Ischemic Susceptibility Research Articles

Cardioprotective O-GlcNAc signaling is elevated in murine female hearts via enhanced O-GlcNAc transferase activity

The post-translational modification of intracellular proteins by O-linked β-N-acetylglucosamine (O-GlcNAc) has emerged as a critical regulator of cardiac function. Enhanced O-GlcNAcylation activates cytoprotective pathways in cardiac models of ischemia reperfusion (I/R) injury; however, the mechanisms underpinning O-GlcNAc-cycling in response to I/R injury have not been comprehensively assessed. The cycling of O-GlcNAc is regulated by the collective efforts of two enzymes: O-GlcNAc transferase (OGT) and O-GlcNAcase (OGA), which catalyze the addition and hydrolysis of O-GlcNAc respectively. It has previously been shown that baseline heart physiology and pathophysiology are impacted by sex. Here, we hypothesized that sex differences in molecular signaling may target protein O-GlcNAcylation both basally and in ischemic hearts. To address this question, we subjected male and female wild type murine hearts to ex vivo ischemia or ischemia-reperfusion injury. We assessed hearts for protein O-GlcNAcylation, abundance of OGT, OGA, and glutamine:fructose-6-phosphate aminotransferase (GFAT2), activity of OGT and OGA, and UDP-GlcNAc levels. Our data demonstrate elevated O-GlcNAcylation in female hearts both basally and during ischemia. We show OGT activity was enhanced in female hearts in all treatments, suggesting a mechanism for these observations. Furthermore, we found ischemia led to reduced O-GlcNAcylation and OGT specific activity. Our findings provide a foundation for understanding molecular mechanisms that regulate O-GlcNAcylation in the heart and highlight the importance of sex as a significant factor when assessing key regulatory events that control O-GlcNAc-cycling. These data suggest the intriguing possibility that elevated O-GlcNAcylation in females contributes to reduced ischemic susceptibility.

Bridging a Century-Old Problem: The Pathophysiology and Molecular Mechanisms of HA Filler-Induced Vascular Occlusion (FIVO)-Implications for Therapeutic Interventions.

Biocompatible hyaluronic acid (HA, hyaluronan) gel implants have altered the therapeutic landscape of surgery and medicine, fostering an array of innovative products that include viscosurgical aids, synovial supplements, and drug-eluting nanomaterials. However, it is perhaps the explosive growth in the cosmetic applications of injectable dermal fillers that has captured the brightest spotlight, emerging as the dominant modality in plastic surgery and aesthetic medicine. The popularity surge with which injectable HA fillers have risen to in vogue status has also brought a concomitant increase in the incidence of once-rare iatrogenic vaso-occlusive injuries ranging from disfiguring facial skin necrosis to disabling neuro-ophthalmological sequelae. As our understanding of the pathophysiology of these injuries has evolved, supplemented by more than a century of astute observations, the formulation of novel therapeutic and preventative strategies has permitted the amelioration of this burdensome complication. In this special issue article, we review the relevant mechanisms underlying HA filler-induced vascular occlusion (FIVO), with particular emphasis on the rheo-mechanical aspects of vascular blockade; the thromboembolic potential of HA mixtures; and the tissue-specific ischemic susceptibility of microvascular networks, which leads to underperfusion, hypoxia, and ultimate injury. In addition, recent therapeutic advances and novel considerations on the prevention and management of muco-cutaneous and neuro-ophthalmological complications are examined.

Metformin-Enhanced Cardiac AMP-Activated Protein Kinase/Atrogin-1 Pathways Inhibit Charged Multivesicular Body Protein 2B Accumulation in Ischemia-Reperfusion Injury.

