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Objective To investigate the effect of intestinal Metrnl on dextran sodium sulfate (DSS)-induced ulcerative colitis mouse model and the regulation mechanism of intestinal microbiota. Methods Different concentrations of DSS (3% DSS and 1% DSS) were used to induce ulcerative colitis on C57 mice to determine the experimental conditions. Intestinal epithelial Metrnl specific knockout mice (Metrnl(-/-)) and its control mice (Metrnl(+/+)) were administrated with 3% DSS for 5 d. Then the survival time, body weight, DAI (disease activity index), colon length and pathological changes in colon tissues were observed. 16S ribosomal RNA gene sequencing was used to detect the composition of intestinal microbiota. Results Compared with 1% DSS, 3% DSS could significantly aggravate ulcerative colitis on C57 mice, such as lower survival rate (P<0.05), more weight loss (P<0.05), higher DAI score (P<0.05), shorter colon length (P<0.05) and higher pathology score (P<0.05). After administrated to 3% DSS for 5 d, comparing with Metrnl(+/+) mice, Metrnl(-/-) mice showed more weight loss (P<0.05), higher DAI score (P<0.05), shorter colon length (P<0.05) and higher pathology score (P<0.05). The 16S ribosomal RNA results showed that the diversity of intestinal microbiota in Metrnl(-/-) mice significantly decreased. Furthermore, Bacteroidetes and Proteobacteria significantly decreased, while Firmicutes increased. Conclusion Metrnl could protect the DSS-induced ulcerative colitis mouse through regulating intestinal microbiota.
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The injury of vascular endothelial cell function is the beginning of the pathological process of atherosclerosis. Mitochondrial oxidative stress is closely related to vascular endothelial cell function, which causes the dysfunction of vascular endothelial cell by inducing mitophagy, reducing nitric oxide production, inflammation, cellular metabolic imbalance and apoptosis. Meanwhile, vascular endothelial cell could also maintain their homeostasis by regulating mitochondrial oxidative stress. The molecular signaling pathways of the vascular endothelial cell injury caused by mitochondrial oxidative stress in the pathological process of atherosclerosis were outlined in this review, which provided reference for further research on the molecular mechanism between mitochondrial oxidative stress and endothelial damage.
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METRNL is a recently identified secreted protein with emerging functions. This study is to find major cellular source of circulating METRNL and to determine METRNL novel function. Here, we show METRNL is abundant in human and mouse vascular endothelium and released by endothelial cells using endoplasmic reticulum-Golgi apparatus pathway. By creating endothelial cell-specific Metrnl knockout mice, combined with bone marrow transplantation to produce bone marrow-specific deletion of Metrnl, we demonstrate that most of circulating METRNL (approximately 75%) originates from the endothelial cells. Both endothelial and circulating METRNL decrease in atherosclerosis mice and patients. By generating endothelial cell-specific Metrnl knockout in apolipoprotein E-deficient mice, combined with bone marrow-specific deletion of Metrnl in apolipoprotein E-deficient mice, we further demonstrate that endothelial METRNL deficiency accelerates atherosclerosis. Mechanically, endothelial METRNL deficiency causes vascular endothelial dysfunction including vasodilation impairment via reducing eNOS phosphorylation at Ser1177 and inflammation activation via enhancing NFκB pathway, which promotes the susceptibility of atherosclerosis. Exogenous METRNL rescues METRNL deficiency induced endothelial dysfunction. These findings reveal that METRNL is a new endothelial substance not only determining the circulating METRNL level but also regulating endothelial function for vascular health and disease. METRNL is a therapeutic target against endothelial dysfunction and atherosclerosis.
