Objective To investigate the effects of adenosine 2A receptor (A2AR) activation on oxidative stress in small-forsize liver transplantation. Methods A rat orthotopic liver transplantation model was performed using 40% graft, 18 recipients were given intravenously saline (control group), CGS21680 (A2AR agonist, CGS21680 group) or ZM241385 (A2AR antagonist, CGS21680+ZM241385 group) randomly. Aspartate aminotransferase (AST), enzymatic antioxidants 〔superoxide dismutase (SOD); catalase (CAT); glutathione peroxidase (GSH-Px)〕, non-enzymatic antioxidants 〔ascorbic acid (AA); glutathione (GSH); α-tocopherol (TOC)〕 and lipid oxidant metabolites malondialdehyde (MDA) were measured and analyzed at 6 h after reperfusion. Results Compared with the control group and CGS21680+ZM241385 group, A2AR activation increased the activities of SOD and GSHPx (Plt;0.05), reduced the productions of AST and MDA (Plt;0.05), increased the levels of AA, GSH and TOC (Plt;0.05) in CGS21680 group. But there was no significant change in CAT activity (Pgt;0.05) among 3 groups. Conclusions A2AR activation improves the antioxidant enzyme activities, promotes the production of antioxidants, and slowes down the increase in MDA level, depresses of the increase in AST activity. A2AR activation suppresses oxidative damage and increases the antioxidant capacity which in turn minimizes their harmful effects of ischemia-reperfusion in small-for-size liver transplantation.
ObjectiveTo systematically review the clinical efficacy and effects on pregnancy outcomes of S-adenosy-L-methionine combined with ursodesoxycholic acid in the treatment of intrahepatic cholestasis of pregnancy. MethodsDatabases such as PubMed, The Cochrane Library, CNKI, VIP, WanFang Data were searched for the studies about the clinical efficacy and effects on pregnancy outcomes of S-adenosy-L-methionine combined with ursodesoxycholic acid in the treatment of intrahepatic cholestasis of pregnancy up to December 31st, 2013. Two reviewers independently screened literature, extracted data and evaluated methodological quality. Then meta-analysis was conducted using RevMan 5.0.24 software. ResultsA total of 11 RCTs involving 776 patients were included. The results of meta-analysis showed that, combined medication reduced blood biochemical indexes inlcuding ALT (MD=3.63, 95%CI 0.63 to 6.64, P=0.02), TB (MD=3.70, 95%CI 1.45 to 5.96, P=0.001), and AST (MD=7.61, 95%CI 2.47 to 12.75, P=0.004). Combined therapy significantly decreased the rates of amniotic fluid contamination (OR=0.29, 95%CI 0.19 to 0.45, P=0.000 01), cesarean section (OR=0.53, 95%CI 0.36 to 0.79, P=0.002), postpartum hemorrhage (OR=0.32, 95%CI 0.12 to 0.90, P=0.03), preterm birth (OR=0.36, 95%CI 0.24 to 0.55, P < 0.000 01), fetal distress (OR=0.33, 95%CI 0.19 to 0.58, P=0.000 1) and neonates asphyxia (OR=0.30, 95%CI 0.19 to 0.47, P < 0.000 01). Combined therapy was also beneficial to improving pruritus symptoms (MD=0.20, 95%CI 0.08 to 0.31, P=0.000 08) and benefiting fetus growth (MD=0.45, 95%CI 0.23 to 0.66, P < 0.000 1). ConclusionThe combination of S-adenosy-L-methionine and ursodesoxycholic acid is superior to ursodesoxycholic acid alone in improving clinical symptoms and pregnant outcomes of patients with intrahepatic cholestasis of pregnancy.