Background: Cardiac autophagic flux is impaired during myocardial ischemia/reperfusion (MI/R). Impaired autophagic flux may exacerbate MI/R injury. Charged multivesicular body protein 2B (CHMP2B) is a subunit of the endosomal sorting complex required for transport (ESCRT-III) complex that is required for autophagy. However, the reverse role of CHMP2B accumulation in autophagy and MI/R injury has not been established. The objective of this article is to elucidate the roles of AMP-activated protein kinase (AMPK)/atrogin-1 pathways in inhibiting CHMP2B accumulation in ischemia–reperfusion injury.Methods: Male C57BL/6 mice (3–4 months) and H9c2 cardiomyocytes were used to evaluate MI/R and hypoxia/reoxygenation (H/R) injury in vivo and in vitro, respectively. MI/R was built by a left lateral thoracotomy and occluded the left anterior descending artery. H9c2 cells were firstly treated in 95% N2 and 5% CO2 for 15 h and reoxygenation for 1 h. Metformin (100 mg/kg/d) and CHMP2B (Ad-CHMP2B) transfected adenoviruses were administered to the mice. The H9c2 cells were treated with metformin (2.5 mM), MG-132 (10 μM), bafilomycin A1 (10 nM), and compound C (20 μM).Results: Autophagic flux was found to be inhibited in H/R-treated cardiomyocytes and MI/R mice, with elevated cardiac CHMP2B accumulation. Upregulated CHMP2B levels in the in vivo and in vitro experiments were shown to inhibit autophagic flux leading to the deterioration of H/R-cardiomyocytes and MI/R injury. This finding implies that CHMP2B accumulation increases the risk of myocardial ischemia. Metformin suppressed CHMP2B accumulation and ameliorated H/R-induced autophagic dysfunction by activating AMPK. Activated AMPK upregulated the messenger RNA expression and protein levels of atrogin-1, a muscle-specific ubiquitin ligase, in the myocardium. Atrogin-1 significantly enhanced the interaction between atrogin-1 and CHMP2B, therefore, promoting CHMP2B degradation in the MI/R myocardium. Finally, this study revealed that metformin-inhibited CHMP2B accumulation induced autophagic impairment and ischemic susceptibility in vivo through the AMPK-regulated CHMP2B degradation by atrogin-1.Conclusion: Impaired CHMP2B clearance in vitro and in vivo inhibits autophagic flux and weakens the myocardial ischemic tolerance. Metformin treatment degrades CHMP2B through the AMPK-atrogin-1-dependent pathway to maintain the homeostasis of autophagic flux. This is a novel mechanism that enriches the understanding of cardioprotection.

Intestinal transplantation: an overview of the recent experimental studies

Experimental intestinal transplantation (ITx ) has generated invaluable knowledge that has ultimately benefited the clinical activity. Herein, we appraise the recent publications pertaining to experimental ITx and highlight the main current research topics. During the recent years, ischemia-reperfusion injury (IRI) and Graft-versus-host disease (GVHD) have gradually replaced acute rejection as the main research topic. New additives to established preservation solutions and relatively novel approaches such as luminal interventions during cold storage may prolong the storage time and alleviate IRI. High donor age does not seem to worsen preservation injury. The ischemic susceptibility seems to differ between species, which may impact the translatability of the experimental findings. A new experimental model of modified multivisceral transplantation including the donor spleen may offer a new tool with which to study GVHD, besides the classical Lewis-Brown Norway rat combination. Flushing the graft with fludarabine may mitigate GVHD in rats. T-cell activation inhibitor-mitochondrial was downregulated in the peripheral blood leukocytes before other signs of acute and severe chronic rejection could be observed. Experimental research in ITx has largely shifted focus from acute rejection to IRI and GVHD. Several lines of research have matured toward clinical translation, yet no breakthrough is imminent.

Protective effects of ischemic preconditioning against neuronal apoptosis and dendritic injury in the hippocampus are age-dependent.

