliferation and migration in human hepatocellular carcinoma SMMC-7721 cells are decreased by high expression of miR-15a-5p.  相似文献   

16.
AIM: To observe the changes of neutrophil gelatinase-associated lipocalin (NGAL) level in serum and renal tissues of the patients with primary nephrotic syndrome (PNS) and acute kidney injury (AKI). METHODS: Seventy-two PNS patients were selected in the study and divided into 2 groups according to the pathological results of renal biopsy.The patients in PNS+AKI group included 15 cases of PNS with acute tubular necrosis (ATN), in which there were 10 cases of minimal change disease (MCD) with ATN and 5 cases of mesangial proliferative glomerulonephritis (MsPGN) with ATN. The patients in PNS alone group included 57 cases of PNS without ATN. According to the pathological types, they were divided into MCD group (24 cases), membranous nephropathy (MN) group (23 cases) and MsPGN group (10 cases). Serum samples from 15 healthy persons and 5 cases of normal renal tissues were used as controls. The serum levels of NGAL were detected by ELISA. The distribution and expression of NGAL in the renal tissues were observed by immunohistochemical method. RESULTS: (1) The serum level of NGAL in PNS alone group (including MCD group, MN group and MsPGN group) was significantly higher than that in normal control group. Compared with MCD group and MsPGN group, the serum level of NGAL in MN group increased significantly. The renal tissue positive expression index of NGAL in MCD group, MN group and MsPGN group was significantly higher than that in normal control group. (2) The serum level of NGAL and the renal tissue positive expression index of NGAL in MCD+ATN group and MsPGN+ATN group were significantly increased compared with PNS alone group. (3) The serum level of NGAL was positively correlated with 24 h urine protein, blood urea nitrogen and serum creatinine. No relation with serum albumin and β2 microglobulin was observed. The serum level of NGAL was positively correlated with the NGAL level in the renal tissue. CONCLUSION: The serum level of NGAL can be used as one of the sensitive indexes for early and non-invasive detection of PNS with AKI.  相似文献   

17.
AIM: To explore the relationship between hypermethylation of p15INK4B gene and the pathogenesis of hematopoietic malignances. METHODS: The expression and methylation of p15INK4B gene and the expression of DNA methyltransferase genes (DNMTs) in bone marrow cells from 54 cases with hematopoietic malignances were detected by RT-PCR, Western blot, and methylation-specific PCR. RESULTS: The p15INK4B gene was methylated more often in high-risk myelodysplastic syndrome (MDS) patients, patients at blast phase of chronic myeloid leukemia (CML-BP) and acute leukemia patients than that in low-risk MDS patients (P<0.01). The expression levels of DNMT3A and DNMT3B in acute leukemia patients, high-risk MDS patients, and CML-BP patients were higher than that in low-risk MDS patients (P<0.05). CONCLUSION: The hypermethylation of p15INK4B gene may be one of the most common genetic event in pathogenesis of acute leukemia, high-risk MDS, and blast phase of chronic myeloid leukemia. Furthermore, DNMT3A and DNMT3B are substantially over-expressed in the bone marrow cells of these patients.  相似文献   

18.
AIM:To investigate the effect of Toll-like receptor 7(TLR7) agonist on the anti-tumor activity of peripheral blood mononuclear cells (PBMCs) in the patient with renal cell carcinoma.METHODS:Primary renal cancer cells from the postoperative specimens of the patient were co-cultured with peripheral blood mononuclear cells from the same patient stimulated by TLR7 agonist. The cytokine levels in culture medium were measured by ELISA, and the cell cycle distribution of the renal cell carcinoma cells was analyzed by flow cytometry. The anti-tumor activity of PBMCs was evaluated by[51Cr] release trial. The protein levels of Skp2 and its downstream pathway molecules were determined by Western blot. RESULTS:TLR7 agonist increased the expression of interferon-γ(IFN-γ), tumor necrosis factor-α (TNF-α) and interleukin-2 (IL-2) in the co-cultured medium, which significantly inhibited the proliferation index of the renal cell carcinoma cells. The cytotoxicity of PBMCs to renal cell carcinoma cells was markedly increased (P<0.05). The protein level of Skp2 in renal cell carcinoma cells was decreased significantly after stimulation, which was consistent with the change of cell proliferation index of renal cell carcinoma cells(P<0.05). The protein level of p27 was increased significantly (P<0.05), which was opposite to the change of Skp2. However, no significant difference in the expression of p21 and p53 was observed. CONCLUSION:TLR7 agonist effectively enhances the anti-tumor activity of PBMCs and results in the growth of renal cell carcinoma cells inhibiting. The mechanism may be relate to the cell cycle arrest by inhibiting the Skp2/p27 pathway.  相似文献   

