Metformin promotes angiogenesis and functional recovery in aged mice after spinal cord injury by adenosine monophosphate-activated protein kinase/endothelial nitric oxide synthase pathway

Treatment with metformin can lead to the recovery of pleiotropic biological activities after spinal cord injury.However,its effect on spinal cord injury in aged mice remains unclear.Considering the essential role of angiogenesis during the regeneration process,we hypothesized that metformin activates the adenosine monophosphate-activated protein kinase/endothelial nitric oxide synthase pathway in endothelial cells,thereby promoting microvascular regeneration in aged mice after spinal cord injury.In this study,we established young and aged mouse models of contusive spinal cord injury using a modified Allen method.We found that aging hindered the recovery of neurological function and the formation of blood vessels in the spinal cord.Treatment with metformin promoted spinal cord microvascular endothelial cell migration and blood vessel formation in vitro.Furthermore,intraperitoneal injection of metformin in an in vivo model promoted endothelial cell proliferation and increased the density of new blood vessels in the spinal cord,thereby improving neurological function.The role of metformin was reversed by compound C,an adenosine monophosphate-activated protein kinase inhibitor,both in vivo and in vitro,suggesting that the adenosine monophosphate-activated protein kinase/endothelial nitric oxide synthase pathway likely regulates metformin-mediated angiogenesis after spinal cord injury.These findings suggest that metformin promotes vascular regeneration in the injured spinal cord by activating the adenosine monophosphate-activated protein kinase/endothelial nitric oxide synthase pathway,thereby improving the neurological function of aged mice after spinal cord injury.

Electroacupuncture preconditioning attenuates ischemic brain injury by activation of the adenosine monophosphate-activated protein kinase signaling pathway

Electroacupuncture has therapeutic effects on ischemic brain injury, but its mechanism is still poorly understood. In this study, mice were stimulated by electroacupuncture at the Baihui（GV20） acupoint for 30 minutes at 1 m A and 2/15 Hz for 5 consecutive days. A cerebral ischemia model was established by ligating the bilateral common carotid artery for 15 minutes. At 72 hours after injury, neuronal injury in the mouse hippocampus had lessened, and the number of terminal deoxynucleotide transferase-mediated d UTP nick-end labeling-positive cells reduced after electroacupuncture treatment. Moreover, expression of adenosine monophosphate-activated protein kinase α（AMPKα） and phosphorylated AMPKα was up-regulated. Intraperitoneal injection of the AMPK antagonist, compound C, suppressed this phenomenon. Our findings suggest that electroacupuncture preconditioning alleviates ischemic brain injury via AMPK activation.

AB015.Metabolic stress in glaucoma engages early activation of the energy biosensor adenosine monophosphate-activated protein kinase leading to neuronal dysfunction

Background:Metabolic stress has been proposed to contribute to neuronal damage in glaucoma,but the mechanism driving this response is not understood.The adenosine monophosphate-activated protein kinase(AMPK)is a master regulator of energy homeostasis that becomes active at the onset of energy stress.AMPK is a potent inhibitor of the mammalian target of rapamycin complex 1(mTORC1),which we showed is essential for the maintenance of retinal ganglion cell(RGC)dendrites,synapses,and survival.Here,we tested the hypothesis that AMPK is an early mediator of metabolic stress in glaucoma.Methods:Unilateral elevation of intraocular pressure was induced by injection of magnetic microbeads into the anterior chamber of mice expressing yellow fluorescent protein in RGCs.Inhibition of AMPK was achieved by administration of siRNA or compound C.RGC dendritic trees were 3D-reconstructed and analyzed with Imaris(Bitplane),and survival was assessed by counting Brn3a or RBPMS-labeled soma and axons in the optic nerve.RGC function was examined by quantification of anterograde axonal transport after intraocular administration of cholera toxinβ-subunit.Retinas from glaucoma patients were analyzed for expression of active AMPK.Results:Ocular hypertension triggered rapid upregulation of AMPK activity in RGCs concomitant with loss of mTORC1 function.AMPK inhibition with compound C or siRNA effectively restored mTORC1 activity and promoted an increase in total dendritic length,surface and complexity relative to control retinas.Attenuation of AMPK activity led to robust RGC soma and axon survival.For example,95%of RGCs(2,983±258 RGCs/mm2,mean±S.E.M.)survived with compound C compared to 77%in vehicle-treated eyes(2,430±233 RGCs/mm2)(ANOVA,P<0.001)at three weeks after glaucoma induction(n=8-10/group).Importantly,blockade of AMPK activity effectively restored anterograde axonal transport.Lastly,RGC-specific upregulation of AMPK activity was detected in human glaucomatous retinas relative to age-matched controls(n=10/group).Conclusions:Metabolic stress in glaucoma involves AMPK activation and mTORC1 inhibition promoting early RGC dendritic pathology,dysfunction and neurodegeneration.