Ischemic preconditioning with non-lethal ischemia can be protective against lethal forebrain ischemia. We hypothesized that aging may aggravate ischemic susceptibility and reduce brain plasticity against preconditioning. Magnetic resonance diffusion tensor imaging (DTI) is a sensitive tool to detect brain integrity and white matter architecture. This study used DTI and histopathology to investigate the effect of aging on ischemic preconditioning. In this study, adult and middle-aged male Mongolian gerbils were subjected to non-lethal 5-min forebrain ischemia (ischemic preconditioning) or sham-operation, followed by 3days of reperfusion, and then lethal 15-min forebrain ischemia. A 9.4-Tesla MR imaging system was used to study DTI indices, namely fractional anisotropy (FA), mean diffusivity (MD), and intervoxel coherence (IC) in the hippocampal CA1 and dentate gyrus (DG) areas. In situ expressions of microtubule-associated protein 2 (MAP2, dendritic marker protein) and apoptosis were also examined. The 5-min ischemia did not cause dendritic and neuronal injury and any significant change in DTI indices and MAP2 in adult and middle-aged gerbils. The 15-min ischemia-induced significant delayed neuronal apoptosis and early dendritic injury evidenced by DTI and MAP2 studies in both CA1 and DG areas with more severe injury in middle-aged gerbils than adult gerbils. Ischemic preconditioning could improve neuronal apoptosis in CA1 area and dendritic integrity in both CA1 and DG areas with better improvement in adult gerbils than middle-aged gerbils. This study thus suggests an age-dependent protective effect of ischemic preconditioning against both neuronal apoptosis and dendritic injury in hippocampus after forebrain ischemia.

Abstract 429: Intensive Necroptosis Due to Enhanced RIP3-CaMKII Signaling Contributes to Diabetic Myocardial Ischemic Susceptibility

Diabetes increases the vulnerability of the heart to ischemic injury. Recent advances have shown that necroptosis is crucially involved in myocardial ischemia/reperfusion (I/R) injury. Our objectives were thus to extend our knowledge to whether and how necroptosis renders diabetic hearts more vulnerable to I/R injury, which involves activation of Ca 2+ -calmodulin-dependent protein kinase (CaMKII) by receptor-interacting protein 3 (RIP3). In this study, diabetes was induced by streptozotocin (40mg/kg, i.v.) and after 6 weeks, anesthetized open-chest diabetic and age-matched control C57 mice were subjected to 30-min regional ischemia (occlusion of LAD coronary artery) followed by 4-h reperfusion. Diabetic animals manifested significantly greater cardiac injury than their littermate controls. Notably, diabetic hearts were sensitized to I/R-induced myocardial necrosis, as evidenced by rise in lactate dehydrogenase (LDH) release and Evans blue dye (EBD) penetration. Notably, compared with control mice, the activation pattern of CaMKII in diabetic hearts subjected to I/R is altered by earlier advanced (60 min after reperfusion) phosphorylation and enhanced oxidation (21.5% increase). Then we proved that inhibition of CaMKII in early stage of I/R could effectively reduce myocardial necroptosis. Further, RIP3 expression and RIP3-CaMKII interaction are markedly enhanced in diabetic hearts during I/R. Inhibition of RIP3 markedly suppresses CaMKII activation and reduces necroptosis. Collectively, these findings identify enhanced RIP3-CaMKII pathway as a key regulator that increases myocardial necroptosis in diabetic I/R hearts.

NOVEL FUNCTION OF SGOL1 IN CARDIOVASCULAR AND GASTROINTESTINAL DEVELOPMENT: UNRAVELING THE MECHANISMS OF CAID SYNDROME

C57Black/6 and SV129 mice are widely used for the production of transgenic mutants in molecular stroke research but the ischemic susceptibility of these strains is influenced by differences in vascular anatomy and the responsiveness to excitotoxins and vasodilatory stimuli. To differentiate between these opposing effects on infarct size, the vascular territory of the two strains was correlated with the hemodynamic, metabolic, and morphological consequences of permanent middle cerebral artery (MCA) occlusion. The vascular anatomy was studied by latex infusion, brain infarction by vital staining, the size of the ischemic penumbra by imaging of ATP and protein synthesis, and blood flow by laser–Doppler flowmetry. In C57Black/6 mice the MCA-supplied vascular territory and the size of brain infarcts were significantly larger than in SV129 mice but the size of the penumbra and the residual blood flow in the center of the MCA-supplying territory were similar in both strains. These findings suggest that differences in infarct size in C57Black/6 and SV129 mice are determined mainly by the vascular anatomy and not by differences in collateral vascular responsiveness or excitotoxicity.

