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Objective To study the effect of high mobility group box B1(HMGB1)gene knockout on alleviating a-cute lung injury and inhibiting toll-like receptor 4(TLR4)/nuclear factor-KB(NF-κB)pathway of sepsis mice.Methods Wild-type(WT)mice were divided into WT-Sham group and WT-model group,and HMGB1 knockout(KO)mice were divided into KO-sham group and KO-model group.Sepsis ALI model was established by cecal ligation and perforation in WT-model group and KO-model group.Sham operation was performed in WT-Sham group and KO-Sham group.24 h after modeling,the partial pressure of arterial oxygen(PaO2)was detected,oxy-genation index(OI)was calculated,pathological changes of lung tissue were detected and lung injury score was calculated,the concentrations of tumor necrosis factor-α(TNF-α),interleukin-1 β(IL-1 β),interleukin-6(IL-6),reactive oxygen species(ROS),malondialdehyde(MDA),superoxide dismutase(SOD),in serum and lung tissues and the expression of HMGB1,TLR4 and nuclear NF-κB in lung tissues were detected.Results The PaO2,OI and the concentration of SOD in serum and lung tissue of WT-model group were lower than those of WT-Sham group,the lung injury scores,the concentrations of TNF-α,IL-1 β,IL-6,ROS and MDA in serum and lung tissue,and the expression levels of HMGB1,TLR4 and nuclear NF-κB in lung tissue were higher than those in WT-Sham group(P＜0.05).HMGB1 was not expressed in lung tissue of KO-model group,and the concentrations of PaO2,OI and the concentration of SOD in serum and lung tissue of KO-model group were higher than those of WT-model group,the lung injury scores,the concentrations of TNF-α,IL-1β,IL-6,ROS and MDA in serum and lung tissue,and the expression levels of TLR4 and nuclear NF-κB in lung tissue were lower than those of the WT-model group(P＜0.05).Conclusion HMGB1 gene knockout alleviates acute lung injury of sepsis mice,the re-lated molecular mechanism may be the inhibition of TLR4/NF-κB pathway mediated inflammation and oxidative stress.
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BACKGROUND:Upregulation of hedgehog protein signaling can increase the expression of osteoarthritis markers,Runx2,a disintegrin and metalloproteinase with thrombospondin motifs,collagen type X alpha 1,and matrix metalloproteinase 13,while inhibition of hedgehog proteins attenuates the severity of osteoarthritis.It is speculated that osteoarthritic chondrocytes can influence bone formation by affecting osteoblasts through the Indian hedgehog protein(IHH)signaling pathway. OBJECTIVE:To investigate the effect of human osteoarthritic chondrocytes on subchondral osteoblasts. METHODS:Tibial plateau specimens from patients with osteoarthritis were collected.Chondrocytes were extracted using enzymatic digestion,and osteoblasts were extracted using enzymatic pre-digestion + bone block method.Chondrocytes were identified by toluidine blue staining and immunofluorescence and osteoblasts were identified by alkaline phosphatase staining and immunofluorescence.Chondrocytes were cultured in sodium alginate beads to maintain chondrocyte phenotype and co-cultured with osteoblasts.The co-culture system was added with IHH signaling pathway inhibitor(cyclopamine,10 nmol/L)and activator(purmorphamine,10 nmol/L)separately.After 48 hours of co-culture,osteoblasts from each group were collected,mRNA expressions of Gli1,osteoprotegerin,Runx2,parathyroid hormone-related peptide,alkaline phosphatase,receptor activator of nuclear factor-kB ligand(RANKL)and osteocalcin were detected by qRT-PCR,and protein expressions of GLi1,oseoprotegerin and RANKL in osteoblasts were detected by western blot. RESULTS AND CONCLUSION:The mRNA expression levels of GLi1,osteoprotegerin and RUNX2 in osteoblasts were significantly increased,while the mRNA expression levels of parathyroid hormone-related peptide were decreased(P＜0.05)when co-cultured with human osteoarthritic chondrocytes.The mRNA and protein levels of Gli1 were significantly decreased after the addition of IHH signaling pathway inhibitor(cyclopamine)(P＜0.05),and the mRNA and protein levels of Gli1 were significantly increased after the addition of IHH signaling pathway activator(purmorphamine)(P＜0.05).Osteoprotegerin showed the same trend as Gli1 in the experiment.The osteoprotegerin/RANKL ratio followed the same trend as osteoprotegerin.To conclude,human osteoarthritic chondrocytes can promote the expression of Gli1,osteoprotegerin,Runx2 and other proteins in osteoblasts.The upregulation of osteoprotegerin is related to the IHH signaling pathway.Osteoarthritic chondrocytes can up-regulate the expression of osteoprotegerin in osteoblasts through the IHH signaling pathway and thus up-regulate the osteoprotegerin/RANKL ratio,which will contribute to bone formation in subchondral bone.