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Objective Nicotinamide phosphoribosyltransferase (Nampt) is a new therapeutic target for ischemic stroke. The aim of this study was to investigate protective effect of liver-derived Nampt on ischemic stroke. Methods Liver-specific Nampt knockout mice were generated using the Cre/loxP system. NamptloxP/loxP mice were crossed with liver-specific Cre recombinase expression mice (Alb-Cre), and the progeny genotypes were identified by polymerase chain reaction. Body weight of knockout mice and control mice were measured. Nampt in liver and brain was determined by Western blot assay. Middle cerebral artery occlusion (MCAO), a classical ischemic stroke model, was generated in liver-specific Nampt knockout mice and control mice by electrocoagulation. After 24 h of modeling, neurological deficit scores of each group were evaluated and TTC staining was performed to determine the cerebral infarction volume. The level of plasma Nampt in each group was determined by ELISA. Results Liver-specific Nampt knockout mice with the genotype of NamptloxP/loxPAlb-Cre were successfully constructed. The hepatic Nampt expression in knockout mice was significantly decreased by 74.2% compared to control mice, while there was no significant difference in the expression of brain Nampt protein between the knockout group and the control group. Specific knockout of liver Nampt gene expression had no effect on the body weight of mice. Under normal physiological conditions, there was no significant difference in plasma Nampt levels between liver-specific Nampt knockout mice and control mice of the same gender. 24 h after MCAO modeling, there were no significant differences in neurological deficit scores, cerebral infarct volume and plasma Nampt concentration between liver-specific Nampt knockout group and control group. Conclusion Liver-specific Nampt knockout mice are successfully constructed. Liver-derived Nampt has no significant protective effects on ischemic stroke.
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Objective To explore the expression of CRELD2 at the gene and protein levels of mouse tissues, and to provide a reference for studying the biological function of CRELD2 in various tissues. Methods The expression level of CRELD2 in the liver, pancreas, stomach, and lung of C57BL/6J mice was determined by real-time PCR and Western Blot. Results RT-PCR and WB showed that CRELD2 was expressed in mouse liver, pancreas, stomach, and lung. The relative expression levels of CRELD2 from high to low were pancreas, stomach, liver, and lung at the gene level, and pancreas, liver, stomach, and lung at protein level respectively. The result suggested that the relative expression levels of the CRELD2 gene and protein in different tissues were not completely consistent, suggesting that it is related to transcriptional regulation. Conclusion CRELD2 is expressed in mouse liver, pancreas, stomach, and lung, and the relative expression levels of CRELD2 are not completely parallel at the gene and protein level.
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Objective To investigate the effects of Nicotinamide mononucleotide (NMN) on ulcerative colitis induced by dextran sulfate sodium (DSS) in mice. Methods DSS-induced ulcerative colitis mice were used to evaluate the effects of NMN. After NMN administration, the survival time, weight, disease activity index (DAI), colon tissue length and pathological changes of colon tissue slices were observed. Results NMN did not cause significant changes in the survival time, weight, DAI, and intestinal morphology of ulcerative colitis mice. Conclusion NMN has no significant effect on DSS-induced ulcerative colitis mice.
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Sepsis can cause life-threatening organ dysfunction and is one of the leading causes of death in critically ill patients. Early diagnosis and correct treatment of sepsis are the key to reducing the fatality, however, there is no golden standard for diagnosis at present. The ideal sepsis biomarker can be used for early diagnosis and predicting poor prognosis with good sensitivity and specificity. There are many candidate biomarkers for sepsis. This article reviews the latest developments on acute phase proteins, soluble receptors, non-coding RNAs and other candidate biomarkers of sepsis that attracted more recent attention.
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Objective To study the effect of nicotinamide mononucleotide (NMN) on the mortality of the lipopoly-saccharide (LPS)-induced endotoxic shock mouse model. Methods 10-week-old C57BL/6J male mice were randomly divided into groups, and were injected intraperitoneally (i.p.) with LPS (10 mg/kg) to induce endotoxic shock models. NMN was i.p. injected in three ways: (1) 0.5 h after modeling, doses of 10, 30, 100 and 300 mg/kg; (2) 0.5 h before modeling, doses of 30, 100, 300 and 600 mg/kg; or (3) 0.5 and 12 h after modeling, dose of 300 mg/kg each time. The death times of each group were recorded, and the survival curves were drawn. Results Compared with the solvent control group, NMN at different doses given 0.5 h after or before modeling didn’t improve the survival rate or delay the death time of endotoxic shock mice; But when given at 0.5 and 12 h 300 mg/kg after modeling, NMN accelerated the death of mice and increased the mortality of mice. NMN products by two manufacturers showed similar effects. Conclusion NMN has no therapeutic effect on LPS-induced endotoxic shock, and repeated administration of NMN after endotoxic shock will increase the mortality.