The innovative “Bio-Oil Spread” prevents metabolic disorders and mediates preconditioning-like cardioprotection in rats

Background and aimsObesity is often associated with an increased cardiovascular risk. The food industry and the associated research activities focus on formulating products that are a perfect mix between an adequate fat content and health. We evaluated whether a diet enriched with Bio-Oil Spread (SD), an olive oil-based innovative food, is cardioprotective in the presence of high-fat diet (HFD)-dependent obesity. Methods and ResultsRats were fed for 16 weeks with normolipidic diet (ND; fat: 6.2%), HFD (fat: 42%), and ND enriched with SD (6.2% of fat + 35.8% of SD). Metabolic and anthropometric parameters were measured. Heart and liver structures were analyzed by histochemical examination. Ischemic susceptibility was evaluated on isolated and Langendorff-perfused cardiac preparations. Signaling was assessed by Western blotting. Compared to ND rats, HFD rats showed increased body weight and abdominal obesity, dyslipidemia, and impaired glucose tolerance. Morphological analyses showed that HFD is associated with heart and liver modifications (hypertrophy and steatosis, respectively), lesser evident in the SD group, together with metabolic and anthropometric alterations. In particular, IGF-1R immunodetection revealed a reduction of hypertrophy in SD heart sections. Notably, SD diet significantly reduced myocardial susceptibility against ischemia/reperfusion (I/R) with respect to HFD through the activation of survival signals (Akt, ERK1/2, and Bcl2). Systolic and diastolic performance was preserved in the SD group. ConclusionsWe suggest that SD may contribute to the prevention of metabolic disorders and cardiovascular alterations typical of severe obesity induced by an HFD, including the increased ischemic susceptibility of the myocardium. Our results pave the way to evaluate the introduction of SD in human alimentary guidelines as a strategy to reduce saturated fat intake.

Susceptibility of murine induced pluripotent stem cell-derived cardiomyocytes to hypoxia and nutrient deprivation.

IntroductionInduced pluripotent stem cell-derived cardiomyocytes (iPS-CMs) may be suitable for myocardial repair. While their functional and structural properties have been extensively investigated, their response to ischemia-like conditions has not yet been clearly defined.MethodsiPS-CMs were differentiated and enriched from murine induced pluripotent stem cells expressing enhanced green fluorescent protein (eGFP) and puromycin resistance genes under the control of an α-myosin heavy chain (α-MHC) promoter. iPS-CMs maturity and function were characterized by microscopy, real-time PCR, calcium transient recordings, electrophysiology, and mitochondrial function assays, and compared to those from neonatal murine cardiomyocytes. iPS-CMs as well as neonatal murine cardiomyocytes were exposed for 3 hours to hypoxia (1% O2) and glucose/serum deprivation, and viability, apoptosis markers, reactive oxygen species, mitochondrial membrane potential and intracellular stress signaling cascades were investigated. Then, the iPS-CMs response to mesenchymal stromal cell-conditioned medium was determined.ResultsiPS-CMs displayed key morphological and functional properties that were comparable to those of neonatal cardiomyocytes, but several parameters indicated an earlier iPS-CMs maturation stage. During hypoxia and glucose/serum deprivation, iPS-CMs exhibited a significantly higher proportion of poly-caspase-active, 7-aminoactinomycin D-positive and TUNEL-positive cells than neonatal cardiomyocytes. The average mitochondrial membrane potential was reduced in “ischemic” iPS-CMs but remained unchanged in neonatal cardiomyocytes; reactive oxygen species production was only increased in “ischemic” iPS-CMs, and oxidoreductase activity in iPS-CMs dropped more rapidly than in neonatal cardiomyocytes. In iPS-CMs, hypoxia and glucose/serum deprivation led to upregulation of Hsp70 transcripts and decreased STAT3 phosphorylation and total PKCε protein expression. Treatment with mesenchymal stromal cell-conditioned medium preserved oxidoreductase activity and restored pSTAT3 and PKCε levels.ConclusioniPS-CMs appear to be particularly sensitive to hypoxia and nutrient deprivation. Counteracting the ischemic susceptibility of iPS-CMs with mesenchymal stromal cell-conditioned medium may help enhance their survival and efficacy in cell-based approaches for myocardial repair.