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Objective To investigate the relationship between nuclear factor(NF)-κB signaling pathway and gender differences in alcoholic liver fibrosis. Methods C57BL/6 N mice at 7-8 weeks of age were randomly divided into: male normal group, male model group, female normal group and female model group of 20 mice each. The normal group was fed with control liquid diet for 8 weeks, and the model group was fed with alcoholic liquid diet for 8 weeks combined with 31.5% ethanol gavage (5g/kg twice a week) to establish an alcoholic liver fibrosis model. The mice were executed at the end of 8 weekends, and the alanine aminotransferase (ALT), aspartate aminotransferase (AST) activity, estradiol (E
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Objective To investigate the effects of dexmedetomidine (DEX) on the nuclear factor-KB inhibitor protein kinase (IKK)/nuclear factor-KB inhibitor protein a (IKB(X)/nuclear factor-KB (N F - K B) pathway and cognitive dysfunction in rats with post-traumatic stress disorder (PTSD) . Methods Rats were randomly divided into control group, model group, positive group and DEX group. Except for the control group, the PTSD model was constructed by single prolonged stress method (SPS), and the corresponding drugs were given after the completion the model. Open field test and Morris water maze method were used to detect the autonomous activity, learning and memory ability of rats; HE staining was used to observe the morphological characteristics of cerebral cortex and hippocampus; ELISA and Western blotting were used to detect the contents of interleukin (IL)-1(3, IL-6, tumor necrosis factor a (TNF-a) and the expression levels of IKK, IKB(X, purinergic ligand-gated ion channel 7 receptor (P2X7R), leucine-rich repeat domain protein 3(NALP3) in hippocampus; the NF-KB activity was assessed by electrophoretic mobility shift assay (EMSA). Results Compared with the control group, the cerebral cortex and hippocampal CA1 region of model group were in structural disorders, nuclear pyysis and other pathological changes happend, learning and memory ability of rats decreased (P < 0. 05), contents of IL-lp, IL-6 and TNF-a, expression levels of IKK, IKB(X, P2X7R and NALP3, NF-KB activity in hippocampus increased (P<0. 05); Compared with the model group, the pathological phenomena in cerebral cortex and hippocampal CA1 area of rats in positive group and DEX group were in alleviated, and the changes of the above indexes were opposite to those of model group (P<0. 05) . Conclusion DEX can significantly improve the autonomous activity ability and learning and memory ability in PTSD rats, reduce inflammatory reaction in hippocampus and improve cognitive dysfunction, which may be related to the down-regulation of IKK/TKBQ!/NF-KB pathway.
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Alcoholic liver disease (ALD) is a chronic liver disease in which the internal liver tissues are inflammation damaged caused by long-term excessive drinking. Direct or indirect induction of hepatic inflammatory response by ethanol and its derivatives in the metabolic process may be an important mechanism of ALD, but the underlying cellular and molecular mechanisms of this process are still unclear. Recent study found that interleukin-6 (IL-6) response to ethanol mediated inflammation of the liver cells with dual role. It is involved in an inflammatory process that drives alcohol damage, activate cell apoptosis signaling pathways to stimulate macrophage and lymphocyte acute reactive protein synthesis aggravate the inflammatory response, and can lead to liver cell regeneration, increase anti-inflammatory cytokine levels play anti-inflammatory function to improve the degree of liver injury, and exercise stress can cause muscle source sex IL-6 temporary increased significantly, change the liver oxidation-inflammatory state. Then the body is kept in the adaptive state of long-term anti-inflammation to prevent the inflammatory damage of liver cells. Based on deepening the understanding of ALD inflammation pathological mechanism at the same time, the review on alcoholic liver cell inflammation related factor change and the IL-6 regulation pathway, considering the clinical use of IL-6 joint inflammatory factor pathway of targeted therapy is expected to be a novel therapy, the feasibility for laboratory screening of inflammatory related ALD drug intervention, for the prevention of alcoholic liver disease and treatment to provide new targets and train of thought.
