Small extracellular vesicles from hypoxia-preconditioned bone marrow mesenchymal stem cells attenuate spinal cord injury via miR-146a-5p-mediated regulation of macrophage polarization

Spinal cord injury is a disabling condition with limited treatment options.Multiple studies have provided evidence suggesting that small extracellular vesicles(SEVs)secreted by bone marrow mesenchymal stem cells(MSCs)help mediate the beneficial effects conferred by MSC transplantation following spinal cord injury.Strikingly,hypoxia-preconditioned bone marrow mesenchymal stem cell-derived SEVs(HSEVs)exhibit increased therapeutic potency.We thus explored the role of HSEVs in macrophage immune regulation after spinal cord injury in rats and their significance in spinal cord repair.SEVs or HSEVs were isolated from bone marrow MSC supernatants by density gradient ultracentrifugation.HSEV administration to rats via tail vein injection after spinal cord injury reduced the lesion area and attenuated spinal cord inflammation.HSEVs regulate macrophage polarization towards the M2 phenotype in vivo and in vitro.Micro RNA sequencing and bioinformatics analyses of SEVs and HSEVs revealed that mi R-146a-5p is a potent mediator of macrophage polarization that targets interleukin-1 receptor-associated kinase 1.Reducing mi R-146a-5p expression in HSEVs partially attenuated macrophage polarization.Our data suggest that HSEVs attenuate spinal cord inflammation and injury in rats by transporting mi R-146a-5p,which alters macrophage polarization.This study provides new insights into the application of HSEVs as a therapeutic tool for spinal cord injury.

Neuroprotectants attenuate hypobaric hypoxia-induced brain injuries in cynomolgus monkeys

Hypobaric hypoxia (HH) exposure can cause serious brain injury as well as life-threatening cerebral edema in severe cases. Previous studies on the mechanisms of HH-induced brain injury have been conducted primarily using non-primate animal models that are genetically distant to humans, thus hindering the development of disease treatment. Here, we report that cynomolgus monkeys (Macaca fascicularis) exposed to acute HH developed human-like HH syndrome involving severe brain injury and abnormal behavior. Transcriptome profiling of white blood cells and brain tissue from monkeys exposed to increasing altitude revealed the central role of the HIF-1 and other novel signaling pathways, such as the vitamin D receptor (VDR) signaling pathway, in co-regulating HH-induced inflammation processes. We also observed profound transcriptomic alterations in brains after exposure to acute HH, including the activation of angiogenesis and impairment of aerobic respiration and protein folding processes, which likely underlie the pathological effects of HH-induced brain injury. Administration of progesterone (PROG) and steroid neuroprotectant 5α-androst-3β,5,6β-triol (TRIOL) significantly attenuated brain injuries and rescued the transcriptomic changes induced by acute HH. Functional investigation of the affected genes suggested that these two neuroprotectants protect the brain by targeting different pathways, with PROG enhancing erythropoiesis and TRIOL suppressing glutamate-induced excitotoxicity. Thus, this study advances our understanding of the pathology induced by acute HH and provides potential compounds for the development of neuroprotectant drugs for therapeutic treatment.

Hypoxia inducible factor-1 alpha stabilization for regenerative therapy in traumatic brain injury

Mild traumatic brain injury（TBI）, also called concussion, initiates sequelae leading to motor deficits, cognitive impairments and subtly compromised neurobehaviors. While the acute phase of TBI is associated with neuroinflammation and nitroxidative burst, the chronic phase shows a lack of stimulation of the neurorepair process and regeneration. The deficiency of nitric oxide（NO）, the consequent disturbed NO metabolome, and imbalanced mechanisms of S-nitrosylation are implicated in blocking the mechanisms of neurorepair processes and functional recovery in the both phases. Hypoxia inducible factor-1 alpha（HIF-1α）, a master regulator of hypoxia/ischemia, stimulates the process of neurorepair and thus aids in functional recovery after brain trauma. The activity of HIF-1α is regulated by NO via the mechanism of S-nitrosylation of HIF-1α. S-nitrosylation is dynamically regulated by NO metabolites such as S-nitrosoglutathione（GSNO） and peroxynitrite. GSNO stabilizes, and peroxynitrite destabilizes HIF-1α. Exogenously administered GSNO was found not only to stabilize HIF-1α and to induce HIF-1α-dependent genes but also to stimulate the regeneration process and to aid in functional recovery in TBI animals.

