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Vascular complications from diabetes often result in poor outcomes for patients, even after optimized interventions. Forkhead box protein O1 (FoxO1) is a key regulator of cellular metabolism and plays an important role in vessel formation and maturation. Alterations of FoxO1 occur in the cardiovascular system in diabetes, yet the role of FoxO1 in diabetic vascular complications is poorly understood. In Streptozotocin (STZ)‐induced type 1 diabetic rats, FoxO1 expression was up‐regulated in carotid arteries at 8 weeks of diabetes that was accompanied with adverse vascular remodelling characterized as increased wall thickness, carotid medial cross‐sectional area, media‐to‐lumen ratio and decreased carotid artery lumen area. This adverse vascular remodelling induced by hyperglycaemia in diabetic rats required FoxO1 activation as pharmacological inhibition of FoxO1 with 50mg/kg AS1842856 (AS) reversed vascular remodelling in type 1 diabetic rats. The adverse vascular remodelling in type 1 diabetes mellitus (T1DM) occurred concomitantly with increases in pro‐inflammatory factors, adhesion factors, apoptosis, NOD‐like receptor family protein‐3 inflammasome activation and the phenotypic switch of arterial smooth muscle cells, which were all reversed by AS. In addition, FoxO1 inhibition counteracted the down‐regulation of its upstream mediator PDK1 in T1DM. PDK1 activator reduced FoxO1 nuclear translocation, which serves as the basis for subsequent transcriptional regulation during hyperglycaemia. Taken together, our data suggest that FoxO1 is a critical trigger for type 1 diabetes‐induced vascular remodelling in rats, and inhibition of FoxO1 thus offers a potential therapeutic option for diabetes‐associated cardiovascular diseases.

Forkhead box protein O1 (FOXO1), a nuclear transcription factor, is preferably activated in the myocardium of diabetic mice. However, its role and mechanism in the development of diabetic cardiomyopathy in non‐obese insulin‐deficient diabetes are unclear. We hypothesized that cardiac FOXO1 over‐activation was attributable to the imbalanced myocardial oxidative metabolism and mitochondrial and cardiac dysfunction in type 1 diabetes. FOXO1‐selective inhibitor AS1842856 was administered to streptozotocin‐induced diabetic (D) rats, and cardiac functions, mitochondrial enzymes PDK4 and CPT1 and mitochondrial function were assessed. Primary cardiomyocytes isolated from non‐diabetic control (C) and D rats were treated with or without 1 µM AS1842856 and underwent Seahorse experiment to determine the effects of glucose, palmitate and pyruvate on cardiomyocyte bioenergetics. The results showed diabetic hearts displayed elevated FOXO1 nuclear translocation, concomitant with cardiac and mitochondrial dysfunction (manifested as elevated mtROS level and reduced mitochondrial membrane potential) and increased cell apoptosis (all P

Hypoxia‐inducible factor 1α (HIF‐1α) plays a critical role in the apoptotic process during cardiac ischaemia/reperfusion (I/R) injury. This study aimed to investigate whether post‐treatment with dexmedetomidine (DEX) could protect against I/R‐induced cardiac apoptosis in vivo and in vitro via regulating HIF‐1α signalling pathway. Rat myocardial I/R was induced by occluding the left anterior descending artery for 30 minutes followed by 6‐hours reperfusion, and cardiomyocyte hypoxia/reoxygenation (H/R) was induced by oxygen‐glucose deprivation for 6 hours followed by 3‐hours reoxygenation. Dexmedetomidine administration at the beginning of reperfusion or reoxygenation attenuated I/R‐induced myocardial injury or H/R‐induced cell death, alleviated mitochondrial dysfunction, reduced the number of apoptotic cardiomyocytes, inhibited the activation of HIF‐1α and modulated the expressions of apoptosis‐related proteins including BCL‐2, BAX, BNIP3, cleaved caspase‐3 and cleaved PARP. Conversely, the HIF‐1α prolyl hydroxylase‐2 inhibitor IOX2 partly blocked DEX‐mediated cardioprotection both in vivo and in vitro. Mechanistically, DEX down‐regulated HIF‐1α expression at the post‐transcriptional level and inhibited the transcriptional activation of the target gene BNIP3. Post‐treatment with DEX protects against cardiac I/R injury in vivo and H/R injury in vitro. These effects are, at least in part, mediated via the inhibition of cell apoptosis by targeting HIF‐1α signalling.