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Aim To explore the effect of beta-sitosterol (BS) on liver fibrosis induced by CCL4 in mice and the mechanisms. Methods Fifty C57BL/6 male mice were randomly divided into five groups; control group (CG) , carbon tetrachloride group (CTG), low/medium/high dose of BS group ( BS-L/M/H), with 10 mice in each group. The model of hepatic fibrosis was established by injecting CCL4 in peritoneal cavity, the study lasted 30 days, and different doses of BS were given from 1st day to 15 th day. All mice were sacrificed for the observation of morphological changes and the measurement of liver index. Liver collagenous fibers were observed by HE and Masson staining, the changes of serum ( ALT and AST) were assessed by Elisa, the expressions of a-SMA and Collagen I were detected by Western blot and immunohistochemistry, and the changes of TpRl-Smad2/3 and TNF-a-NF∗kB were detected by Elisa and Western blot. Results Compared to control group, different doses of BS markedly inhibited the increase of liver index, A .T, AST, a-SMA and Collagen I in a dose-dependent n an-ner ( P < 0. 05 or P < 0. 01 ). Liver morphology, inflammatory cell infiltration and collagenous fiber irj BS groups were better than those in CCL4 group, meanwhile BS-M decreased the expression of TgKl, Smad2/3, TNF-a and p-NF-KB (P <0. 01). Conclusions BS dose-dependently inhibits mouse liver f bro-sis induced by CCL4, and its mechanism may be related to inhibiting TpRl-Smad2/3 and TNF-a-N •-kB signaling pathways.
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Objective@#Hyperbaric oxygen treatment (HBOT) has demonstrated efficacy in improving hearing levels of patients with idiopathic sudden sensorineural hearing loss (ISSHL); however, the underlying mechanisms are not well understood. HBOT alleviates the inflammatory response, which is mediated by Toll-like receptor (TLR) 4 and nuclear factor (NF)-κB. In this study we investigated whether HBOT attenuates inflammation in ISHHL patients alteration of TLR4 and NF-κB expression.@*Methods@#ISHHL patients ( = 120) and healthy control subjects ( = 20) were enrolled in this study. Patients were randomly divided into medicine group treated with medicine only ( = 60) and HBO group receiving both HBOT and medicine ( = 60). Audiometric testing was performed pre- and post-treatment. TLR4, NF-кB, and TNF-α expression in peripheral blood of ISSHL patients and healthy control subjects was assessed by ELISA before and after treatment.@*Results@#TLR4, NF-κB, and TNF-α levels were upregulated in ISSHL patients relative to healthy control subjects; the levels were decreased following treatment and were lower in the HBO group than that in the medicine group post-treatment ( < 0.05 and < 0.01).@*Conclusion@#HBOT alleviates hearing loss in ISSHL patients by suppressing the inflammatory response induced by TLR4 and NF-κB signaling.
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Adolescent , Adulte , Sujet âgé , Femelle , Humains , Mâle , Adulte d'âge moyen , Jeune adulte , Chine , Surdité neurosensorielle , Thérapeutique , Perte auditive soudaine , Thérapeutique , Oxygénation hyperbare , Inflammation , Génétique , Thérapeutique , Sous-unité p50 de NF-kappa B , Génétique , Métabolisme , Récepteur de type Toll-4 , Génétique , MétabolismeRÉSUMÉ
BACKGROUND: Currently, studies have focused on the role and mechanism of nuclear factor-kappa B pathway in the pathological process of acute lung injury in burned rats, such as the targeting inhibition of kB kinase by miR-155, which further weakens the activity of nuclear factor-KB and plays a role in acute lung injury in burned rats. However, there are still some pathological mechanisms to be studied and confirmed. OBJECTIVE: To investigate the effect of miR-155 on acute lung injury in burned rats through nuclear factor-KB pathway. METHODS: The rat models of acute lung injury were established by warm water bath simulating bum injury. The burned rats were divided into acute lung injury, miR-155-mimics and miR-155-inhibitor groups. After fluid resuscitation, the rats in the miR-155-mimics and miR-155-inhibitor groups were injected into the tail vein of 5 |_iL of miR-155-mimics and miR-155-inhibitions, respectively. The expression levels of tumor necrosis factor-a and interleukin-1 p in bronchoalveolar lavage fluid were detected by ELISA. The lung morphology in the three groups was observed by hematoxylin-eosin staining. The protein expression levels of nuclear factor-KB and cyclooxygenase 2 were evaluated by western blot assay. The nuclear factor-KB protein in lung tissues was detected by immunohistochemistry. RESULTS AND CONCLUSION: (1) The results of hematoxylin-eosin staining showed that the severity of lung injury in the miR-155-inhibitor group, acute lung injury group and the miR-155-mimics group was increased gradually (P < 0.05). (2) ELISA results showed that compared with the acute lung injury group, the expression levels of tumor necrosis factor-a and interleukin-1 p were increased in the miR-155-mimics group (P < 0.05), and decreased in the miR-155-inhibitor group (P < 0.05). (3) Western blot assay results showed that compared with the acute lung injury group, the expression levels of nuclear factor-KB and cyclooxygenase 2 proteins were increased in the miR-155-mimics group (P < 0.05), and decreased in the miR-155-inhibitor group (P < 0.05). (4) Immunohistochemical results showed that the expression level of nuclear factor-KB was increased in the miR-155-inhibitor group, which was dark brown. The expression of nuclear factor-KB in cytoplasm and nucleus of neutrophils, mononuclear macrophages, alveolar epithelial cells was the most obvious. (5) These results indicate that in lung tissue cells, decreased miR-155 can down-regulate nuclear factor-KB activity, which reduces the inflammatory response of the lung between the damaged tissue. The study was approved by the Laboratory Animal Ethics Committee of the First People’s Hospital of Neijiang, approval No. 1801270.