Objective To investigate the effects of wedelolactone (WEL) on lipopolysaccharide (LPS)-induced pyroptosis of alveolar epithelial cells and AMP-activated protein kinase/nucleotide binding oligomeric domain like receptor 3 (NLRP3)/cysteinyl aspartate specific proteinase-1 (Caspase-1) signaling pathway. Methods Human lung epithelial cells BEAS-2B were treated with 5 - 200 μmol/L wedelolactone, and cell activity was detected using MTT assay. The alveolar epithelial cells were divided into control group, lipopolysaccharide group (LPS group), 10 μmol/L wedelolactone group (WEL-L group), 20 μmol/L wedelolactone group (WEL-M group), 40 μmol/L wedelolactone group (WEL-H group), 40 μmol/L wedelolactone+10 μmol/L AMPK inhibitor Compound C group (WEL-H+Compound C group), and 20 μmol/L Caspase-1 inhibitor Z-YVAD-FMK group (Z-YVAD-FMK group). Transmission electron microscopy was applied to observe the microstructure of cells. ELISA was applied to detect levels of inflammatory factors such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-8 (IL-8). Immunofluorescence was applied to detect Caspase-1 and gasdermin family proteins (DGSDMD). Western blot was applied to detect protein expression levels of AMPK, NLRP3, and Caspase-1. Results Wedelolactone concentrations of 10, 20 and 40 μmol/L were selected for follow-up experiments. Compared with Control group, LPS group showed decreased cell activity, severe damage, cell contraction, mitochondrial ridge breakage and decreased number, increased levels of TNF-α, IL-1β, IL-8 and GSDMD, NLRP3, Caspase-1 expression, and decreased p-AMPK/AMPK expression (P<0.05). Wedelolactone treatment could significantly improve LPS-induced pyrosis of alveolar epithelial cells (P<0.05). Compound C could partially reverse the effect of wedelactone on LPS-induced pyrodeath of alveolar epithelial cells (P<0.05). Z-YVAD-FMK treatment also significantly improved LPS-induced pyroptosis of alveolar epithelial cells (P<0.05). Conclusion Wedelolactone can inhibit LPS-induced pyroptosis of pulmonary alveolar epithelial cells by inhibiting AMPK/NLRP3/Caspase-1 signaling pathway.
Objective To investigate the mechanism of AMP-activated protein kinase (AMPK) in hepatic ischemia-reperfusion injury (HIRI). Methods ① Grouping. Forty-two mice were randomly divided into Sham group, 4 ischemia reperfusion (IR) group of different times (2, 6, 12, and 24 h), Compound C group, and Compound C+repamycin (Rapa) group, each group enrolled in 6 mice. Compound C group: mice were modeled at 1 h after intraperitoneal injection of Compound C (25 mg/kg). Compound C+Rapa group: mice were modeled at 1 h after intraperitoneal injection of rapamycin (1 mg/kg) and Compound C (25 mg/kg). Mice of 4 IR groups, Compound C group, and Compound C+Rapa group were used to prepare HIRI model. Mice of Sham group were treated only for laparotomy, freeing the first portal hepatis and closing peritoneal. ② To filter the best IR time. The levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in the serum of mice in Sham group and IR groups of 4 different reperfusion time points were measured. The pathological changes of liver tissues were observed by HE staining, and the expressions of related proteins in liver tissue of mice were detected by Western blot. Considering the results of blood biochemical test, HE staining, and Western blot together to determine the best IR point. ③ The exploration of signal pathway for AMPK. The expressions of proliferating cell nuclear antigen (PCNA) were observed by immunohistochemical staining in the liver tissues of IR-12 h group, Compound C group (12 h after IR) and compound C+Rapa group (12 h after IR). The mitochondrial damage was observed by rhodamine 123 staining, and the apoptotic status of liver cells was detected by terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay (TUNEL). Results ① The 12 h after IR