Dynamic changes in cerebral microcirculation and hypoxia in the early stages of diffuse axonal injury

This study demonstrated that damage to the cerebral microvasculature, the formation of microthrombi and swelling of vascular endothelial cells occur early and peak 12 hours after injury in a rat model of diffuse axonal injury. Moreover, these pathological changes were most evident in the cerebral cortex. Cerebral microcirculatory dysfunction peaked later and had a shorter duration than axonal injury. In addition, the radioactive imaging agent, 99Tcm-4, 9-diaza-2, 3, 10, 10- tetramethyldodecan-2, 11 -dione dioxime, was used to visualize the dynamic changes that occur in tissue with cerebral hypoxia. The results demonstrated that cerebral hypoxia occurs at an early stage in diffuse axonal injury. Cerebral hypoxia was evident 12 hours after injury and declined slightly 24 hours after injury, but was significantly higher than in the control group. The pathological changes that underpin microcirculatory dysfunction did not occur at the same time as axonal injury, but did occur simultaneously with neuronal injury. Cerebral hypoxia plays a key role in promoting the secondary brain injury that occurs after diffuse axonal injury.

Protective effects of erythropoietin pretreatment on myocardium with hypoxia/reoxygenation injury in rats

Objective: To establish the rat model with myocardial hypoxia/reoxygenation (H/R) injury, and investigate the protective effect of EPO pretreatment on the myocardium. Methods: Sixty male adult Wistar rats were randomly divided into 3 groups: control group, H/R group, and EPO group, 20 in each group. The rats in EPO group accepted injection of 5 000 U/kg recombinant human erythropoietin (RHuEPO) through vein, and the other rats accepted the injection of the same volume of saline. Twenty-four hours after the injection, rats in the EPO and H/R groups were put into the hypoxia environment for 12 h and then returned to the normoxic environment for 2 h, and then the samples of blood and myocardium were collected. Serum myocardial enzyme activity, apoptosis, ultrastructure, myocardial MDA contents, EPO receptor (EPOR) expression in cardiac myocytes and cardiac functions were tested. Results: EPOR expression was positive in cardiac myocytes of adult rat according to the result of immunonistochemitry assaying. Compared to those in H/R group, rats in EPO group presented lighter injury of myocardial ultrastructure, the reduction of serum myocardial enzyme activity, inhibition of apoptosis, the better recovery of cardiac functions, and the less production of oxygen-derived free radicals. Conclusion: Adult rat cardiac myocytes could express EPOR, and EPO pretreatment produced protective effects on myocardium with H/R injury.

Effects of altitude hypoxia on contents of TXB_2, 6-keto-PGF_(1α) and LENK and their correlations with lung injury induced by pyosepticemia

Objective: To explore the effects of altitude hypoxia on septic pulmonary injury (3080 m above sea level). Methods : A model of pyosepticemia was established by cecal ligation and puncture. Thromboxane A2 (TXA2),prostacyclin (PGI2) of the venous plasma and leucine-enkephalins (L-ENK) in the arterial walls were measured using radioiizimunoassay in 31 rabbits. Mouse morrality was observed. Results: 24 h mouse accumulative mortality (97. 1% )was greater than that of control (35%). TXB2 content was 4 times as much as in control group and 6-keto-PGF1α level was elevated too. These changes of arachidonic metabolism could be blorked by indomethacin. The contents of LENK in arterial walls of puhiionary, mesenteric and renal arteries dropped significantly in comparison with control group (P<0. 01 ). The albumin content in the bronchoalveolar lavage fluid rase significantly in comparison with control group (P<0. 01 ). Conclusion : The results demonstrate that the metabolites of cyclcoxygenase are not the major mediator of puhiionary permeability in the experiment. The alteration of TXA2 and L-ENK contents may be one of the mechanisms, by which the puhiionary artery pressure and pulmonary vascular resistance are increased during sepsis.