Metformin attenuates motility,contraction,and fibrogenic response of hepatic stellate cells in vivo and in vitro by activating AMP-activated protein kinase

AIM To investigate the effect of metformin on activated hepatic stellate cells(HSCs) and the possible signaling pathways involved. METHODS A fibrotic mouse model was generated by intraperitoneal injection of carbon tetrachloride(CCl_4) and subsequent treatment with or without metformin. The level of fibrosis was detected by hematoxylin-eosin staining, Sirius Red staining, and immunohistochemistry. The HSC cell line LX-2 was used for in vitro studies. The effect of metformin on cell proliferation(CCK8 assay),motility(scratch test and Transwell assay), contraction(collagen gel contraction assay), extracellular matrix(ECM) secretion(Western blot), and angiogenesis(ELISA and tube formation assay) was investigated. We also analyzed the possible signaling pathways involved by Western blot analysis.RESULTS Mice developed marked liver fibrosis after intraperitoneal injection with CCl_4 for 6 wk. Metformin decreased the activation of HSCs, reduced the deposition of ECM, and inhibited angiogenesis in CCl_4-treated mice. Platelet-derived growth factor(PDGF) promoted the fibrogenic response of HSCs in vitro, while metformin inhibited the activation, proliferation, migration, and contraction of HSCs, and reduced the secretion of ECM. Metformin decreased the expression of vascular endothelial growth factor(VEGF) in HSCs through inhibition of hypoxia inducible factor(HIF)-1α in both PDGF-BB treatment and hypoxic conditions, and it down-regulated VEGF secretion by HSCs and inhibited HSC-based angiogenesis in hypoxic conditions in vitro. The inhibitory effects of metformin on activated HSCs were mediated by inhibiting the Akt/mammalian target of rapamycin(m TOR) and extracellular signal-regulated kinase(ERK) pathways via the activation of adenosine monophosphate-activated protein kinase(AMPK).CONCLUSION Metformin attenuates the fibrogenic response of HSCs in vivo and in vitro, and may therefore be useful for the treatment of chronic liver diseases.

The potential of herbal drugs to treat heart failure:The roles of Sirt1/AMPK

Heart failure(HF)is a highly morbid syndrome that seriously affects the physical and mental health of patients and generates an enormous socio-economic burden.In addition to cardiac myocyte oxidative stress and apoptosis,which are considered mechanisms for the development of HF,alterations in cardiac energy metabolism and pathological autophagy also contribute to cardiac abnormalities and ultimately HF.Silent information regulator 1(Sirt1)and adenosine monophosphate-activated protein kinase(AMPK)are nicotinamide adenine dinucleotide(NAD+)-dependent deacetylases and phosphorylated kinases,respectively.They play similar roles in regulating some pathological processes of the heart through regulating targets such as peroxisome proliferator-activated receptorγcoactivator 1α(PGC-1α),protein 38 mitogen-activated protein kinase(p38 MAPK),peroxisome proliferator-activated receptors(PPARs),and mammalian target of rapamycin(mTOR).We summarized the synergistic effects of Sirt1 and AMPK in the heart,and listed the traditional Chinese medicine(TCM)that exhibit cardioprotective properties by modulating the Sirt1/AMPK pathway,to provide a basis for the development of Sirt1/AMPK activators or inhibitors for the treatment of HF and other cardiovascular diseases(CVDs).

