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Objective To explore the mechanism of gastrointestinal dysfunction caused by chronic renal failure (CRF), and to determine whether colon is involved in the activation of oxidative stress (OS) in CRF. Methods Thirty male SD rats were randomly divided into control group (n＝10) and CRF group (n＝20). The rats in the CRF group were treated with 5/6 nephrectomy to establish CRF model, and the rats in the control group were only sutured after opening renal capsule. The rats were sacrificed at 10 weeks after model administration, and the serums and colon tissues near ileocecal valve were collected. Blood urea nitrogen (BUN) and serum creatinine (SCr) were measured to evaluate the success of the model. Malonodialdehyde (MDA), superoxide dismutase (SOD), total antioxidant capacity (TAC) and 8-hydroxy deoxyguanosine (8-OHdG) in the serum and colon tissues were detected to evaluate the level of OS. The ubiquinol cytochrome C reductase core protein(UQCRC1) was tested for the evaluation of mitochondrial function. Results Compared with the control group, the levels of BUN and SCr in serum of the rats in the CRF group were increased, suggesting that the model was successfully established. Compared with the control group, serum and colonic MDA levels were significantly increased in the CRF group (P0.05); however, there were no significant differences in 8-OHdG or anti-oxidative markers (SOD, TAC) in serum or colon tissues between the two groups (P0.05). The protein level of UQCRC1 in colon tissues was significantly reduced in the CRF group compared with the control group (P0.05). However, there was no significant difference in the mRNA level of UQCRC1 in colon tissues between the control and CRF groups (P0.05). Conclusion There is an imbalance between oxidation and antioxidation in the colonic tissues of CRF rats, which may be related to mitochondrial dysfunction.
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Objective To confirm whether fasudil can block C2C12 myoblasts respiration dysfunction triggered by Rho-associated coiled-coil containing protein kinase 1 (ROCK1), and whether it can block the occurrence of muscle atrophy. Methods C2C12 myoblasts were cultured in vitro, and 2% horse serum was used to induce cell differentiation and maturation. The obtained mature muscle tubule cells were divided into four groups according to the different stimuli: Ad-GFP group, only transfected GPT-adenovirus vector (Adv) in C2C12 myoblasts; Ad-ROCKl group, transfected ROCKl-Adv in C2C12 myoblasts to induce ROCK1 overexpression; Ad-GFPF group, transfected GFP-Adv and given 10 μmol/L fasudil in C2C12 myoblasts; and Ad-ROCKIF group, transfected ROCKl-Adv and given 10 μmol/L fasudil in C2C12 myoblasts. The oxygen consumption rate (OCR) and extracelluar acidification rate (ECAR) of C2C12 myoblasts under different stimulation conditions were evaluated by cell energy metabolism analyzer (Seahorse), so as to determine the effect of ROCK1 overexpression and fasudil stimulation on the respiratory function of C2C12 myoblasts. Mitochondrial fission was measured by MitoTracker® red fluorescent probes. The expressions of ROCK1, mitochondrial related protein 1 (Drpl) and phosphorylated p-Drpl, E3 ubiquitin ligase muscle RING finger-1 protein (MuRFl) and muscle atrophy F-box (MAFbx, Atrogin 1) was measured by Western blotting analysis. Results Seahorse analysis showed that the OCR, ECAR, basal respiration, maximal respiration and respiration required for coupling ATP of C2C12 myoblasts in the Ad ROCK1 group were significantly increased compared with those in the Ad-GFP group (P<0.01); Meanwhile, MitoTracker® staining showed that the mitochondrial fission was increased and the mitochondrial size frequency distribution shifted left in the Ad-ROCK1 group. After exposed to fasudil. the OCR and EACR of C2C12 myoblasts in the Ad-ROCKIF group were significantly decreased versus the Ad-ROCK1 group, and the basal respiration and maximal respiration were significantly increased (P<0.05). Western blotting analysis showed that p-Drpl/Drpl ratio, and the expressions of ROCK1, MuRFl and Atroginl in Ad-ROCKIF group were significantly reduced compared with Ad-ROCKl group (P<0.05). Conclusion Fasudil, an inhibitor of ROCK1, can block the abnormal cell respiration of C2C12 myoblasts caused by overexpressed ROCK 1 in vitro, and can reduce the activity of mitochondrial kinetic protein and the expression of muscle atrophy-related proteins.
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The discovery of microRNAs (miRNAs) has introduced a new paradigm into gene regulatory systems. Since inception, computational methods have been an invaluable tool complementing experimental approaches, and many discoveries have been obtained through combination of experimental and computational approaches. The knowledge that has been accumulated about the principles of miRNAs and target recognition were reviewed. The currently available computational methodologies and software for prediction of miRNA and their target genes also have been discussed.
RESUMO
The discovery of microRNAs (miRNAs) has introduced a new paradigm into gene regulatory systems. Since inception, computational methods have been an invaluable tool complementing experimental approaches, and many discoveries have been obtained through combination of experimental and computational approaches. The knowledge that has been accumulated about the principles of miRNAs and target recognition were reviewed. The currently available computational methodologies and software for prediction of miRNA and their target genes also have been discussed.