was the best observation time point. Compared with IR-12 h group, the levels of ALT and AST in Sham group, IR-2, 6, and 24 h groups were lower (P<0.05). HE staining showed that liver tissue destruction in IR-12 h group was the most severe. Western blot showed that, expressions of AMPKα, phosphorylated adenylate activated protein kinase α (p-AMPKα), Nip3-like protein X (Nix), BCL-2 homologous water-soluble protein (Bax), as well as ratio of autophagy microtubule-associated protein light chain 3 (LC3)Ⅱto LC3Ⅰof Sham group, IR-2, 6, and 24 h group were all lower than those of IR-12 h group (P<0.05), but the expressions of phosphorylated mammalian target of Rapa (p-mTOR) of Sham group, IR-2, 6, and 24 h group were all higher (P<0.05). Therefore, 12 h after IR was the best time to observe. ② Compared with IR-12 h group, the expression level of PCNA protein in liver tissue of Compound C group was lower (P<0.05), the mitochondrial luminescence intensity was weaker and the apoptotic cells were more. Compared with Compound C group, the expression of PCNA protein in the liver tissue of the Compound C+Rapa group was higher (P<0.05), the mitochondrial intensity was stronger and the apoptotic cells were less. ③ Compared with IR-12 h group, the expressions of Nix and p-AMPKα, and ratio of LC3Ⅱ to LC3Ⅰ in liver tissue of Compound C group decreased (P<0.05), while the expressions of p-mTOR, Caspase-3, and Cleaved Caspase-3 increased (P<0.05). Compared with Compound C group, the expressions of p-AMPKα and Nix in the liver tissue of Compound C+Rapa group increased (P<0.05), while the expressions of p-mTOR, Caspase-3, and Cleaved Caspase-3 decreased (P<0.05). Conclusion During the HIRI in mouse, AMPK regulates mitophagy and apoptosis through the mTOR/Nix pathway.
The Dacron grafts seeded with autologous venous fragments were implanted into IVC of 13 canines as seeded group and the control grafts (8 cases), which were only preclotted with fresh blood. The amounts of cAMP and cGMP in serum and within platelet were measured. All of the specimens explanted at exsaguination were observed morphologically. The results shown that the total patency rate were 61.5% in seeded group, but 25.0% in control one and new endothelial lining formed at two weeks after implantation of the seeded grafts. The amounts of cAMP in serum and within platelet were higher in seeded group, but the amounts of cGMP were lower in serum and within platelet. These were in accordance with the results that the endothelialization of the grafts were complete in seeded group but not complete in control one. The results indicate that seeding Dacron with autologous venous fragment makes new endothelium formed at two weeks after implantation, increases the amounts of cAMP in serum and within platelet, but reduces the amounts of cGMP and thus improves graft patency rate.
Objective To evaluate the inhibiting effect of adenosine on rat retinal ganglion cells (RGC) death induced by P2X7 and N-methyl-D-aspartate (NMDA) receptor. Methods (1) Long-Evan neonatal rats were back labeled with aminostilbamidine to identify RGC. The viability of RGC affected by P2X7 excitomotor BzATP (50 mu;mol/L), glutamate receptor excitomotor NMDA (100 mu;mol/L) and adenosine (300 mu;mol/L) was detected. (2) RGC from the retinae of unlabeled neonatal rats were cultured in vitro. After labeled with Fura-2 methyl acetate, an intracellular calcium indicator, the effect of BzATP, NMDA and adenosine on intracellular Ca2+ level was detected byCa2+ imaging system. Results Both BzATP (50 mu;mol/L) and NMDA(100 mu;mol/L) could kill about 30% of the RGC. Cell death was prevented by adenosine (300 mu;mol/L) with the cell viability increased from (68.9plusmn;2.3)% and (69.9plusmn;3.2)% to (91.2plusmn;3.5)% (P＜0.001) and (102.1plusmn;3.9)% (P＜0.001), respectively. BzATP (50 mu;mol/L) led to a large, sustained increase of intracellular Ca2+ concentration to (1183plusmn;109) nmol/L. After the adenosine intervened, Ca2+ concentration increased slightly to (314plusmn;64) nmol/L (P＜0.001). Conclusion Adenosine may prevent RGC death and increase of intracellular Ca2+ concentration from P2X7and NMDA receptor stimulation. (Chin J Ocul Fundus Dis, 2007, 23: 133-136)