Hepatic ischaemia/reperfusion (HIR) induces severe damage on hepatocyte cell membrane, which leads to hepatocyte death and the subsequent HIR injury. In this study, we investigated the role and the mechanism of mitsugumin‐53 (MG53), a novel cell membrane repair protein, in protecting the liver against HIR injury. Rats were subjected to sham operation or 70% warm HIR with or without recombined MG53 (rhMG53), caudal vein‐injected 2 hrs before inducing HIR. In vitro, cultured hepatocyte AML12 cells were subjected to hypoxia/reoxygenation (H/R) in the presence of rhMG53 and/or dysferlin gene shRNAs or adenovirus transfection. HIR resulted in severe liver injury manifested as severe liver histological changes and increased AST and ALT release. Post‐ischaemic hepatic oxidative stress was significantly enhanced demonstrated by elevated dihydroethidium level, increased 4‐hydroxynonenal, enhanced 15‐F2t‐isoprostane and decreased SOD activity. rhMG53 administration attenuated post‐HIR liver injury, decreased liver oxidative stress and further enhanced dysferlin protein expression and its colocalization with MG53. Similarly, H/R induced AML12 cell injury and oxidative stress, which were abolished by either rhMG53 or dysferlin overexpression but were exacerbated by dysferlin gene knockdown. Dysferlin overexpression further increased H/R‐induced increased colocalization of MG53 and dysferlin. In conclusion, MG53 was anchored by dysferlin to reduce oxidative stress and cell death and attenuate HIR injury.

Background Ischemic heart disease is the leading cause of mortality in patients with diabetes mellitus. Although remote ischemic preconditioning achieved by non-invasively occluding the brachial ar...

Bupivacaine, a commonly used local anesthetic, has potential neurotoxicity through diverse signaling pathways. However, the key mechanism of bupivacaine-induced neurotoxicity remains unclear. Cultured human SH-SY5Y neuroblastoma cells were treated (bupivacaine) or untreated (control) with bupivacaine for 24 h. Compared to the control group, bupivacaine significantly increased cyto-inhibition, cellular reactive oxygen species, DNA damage, mitochondrial injury, apoptosis (increased TUNEL-positive cells, cleaved caspase 3, and Bcl-2/Bax), and activated autophagy (enhanced LC3II/LC3I ratio). To explore changes in protein expression and intercommunication among the pathways involved in bupivacaine-induced neurotoxicity, an 8-plex iTRAQ proteomic technique and bioinformatics analysis were performed. Compared to the control group, 241 differentially expressed proteins were identified, of which, 145 were up-regulated and 96 were down-regulated. Bioinformatics analysis of the cross-talk between the significant proteins with altered expression in bupivacaine-induced neurotoxicity indicated that phosphatidyl-3-kinase (PI3K) was the most frequently targeted protein in each of the interactions. We further confirmed these results by determining the downstream targets of the identified signaling pathways (PI3K, Akt, FoxO1, Erk, and JNK). In conclusion, our study demonstrated that PI3K may play a central role in contacting and regulating the signaling pathways that contribute to bupivacaine-induced neurotoxicity.