Hypoxia in acute cardiac injury of coronavirus disease 2019:lesson learned from pathological studies

Coronavirus disease 2019(COVID-19)is an infectious respiratory disease caused by the severe acute respiratory syndrome coronavirus 2(SARS-CoV-2),which has infected 972,303 people and caused 50,322 deaths all over the world according to the latest WHO report.[1]As a highly contagious disease,COVID-19 has killed more people than severe acute respiratory syndrome(SARS)and middle east respiratory syndrome(MERS)combined,despite an relatively low case-fatality rate.[2,3]Although it mainly attacks respiratory system,other systems including cardiovascular system are also influenced by COVID-19.Acute cardiac injury(ACI)is also one of the noteworthy issues which researchers have noticed in several studies.[4–7] .

Aldehyde dehydrogenase 2 preserves mitochondrial morphology and attenuates hypoxia/reoxygenationinduced cardiomyocyte injury

BACKGROUND:Disturbance of mitochondrial fi ssion and fusion(termed mitochondrial dynamics)is one of the leading causes of ischemia/reperfusion(I/R)-induced myocardial injury.Previous studies showed that mitochondrial aldehyde dehydrogenase 2(ALDH2)conferred cardioprotective effect against myocardial I/R injury and suppressed I/R-induced excessive mitophagy in cardiomyocytes.However,whether ALDH2 participates in the regulation of mitochondrial dynamics during myocardial I/R injury remains unknown.METHODS:In the present study,we investigated the effect of ALDH2 on mitochondrial dynamics and the underlying mechanisms using the H9c2 cells exposed to hypoxia/reoxygenation(H/R)as an in vitro model of myocardial I/R injury.RESULTS:Cardiomyocyte apoptosis was significantly increased after oxygen-glucose deprivation and reoxygenation(OGD/R),and ALDH2 activation largely decreased the cardiomyocyte apoptosis.Additionally,we found that both ALDH2 activation and overexpression significantly inhibited the increased mitochondrial fission after OGD/R.Furthermore,we found that ALDH2 dominantly suppressed dynamin-related protein 1(Drp1)phosphorylation(Ser616)and adenosine monophosphate-activated protein kinase(AMPK)phosphorylation(Thr172)but not interfered with the expression levels of mitochondrial shaping proteins.CONCLUSIONS:We demonstrate the protective effect of ALDH2 against cardiomyocyte H/R injury with a novel mechanism on mitochondrial fission/fusion.

Effects of Crocin on Nox2 Expression and ROS Level of Hypoxia/Reoxygenation-induced Injury of Cardiomyocytes

[Objectives]To explore the protection mechanism of crocin against ischemia-reperfusion injury of myocardial cells.[Methods]Newborn male SD rats were selected,left ventricular cardiomyocytes(CMs)were isolated,and a hypoxia/reoxygenation model of CMs was established to simulate the process of ischemia/reperfusion injury.The cells were randomly divided into four groups:normal cell group(control group),crocin group),hypoxia/reoxygenation group(H/R group),hypoxia/reoxygenation+crocin group(H/R+crocin group).H/R+crocin group selected the concentration of crocin 1,10,and 100μmol/L,and determined the optimal concentration of crocin by detecting the cell proliferation ability.After the cells were pretreated using the optimal concentration of crocin,the levels of superoxide anion,cell proliferation,apoptosis and Nox2 levels in each group of cells were detected.[Results]Compared with the control group,the proliferation ability of CMs after hypoxia-reoxygenation injury was reduced(P<0.05),while cell apoptosis and intracellular superoxide anion levels were significantly increased(P<0.01);the CMs pretreated with crocin can reduce the level of Nox2(P<0.01),increase the cell proliferation ability of CMs,reduce cell apoptosis,and accordingly reduce the level of superoxide anion in the cell(P<0.05).[Conclusions]Crocin protects CMs from hypoxia/reoxygenation injury through down-regulating the level of Nox2 and reducing oxidative stress injury.