Objective To assess the efficacy and safety of S-adenosyl-l-methionine (SAMe) for outcome improvement of intrahepatic cholestasis of pregnancy. Methods Randomized controlled trials (RCT) and quasi-randomized controlled trials were identified from MEDLINE (1983 to 2003), The Cochrane Library (Issue 4,2003), EMBASE (1980 to 2003), China Hospital Digital Library (CHDL) and Wanfang data (1994 to 2003). We also handsearched the relative references. Two researchers evaluated the quality of the trials and extracted the data independently. RevMan software 4.2 was used for meta-analysis. Results Eight studies involving 424 pregnant women were included. The following data were the results of meta-analysis of SAMe for improvements: ① Reducing cesarean-section ratio: no significant difference was seen between SAMe and placebo groups with OR 1.00, 95%CI 0.23 to 4.33 and P= 1.00; significant differences were seen SAMe versus dexamethasone and SAMe versus Dianglining with OR 0.44, 95%CI 0.23 to 0.85 and P=0.01; OR 0.28 95%CI 0.10 to 0.75 and P=0.01 respectively。② Prolonging the period of pregnancy: SAMe had no significant difference compared with placebo groups with WMD=0.70, 95%CI －0.69 to 2.10, P=0.32. SAMe was more effective than dexamethasone, Ganyinling and Qianglining on prolonging the period of pregnancy with WMD=1.10，95%CI 0.46 to 1.74, P=0.000 07; WMD=2.50，95%CI 1.86 to 3.14, P≤0.000 01; WMD=2.20，95%CI 1.61 to 2.79, P≤0.000 01 respectively；③ Increasing the weight of the newborn: meta-analysis showed that SAMe group had not significant difference compared with placebo group on increasing the weight of the newborn with WMD=-26.27，95%CI -338.35 to 285.82, P=0.87. Significant differences were seen between SAMe and dexamethasone, SAMe and Ganyiling, SAMe and Qiangling with WMD=386.86，95%CI 134.41 to 603.31, P=0.002; WMD=410.00,95%CI 321.10 to 498.90, P≤0.000 01 respectively. ④ Fetal distress: There was no significant difference compared with dexamethasone and Kuhuang groups on decreasing the fetal distress with OR=0.47, 95%CI 0.14 to 1.16, P=0.23; OR=0.44, 95%CI 0.10 to 1.97, P=0.29 respectively; ⑤ Decreasing pollution of amniotic fluid: no significant differences were seen in SAMe versus dexamethasone, SAMe versus ursoddeoxycholic and SAMe versus Kuhuang with OR=0.46, 95%CI 0.21 to 1.02, P=0.06; OR=0.68, 95%CI 0.20 to 2.31, P=0.53; OR=0.82 95%CI 0.24 to 2.81,P=0.75 recpectively. ⑥ Newborn stifile: SAMe group had no significant difference compared with dexamethasone and Kuhuang groups on decreasing the Newborn stifile with OR=0.19, 95%CI 0.01 to 4.06, P=0.29; OR=0.31, 95%CI 0.08 to 1.13, P=0.08 respectively. Compared with Qianglining group, SAMe group had better effect on reducing ratio of newborn stifile with OR=0.09, 95%CI 0.02 to 0.42, P=0.002. ⑦ Improving Apgar scores: no significant differences were seen between SAMe and placebo, dexamethasone and ursoddeoxycholic with OR=0.25, 95%CI 0.02 to 3.04, P=0.28; OR=2.09, 95%CI 0.70 to 6.27, P=0.19; OR=1.22, 95%CI 0.35 to 4.19, P=0.75 respectively. Six RCTs mentioned the side effects of S-adenosy-l-methionine, only one RCT reported mild gastrointestinal irritation. Conclusions SAMe is partly effective on improving the pregnancy outcomes of intrahepatic choletasis of pregnancy, such as reducting cesarean-section ratio, prolonging the period of pregnancy and increasing the weight of the newborn. The specified efficacy and safety of SAMe require rigorously designed, randomized, double-blind and placebo-controlled trials to offer evidence.