Abstract W MP62: Vitamin D Status, Cerebral Ischemic Susceptibility, and Post-stroke Outcome

Background: Prevalence of vitamin D insufficiency is high in western countries and low vitamin D has been associated with neurovascular injury. To better understand how vitamin D status affects cerebral ischemic susceptibility we assess whether low serum 25-hydroxyvitamin D (25[OH]D), a marker of vitamin D status, is predictive of the ischemic infarct volume and whether it relates to a worse outcome. Methods: We retrospectively analyzed prospective consecutive acute ischemic stroke patients that were evaluated from January 2013 to January 2014 at a tertiary referral center. All patients (n=96) had MRI-proven acute ischemic stroke. Multivariable linear and logistic regression analyses were used to test whether vitamin D represents an independent predictors of infarct volume and poor 90-day outcome (modified Rankin Scale score of >2). Results: There was a significant inverse correlation between infarct volume and serum 25(OH)D concentration in patients with lacunar (r=-.438; p=0.025) as well as non-lacunar (r=-.399; p=0.001) infarcts. Patients with ≥30 ng/mL had smaller infarct volumes than those with <30 ng/mL (Figure). The association of 25(OH)D with ischemic infarct volume was independent of other known predictors of the infarct extent (p<0.001). After adjustment, lower vitamin D (trichotomized as ≥30 vs 29-20 vs <20 ng/mL) was associated with a poor 90-day outcome (OR 9.46, 95%-CI 1.90-47.11, p=0.006). Conclusion: Low vitamin D independently predicts larger infarct volumes and a poor outcome following ischemic stroke. Though causality remains to be proven, our results highlight the need to further explore vitamin D as a promising biomarker for cerebral ischemic vulnerability and potential target for stroke prevention.

Abstract W P110: Preexisting Cerebral Small Vessel Disease Increases Cortical Infarct Volume after Distal Middle Cerebral Artery Occlusion

Background: Cerebral small vessel disease (cSVD) predominantly affects the subcortical white matter. However, little is known about its impact on cortical ischemic susceptibility. We sought to assess whether preexisting cSVD is predictive of the cortical infarct volume following middle cerebral artery (MCA) branch occlusion and whether it relates to a worse 90-day outcome. Methods: We analyzed data from 117 consecutive patients with MCA branch occlusion as documented by admission computed tomography (CT)-angiography. To avoid potential confounding, we excluded patients with subcortical infarction. Baseline clinical, laboratory, and outcome data as well as final cortical infarct volumes on follow-up imaging were retrospectively analyzed from a prospectively collected database. Severity of cSVD was assessed on admission CT using the van Swieten scale (VSS), grading the white matter lesions on a 3-point scale from 0 (absent) to 4 (severe). We also dichotomized the degree of cSVD (VSS 0-2 [absent to moderate] vs. 3-4 [severe]) for statistical purposes. Multivariable linear and logistic regression analyses were used to identify independent predictors of the final infarct volume. Results: A final infarct volume of ≤27 mL best predicted a favorable 90-day outcome (mRS≤2) as determined by receiver operating characteristic curves. Severe cSVD (OR 7.434; 95%-CI 1.549-35.676; p=0.012) was independently associated with infarct volume >27 mL. Patients with severe cSVD had significantly larger cortical lesion volumes compared to patients without (Figure1A). Severe cSVD (OR 3.027; 95%-CI 1.013-9.048; p=0.047) and infarct volume >27 mL (OR 9.038; 95%-CI 1.997-40.899; p=0.004) were independent predictors of a poor 90-day outcome (mRS 3-6; Figure 1B). Conclusion: Severe subcortical cSVD contributes to larger cortical infarct volumes and worse functional outcome adding to the notion that the brain is negatively affected beyond cSVD ‘s macroscopic boundaries.