Beneficial effects of metformin on primary cardiomyocytes via activation of adenosine monophosphate-activated protein kinase

Background Metformin has become a cornerstone in the treatment of patients with type-2 diabetes. Accumulated evidence suggests that metformin supports direct cardiovascular effects. The present study aimed to investigate if metformin has beneficial effects on primary cardiomyocytes damaged by H2O2, and reveal the potential mechanism of action of metformin. Methods Cardiomyocytes were incubated in the presence of 100μmol/L H2O2 for 12 hours. Cardiomyocytes were pretreated with metformin at different concentrations and time and with aminoimidazole carboxamide ribonucleotide （AICAR） （500μmol/L）, an adenosine monophophate （AMP）-activated protein kinase （AMPK） agonist for 60 minutes before the addition of H2O2. Other cells were preincubated with compound C （an AMPK antagonist, 20μmol/L） for 4 hours. The viability and apoptosis of cells were analyzed. AMPK, endothelial nitric oxide synthase （eNOS）, and transforming growth factor （TGF）-β1 were analyzed using immunblotting. Results Metformin had antagonistic effects on the influences of H2O2 on cell viability and attenuated oxidative stress-induced apoptosis. Metformin also increased phosphorylation of AMPK and eNOS, and reduced the expression of TGF-β1, basic fibroblast growth factor （bFGF）, and tumor necrosis factor （TNF）-α. Conclusions Metformin has beneficial effects on cardiomyocytes, and this effect involves activation of the AMPK-eNOS pathway. Metformin may be potentially beneficial for the treatment of heart disease.

Metformin inhibits nuclear factor-κB activation and inflammatory cytokines expression induced by high glucose via adenosine monophosphate-activated protein kinase activation in rat glomerular mesangial cells in vitro

Background The renoprotective mechanisms of adenosine monophosphate （AMP）-activated protein kinase （AMPK） agonist-metformin have not been stated clearly.We hypothesized that metformin may ameliorate inflammation via AMPK interaction with critical inflammatory cytokines The aim of this study was to observe the effects of metformin on expression of nuclear factor-κB （NF-κB）,monocyte chemoattractant protein-1 （MCP-1）,intercellular adhesion molecule-1 （ICAM-1） and transforming growth factor-beta 1 （TGF-β1） induced by high glucose （HG） in cultured rat glomerular mesangial cells （MCs）.Methods MCs were cultured in the medium with normal concentration glucose （group NG,5.6 mmol/L）,high concentration glucose （group HG,25 mmol/L） and different concentrations of metformin （group M1,M2,M3）.After 48-hour exposure,the supernatants and MCs were collected.The expression of NF-κB,MCP-1,ICAM-1,and TGF-β1 mRNA was analyzed by real time polymerase chain reaction.Westem blotting was used to detect the expression of AMPK,phospho-Thr-172 AMPK （p-AMPK）,NF-κB p65,MCP-1,ICAM-1,and TGF-β1 protein.Results After stimulated by HG,the expression of NF-κB,MCP-1,ICAM-1,TGF-β1 mRNA and protein of MCs in group HG increased significantly compared with group NG （P ＜0.05）.Both genes and protein expression of NF-κB,MCP-1,ICAM-1,TGF-β1 of MCs induced by high glucose were markedly reduced after metformin treatment in a dose-dependent manner （P ＜0.05）.The expression of p-AMPK increased with the rising of metformin concentration,presenting the opposite trend,while the level of total-AMPK protein was unchanged with exposure to HG or metformin.Conlusion Metformin can suppress the expression of NF-κB,MCP-1,ICAM-1 and TGF-β1 of glomerular MCs induced by high glucose via AMPK activation,which may partlv contribute to its reno-protection.

Adenosine Monophosphate-Activated Protein Kinase,Oxidative Stress,and Diabetic Endothelial Dysfunction

Endothelial dysfunction characterized by impaired endothelium-dependent vaso-relaxation is one of the earliest detectable pathological events in smoking,diabetes,and many cardiovascular diseases including hypertension,atherosclerosis.Overwhelming data from human and animals demonstrate that the endothelial dysfunction associated with diabetes is due to the local formation of oxidants and free radicals.However,the mechanisms by which diabetes instigates oxidative stress,and those by which oxidative stress perpetuates endothelial dysfunction are the subjects of intensive research in the last 3 decades.The studies from us and others have demonstrated that adenosine monophosphate-activated protein kinase(AMPK),a well-characterized energy sensor and modulator,serves as a highly efficient sensor as AMPK can be activated by very low levels of reactive oxygen species(ROS)and reactive nitrogen species(RNS)generated by physiological,pharmacological,and pathologic stimuli(redox sensor).Interestingly,oxidants-activated AMPK feedback lowers the levels of ROS by either suppressing ROS/RNS from reduced nicotinamide adenine dinucleotide phosphate(NADPH)oxidase and mitochondria or by increasing the levels of antioxidant enzymes(redox modulator).Further,our studies demonstrate that AMPK’s functions as a redox sensor and modulator are vital to maintain endothelial cell function under physiological conditions.Finally,we discover that under chronic oxidative stress or large influx of ROS,AMPK is particularly susceptible to inhibition by ROS.We conclude that oxidative inactivation of AMPK in diabetes perpetuates oxidative stress and accelerates atherosclerosis in diabetes.