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BACKGROUND: There is an inflammatory response in the lesion tissue of ischemic cerebral infarction, and the expression of miR-150-5p is significantly decreased. Whether miR-150-5p inhibits the release of inflammatory factors and alleviates the injury of ischemic cerebral infarction tissue through the Toll-like receptor-5/nuclear factor-KB pathway remains unclear. OBJECTIVE: To investigate the role and preliminary mechanism of miR-150-5p in ischemic cerebral infarction in rats. METHODS: (1) The rat models of middle cerebral artery occlusion were constructed and the rat models were divided into five groups: Control, miR-150-5p agomir, agomir control, miR-150-5p antagomir and antagomir control groups. (2) The rats in the control group was given the intracerebroventricular injection of normal saline, and the rats in the latter four groups were given the intracerebroventricular injection of miR-150-5p agomir (miR-150-5p agonist), agomir negative control, miR150-5p antagomir (miR150-5p inhibitor) and antagomir negative control, respectively. (3) After 7 days, the brain was graded by modified neurological severity score, the cerebral infarct volume was measured by MRI, and the histopathological changes were observed by hematoxylin-eosin staining. The expression levels of miR-150-5p, interleukin-6, tumor necrosis factor-a, Toll-like receptor-5 and nuclear factor-KB p65 in brain tissues were detected by qRT-PCR, ELISA and western blot assay, respectively. The target relationship between miR150-5p and Toll-like receptor-5 was verified by luciferase assay by retrieving the bioinformatics website Targetscan to predict the binding sites of miR-150-5p and Toll-like receptor-5. RESULTS AND CONCLUSION: (1) Compared with the control group, the modified neurological severity score, and levels of interleukin-6, tumor necrosis factor-a, Toll-like receptor-5 and nuclear factor-KB p65 proteins were significantly decreased in the miR-150-5p agomir group (P 0.05). (3) TargerScan website prediction results and luciferase reporter gene analysis results showed that miR-150-5p and Toll-like receptor-5 had a targeted binding site. (4) These results imply that miR-150-5p can inhibit the inflammatory signaling pathway of Toll-like receptor-5/nuclear factor-KB p65 in brain injury caused by ischemia and reduce the inflammatory response, thereby alleviating the damage of nerve function and playing a protective role.
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Objective To explore the effect and probable mechanism of rosuvastatin preconditioning on inflammatory cytokines interleukin (IL) -1β, IL-6 and tumor necrosis factor a (TNF-α) release in vascular smooth muscle cells (VSMCs) of middle cerebral artery after ischemia-reperfusion. Methods Thirty-six healthy SD rats were randomly assigned into three groups; sham operation group, focal cerebral ischemia-reperfusion group and rosuvastatin preconditioning group. There were 12 rats in each group. At the 24th hour of reperfusion after middle cerebral artery occlusion (MCAO) for 2 hours, the mRNA and protein expression of IL-1β, IL-6 and TNF-α release in VSMCs of middle cerebral artery were detected by Real-time PCR and Western blotting, respectively. Also, the mRNA and protein expression of nuclear factor κB (NF-κB) were measured by Real-time PCR and Western blotting. Results At the 24th hour of reperfusion after MCAO for 2 hours, the mRNA and protein expression of IL-Iβ, IL-6 and TNF-ot were markedly up-regulated in rats of model group; rosuvastatin preconditioning can significantly inhibited overexpression of IL-1β, IL-6 and TNF-ot at the mRNA and protein levels. And the decreasing of mRNA and protein expression of NF-κB was also found in this study. Conclusion Rosuvastatin preconditioning can decrease the release of inflammatory cytokines IL-1β, IL-6 and TNF-α in VSMCs of MCA. It can relieve the inflammatory injury after ischemia-reperfusion in brain. The effect of rovastatin on IL-1β,IL-6 and TNF-ot may be related to the reduction of the expression of NF-κB in VSMCs.