Longitudinal MR imaging study in the prediction of ischemic susceptibility after cerebral hypoperfusion in rats: Influence of aging and hypertension

Our previous study has shown that aging and hypertension may alter apparent diffusion coefficient (ADC) and cerebral blood flow (CBF) and increase ischemic susceptibility in the non-ischemic rat brain. The present study wishes to further investigate whether aging and hypertension may influence cerebral diffusion/perfusion and increase ischemic susceptibility in the ischemic brain. Brain magnetic resonance (MR) imaging was examined 1day before and 1 and 7days after bilateral common carotid artery occlusion. Young and middle-aged normotensive Wistar-Kyoto rats and young and middle-aged spontaneously hypertensive rats (SHRs) were studied. Infarction occurred mainly in the parietal cortex and was larger in middle-aged SHRs than the other three groups (P<0.05). In pre-operation, ADC was higher and CBF was lower in middle-aged/hypertensive rats than young/normotensive rats (P<0.05). The ADC was higher in the parietal cortex of the rats with infarction at 7days when compared to the rats without infarction [receiver operating characteristic curve (ROC), P=0.001; binary logistic regression (BLR), P=0.006]. However, there was no difference in the hippocampus and thalamus. At day 1 post-operation, CBF reduced and ADC/CBF ratio elevated significantly in the parietal cortex of the rats with infarction when compared to the rats without infarction (CBF: ROC, P=0.002; BLR, P=0.017. ADC/CBF ratio: ROC, P=0.001; BLR, P=0.018). Our results demonstrated that pre-operation ADC and post-operation CBF and ADC/CBF ratio can be used as good MR markers in the prediction of ischemic susceptibility after cerebral hypoperfusion.

Atlas-Based Classification of Hyperintense Regions from MR Diffusion-Weighted Images of the Brain: Preliminary Results

The study of subjects with acquired brain damage in a specific location is important in exploring human brain function. Description of lesion locations within and across subjects is a crucial methodological component that usually involves the distinction of normal from damaged tissue (lesion segmentation) in relation to lesion locations in terms of a standard anatomical reference space (lesion mapping). Our study provides an atlas-based, computer-aided methodology for classification of hyperintense regions on diffusion-weighted images of the brain, representing either ischemic lesions or susceptibility artifacts. We applied a leave-one-out method of cross-validation that computed probabilistic atlases of true lesions and artifacts, based on training data. Our approach accurately classifies lesions and artifacts, but leaves a significant number of regions unclassified, due to the relatively small number of training samples. An initial segmentation step based on a larger sample of data sets is required to automate discrimination of lesions and artifacts.

Effects of aging and hypertension on cerebral ischemic susceptibility: Evidenced by MR diffusion–perfusion study in rat

Aging is the most significant non-modifiable risk factor and hypertension is the most significant modifiable risk factor for ischemic stroke. We used magnetic resonance imaging (MRI) to investigate the evidence of ischemic susceptibility after aging and hypertension. Four groups of rat were studied: young normotensive Wistar-Kyoto (WKY) rat, aged normotensive WKY rat, young spontaneously hypertensive rat (SHR) and aged SHR. Brain images were acquired at a 3.0T Tim-Trio MRI system. For diffusion-weighted images, apparent diffusion coefficient (ADC) was measured. Relative cerebral blood flow (CBF) was also calculated. Cerebral ischemic susceptibility was examined by using ischemic model of bilateral common carotid artery occlusion. In the MRI study of non-ischemic rat, aged SHR had significantly higher ADC ( P < 0.01) and significantly lower CBF ( P < 0.01) in the parietal cortex, but aged WKY rat had only significantly lower CBF ( P < 0.01) when compared with young WKY rat. The ADC/CBF ratio in the parietal cortex was significantly higher in aged SHR when compared with young WKY rat, young SHR and aged WKY rat ( P < 0.01, P < 0.05, P < 0.05, respectively) suggesting a significant diffusion–perfusion disparity in aged SHR. After bilateral common carotid artery occlusion, there was significantly larger damage in the parietal cortex of aged SHR when compared with young WKY rat, young SHR and aged WKY rat (all P < 0.05), but not in the hippocampus and thalamus ( P > 0.05). Our study demonstrated a significantly increased ADC value and reduced CBF in the ischemia-vulnerable cortical area. This cerebral diffusion–perfusion disparity is seen mainly in aged rat and can be more evident if associated with hypertension indicating an increased susceptibility of brain tissue to ischemic injury.