LncRNA PFL通过AMPK-PPARα信号通路改善心肌纤维化

目的研究促纤维化长链非编码RNA(LncRNA PFL)改善压力负荷诱导的心肌纤维化同腺苷酸活化蛋白激酶(AMPK)-过氧化物酶增殖激活的α亚型受体(PPAR)α信号通路的激活是否相关。方法将60只大鼠随机分为A(假手术)组、B(压力负荷诱导的心肌纤维化模型)组、C(压力负荷诱导的心肌纤维化模型+LncRNA PFL抑制剂)组。采用苏木素-伊红(HE)染色检测心肌组织的病理形态和纤维化程度,羟脯氨酸(HYP)检测心肌质量,Western印迹测定心肌组织中AMPK、PPARα蛋白水平。结果与A组比较,B组和C组心脏重量指数(HWI)和左心室重量指数(LVWI)均显著升高(P<0.01);与B组比较,C组显著下降(P<0.01)。HE染色结果:与A组比较,B组和C组心肌细胞的炎症浸润和细胞间胶原纤维均显著增加,神经元细胞损伤程度加重(P<0.05);与B相比,C组显著好转(P<0.05)。免疫荧光结果:B组AMPK及PPARα蛋白水平明显低于A组和C组,且C组明显低于A组。RT-qPCR及Western印迹结果:B组心肌组织中AMPK和PPARαmRNA及蛋白表达显著低于A组和C组,且C组显著高于A组(P<0.05)。结论LncRNA PFL表达下调改善压力负荷诱导的心肌纤维化与激活AMPK-α信号通路有关。

橙花叔醇对脓毒症急性肺损伤大鼠炎症反应及AMPK/SIRT1通路的影响

【目的】探讨橙花叔醇对脓毒症急性肺损伤大鼠的治疗作用及机制。【方法】采用脂多糖(LPS)股静脉注射法构建脓毒症急性肺损伤大鼠模型,应用橙花叔醇进行预防性治疗。测试用力呼出25%~75%肺活量时的呼气流量(FEF25%-75%)和20毫秒用力呼气量/用力肺活量(FEV20/FVC)比值评估大鼠肺功能。收集大鼠支气管肺泡灌洗液(BALF)和肺组织,采用苏木素-伊红(HE)染色法观察肺组织病理形态,应用血细胞计数器计数BALF中总白细胞数和中性粒细胞数,采用酶联免疫吸附分析(ELISA)检测BALF中炎症因子白细胞介素(IL)-1、IL-10和肿瘤坏死因子(TNF)-α水平,采用Western Blot法检测肺组织单磷酸腺苷活化蛋白激酶(AMPK)/SIRT1通路相关蛋白表达。继而给予AMPK抑制剂Dorsomorphin进行干预,进一步观察橙花叔醇通过激活AMPK/SIRT1通路对LPS诱导的脓毒症急性肺损伤大鼠的影响。【结果】橙花叔醇可减轻LPS诱导的脓毒症急性肺损伤大鼠肺组织病理及肺功能损伤,降低BALF中总白细胞数、中性粒细胞数及炎症因子IL-1和TNF-α水平,提高BALF中IL-10水平,上调肺组织磷酸化AMPK(p-AMPK)、SIRT1蛋白表达水平。【结论】橙花叔醇可通过激活AMPK/SIRT1通路及减轻炎症反应改善脓毒症急性肺损伤大鼠。