Background Xanthine oxidase-derived superoxide production and oxidative stress contribute to the development of diabetic complications including diabetic cardiomyopathy. We hypothesized that xanthine oxidase-inhibitor allopurinol (ALP) may decrease hyperglycemia-induced oxidative stress, ameliorate cardiomyocyte hypertrophy and fibrosis, and attenuate the development of left ventricle (LV) diastolic dysfunction in rats with streptozotocin (STZ)-induced diabetes. Methods Control Sprague Dawley (C) or streptozotocin-induced diabetic (D) rats were either untreated or treated with allopurinol (100 mg/kg/day) for 4 weeks starting at 1 week after streptozotocin injection. Free 15-F2t-isoprostane, a specific indicator of oxidative stress was measured by enzymatic immunoassay. The cardiomyocyte cross-sectional area was assessed by hematoxylin and eosin-stained paraffin-embedded sections of LVs. Myocardial collagens I and III were assessed by immunol histochemistry and Western blotting. Echocardiography was performed to characterize cardiac structure and function. Results In diabetic rats, both plasma and cardiac tissue levels of free 15-F2t-isoprostane were increased (p

Summary 1. Indole-3-carbinol (I3C), a major indole compound found in high levels in cruciferous vegetables, shows a broad spectrum of biological activities. However, few studies have reported the effect of I3C on alcoholic liver injury. In the present study, we investigated the protective effect of I3C on acute ethanol-induced hepatotoxicity and acetaldehyde-stimulated hepatic stellate cells (HSC) activation using precision-cut liver slices (PCLS). 2. Rat PCLS were incubated with 50 mmol/L ethanol or 350 μmol/L acetaldehyde, and different concentrations (100–400 μmol/L) of I3C were added into the culture system of these two liver injury models, respectively. Hepatotoxicity was assessed by measuring enzyme leakage and malondialdehyde (MDA) content in tissue. Activities of alcoholic enzymes were also determined. α-Smooth muscle actin (α-SMA), transforming growth factor (TGF-β1) and hydroxyproline (HYP) were used as indices to evaluate the activation of HSC. In addition, matrix metalloproteinase-1 (MMP-1) and the tissue inhibitor of metalloproteinase (TIMP-1) were observed to estimate collagen degradation. 3. I3C significantly reduced the enzyme leakage in ethanol-treated slices. In I3C groups, cytochrome P450 (CYP) 2E1 activities were inhibited by 40.9–51.8%, whereas alcohol dehydrogenase (ADH) activity was enhanced 1.6-fold compared with the ethanol-treated group. I3C also showed an inhibitory effect against HSC activation and collagen production stimulated by acetaldehyde. After being incubated with I3C (400 μmol/L), the expression of MMP-1 was markedly enhanced, whereas TIMP-1 was decreased. 4. These results showed that I3C protected PCLS against alcoholic liver injury, which might be associated with the regulation of ethanol metabolic enzymes, attenuation of oxidative injury and acceleration of collagen degradation.

Intestinal ischemia/reperfusion (I/R) injury is a critical condition associated with high morbidity and mortality. Studies show that ischemic preconditioning (IPC) can protect the intestine from I/R injury. However, the underlying molecular mechanisms of this event have not been fully elucidated. In the present study, 2-DE combined with MALDI-MS was employed to analyze intestinal mucosa proteomes of rat subjected to I/R injury in the absence or presence of IPC pretreatment. The protein content of 16 proteins in the intestinal mucosa changed more than 1.5-fold following intestinal I/R. These proteins were, respectively, involved in the cellular processes of energy metabolism, anti-oxidation and anti-apoptosis. One of these proteins, aldose reductase (AR), removes reactive oxygen species. In support of the 2-DE results, the mRNA and protein expressions of AR were significantly downregulated upon I/R injury and enhanced by IPC as confirmed by RT-PCR and western blot analysis. Further study showed that AR-selective inhibitor epalrestat totally turned over the protective effect of IPC, indicating that IPC confers protection against intestinal I/R injury primarily by increasing intestinal AR expression. The finding that AR may play a key in intestinal ischemic protection might offer evidences to foster the development of new therapies against intestinal I/R injury.