Hypoxia inducible factor 1α promotes interleukin-1 receptor antagonist expression during hepatic ischemia-reperfusion injury

BACKGROUND Ischemia-reperfusion injury(IRI) is a major risk associated with liver surgery and transplantation,and its pathological mechanism is complex.Interleukin-1 receptor antagonist(IL-1ra) can protect the liver from IRI.However,the regulatory mechanism of IL-1ra expression is still unclear.AIM To identify the mechanism that could protect the liver in the early stage of IRI.METHODS To screen the key genes in hepatic IRI,we performed RNA sequencing and gene enrichment analysis on liver tissue from mice with hepatic IRI.Subsequently,we verified the expression and effect of IL-1ra in hepatic IRI.We also used promoter mutagenesis and chromatin immunoprecipitation assay to search for the transcriptional regulatory sites of hypoxia-inducible factor(HIF)-1α.Finally,to explore the protective mechanism of ischemic preconditioning(IP),we examined the expression of HIF-1α and IL-1ra after IP.RESULTS We identified IL-1ra as a key regulator in hepatic IRI.The expression of IL-1ra was significantly upregulated after hepatic IRI both in vivo and in vitro.Furthermore,we found that HIF-1αregulated Il-1ra transcription in response to hypoxia.Increased HIF-1α accumulation promoted IL-1ra expression,whereas inhibition of HIF-1α exhibited the opposite effect.We also confirmed a predominant role for hypoxia response element in the regulation of Il1ra transcription by HIF-1αactivation.Of note,we demonstrated that IP protects against hepatic IRI by inducing IL-1ra expression,which is mediated through HIF-1α.CONCLUSION We demonstrated that ischemia or hypoxia leads to increased expression of IL-1ra through HIF-1α.Importantly,IP protects the liver from IRI via the HIF-1α–IL-1ra pathway.

Hypobaric Hypoxia Aggravates Renal Injury by Inducing the Formation of Neutrophil Extracellular Traps through the NF-κB Signaling Pathway

Objective The hypersensitivity of the kidney makes it susceptible to hypoxia injury.The involvement of neutrophil extracellular traps(NETs)in renal injury resulting from hypobaric hypoxia(HH)has not been reported.In this study,we aimed to investigate the expression of NETs in renal injury induced by HH and the possible underlying mechanism.Methods A total of 24 SD male rats were divided into three groups(n=8 each):normal control group,hypoxia group and hypoxia+pyrrolidine dithiocarbamate(PDTC)group.Rats in hypoxia group and hypoxia+PDTC group were placed in animal chambers with HH which was caused by simulating the altitude at 7000 meters(oxygen partial pressure about 6.9 kPa)for 7 days.PDTC was administered at a dose of 100 mg/kg intraperitoneally once daily for 7 days.Pathological changes of the rat renal tissues were observed under a light microscope;the levels of serum creatinine(SCr),blood urea nitrogen(BUN),cell-free DNA(cf-DNA)and reactive oxygen species(ROS)were measured;the expression levels of myeloperoxidase(MPO),citrullinated histone H3(cit-H3),B-cell lymphoma 2(Bcl-2),Bax,nuclear factor kappa B(NF-κB)p65 and phospho-NF-κB p65(p-NF-κB p65)in rat renal tissues were detected by qRT-qPCR and Western blotting;the localization of NF-κB p65 expression in rat renal tissues was observed by immunofluorescence staining and the expression changes of NETs in rat renal tissues were detected by multiplex fluorescence immunohistochemical staining.Results After hypoxia,the expression of NF-κB protein in renal tissues was significantly increased,the levels of SCr,BUN,cf-DNA and ROS in serum were significantly increased,the formation of NETs in renal tissues was significantly increased,and a large number of tubular dilatation and lymphocyte infiltration were observed in renal tissues.When PDTC was used to inhibit NF-κB activation,NETs formation in renal tissue was significantly decreased,the expression level of Bcl-2 in renal tissues was significantly increased,the expression level of Bax was significantly decreased,and renal injury was significantly alleviated.Conclusion HH induces the formation of NETs through the NF-κB signaling pathway,and it promotes apoptosis and aggravates renal injury by decreasing Bcl-2 and increasing Bax expression.