加味当归补血汤对糖尿病肾病大鼠AMPK及PGC-1α的影响及相关作用机制

目的:观察加味当归补血汤(MDBT)对糖尿病肾病(DKD)大鼠单磷酸腺苷激活蛋白激酶(AMPK)及过氧化物酶体增殖物激活受体γ辅激活因子1α(PGC-1α)活性的影响,探讨其治疗DKD的可能机制。方法:52只SD雄性大鼠随机分为正常组(CON)8只和造模组44只。后随机将40只成模大鼠分为模型组(MOD)、厄贝沙坦组(IRB)、加味当归补血汤高剂量组(MDBTH)、加味当归补血汤中剂量组(MDBTM)、加味当归补血汤低剂量组(MDBTL),8只/组。药物组灌相应药物,CON组予等体积生理盐水,1次/d,持续20周。检测各组大鼠24 h尿蛋白(24 h-UTP)水平、血清锰超氧化物歧化酶(MnSOD)及丙二醛(MDA)活性;观察大鼠肾组织病理变化;免疫组化法(IHC)及Western blot法检测肾组织磷酸化AMPK(p-AMPK)、PGC-1α蛋白表达。结果:与CON组比较,MOD组24 h-UTP及血清MDA水平显著升高,MnSOD活性显著降低(P<0.01);病理表现为肾小管和肾小囊严重分离,肾小球内基底膜均匀性增厚,球内糖原沉积;肾组织中p-AMPK、PGC-1α蛋白表达水平显著降低(P<0.01)。与MOD组比较,MDBTH与IRB组24 h-UTP及血清MDA水平显著下降,MnSOD活性显著提高(P<0.01);肾组织病理表现明显改善;p-AMPK、PGC-1α在肾组织的表达显著增加(P<0.01)。结论:加味当归补血汤可能通过激活AMPK及PGC-1α的表达改善DKD大鼠氧化应激,降低肾脏病理损害程度,减少蛋白尿,从而有效保护肾脏,缓解DKD进展。

Effects of Ginsenoside Rb1 on Skeletal Muscle Insulin Resistance and Adenosine Monophosphate?activated Protein Kinase Signaling Pathway in Obese Mice

Objectives: The objective of the study is to observe the effects of ginsenoside Rb1 on indexes of body weight, body composition, blood lipid, skeletal muscle endurance, and insulin sensitivity in obese mice, probe into its pharmacological action, and further explore its effects on adenosine monophosphate-activated protein kinase(AMPK) signaling pathway in skeletal muscle. Materials and Methods: Eight-week-old C57 BL/6 J mice were fed with high-fat diet for 12 weeks to establish obese mouse model. The model-establishment obese mice were randomly divided into three groups including model control group, metformin group, and ginsenoside Rb1 group. In the normal control group, normal diet was administered. The intervention period was 8 weeks. Body weight and food intake of the mice were measured regularly every week. The treadmill test was performed at weeks 3 and 7, and the oral glucose tolerance test was carried out at weeks 4 and 8. Body composition of the mice was detected by applying NMR Animal Body Composition Analyzer at week 8. Four parameters of blood lipids and free fatty acid(FFA)levels were detected. The m RNA expression of AMPKα and proliferator-activated receptor gamma coactivator-1α(PGC-1α) in skeletal muscle was examined by real-time fluorescence quantitative polymerase chain reaction, and the influence of ginsenoside Rb1 on protein expression of AMPKα, p-AMPKα, and PGC-1α was observed by western blotting. Results: The body weight(since the 5 th week of drug administration)and food intake of the mice in the ginsenoside Rb1 group were significantly lower than those in the model control group(P < 0.05) in a time-dependent manner. Ginsenoside Rb1 could significantly reduce the levels of triglyceride and low-density lipoprotein cholesterol, while increase the high-density lipoprotein cholesterol level(P < 0.05). In addition, ginsenoside Rb1 could reduce the serum FFA level(P < 0.05).After the administration of ginsenoside Rb1 for 8 weeks, the body fat mass of obese mice decreased and the lean mass increased(P < 0.05).The skeletal muscle endurance and the oral glucose tolerance of the obese mice improved using ginsenoside Rb1. At the molecular level,ginsenoside Rb1 could up-regulate the mRNA and protein expression of AMPKα in skeletal muscle, and increase the content of p-AMPK protein significantly(P < 0.01). At the same time, the mRNA and protein level of PGC-1α was also un-regulated, correspondingly(P < 0.01).Conclusion: Ginsenoside Rb1 exerts effects on reducing body weight, decreasing blood lipid levels, enhancing the skeletal muscle endurance,and increasing the insulin sensitivity in obese mice by activating the related proteins in AMPK signaling pathway in skeletal muscle.