Background: Intrathecal morphine pre-conditioning attenuates cardiac ischemia―reperfusion injury via activation of central opioid receptors. We hypothesized that intrathecal morphine also post-conditions the myocardium in the rat. Methods: Intrathecal morphine at 0.3μg/kg (LMPC), 3μg/kg (MMPC) or 30μg/kg (HMPC) was administered for 5 min before 120-min reperfusion following 30-min ischemia. Infarct size as a percentage of area at risk (IS/ AAR) was determined using triphenyltetrazolium staining. MMPC was repeated following the intrathecal administration of nor BNI, NTD, CTOP, or naloxone methiodide (NM), kappa, delta, mu and non-specific opioid receptor antagonists, respectively. The role of peripheral opioid, adenosine and calcitonin gene-related peptide (CGRP) receptors was examined by the intravenous administration of NM, 8-p-sulfophenyl theophylline (8-SPT) and human CGRP fragment (CGRP 8 ― 37 ), respectively. Results: Morphine post-conditioning at all three doses was cardioprotective (IS/AAR of LMPC = 37 ± 4%, MMPC = 35 ± 5%, HMPC = 32 ± 4%, control = 50 ± 5%, P < 0.01). The prior administration of opioid receptor antagonists intrathecally as well as intravenous 8-SPT and CGRP 8 ― 37 receptor antagonists, abolished this effect (nor BNI+MMPC = 47 ± 7%, NTD+MMPC = 49 ± 7%, CTOP+MMPC = 45 ± 9%, NM+MMPC = 47 ± 6% 8-SPT+ MPC = 46 ± 5% & CGRP 8 ― 37 +MPC = 53 ± 6%, P = 0.63). However, the intravenous administration of NM did not prevent the protective effect (34 ± 4%, P

Dexmedetomidine (DEX), a highly selective α2-adrenoceptor agonist, produces both sedative and antinociceptive action. However, the mechanism for DEX analgesia remains unclear. Opening of G protein-...

Ischemic postconditioning (IPo) protects against myocardial ischemic reperfusion (IR) injury by activating signal transducer and activator of transcription 3 (STAT3) which requires adiponectin (APN) (FASEB, 2014, 28: 663.7), but how APN activates STAT3 during IPo is unclear. Caveolin-3 (Cav3), a protein of the lipid rafts/caveolae, is essential in APN receptor 1 (AdipoR1) transmembrane signaling. We postulated that IPo activates mitochondrial STAT3 (mitoSTAT3) via Cav3 AdipoR1 signaling. Wild type (WT) and APN knockout (KO) mice were subjected to sham operation or IR (30min of coronary artery occlusion and 2 hours of reperfusion) with or without IPo (3 cycles of 10s of reperfusion and 10s of reocclusion). KO mice exhibited more severe myocardial injury (increased post-ischemic infarction and poorer functional recovery; P

Protein tyrosine phosphatase 1B (PTP1B) acts as a physiological negative regulator of insulin signaling by dephosphorylating the activated insulin receptor (IR). Here we examine the role of PTP1B in the insulin-sensitizing action of rosiglitazone (RSG) in skeletal muscle and liver. Fat-fed, streptozotocin-treated rats (10-week-old), an animal model of type II diabetes, and age-matched, nondiabetic controls were treated with RSG (10 micromol kg(-1) day(-1)) for 2 weeks. After RSG treatment, the diabetic rats showed a significant decrease in blood glucose and improved insulin sensitivity. Diabetic rats showed significantly increased levels and activities of PTP1B in the skeletal muscle (1.6- and 2-fold, respectively) and liver (1.7- and 1.8-fold, respectively), thus diminishing insulin signaling in the target tissues. We found that the decreases in insulin-stimulated glucose uptake (55%), tyrosine phosphorylation of IRbeta-subunits (48%), and IR substrate-1 (IRS-1) (39%) in muscles of diabetic rats were normalized after RSG treatment. These effects were associated with 34 and 30% decreases in increased PTP1B levels and activities, respectively, in skeletal muscles of diabetic rats. In contrast, RSG did not affect the increased PTP1B levels and activities or the already reduced insulin-stimulated glycogen synthesis and tyrosine phosphorylation of IRbeta-subunits and IRS-2 in livers of diabetic rats. RSG treatment in normal rats did not significantly change PTP1B activities and levels or protein levels of IRbeta, IRS-1, and -2 in diabetic rats. These data suggest that RSG enhances insulin activity in skeletal muscle of diabetic rats possibly by ameliorating abnormal levels and activities of PTP1B.