Increased susceptibility to ischemia and macrophage activation in STZ-diabetic rat nerve

Ischemic vulnerability in diabetic nerve plays a paramount role in the development of diabetic neuropathy, yet little is known of the underlying mechanism. Diabetes enhances the inflammatory response to ischemia and reperfusion. We investigated pathological characteristics of nerve fibers and endoneurial macrophages along the length of sciatic–tibial nerves before and after ischemia (60 to 90min) and reperfusion (6h to 7days) in 8weeks of STZ-induced diabetic rats. Without ischemia, diabetic nerves revealed significantly increased the density of Iba-1-positive endoneurial macrophages when compared with controls. Most of macrophages appeared slim and triangular in shape, but in diabetic nerves, some were rounded with bromodeoxyuridine (BrdU) incorporation, suggesting proliferating macrophages. Seventy-five minutes of ischemia is the minimal ischemic time to cause pathological changes in diabetic nerves. Following 90min of ischemia and 6h of reperfusion in diabetic rats, the number of Iba-1-positive endoneurial macrophages was increased significantly at the thigh level of sciatic nerve when compared with those before ischemia. Endoneurial macrophages in diabetic nerves increased in number further significantly after 24 and 48h of reperfusion and underwent morphological alterations; swollen and rounded including phagocytosis. After 90min of ischemia and 7days of reperfusion, severe pathological alterations, e.g., demyelination and endoneurial edema at proximal nerves and axonal degeneration distally, were observed in diabetic nerves, while control nerves showed normal morphology. We conclude that macrophage proliferation occurs in STZ-diabetic nerves. The acute inflammatory response after ischemia and reperfusion was intensified in diabetic nerves. Activation of resident macrophages and infiltration by recruited macrophages could be casually linked to ischemic susceptibility in diabetic nerve.

Cardiac Remodeling: Mechanism and Treatment

Barry Greenberg, MD, ed 569 pages. New York, NY: Taylor and Francis; 2006. $199.95. ISBN 0-8247-2387-2. The appreciation that the myocardium changes shape and structure in response to disease has been appreciated for more than a century. The German anatomists including Krehl, Romberg, Aschoff, and Tawara debated the morphological changes occurring in the heart related to function at the end of the last century. The seminal work of Linzbach1 described the changes in myocyte size in response to aging and to hypertension, valvular heart disease, and heart failure. Because the cardiomyocyte is considered to be “terminally differentiated,” it must respond to disease by adapting to change by modifying its molecular and …

Excitability differences in lower‐limb motor axons during and after ischemia

Neuropathic diseases typically begin distally and spread proximally. Irrespective of the etiology, pathological investigations often indicate changes consistent with ischemia. In the present study, threshold tracking was used to investigate length-dependent differences in ischemic susceptibility of lower-limb axons in 6 healthy volunteers, with ischemia induced by a sphygmomanometer cuff inflated to 200 mm Hg and maintained for 13 minutes. Following stimulation of the peroneal nerve at the fibula neck, compound muscle action potentials were recorded proximally from tibialis anterior (TA) and distally from extensor digitorum brevis (EDB). During ischemia, excitability changes were consistent with nerve depolarization, with a greater reduction in threshold in EDB than TA. This reduction in threshold was associated with an increase in refractoriness, decrease in superexcitability, and prolongation of strength-duration time constant, consistent with axonal depolarization. With release of ischemia, reversal of these changes was associated with an increase in threshold, greater in EDB than TA, indicating axonal hyperpolarization. The rate of recovery of threshold was similar proximally and distally, arguing against a gradient in Na(+)/K(+) pump function along the peroneal nerve. The greater changes in threshold in EDB during and after ischemia suggest an increased susceptibility of more distal axons to ischemia and are likely to contribute to the length-dependent development of neuropathy.

Left Ventricular Mechanical Unloading Restores Beta-2-Adrenergic Receptor mRNA Expression and Decreases Susceptibility to Ischemia and Reperfusion in the Failing Heart