AMPK细胞能量感受器研究进展

腺苷酸活化蛋白激酶(AMPK)作为体内能量感受器能感知能量改变情况.阐述了AMPK在外周组织、肝脏组织、骨骼肌和脂肪组织中的能量代谢调节及在下丘脑中枢中的神经调节等相关作用机制.

二甲双胍通过激活腺苷酸活化蛋白激酶(AMPK)的抗肿瘤机制

二甲双胍是一种传统的口服降糖药,临床上普遍用于2型糖尿病的治疗。近年来大量流行病学研究报道二甲双胍能够降低2型糖尿病患者的肿瘤发病率,亦有研究发现二甲双胍能在代谢途径、细胞周期、氧化应激、肿瘤干细胞转化等方面通过激活腺苷酸活化蛋白激酶(adenosin emonophosphate-activated protein kinase,AMPK)信号通路,从而抑制肿瘤细胞的生长、增殖以及转化。但二甲双胍通过激活AMPK的抗肿瘤机制仍存在着争议,其确切的作用机制有待进一步深入的研究,同时亟需大规模的临床试验来证实。

小檗碱的降糖作用独立于AMPK的信号通路

目的：研究小檗碱的降糖作用是否依赖于单磷酸腺苷激活蛋白激酶（AMPK）信号途径。方法培养HepG2细胞和C2C12细胞，给予不同浓度小檗碱处理。葡萄糖消耗实验和乳酸生成实验用于检测小檗碱的降糖以及刺激糖酵解的作用。AMPK抑制剂化合物C（Compound C，CC）和显性失活突变型AMPK，即腺病毒负显性AMPK（Ad-DN-AMPK）腺病毒用于抑制AMPK的表达和活性。Western印迹法用于检测AMPK以及乙酰辅酶A羧化酶（ACC）磷酸化水平，以评估AMPK通路的活性。结果小檗碱显著刺激了HepG2细胞和C2C12细胞的葡萄糖消耗和乳酸生成，并表现出剂量依赖性的药物作用。5和10μmol/L的小檗碱显著增加AMPK及其下游蛋白ACC的磷酸化水平。CC和Ad-DN-AMPK腺病毒转染能明显抑制细胞内AMPK信号通路的活性。然而，在AMPK活性被抑制的条件下，小檗碱依然能够显著增加细胞的葡萄糖消耗和乳酸生成。结论小檗碱通过刺激糖酵解而上调细胞的糖代谢，该作用无需AMPK信号通路的参与。即使在AMPK的表达或者活性被抑制的情况下，小檗碱依然能够发挥显著的降糖作用。

基于气机升降理论观察旋复代赭汤及其拆方对RE模型大鼠AMPK的影响

目的:观察旋复代赭汤及其拆方对反流性食管炎(RE)模型大鼠腺苷酸活化蛋白激酶(AMPK)含量的影响。方法:将84只雄性Wistar大鼠,随机分成7组,即正常对照组、模型对照组、旋复代赭汤全方组及拆方各组(包括苦降组、甘升组、升降相因组)、西药组(兰索拉唑+莫沙必利),每组12只大鼠。对造模组大鼠采用"4.2 mm幽门夹+胃底2/3结扎术"制备反流性食管炎动物模型。从造模术后7天开始,正常对照组、模型对照组给予生理盐水灌胃,旋复代赭汤全方组及拆方各组分别给予相应药液,西药组给予(兰索拉唑+莫沙必利)灌胃,干预14天后,处死全部大鼠后,其中每组分别送2份大鼠食管组织制作线粒体超微结构切片,应用电镜观察食管下段黏膜组织线粒体超微结构变化,其余食管组织匀浆后应用ELISA测定AMPK含量的表达情况。结果:食管组织线粒体超微结构:模型对照组大鼠电镜下食管黏膜线粒体稀少,形态、大小不一,内膜、外膜及嵴断续模糊,部分线粒体呈肿胀、空泡化,偶见巨线粒体;旋复代赭汤全方组、西药组较模型对照组改善(P<0.05),甘升组、升降相因组较模型对照组也明显改善(P<0.05);各组AMPK含量:旋复代赭汤全方组、西药组较模型对照组降低(P<0.05);甘升组、升降相因组较模型组也降低(P<0.05)。结论:旋复代赭汤及其药物通过调节脾胃气机,降低食管组织AMPK含量,促进线粒体能量三磷酸腺苷(ATP)的生成代谢,增加线粒体数量,减少线粒体结构及食管黏膜损伤,调节线粒体能量代谢,治疗RE。