Endothelin-1 (ET-1) is a 21 amino acid vasoconstrictor and implicated in the regulation of hypertention. Recently, our colleagues found that transgenic mice (TET-1) with endothelial ET-1 over-expression developed hypertention in young mice by changing vascular properties. However, the effects of over-expressing endothelial ET-1 on cardiovascular system in aged mice are still unclear. Therefore, this study investigated the blood pressure and heart rate of TET-1 aged mice and non-transgenic (NTg) aged mice (1.5-2 years old). A non-invasive tail cuff measurement showed that a significant elevation of both systolic (115.1 ± 2.935 vs 103.5 ± 1.846 mmHg, p

C-X-C chemokine receptor type 4 (CXCR4) is an emerging area of pain hypersensitivity in the peripheral nervous system; however, its role in the central nervous system (CNS) for pain processing remains unclear. This study aimed at exploring the function of central CXCR4 in pain processing in vivo using its specific antagonist AMD3100 and partial sciatic nerve ligation (pSNL) model of peripheral neuropathic pain in C57BL/6 mice. A single intrathecal (central) administration of AMD3100 (intrathecal AMD3100, 5 μg) was found to prevent the development of pSNL-induced allodynia. Intrathecal AMD3100 (1 μg, 5 μg and 25 μg) also reversed the established pSNL-induced mechanical allodynia in a dose-dependent way on post-operative day 7 (POD 7). In rotarod test, intrathecal AMD3100 of different dosages (1 μg, 5 μg and 25 μg) did not impair the motor function of mice. Among mitogen-activated protein kinases (MAPKs) pathways, intrathecal AMD3100 was found to downregulate the activation of JNK1 and p38 pathway signaling...

Ischemic postconditioning (IPostC) protects the heart against ischemia reperfusion injury (IRI) by activating signal transducer and activator of transcription 3 (STAT3). Adiponectin (APN), a protein with anti-IRI property, activates STAT3 (Cell Metab. 2011, 13: 401-12), but its role in IPostC is unknown. We postulated that APN is essential for IPostC to confer cardioprotection. Wild type (WT) and APN knockout (KO) mice were subjected to 30 min coronary occlusion followed by 120 min reperfusion without or with IPostC achieved by 3 cycles of 10s of ischemia/10s of reperfusion. Cardiac function was assessed by pressure-volume (PV) loop system. At the end of reperfusion, KO mice displayed larger infarct size and higher plasma creatine kinase-MB, but lower end-systolic PV relation (a reliable measure of ventricular function) and dP/dt (all P

Dexmedetomidine(DEX) has been shown to attenuate neuropathic pain (NP), but the mechanism is unclear. Astrocytes connexin 43(Cx43) is increased in subjects with NP and plays an important role in the development and maintenance of NP (Glia. 2012;60(11):1660-70). We thus investigated the role of astroglia Cx43 in DEX-induced anti-nociceptive effects in rats with NP. Peripheral NP was induced by partial sciatic nerve ligation (PSNL) in SD rats. After 10 days of inducing NP, DEX was intrathecally administrated for 5 days and ipsilateral mechanical allodynia and thermal hyperalgesia were examined by Von Frey Test and Hargreaves Test. Compared with control rats, PSNL induced significant ipsilateral mechanical allodynia and thermal hyperalgesia (P

Dexmedetomidine(DEX), an alpha-2 adrenergic agonist which can affect cell-to-cell gap junctions(GJ), is used as an adjuvant analgesic and sedatives in patients with cancer pain, especially in neura...