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Catalpol is the main active ingredient of an extract from Radix rehmanniae, which in a previous study showed a protective effect against various types of tissue injury. However, a protective effect of catalpol on uterine inflammation has not been reported. In this study, to investigate the protective mechanism of catalpol on lipopolysaccharide (LPS)-induced bovine endometrial epithelial cells (bEECs) and mouse endometritis, in vitro and in vivo inflammation models were established. The Toll-like receptor 4 (TLR4)/nuclear factor-κB (NF-κB) signaling pathway and its downstream inflammatory factors were detected by enzyme-linked immunosorbent assay (ELISA), quantitative real-time polymerase chain reaction (qRT-PCR), western blot (WB), and immunofluorescence techniques. The results from ELISA and qRT-PCR showed that catalpol dose-dependently reduced the expression of pro-inflammatory cytokines such as tumor necrosis factor α (TNF-α), interleukin (IL)-1β, and IL-6, and chemokines such as C-X-C motif chemokine ligand 8 (CXCL8) and CXCL5, both in bEECs and in uterine tissue. From the experimental results of WB, qRT-PCR, and immunofluorescence, the expression of TLR4 and the phosphorylation of NF-κB p65 were markedly inhibited by catalpol compared with the LPS group. The inflammatory damage to the mouse uterus caused by LPS was greatly reduced and was accompanied by a decline in myeloperoxidase (MPO) activity. The results of this study suggest that catalpol can exert an anti-inflammatory impact on LPS-induced bEECs and mouse endometritis by inhibiting inflammation and activation of the TLR4/NF-κB signaling pathway.
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@#AIM: To investigate the expression levels of serum miR-23a and miR-34a in patients with age-related macular degeneration(ARMD)and its relationship with the development of ARMD. <p>METHODS: Totally 102 patients with ARMD who were treated in our hospital from May 2015 to February 2018 were enrolled in the case group, and 70 healthy subjects in the same period were used as control group. The relative expression levels of miR-23a and miR-34a in serum were detected by RT-PCR, and the levels of serum tumor necrosis factor alpha(TNF-α)and nuclear factor kB(NF-kB)were detected by enzyme linked immunosorbent assay(ELISA). Analysis on the relationship of miR-23a, miR-34a expression levels with TNF-a, NF-kB in patients with ARMD and its diagnostic value for ARMD were taken.<p>RESULTS: The relative expression levels of serum miR-23a and miR-34a in the case group were significantly higher than those in the control group(<i>P</i><0.01). The relative expression levels of serum miR-23a and miR-34a in the advanced group were significantly higher than those in the middle and early stage(<i>P</i><0.01). The relative expression levels of serum miR-23a and miR-34a in the middle term patients were significantly higher than those in the early stage(<i>P</i><0.01). The serum levels of TNF- α and NF-κB in the case group were significantly higher than those in the control group(<i>P</i><0.01). There was a significant positive correlation between serum miR-23a and TNF-α and NF-kB in the case group(<i>r</i>=0.798, 0.720, both <i>P</i><0.01), and serum miR-34a was significantly positively correlated with TNF-α and NF-kB(<i>r</i>=0.814, 0.740, both <i>P</i><0.01). The area under the ROC curve(AUC)of serum miR-23a and miR-34a for diagnosis of ARMD was 0.831 and 0.867, respectively.<p>CONCLUSION: The expression of miR-23a and miR-34a in serum of ARMD patients is up-regulated, which may be involved in the development and progression of ARMD by promoting inflammation and oxidative stress. Detection of serum miR-23a and miR-34a may be helpful for early diagnosis and prevention of ARMD.
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Objective To observe the changes in the expression of RANK/RANKL in rat kidney treated by tripterygium wilfordii polyglucosides (TWP) in STZ induced type 2 diabetic kidney disease (DKD) and to explore its possible renoprotective mechanism.Methods T2DM animal model was established by high glucose and high fat diet plus intraperitoneal injection of STZ.The modeled rats were randomly divided into DKD group(DKD,n=8) and TWP treatment group(DT,n=8).Normal rats were taken as control group(NC,n=8).DT rats were lavaged with TWP in a dose of 50 mg/kg · d,while the NC group and DKD group were lavaged with equal volume of normal saline every day.The indicators of FPG,FIns,UAlb,BUN,Scr,and Ucr were measured before and after 12-week intervention.PAS staining was used to evaluate the pathological change of the kidney.Immunohistochemistry and Western-blot were used to observe the protein expressions of RANK,RANKL,and nephrin.Results Compared with NC group,kidney pathological changes of DN group were aggravated with higher levels of FPG,UAlb,Ccr and BUN at 12th week.The expressions of RANK[(0.27±0.05) vs (0.68±0.11)] and RANKL[(0.23± 0.07) vs (0.62±0.08)] were prominently increased in kidney in DN group than those in NC group,while the expressions of nephrin were decreased(P＜0.01).Compared with DKD group,the above indexes and renal pathological changes were improved in DT group.The expressions of RANK[(0.45 ± 0.09) vs (0.68±0.11)],and RANKL[(0.39±0.06) vs(0.62±0.08)],were markedly inhibited in DT group,while nephrin expressions were increased(P＜0.01).Conclusion TWP can protect the kidney in rats with DKD by inhibiting the expression of RANK/RANKL.