Targeted metabolomics reveals the aberrant energy status in diabetic peripheral neuropathy and the neuroprotective mechanism of traditional Chinese medicine JinMaiTong

Diabetic peripheral neuropathy (DPN) is a common and devastating complication of diabetes, for which effective therapies are currently lacking. Disturbed energy status plays a crucial role in DPN pathogenesis. However, the integrated profile of energy metabolism, especially the central carbohydrate metabolism, remains unclear in DPN. Here, we developed a metabolomics approach by targeting 56 metabolites using high-performance ion chromatography-tandem mass spectrometry (HPIC-MS/MS) to illustrate the integrative characteristics of central carbohydrate metabolism in patients with DPN and streptozotocin-induced DPN rats. Furthermore, JinMaiTong (JMT), a traditional Chinese medicine (TCM) formula, was found to be effective for DPN, improving the peripheral neurological function and alleviating the neuropathology of DPN rats even after demyelination and axonal degeneration. JMT ameliorated DPN by regulating the aberrant energy balance and mitochondrial functions, including excessive glycolysis restoration, tricarboxylic acid cycle improvement, and increased adenosine triphosphate (ATP) generation. Bioenergetic profile was aberrant in cultured rat Schwann cells under high-glucose conditions, which was remarkably corrected by JMT treatment. In-vivo and in-vitro studies revealed that these effects of JMT were mainly attributed to the activation of adenosine monophosphate (AMP)-activated protein kinase (AMPK) and downstream peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α). Our results expand the therapeutic framework for DPN and suggest the integrative modulation of energy metabolism using TCMs, such as JMT, as an effective strategy for its treatment.

Fusobacterium nucleatum-induced imbalance in microbiome-derived butyric acid levels promotes the occurrence and development of colorectal cancer

BACKGROUND Colorectal cancer(CRC)ranks among the most prevalent malignant tumors globally.Recent reports suggest that Fusobacterium nucleatum(F.nucleatum)contributes to the initiation,progression,and prognosis of CRC.Butyrate,a short-chain fatty acid derived from the bacterial fermentation of soluble dietary fiber,is known to inhibit various cancers.This study is designed to explore whether F.nucleatum influences the onset and progression of CRC by impacting the intestinal metabolite butyric acid.AIM To investigate the mechanism by which F.nucleatum affects CRC occurrence and development.METHODS Alterations in the gut microbiota of BALB/c mice were observed following the oral administration of F.nucleatum.Additionally,DLD-1 and HCT116 cell lines were exposed to sodium butyrate(NaB)and F.nucleatum in vitro to examine the effects on proliferative proteins and mitochondrial function.RESULTS Our research indicates that the prevalence of F.nucleatum in fecal samples from CRC patients is significantly greater than in healthy counterparts,while the prevalence of butyrate-producing bacteria is notably lower.In mice colonized with F.nucleatum,the population of butyrate-producing bacteria decreased,resulting in altered levels of butyric acid,a key intestinal metabolite of butyrate.Exposure to NaB can impair mitochondrial morphology and diminish mitochondrial membrane potential in DLD-1 and HCT116 CRC cells.Consequently,this leads to modulated production of adenosine triphosphate and reactive oxygen species,thereby inhibiting cancer cell prolif-eration.Additionally,NaB triggers the adenosine monophosphate-activated protein kinase(AMPK)signaling pathway,blocks the cell cycle in HCT116 and DLD-1 cells,and curtails the proliferation of CRC cells.The combined presence of F.nucleatum and NaB attenuated the effects of the latter.By employing small interfering RNA to suppress AMPK,it was demonstrated that AMPK is essential for NaB’s inhibition of CRC cell proliferation.CONCLUSION F.nucleatum can promote cancer progression through its inhibitory effect on butyric acid,via the AMPK signaling pathway.