Objective To investigate the effects of sodium valproate (VPA) in inhibiting Erastin-induced ferroptosis in bone marrow mesenchymal stem cells (BMSCs) and its underlying mechanisms. Methods BMSCs were isolated from bone marrow of 8-week-old Spragur Dawley rats and identified [cell surface antigens CD90, CD44, and CD45 were analyzed by flow cytometry, and osteogenic and adipogenic differentiation abilities were assessed by alizarin red S (ARS) and oil red O staining, respectively]. Cells of passage 3 were used for the Erastin-induced ferroptosis model, with different concentrations of VPA for intervention. The optimal drug concentration was determined using the cell counting kit 8 assay. The experiment was divided into 4 groups: group A, cells were cultured in osteogenic induction medium for 24 hours; group B, cells were cultured in osteogenic induction medium containing optimal concentration Erastin for 24 hours; group C, cells were cultured in osteogenic induction medium containing optimal concentration Erastin and VPA for 24 hours; group D, cells were cultured in osteogenic induction medium containing optimal concentration Erastin and VPA, and 8 μmol/L EX527 for 24 hours. The mitochondrial state of the cells was evaluated, including the levels of malondialdehyde (MDA), glutathione (GSH), and reactive oxygen species (ROS). Osteogenic capacity was assessed by alkaline phosphatase (ALP) activity and ARS staining. Western blot analysis was performed to detect the expressions of osteogenic-related proteins [Runt-related transcription factor 2 (RUNX2) and osteopontin (OPN)], ferroptosis-related proteins [glutathione peroxidase 4 (GPX4), ferritin heavy chain 1 (FTH1), and solute carrier family 7 member 11 (SLC7A11)], and pathway-related proteins [adenosine monophosphate-activated protein kinase (AMPK) and Sirtuin 1 (SIRT1)]. Results The cultured cells were identified as BMSCs. VPA inhibited Erastin-induced ferroptosis and the decline of osteogenic ability in BMSCs, acting through the activation of the AMPK/SIRT1 pathway. VPA significantly reduced the levels of ROS and MDA in Erastin-treated BMSCs and significantly increased GSH levels. Additionally, the expression levels of ferroptosis-related proteins (GPX4, FTH1, and SLC7A11) significantly decreased. VPA also upregulated the expressions of osteogenic-related proteins (RUNX2 and OPN), enhanced mineralization and osteogenic differentiation, and increased the expressions of pathway-related proteins (AMPK and SIRT1). These effects could be reversed by the SIRT1 inhibitor EX527. ConclusionVPA inhibits ferroptosis in BMSCs through the AMPK/SIRT1 axis and promotes osteogenesis.
Adenosine activated protein kinase (AMPK) is a serine/threonine protein kinase that can sense the change of intracellular energy. AMPK plays a critical part in the occurrence and development of tumors. According to the difference of AMPK catalytic subunits, it is divided into AMPKα1 and AMPKα2. The AMPKα1 subunit is the catalytic subunit of AMPK and is extensively distributed in the various tissues and organs. This review focuses on the structural, activated and functional aspects of AMPKα1 and the involvement of AMPKα1 in the regulation of intracellular substance metabolism, and summarizes the respective performances of AMPKα1 in different cancers and the corresponding potential applications of AMPKα1 as a drug target in the relevant cancers. AMPKα1 can be used as a diagnostic marker or drug target for cancer diagnosis and therapy, providing an idea for cancer treatment, which has importance clinical significance.