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Objective To investigate the effects of oxidative stress and lipoic acid(antioxidant)on bone metabolism and explore the underlying mechanism. Methods A total of 24 Wistar rats aged 8 weeks were randomly divided into three groups. Osteoporosis rats model was established by bilateral ovaries deleted. Rat in lipoic acid group was injected with lipoic acid(60 mg/kg)for 8 weeks. The bone mineral density(BMD),steo?calcin,ALP,Ca,P,MDA,SOD and GSH?Px were detected. The levels of OPG and RANKL in serum were measured by Western blotting. OPG and RANKL mRNA were detected by real?time PCR. Results The level of BMD level in blood,SOD,GSH?Px,OPG mRNA and protein level in femur of osteoporosis group were significantly lower than the control group(P<0.05). On the other hand,steocalcin,ALP,MDA,RANKL mRNA and protein level were significantly higher than the control group(P<0.05). The level of BMD level in blood,SOD,GSH?Px,OPG mRNA and protein level of lipoic acid group were significantly higher than the osteoporosis group(P<0.05). The steocalcin,ALP,MDA,RANKL mRNA and protein level were significantly lower than the osteoporosis group(P<0.05). Conclusion Oxidative stress may increase osteoporosis through the upregulation of OPG/RANKL pathway in rats ,and antioxidant lipoic acid can alleviate the progress of osteoporosis.
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Lesão renal por isquemia/reperfusão (I/R) e acidose metabólica (AM) são duascondições críticas que ocorrem frequentemente na prática clínica. O resultado dessacombinação pode ser prejudicial para os rins, mas esta questão não tem sidoexaustivamente estudada até hoje. O presente estudo avaliou em ratos a influênciado baixo pH sistêmico em vários parâmetros da função renal mediante lesão renalpor I/R. A acidose metabólica foi induzida em ratos Wistar machos através daingestão de cloreto de amônio (NH4CI) dissolvido na água de beber, iniciando 2 diasantes da indução de lesão renal por isquemia/reperfusão e mantida durante todo oestudo. Isquemia/reperfusão renal foi induzida por clampeamento bilateral dasartérias renais durante 45 min, seguido por 48 h de reperfusão. Ao final do estudo,foram obtidas amostras de sangue arterial, urina e tecido renal. Os animais foramdivididos em quatro grupos: controle (submetido à cirurgia sham, n = 8), I/R (n = 8),acidose metabólica (AM; solução de NH4CI 0,28 M + cirurgia sham, n = 6), e AM+I/R(solução de NH4CI 0,28 M + I/R, n = 9). Em comparação com grupo I/R, ratosAM+I/R apresentaram maior mortalidade (50% vs. 11%), redução significativa de pHsanguíneo (7,00 ± 0,04 vs. 7,35 ± 0,03), bicarbonato plasmático (pBic; 9,0 ± 1,4 vs.21,4 ± 0,9 mmol/L), e excesso de base (SBE; -23,8 ± 1,5 vs. -2,7 ± 0,9 mmol/L), comdeclínio no ritmo de filtração glomerular (0,05 ± 0,02 vs. 0,14 ± 0,03 mL/min/100 g) efunção tubular...