AMPK激活剂减轻重症急性胰腺炎模型大鼠胰腺损伤

目的探讨磷酸腺苷活化蛋白激酶(AMPK)激活剂5-氨基-4-咪唑羧基酰胺核苷(AICAR)通过抑制炎性反应和氧化应激,对重症急性胰腺炎(SAP)大鼠胰腺损伤的保护作用。方法将大鼠随机分为假手术组(NC组)、模型组(SAP组)和实验组[(SAP+AICAR)组,造模前2 h给予AICAR 400 mg/kg干预],5%牛黄胆酸钠按0.1 mL/kg泵入胆管造模,每组6只大鼠。造模24 h后异氟烷麻醉大鼠,取腹主动脉血5~7 mL,并剪除心脏使大鼠安乐死。Western blot测定胰腺磷酸化p-AMPK/AMPK比值;HE染色观察胰腺病理损伤和水肿程度;免疫组织化学染色测定胰腺单核细胞趋化蛋白1(MCP-1)表达量和髓过氧化物酶(MPO)阳性细胞数量;ELISA测定血清淀粉酶、脂肪酶、IL-6和TNF-α含量,并测定胰腺组织丙二醛(MDA)含量和超氧化物歧化酶(SOD)活性。结果SAP组胰腺磷酸化AMPK水平较NC组明显降低(P<0.05),实验组磷酸化AMPK水平较SAP组明显升高(P<0.01);SAP组胰腺的病理评分、胰腺水分含量均明显高于NC组(P<0.05),实验组胰腺的病理评分、胰腺水分含量较SAP组明显降低(P<0.01);SAP组MCP-1表达量、MPO阳性细胞数量、血清淀粉酶含量、血清脂肪酶含量、血清IL-6和TNF-α含量以及胰腺组织MDA含量和SOD活性较NC组明显升高(P<0.05),而实验组MCP-1表达量、MPO阳性细胞数量、血清淀粉酶含量、血清脂肪酶含量、血清IL-6和TNF-α含量以及胰腺组织MDA含量和SOD活性较SAP组明显降低(P<0.01)。结论AMPK激活剂抑制SAP大鼠炎性反应和氧化应激反应,减轻胰腺损伤和胰腺水肿。

α-硫辛酸通过激活AMPK/mTOR通路改善2型糖尿病大鼠肝脏病变

目的:探讨α-硫辛酸是否通过激活腺苷酸活化蛋白激酶(AMPK)/雷帕霉素靶蛋白(mTOR)通路改善2型糖尿病(T2DM)大鼠肝损伤。方法:应用高糖高脂饮食联合链脲佐菌素27.5 mg/(kg·d)腹腔注射法制备T2DM大鼠模型。将32只成模大鼠随机分为4组:T2DM组、α-硫辛酸组、Compound C(AMPK抑制剂)组和α-硫辛酸+Compound C组,每组各8只。另8只健康Sprague-Dawlay(SD)大鼠作为正常对照组。α-硫辛酸注射液100 mg/(kg·d)腹腔注射,Compound C 20 mg/(kg·d)腹腔注射,药物干预共持续8周。检测相关生化指标;计算肝脏质量指数;进行光镜、电镜观察;采用Western blot方法检测大鼠肝脏中AMPK、p-AMPK、mTOR、p-mTOR蛋白的表达水平。结果:与正常对照组相比,T2DM组大鼠肝脏质量指数、胰岛素抵抗指数、空腹血糖、谷丙转氨酶、谷草转氨酶、γ–谷氨酰转肽酶、甘油三酯水平均升高(P均<0.05);肝组织结构损伤,脂肪变明显;肝脏组织中p-AMPK表达显著减少(P<0.05),p-mTOR表达显著增多(P<0.05)。α-硫辛酸可逆转上述改变,改善大鼠的胰岛素抵抗(P<0.05),保护肝脏的结构和功能,同时激活肝细胞内的AMPK/mTOR通路(P均<0.05)。应用Compound C抑制AMPK活性后,α-硫辛酸的上述保护作用受到抑制(P<0.05)。结论:α-硫辛酸可通过激活AMPK/mTOR信号通路发挥对T2DM大鼠的肝脏保护作用。