19.
AIM: To explore the role of DNA methylation of microRNA-30a-5p(miR-30a-5p) promoter region in hepatic injury. METHODS: Four-week-old normal mice and cystathionine β-synthase (CBS) single gene knockout mice were used and divided into normal (CBS+/+, n=12) group and single gene knockout (CBS+/-, n=12) group, and the mice were fed with high methionine diet for 8 weeks. HL-7702 hepatic cells were routinely cultured in vitro and divided into control group, homocysteine (Hcy) group and Hcy+5-azacytidne (AZC) group. Serum Hcy, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels were measured by automatic biochemical analyzer. The levels of ALT and AST in the cells culture medium were determined by the microplate method. Hepatic injury in the mice were observed with HE staining. Cell viability staining was used to measure the viability of hepatocytes. RT-qPCR was used to detect the expression of miR-30a-5p in the liver tissues and hepatocytes. The correlation between the expression of miR-30a-5p and serum ALT and AST levels was analyzed by Pearson correlation analysis. DNA methylation level of miR-30a-5p promoter region in the liver tissues and hepatocytes was detected by nested landing methylation-specific PCR (nMS-PCR). RESULTS: Compared with the CBS+/+ mice, the serum levels of Hcy, ALT and AST in the CBS+/- mice were significantly increased (P < 0.05). HE staining showed the hepatocyte swelling and nuclear fragmentation and dissolution. The expression level of miR-30a-5p in the liver tissues was decreased (P < 0.01). Besides, the expression level of miR-30a-5p in the mice was negatively correlated with serum ALT and AST levels (r2=0.4557, P=0.0003, r2=0.4626, P=0.0003), and the DNA methylation of miR-30a-5p promoter region was increased (P < 0.01). In the HL-7702 cells, compared with control group,the ALT and AST levels were increased in Hcy group (P < 0.05, P < 0.01), and the cell viability was remarkablely decreased. DNA methylation of miR-30a-5p promoter region was increased (P < 0.01), which decreased after treated the cells with AZC (P < 0.05), while the expression level of miR-30a-5p in the cells was increased (P < 0.05). CONCLUSION: Hypermethylation of miR-30a-5p promoter region may play an important role in hepatic injury.  相似文献   

20.
AIM: To study the effects of over-expression of tricarboxylic acid cycle intermediates transporter NaDC3 (high affinity sodium-dependent dicarboxylate transporter) on energy metabolism and ROS generation in human renal tubular cells. METHODS: Recombinant retrovirus vector containing NaDC3 gene was constructed and used for infecting human renal tubular epithelial cell HKC. Control vector containing Neo gene was also constructed and infected cells. Liquid scintillation method was used to determine the level of ［3H］-succinate (as a transport substrate of NaDC3) in the cells. Clark electrode method and reverse phase HPLC were used to detect oxygen consumption and ATP content intracellularly, respectively. Mitochondrial membrane potential and reactive oxygen species (ROS) content in HKC were determined with laser confocal microscope after treatment with fluorescent probe JC-1 and CM-H2DCFDA, respectively. RESULTS: Western blotting analysis showed that the expression of NaDC3 protein in uninfected- and control vector-infected cells was at lower level. After infection with recombinant NaDC3 vector, expression level of NaDC3 protein in HKC cells was increased markedly. Transport assay revealed that the level of ［3H］-succinate in NaDC3-infected cells was noticeably increased. Oxygen consumption and ATP content in NaDC3-infected HKC were significantly higher than those in uninfected- and control vector-infected cells. Laser confocal analysis revealed that mitochondrial membrane potential and ROS level in NaDC3-infected HKC were increased, compared with uninfected- and control vector-infected cells. CONCLUSION: Over-expression of NaDC3 accelerates the speed rate of energy metabolism and increases intracellular ROS generation by transporting an overdose of tricarboxylic acid cycle intermediates in human renal tubular epithelial cells.  相似文献   

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