Relationship of serum folic acid and vitamin B12 contents with maternal endothelial injury and placental ischemia hypoxia in patients with preeclampsia

Objective:To investigate the relationship of serum folic acid and vitamin B12 contents with maternal endothelial injury and placental ischemia hypoxia in patients with preeclampsia. Methods:A total of 76 puerperae with preeclampsia who gave birth in this hospital between March 2016 and October 2017 were selected as the preeclampsia group, and 100 healthy puerperae who gave birth in this hospital during the same period were selected as the normal control group. The differences in serum contents of folic acid, vitamin B12, endothelial injury indexes and oxidative stress indexes were compared between the two groups 24 h before delivery. Pearson test was used to assess the intrinsic relationship of serum folic acid and vitamin B12 contents with the disease severity in puerperae with preeclampsia.Results:Serum folic acid and vitamin B12 contents of preeclampsia group were lower than those of control group;endothelial injury markers sEng, sFlt-1, ET-1 and TM contents were higher than those of control group whereas ADM content was lower than that of control group;oxidative stress indexes AOPPs and LHP contents were higher than those of normal control group whereas T-AOC content was lower than that of control group. The Pearson test showed that serum folic acid and vitamin B12 contents in puerperae with preeclampsia were directly correlated with vascular endothelial injury and placental ischemia hypoxia.Conclusion: There is folic acid and vitamin B12 deficiency in patients with preeclampsia, and it can aggravate the vascular endothelial injury and placental ischemia hypoxia.

Nicotinamide adenine dinucleotide treatment confers resistance to neonatal ischemia and hypoxia:effects on neurobehavioral phenotypes

Neonatal hypoxic-ischemic brain injury is the main cause of hypoxic-ischemic encephalopathy and cerebral palsy.Currently,there are few effective clinical treatments for neonatal hypoxic-ischemic brain injury.Here,we investigated the neuroprotective and molecular mechanisms of exogenous nicotinamide adenine dinucleotide,which can protect against hypoxic injury in adulthood,in a mouse model of neonatal hypoxic-ischemic brain injury.In this study,nicotinamide adenine dinucleotide(5 mg/kg)was intraperitoneally administered 30 minutes befo re surgery and every 24 hours thereafter.The results showed that nicotinamide adenine dinucleotide treatment improved body weight,brain structure,adenosine triphosphate levels,oxidative damage,neurobehavioral test outcomes,and seizure threshold in experimental mice.Tandem mass tag proteomics revealed that numerous proteins were altered after nicotinamide adenine dinucleotide treatment in hypoxic-ischemic brain injury mice.Parallel reaction monitoring and western blotting confirmed changes in the expression levels of proteins including serine(or cysteine)peptidase inhibitor,clade A,member 3N,fibronectin 1,5'-nucleotidase,cytosolic IA,microtubule associated protein 2,and complexin 2.Proteomics analyses showed that nicotinamide adenine dinucleotide ameliorated hypoxic-ischemic injury through inflammation-related signaling pathways(e.g.,nuclear factor-kappa B,mitogen-activated protein kinase,and phosphatidylinositol 3 kinase/protein kinase B).These findings suggest that nicotinamide adenine dinucleotide treatment can improve neurobehavioral phenotypes in hypoxic-ischemic brain injury mice through inflammation-related pathways.

Ultrasonic evaluation of uterine spiral artery characteristics in patients with gestational hypertension and their correlation with the placental hypoxia injury

Objective: To investigate the ultrasonic evaluation of uterine spiral artery characteristics in patients with gestational hypertension and their correlation with the placental hypoxia injury. Methods: A total of 128 puerperae with gestational hypertension who gave birth in this hospital between December 2014 and February 2017 were collected as gestational hypertension group, and 100 healthy puerperae who gave birth in this hospital during the same period were collected as normal control group. The differences in uterine spiral artery parameter levels as well as the contents of angiogenesis indexes, oxidative stress indexes and inflammatory mediators in the placental grinding fluid were compared between the two groups 1 week before delivery. Pearson test was used to evaluate the correlation between uterine spiral artery characteristics and placental hypoxia injury in patients with gestational hypertension. Results: Uterine spiral artery RI, PI and S/D levels in gestational hypertension group were higher than those in normal control group;angiogenesis indexes HIF-1α, VEGF and sFlt-1 contents in placental grinding fluid of gestational hypertension group were higher than those of normal control group;anti-oxidation indexes CTA and SOD contents were lower than those of control group whereas LHP content was higher than that of normal control group;inflammatory mediators IL-6, TNF-α, NF-κB and HSP70 contents were higher than those of normal control group. Pearson test showed that the uterine spiral artery RI, PI and S/D levels in puerperae with gestational hypertension were positively correlated with the degree of angiogenesis, oxidative stress and inflammatory response in the placenta. Conclusion: There are abnormal changes of uterine spiral artery flow parameters in puerperae with gestational hypertension, and the abnormality can objectively reflect the degree of placental hypoxia injury.