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Objective To study the effects of atherosclerotic high-fat diet on body weight, blood glucose, blood lipid levels and atherosclerotic plaque formation in mice, and to determine the effect of fasting time on the results of blood lipid testing. Methods 10-week-old male ApoE knockout (ApoE-/-) mice were given high-fat diet and normal diet. The atherosclerotic plaques were observed four months later. 10 week old C57BL/6J male mice were given regular diet for 4 weeks, regular diet for 2 weeks + high-fat diet for two or four weeks. Body weight、liver、glucose, and the serum lipid levels were examined. The influence of fasting for 12 h, 6 h or no fasting on blood lipid detection results before sacrificing were studied. Results The atherosclerotic plaque area of ApoE-/- mice given high-fat diet increased significantly (P<0.01). C57BL/6J mice given high-fat diet gained weight (P<0.01). The glucose, TC, LDL-c and HDL-c were also increased in C57BL/6J mice with liver fat accumulation while the level of TG was significantly decreased(P<0.01). Compared with the fasting 12 h group, serum triglyceride (TG) was significantly increased (P<0.01）in fasting 6 h and no fasting groups. Conclusion The atherosclerotic high-fat diet can accelerate the formation of atherosclerotic plaques in ApoE-/- mice, significantly increase blood sugar, TC and LDL-c levels, but significantly reduce TG values.. The fasting time can affect serum triglyceride (TG) level.
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Objective To establish and optimize a mouse myocardial infarction (MI) model, and to use twice limb lead ECGs immediately after coronary ligation and 4 h after surgery to evaluate the occurrence of myocardial infarction. Methods Twenty-nine male C57BL/6J mice were anesthetized with isoflurane. then a myocardial infarction model was established by ligating the left anterior descending (LAD) coronary artery through the third/fourth intercostal space of left anterior chest. Immediate and 4 h postoperative limb lead ECGs were performed. Twenty-four hours after surgery, the chest was opened and the occurrence of myocardial infarction was evaluated. The heart samples were taken for TTC staining to determine the infarct area and calculate the infarct area. Results During the mice underwent coronary artery ligation the intraoperative mortality was 6.8% (2/29), and the early postoperative (<4 h) mortality was 10.3% (3/29). The 24 h survival rate was 82.8% (24/29). 24 hours after TTC staining confirmed the occurrence of infarction, the myocardial infarction model was established. The success rate of the model was 79.3% (23/29), and the average infarct size (infarcted myocardial weight / whole ventricular weight) was (28 ± 6)%; The mice successfully established by the model showed obvious ST-T changes in the ECG at 4 hours after surgery, suggesting that a myocardial infarction has occurred. Conclusions The mouse myocardial infarction model was successfully established. The combined use of ECG immediately after surgery and 4 h after surgery could be used as a rapid and non-invasive evaluation method for mouse myocardial infarction.
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Objective To construct and explore the in vivo and in vitro D-galactose induced cognitive impairment models and evaluate the application value of the combined models in the study of cognitive impairments.Methods The cognitive impairment mice model induced by D-gal was prepared by continuous intraperitoneal injection of D-gal saline solution for 8weeks, followed by detection of learning and memory functions with Morris water maze.The related molecular markers in the brain tissue were assayed to evaluate the effect and application value.D-gal cell model was prepared by adding D-gal in different concentrations into the cell cultural medium of neurons harvested from the hippocampus of young mice.The effect and application value were evaluated by detecting the molecular markers related to the level of cell injury.Results The Morris water maze on the D-gal model showed that the learning and memory functions of mice in the model group were significantly lower than those in the control group.Meanwhile, the levels of apoptosis and oxidative stress in the model group were significantly higher than those in the control group.In the hippocampal neuron model of D-gal, the neurons showed a dose-dependent morphologic and functional change with the increase of D-gal dose and the levels of apoptosis and oxidative stress were significantly higher than those in the negative control.Conclusion D-galactose can be successfully used to induce cognitive impairment models both in vivo and in vitro through the decrease of the learning and memory functions of mice and induction of apoptosis and oxidative stress in neurons.Combined application of the two models of D-gal can be one of effective and promising tools for the study of cognitive impairment and pharmacodynamic evaluation.