Ischemia/reperfusion (I/R) injury and metabolic acidosis (MA) are two criticalconditions that frequently occur in the clinical practice. The result of this combinationcan be harmful to the kidneys, but this issue has not been thoroughly investigatedhitherto. The present study evaluated the influence of low systemic pH on severalkidney function parameters in rats subjected to experimental model of renal I/Rinjury. Metabolic acidosis was induced in male Wistar rats by ingesting ammoniumchloride (NH4Cl) in tap water, beginning 2 days before ischemic insult and maintainedduring the entire study. Ischemia/reperfusion was induced by clamping both renalarteries for 45 min, followed by 48 h of reperfusion. At the end of the study, arterialblood samples and urine were collected and left kidneys were harvested. Fourgroups were studied: control (subjected to sham surgery, n = 8), I/R (n = 8),metabolic acidosis (MA; 0.28 M NH4Cl solution and sham surgery, n = 6), andMA+I/R (0.28 M NH4Cl solution plus I/R, n = 9). Compared with I/R rats, MA+I/R ratsexhibited higher mortality (50% vs. 11%), significant reduction of blood pH (7.00 ±0.04 vs. 7.35 ± 0.03), plasma bicarbonate (pBic; 9.0 ± 1.4 vs. 21.4 ± 0.9 mmol/L), andstandard base excess (SBE; -23.8 ± 1.5 vs. -2.7 ± 0.9 mmol/L), with a severe declinein the glomerular filtration rate (0.05 ± 0.02 vs. 0.14 ± 0.03 mL/min/100 g) and tubularfunction. In addition, tubular changes were more intense determining higher scores oftubular injury...
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Humains , Cétose , Heme oxygenase-1 , Facteur de transcription NF-kappa B , Atteinte rénale aigüeRÉSUMÉ
Objective To explore the changes in expression of Klotho, an aging-suppression protein, and neutrophil gelatinase associated lipocalin ( NGAL) and their relationship with rat mesangial cells ( RMCs) cultured with high glucose in vitro, and to explore the role played by Toll-like receptor-4 (TLR4) / nuclear factor-kB(NF-kB) p65 pathways in this process. Methods Three NGAL-siRNA sequences were designed and synthesized. The effective sequence in subsequent experiments was chosen. RMCs were preincubated with pyrrolidinedithiocarbamate (PDTC)or exogenously added Klotho prior to high glucose treatment. Realtime PCR was used to analyze Klotho, TLR4, NGAL mRNA expressions. Western blot was used to observe Klotho, TLR4,NF-kB p65, NGAL,fibronectin (FN), and connective tissue growth factor ( CTGF) protein expression. ELISA assay was used to detect monocyte chemoattractant protein-1 ( MCP-1) and CXCL5 secretions. Results High glucose suppressed Klotho expression significantly(P<0. 05) and activated TLR4 / NF-kB p65 pathway. Meanwhile,the levels of NGAL,FN,CTGF, MCP-1, and CXCL5 were highly expressed ( P < 0. 01). NGAL gene silencing obviously down-regulated the increased expressions of FN, CTGF, MCP-1, and CXCL5 ( P < 0. 01). After PDTC treatment the overexpression of NGAL protein was markedly lowered(P<0. 01). In addition, Klotho treatment significantly inhibited the activity of TLR4 /NF-kB p65 pathways and down-regulated the expressions of NGAL, FN, CTGF, MCP-1 and CXCL5 stimulated by high glucose(P<0. 01). Conclusion Klotho inhibits the activity of TLR4 / NF-kB p65 pathways and thus inhibits NGAL expression in RMCs cultured with high glucose in vitro. And then it suppresses the expressions of FN, CTGF, MCP-1, and CXCL5. This provides a new basis to illustrate the protection mechanism of the anti-aging protein Klotho in diabetic nephropathy, and may provide new ideas and therapeutic targets for prevention and treatment.
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Objective: To investigate the relationship between non-alcoholic fatty liver disease (NAFLD) and hepatocellular carcinoma (HCC), and to explore the role of nuclear factor-kappa B (NF-KB): and interleukin-6 (IL-6): in mechanism of hepatocarcinogenesis. Methods: Mice were divided into vacuity contrast group, normal diet group and high fat diet group. After feeding for 15 weeks, diethylnitrosamine (DEN, 45mg/kg): was weekly administrated by intraperitoneal injection into mice in both normal diet and high fat diet groups to gradually establish a model of HCC. The mice in blank control group were not treated with any drugs through the whole experiment. Livers and serum were individually collected from five mice in normal diet and high fat diet groups, respectively, and three mice in the blank control group in week 15, 20, 25 and 30. All the remaining mice were executed in week 36, and serum and liver pathologic gross were collected. The pathological changes of liver tissues were observed with HE staining. The differences in hepatocarcinogenesis among three groups were also analyzed. The levels of serum triglyceride (TC), cholesterol (TC), alanine aminotransferase (ALT): and aspartate aminotransferase (AST): were detected. The levels of serum NF-KB and IL-6 were assayed by ELISA. Results: Animal models of NAFLD were successfully established after 15 weeks of high-fat diet feeding. In the week 36, no mice in the blank control group developed HCC. Eight of 20 (40.0%): mice in normal diet group developed HCC. Thirteen of 18 (83.3%): mice in high fat diet group developed HCC. The levels of serum NF-KB and IL-6 in high fat diet group were significantly higher than those in the normal diet group in each period during study (all P < 0.01). Conclusion: The results suggest that NAFLD can promote hepatocarcinogenesis in the presence of DEN. The abnormal levels of NF-KB and IL-6 play important roles during the development of HCC.