Over-starvation aggravates intestinal injury and promotes bacterial and endotoxin translocation under high-altitude hypoxic environment

AIM:To study whether over-starvation aggravates intestinal mucosal injury and promotes bacterial and endotoxin translocation in a high-altitude hypoxic environment.METHODS:Sprague-Dawley rats were exposed to hy-pobaric hypoxia at a simulated altitude of 7000 m for 72 h.Lanthanum nitrate was used as a tracer to detect intestinal injury.Epithelial apoptosis was observed with terminal deoxynucleotidyl transferase dUTP nick end labeling staining.Serum levels of diamino oxidase(DAO),malondialdehyde(MDA),glutamine(Gln),superoxide dismutase(SOD) and endotoxin were measured in intestinal mucosa.Bacterial translocation was detected in blood culture and intestinal homogenates.In addition,rats were given Gln intragastrically to observe its protective effect on intestinal injury.RESULTS:Apoptotic epithelial cells,exfoliated villi and inflammatory cells in intestine were increased with edema in the lamina propria accompanying effusion of red blood cells.Lanthanum particles were found in the intercellular space and intracellular compartment.Bacterial translocation to mesenteric lymph nodes(MLN) and spleen was evident.The serum endotoxin,DAO and MDA levels were significantly higher while the serum SOD,DAO and Gln levels were lower in intestine(P< 0.05).The bacterial translocation number was lower in the high altitude hypoxic group than in the high altitude starvation group(0.47±0.83 vs 2.38±1.45,P<0.05).The bacterial translocation was found in each organ,especially in MLN and spleen but not in peripheral blood.The bacterial and endotoxin translocations were both markedly improved in rats after treatment with Gln.CONCLUSION:High-altitude hypoxia and starvation cause severe intestinal mucosal injury and increase bacterial and endotoxin translocation,which can be treated with Gln.

An animal model of cerebral palsy induced by prenatal exposure to lipopolysaccharide and hypoxia

BACKGROUND： Neonatal cerebral palsy is mainly caused by prenatal factors. At present, an animal model of prenatal infection and early postnatal hypoxia does not exist. OBJECTIVE： To observe morphology and motor performance following prenatal infection and hypoxic insult-induced brain damage of neonatal rats to verify the feasibility to establish a model of cerebral palsy. DESIGN, TIME AND SETTING： A randomized, controlled, animal experiment was performed at the Laboratories of Xinjiang Center for Disease Control and Prevention from September 2007 to June 2008. MATERIALS： The hypoxic incubator was purchased from Shanghai Pediatric Medical Institute, China. Lipopolysaccharide （LPS, Escherichia coil, 055： B5） was purchased from Sigma-Aldrich （St. Louis, MO, USA）. METHODS： A total of 27 Wistar rats, aged 7 days, were randomly assigned to sham-surgery group （n = 15） with no carotid artery incision or hypoxia treatment, hypoxia/ischemia （H/I） group （n = 12） undergoing ligature of the right common carotid artery followed by exposure to hypoxia at postnatal day 7 （P7）, and LPS/H group （n = 19）, in which pregnant rats were exposed in utero to LPS followed by prenatal hypoxia at embryonic day 16. MAIN OUTCOME MEASURES： Behavior, compound muscle action potential, and pathological changes were observed in 28-day-old rats. RESULTS： The footprint repeat space showed that left limb footprint repeatability in the H/I and LPS/H groups was lower than in the sham-surgery group （P 〈 0.05）. The space between the footprints was larger and unstable. Hind limb quadricep compound muscle action potential in the H/I and LPS/H groups showed lower wave amplitude compared with the sham-surgery group （P〈 0.05） Hematoxylin and eosin staining showed irregular cells around the ventricle, as well as periventricular leukomalacia. CONCLUSION： An animal model of cerebral palsy was established, which simulated the human condition most likely associated with occurrence of this disease. This model could be used for experimental studies related to cerebral palsy.