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Monoclonal antibodies have become an important part of biotechnology and pharmaceutical industry.Currently chimeric antibody, humanized antibody and fully human antibody are generally prepared by genetic engineering.With constant improvement in the antibody preparation technology, the antibody applications are rapidly increasing, especially in the development of antibody medications.In recent years, there have been many significant antibody drugs (such as PD-1 antibody, IL-17 antibody, IL-5 antibody, PCSK9 antibody) for the treatment of various diseases and solved a number of clinical problems.In modern medicine, monoclonal antibodies have become effective diagnostic and therapeutic tools.This article is a brief review of the recent antibody development process, clinic applications and significant antibody drugs approved by FDA.
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Objective To prepare monoclonal antibodies (mAb) against mouse Metrnl and identify its specificity . Methods Mouse Metrnl polypeptide fragments and full-length protein were prepared as antigens to immunize mice .Then mice spleen cells were fused with SP2/0 myeloma cells to obtain hybridoma cells which were screened for positive clone in order to subclone for stable cell lines .After ascites were prepared ,ELISA method was used to detect the antibodies titer .Western blot method was applied to identify their specificity .Results No effective antibodies were identified from the ascites derived from 14 polypeptide antigens .Among the 25 antibodies derived from the full-length protein ,12 monoclonal antibodies can be used to identify the recombinant Metrnl protein .Conclusion 12 monoclonal antibodies were successfully prepared to identify mouse Metrnl protein .
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Intracerebral hemorrhage (ICH) refers to bleeding within the brain parenchyma due to the rupture of blood vessels ,and is a highly lethal stroke subtype ,accounting for nearly 10%-15% of all strokes .The morbidity and mortality of ICH are very high and its pathophysiological mechanisms are currently not fully understood .Therefore ,based on current clini-cal evidence-based medicine ,there is no definite and effective medical treatment that can improve the prognosis and survival of patients available yet .As an important tool for basic research ,the development and application of the ICH animal model pro-moted the understanding of the pathophysiological mechanisms and molecular mechanisms leading to brain injury by ICH .Re-cently ,the ICH animal model studies contributed to a number of proposed potential therapeutic strategies ,such as the inhibi-tion of thrombin and the reduction of pro-inflammatory pathways .In addition ,recent research of stem cells suggested that cell transplantation therapy for the treatment of ICH may also have good prospects .In this review ,we discuss the development and application of animal models for studies on ICH and the advances regarding the potential therapeutic strategies for ICH .
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As the first most common cause of death in China,stroke has become a public health problem that seriously affects national economy and people′s livelihood. Unfortunately,only 3% to 5% of stroke patients receive tissue plasminogen activator(tPA)treatment,the only pharmacological therapy ap?proved for ischemic stroke,and no drug is available for hemorrhagic stroke. Therefore,there is an ur?gent need to develop new drugs for stroke therapy. Despite the awareness that neuroprotective agents could be a common strategy for the treatment of both ischemic and hemorrhagic stroke,numerous neu?roprotective agents have showed failure in clinical trials. Combined with the current therapeutic strategies and drug development of stroke,this paper elaborated the stroke injury mechanisms and corresponding clinical drug research targeting excitotoxicity,oxidative and nitrosative stress,and inflammation. From a new perspective,this paper has proposed a novel therapeutic strategy targeting inherent defense mechanisms against stroke,with nicotinamide phosphoribosyltransferase(Nampt)- nicotinamide ade?nine dinucleotide defense system as an example to present our experimental evidence that Nampt can serve as an anti-stroke target and nicotinamide mononucleotide as an anti-stroke agent under development. It is hoped that the bottleneck of stroke therapy can be overcome with unremitting efforts so as to reduce the financial burden and mental stress,and bring benefits to people around the world.