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Diabetic rat model was established by peritoneal injection of streptozocin.At the end of 2 weeks,oxidized low-density lipoprotein (oxLDL) level in diabetic rats was raised [ ( 2.87 ± 0.40 vs 2.27 ± 0.36 ) μg/dl,P＜0.05 ] and endothelium-dependent relaxation was sluggish compared with normal rats.At the end of 6 weeks,oxLDL level continued to increase [ 4.32 ±0.66 ) μg/dl,P＜0.01] and endothelium-dependent maximum relaxation ( Rmax ) was decreased obviously ( P ＜0.01 ).Meanwhile,the protein and mRNA expressions of lectin-like oxidized lowdensity lipoprotein receptor-1 ( LOX-1 ),NF-kB,and ICAM-1 on vessel wall of diabetic rats were higher than those in normal rats,and LOX-1 mRNA was positively correlated with the levels of oxLDL,NF-kB,and ICAM-1 mRNA,while negatively correlated with Rmax,indicating that OxLDL/LOX-1 system may cause early endothelial dysfunction in diabetes via activating NF-kB and up-regulating ICAM-1 expression.
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[Objective]To investigate the effect of adenovirus-mediated glucose transporter 3 (Ad-GLUT3) infection on PC 12 cells of rats after hypoxia-reperfusion injury.[Methods]PC 12 cells treated with hypoxia and re--oxygenation were prepared;after PC12 cells being infected by Ad-GLUT3 (experimental group),adenovirus encoding green fluorescent protein (Ad-GFP group) and blank plasmid (control group),respectively,flow cytometry (FC) and TUNEL assay were used to analyze the apoptosis of PC12 cells;the glucose uptake rate was detected by isotopic-scintiscan,and the expression of nuclear factor (NF-kB) was detected by Western blotting.[Results] FC indicated that the apoptotic rate in the experimental group (4.7％±1.75％) was significantly lower as compared with that in the Ad-GFP group (14.25％±3.46％) and control group (13.1％±2.98％,P＜0.05);TUNEL assay showed that the apoptotic rate in the experimental group (7.7％±3.31％) was significantly lower as compared with that in the Ad-GFP group (17.1％±3.10％) and control group (15.2％±2.70％,P＜0.05).And the glucose uptake rate in the experimental group (603％±13.1％) was obviously higher as compared with that in the Ad-GFP group (300％±8.77％) and control group (319％±11.2％,P＜0.05);and the NF-k＜B level in the experimental group (0.92±0.17) was significantly lower as compared with that in the Ad-GFP group (1.02±0.24) and control group (1.06±0.21,P＜0.05).[Conclusion] After being infected by Ad-GLUT3,the PC12 cells treated with hypoxia and re-oxygenation enjoy elevated glucose uptake rate and inhibited apoptotic rate.
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OBJECTIVES: To investigate the relationship between single nucleotide polymorphism (SNP)s in Wnt antagonist genes, and production of osteoprotegerin (OPG) and soluble receptor activator of NF-kappaB ligand (sRANKL) by whole blood cells after hormone therapy (HT) in postmenopausal Korean women. MATERIALS AND METHODS: The Dkk1 c.318A>G, Dkk2 c.437G>A, Dkk3 c.1003A>G polymorphisms and sFRP3 c.970C>G, sFRP4 c.958C>A, and c.1019G>A polymorphisms, and sFRP5 c.20G>C polymorphisms were analyzed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), direct sequencing, and Taqman assay in 75 postmenopausal Korean women receiving estrogen-progestogen therapy. The production of OPG and sRANKL by lipopolysaccharide-stimulated whole blood cells (WBC) before and after HT of 6 months were also measured. RESULTS: Changes in the production of OPG and sRANKL by lipopolysaccharide-stimulated WBC, and in ratios of sRANKL(x1,000)/OPG after HT of 6 months were not different according to SNPs in Wnt signal pathway genes except Dkk1 c.318A>G SNP. The AA genotype of Dkk1 c.318A>G SNP showed significantly higher changes (pA, and c.1019G>A polymorphisms after HT. CONCLUSIONS: Dkk1 c.318A>G SNP are related with changes in ratios of sRANKL(x1,000)/OPG in terms of the production of OPG and sRANKL by lipopolysaccharide-stimulated whole blood cells after HT.