Effect of lentiviral vector-mediated overexpression of hypoxia-inducible factor 1 alpha delivered by pluronic F-127 hydrogel on brachial plexus avulsion in rats

Brachial plexus avulsion often results in massive motor neuron death and severe functional deficits of target muscles. However, no satisfactory treatment is currently available. Hypoxia-inducible factor 1α is a critical molecule targeting several genes associated with ischemia-hypoxia damage and angiogenesis. In this study, a rat model of brachial plexus avulsion-reimplantation was established, in which C5–7 ventral nerve roots were avulsed and only the C6 root reimplanted. Different implants were immediately injected using a microsyringe into the avulsion-reimplantation site of the C6 root post-brachial plexus avulsion. Rats were randomly divided into five groups: phosphate-buffered saline, negative control of lentivirus, hypoxia-inducible factor 1α(hypoxia-inducible factor 1α overexpression lentivirus), gel(pluronic F-127 hydrogel), and gel + hypoxia-inducible factor 1α(pluronic F-127 hydrogel + hypoxia-inducible factor 1α overexpression lentivirus). The Terzis grooming test was performed to assess recovery of motor function. Scores were higher in the hypoxia-inducible factor 1α and gel +hypoxia-inducible factor 1α groups(in particular the gel + hypoxia-inducible factor 1α group) compared with the phosphate-buffered saline group. Electrophysiology, fluorogold retrograde tracing, and immunofluorescent staining were further performed to investigate neural pathway reconstruction and changes of neurons, motor endplates, and angiogenesis. Compared with the phosphate-buffered saline group, action potential latency of musculocutaneous nerves was markedly shortened in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor1α groups. Meanwhile, the number of fluorogold-positive cells and ChAT-positive neurons, neovascular area(labeled by CD31 around av ulsed sites in ipsilateral spinal cord segments), and the number of motor endplates in biceps brachii(identified by α-bungarotoxin) were all visibly increased, as well as the morphology of motor endplate in biceps brachil was clear in the hypoxia-inducible factor 1α and gel + hypoxia-inducible factor 1α groups. Taken together, delivery of hypoxia-inducible factor 1α overexpression lentiviral vectors mediated by pluronic F-127 effectively promotes spinal root regeneration and functional recovery post-brachial plexus avulsion. All animal procedures were approved by the Institutional Animal Care and Use Committee of Guangdong Medical University, China.

Acute high-altitude hypoxic brain injury Identification of ten differential proteins

Hypobaric hypoxia can cause severe brain damage and mitochondrial dysfunction, and is involved in hypoxic brain injury. However, little is currently known about the mechanisms responsible for mi- tochondrial dysfunction in hypobaric hypoxic brain damage. In this study, a rat model of hypobaric hypoxic brain injury was established to investigate the molecular mechanisms associated with mi- tochondrial dysfunction. As revealed by two-dimensional electrophoresis analysis, 16, 21, and 36 differential protein spots in cerebral mitochondria were observed at 6, 12, and 24 hours post-hypobaric hypoxia, respectively. Furthermore, ten protein spots selected from each hypobaric hypoxia subgroup were similarly regulated and were identified by mass spectrometry. These de- tected proteins included dihydropyrimidinase-related protein 2, creatine kinase B-type, is- ovaleryI-CoA dehydrogenase, elongation factor Ts, ATP synthase beta-subunit, 3-mercaptopyruvate sulfurtransferase, electron transfer flavoprotein alpha-subunit, Chain A of 2-enoyI-CoA hydratase, NADH dehydrogenase iron-sulfur protein 8 and tropomyosin beta chain. These ten proteins are all involved in the electron transport chain and the function of ATP synthase. Our findings indicate that hypobaric hypoxia can induce the differential expression of several cerebral mitochondrial proteins, which are involved in the regulation of mitochondrial energy production.