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Objective:It has been proposed that blood pressure variability(BPV) is positively related to end-organ damage(EOD) in hypertension.The present work was designed to observe the effects of long-term treatment with nitrendipine and hydralazine on BPV and EOD in spontaneously hypertensive rats(SHR),to examine the hypothesis that lowering BPV with an antihypertensive drug is an important factor in organ protection.Design and methods:Drugs were mixed in rat chow.After 4 months of drug administration,blood pressure was recorded continuously in conscious freely moving rats for 24 h.The heart,kidneys,and brain were then isolated and examined.Results:It was found that nitrendipine significantly decreased blood pressure and BPV,and significantly decreased EOD score in SHR.Hydralazine decreased blood pressure,but did not lower BPV.No effect on EOD was found in hydralazine-treated rats.In control rat(n=38),EOD score was weakly related to systolic blood pressure(r=0.331,P<0.05) and closely related to long-term systolic BPV(r=0.551,P<0.01).In nitrendipine-treated rats,EOD score was closely related to long-term systolic BPV(r=0.602,P<0.01),but not to blood pressure level(r=0.174,P>0.05).Conclusion:BPV plays an important role in the organ-protecting effects of nitrendipine.
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Objective:Glucose-insulin-potassium(GIK) is clinically used for reducing mortality in acute myocardial infarction(MI). It is known that ventricular arrhythmia, left ventricular dysfunction and impaired baroreflex sensitivity(BRS) are the three major determinants for predicting the mortality after acute MI. The present work was designed to study the effects of GIK on BRS, ventricular arrhythmia, and left ventricular function in rats with coronary artery ligature. Sprague-Dawley rats were used and the myocardial infarction was produced by ligature of the left anterior descending artery. Five weeks after coronary artery ligation, BRS was measured in conscious state with a computerized blood pressure monitoring system and left ventricular function and electrocardiogram were determined in the anaesthetized state in the subacute phase of myocardial infarction. It was found that GIK did not affect the blood pressure and heart period in both conscious and anaesthetized rats. GIK did not enhance BRS, but reduced ventricular arrhythmia and improved left ventricular function by reducing left ventricular end diastolic pressure in anaesthetized rats with MI. It is proposed that reducing ventricular arrhythmia and improving left ventricular function contribute to the effect of GIK on reducing the mortality after MI.
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It is well known that the arterial baroreflex(ABR)plays a key role in the regulation of heart rate and stabilization of blood pressure.Currently,it appears that ABR dysfunction is involved in the pathophysiology of cardiovascular disease states.Since the mid-1990s,a number of studies have been carried out in our laboratory to explore the pathological significance of ABR function in cardiovascular damage.This minireview summarizes our research work on the topic of ABR and left ventricular hypertrophy(LVH).On the basis of discussion concerning the importance of ABR dysfunction in hypertensive LVH and sinoaortic denervation-induced LVH,we advance a new strategy for reversal of LVH,that is,restoration of impaired ABR function.We tested this hypothesis in animal models with ABR deficiency.It was found that improvement of impaird ABR function with long-term treatment of ketanserin or candesartan was accompanied by reversal of LVH.The preliminary results indicate that it is feasible to target ABR for treatment of LVH.
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AIM To investigate the structural and functional remodeling of thoracic aortae in sinoaortic- denervated (SAD) rats. METHODS SD rats underwent either SAD or sham-operation at the age of 10 weeks. Sixteen weeks after operation, the contraction and relaxation of the thoracic aortae were measured in isolated preparations; The morphological changes of arteries were examined by using histopathological method and computer image analysis. RESULTS The NE-induced contraction was increased and Ach-induced relaxation of aortic rings was depressed in SAD rats; The structural remodeling of thoracic aortae was characterized by medial VSMC hypertrophy and matrix accumulations. CONCLUSION Vascular functional and structural remodeling can be found in sinoaortic-denervated rats.
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AIM: To investigate long-term effects of sinoaortic denervation (SAD) on the contraction and relaxation of thoracic aortae in rats.METHODS:SD rats underwent either SAD or sham operation at the age of 10 weeks. At 4,8, 16, and 32 weeks after operation, the contraction and relaxation of the thoracic aortae were measured by isolated artery technique.RESULTS:The NE-induced contraction and ACh-induced relaxation of aortic rings were progressively increased and depressed respectively after SAD.CONCLUSION:Vascular functional changes induced by purely blood pressure variability is similar to those observed in well-established experimental hypertensive states.