Objectives: Left ventricular (LV) mechanical unloading is known to reduce the hemodynamic demands of failing LV, resulting in improved myocyte contractility. This study was designed to examine effects of LV unloading on β-adrenergic receptor (BAR) expression and ischemic susceptibility to ischemia reperfusion. Methods: Five groups were studied: group 1 [unloading myocardial infarction (MI), n = 6], MI hearts 2 weeks after coronary ligation subjected to LV unloading by heterotopic heart transplantation for 2 weeks; group 2 (2-week MI, n = 6), MI hearts left for 2 weeks without unloading; group 3 (4-week MI, n = 6), MI hearts left for 4 weeks without unloading; group 4 (control, n = 6), normal (non-MI) hearts as a control with no interventions, and group 5 (unloading control, n = 5), normal (non-MI) hearts subjected to LV unloading for 2 weeks.Then, all hearts were isolated and subjected to 20 min of global ischemia and 60 min of reperfusion on Langendorff apparatus. LV pressures and coronary flow were measured throughout the experiment. Either total BAR density or β₂-adrenergic receptor (B2AR) mRNA expression in the noninfarcted myocardium was determined by radioligand binding assays or real-time quantitative RT-PCR, respectively. Results: LV unloading improved postischemic functional recovery (unloading MI vs. 2-week MI vs. 4-week MI: 74 ± 6 vs. 54 ± 5 vs. 51 ± 4%; p < 0.05 vs. unloading MI). LV unloading restored B2AR mRNA expression (unloading MI vs. 2-week MI vs. 4-week MI: 4.78 ± 0.21 vs. 2.80 ± 0.19 vs. 2.24 ± 0.17 × 10⁷ copy/µg total RNA; p < 0.05 vs. unloading MI). Conclusion: LV mechanical unloading restored B2AR mRNA expression and improved postischemic functional recovery.

Expression of human apolipoprotein E downregulates amyloid precursor protein-induced ischemic susceptibility.

Epidemiological findings and experimental data on transgenic mice show that Alzheimer's disease-related changes render the brain more susceptible to ischemic damage. We studied whether the previously observed vulnerability in mice overexpressing the 751-amino-acid isoform of human amyloid precursor protein (APP751) is regulated by human apolipoprotein E (apoE) alleles, which determine the relative risk for Alzheimer's disease and the susceptibility to various forms of acute brain damage. Aged apoE knock out (KO) mice, mice overexpressing APP751 in the apoE KO background and mice expressing either human apoE3 or apoE4 and APP751 in the apoE KO background were exposed to permanent occlusion of the middle cerebral artery (MCA). Infarct volumes were quantified from T2-weighted magnetic resonance images 24 hours after the MCA occlusion. Local cortical blood flow was monitored by laser Doppler flowmetry. Ischemia-induced microgliosis was detected by immunohistochemistry. Overexpression of human APP751 significantly increased the infarct volumes in apoE KO mice. Furthermore, this APP751-induced ischemic vulnerability was attenuated by the coexpression of either human apoE isoform. MCA occlusion resulted in a similar relative reduction in cortical blood flow in all mouse groups. Vascular anatomy showed no variation in the MCA territory between the groups. Instead, the expression of human apoE isoforms reduced the ischemia-induced microgliosis. Expression of either the human apoE3 or apoE4 isoform protects against the increased ischemic vulnerability observed in aged mice overexpressing human APP751, probably by modulating the inflammatory response induced by MCA occlusion.

Regional Metabolic Disturbances and Cerebrovascular Anatomy after Permanent Middle Cerebral Artery Occlusion in C57Black/6 and SV129 Mice

C57Black/6 and SV129 mice are widely used for the production of transgenic mutants in molecular stroke research but the ischemic susceptibility of these strains is influenced by differences in vascular anatomy and the responsiveness to excitotoxins and vasodilatory stimuli. To differentiate between these opposing effects on infarct size, the vascular territory of the two strains was correlated with the hemodynamic, metabolic, and morphological consequences of permanent middle cerebral artery (MCA) occlusion. The vascular anatomy was studied by latex infusion, brain infarction by vital staining, the size of the ischemic penumbra by imaging of ATP and protein synthesis, and blood flow by laser–Doppler flowmetry. In C57Black/6 mice the MCA-supplied vascular territory and the size of brain infarcts were significantly larger than in SV129 mice but the size of the penumbra and the residual blood flow in the center of the MCA-supplying territory were similar in both strains. These findings suggest that differences in infarct size in C57Black/6 and SV129 mice are determined mainly by the vascular anatomy and not by differences in collateral vascular responsiveness or excitotoxicity.