Increased susceptibility of aging gastric mucosa to injury:The mechanisms and clinical implications

This review updates the current views on aging gastric mucosa and the mechanisms of its increased susceptibility to injury. Experimental and clinical studies indicate that gastric mucosa of aging individuals-&#x0201c;aging gastropathy&#x0201d;-has prominent structural and functional abnormalities vs young gastric mucosa. Some of these abnormalities include a partial atrophy of gastric glands, impaired mucosal defense (reduced bicarbonate and prostaglandin generation, decreased sensory innervation), increased susceptibility to injury by a variety of damaging agents such as ethanol, aspirin and other non-steroidal anti-inflammatory drugs (NSAIDs), impaired healing of injury and reduced therapeutic efficacy of ulcer-healing drugs. Detailed analysis of the above changes indicates that the following events occur in aging gastric mucosa: reduced mucosal blood flow and impaired oxygen delivery cause hypoxia, which leads to activation of the early growth response-1 (egr-1) transcription factor. Activation of egr-1, in turn, upregulates the dual specificity phosphatase, phosphatase and tensin homologue deleted on chromosome ten (PTEN) resulting in activation of pro-apoptotic caspase-3 and caspase-9 and reduced expression of the anti-apoptosis protein, survivin. The imbalance between pro- and anti-apoptosis mediators results in increased apoptosis and increased susceptibility to injury. This paradigm has human relevance since increased expression of PTEN and reduced expression of survivin were demonstrated in gastric mucosa of aging individuals. Other potential mechanisms operating in aging gastric mucosa include reduced telomerase activity, increase in replicative cellular senescence, and reduced expression of vascular endothelial growth factor and importin-&#x003b1;-a nuclear transport protein essential for transport of transcription factors to nucleus. Aging gastropathy is an important and clinically relevant issue because of: (1) an aging world population due to prolonged life span; (2) older patients have much greater risk of gastroduodenal ulcers and gastrointestinal complications (e.g., NSAIDs-induced gastric injury) than younger patients; and (3) increased susceptibility of aging gastric mucosa to injury can be potentially reduced or reversed pharmacologically.

Hypoxia inducible factor-1α mediates protective effects of ischemic preconditioning on ECV-304 endothelial cells

AIM: To investigate whether hypoxia inducible factor-1α (HIF-1α) is linked to the protective effects of ischemic preconditioning (IP) on sinusoidal endothelial cells against ischemia/reperfusion injury. METHODS: Sinusoidal endothelial cell lines ECV-304 were cultured and divided into four groups: control group, cells were cultured in complete DMEM medium; cold anoxia/warm reoxygenation (A/R) group, cells were preserved in a 4℃ UW solution in a mixture of 95% N2 and 5% CO2 for 24 h; anoxia-preconditioning (APC) group, cells were treated with 4 cycles of short anoxia and reoxygenation before prolonged anoxia- preconditioning treatment; and anoxia-preconditioning and hypoxia inducible factor-1α (HIF-1α) inhibitor (I-HIF-1) group, cells were pretreated with 5 μm of HIF-1α inhibitor NS398 in DMEM medium before subjected to the same treatment as group APC. After the anoxia treatment, each group was reoxygenated in a mixture of 95% air and 5% CO2 incubator for 6 h. Cytoprotections were evaluated by cell viabilities from Trypan blue, lactate dehydrogenase (LDH) release rates, and intracellular cell adhesion molecule-1 (ICAM-1) expressions. Expressions of HIF-1α mRNA and HIF-1α protein from each group were determined by the RT-PCR method and Western blotting, respectively. RESULTS: Ischemia preconditioning increased cell viability, and reduced LDH release and ICAM-1 expressions. Ischemia preconditioning also upregulated the HIF-1α mRNA level and HIF-1α protein expression. However, all of these changes were reversed by HIF-1α inhibitor NS398.CONCLUSION: Ischemia preconditioning effectively inhibited cold hypoxia/warm reoxygenation injury to endothelial cells, and the authors showed for the first time HIF-1α is causally linked to the protective effects of ischemic preconditioning on endothelial cells.