Duration Of Reperfusion Research Articles

Transcriptional responses in a mouse model of silicone wire embolization induced acute retinal artery ischemia and reperfusion.

Acute retinal ischemia and ischemia-reperfusion injury are the primary causes of retinal neural cell death and vision loss in retinal artery occlusion (RAO). The absence of an accurate mouse model for simulating the retinal ischemic process has hindered progress in developing neuroprotective agents for RAO. We developed a unilateral pterygopalatine ophthalmic artery occlusion (UPOAO) mouse model using silicone wire embolization combined with carotid artery ligation. The survival of retinal ganglion cells and visual function were evaluated to determine the duration of ischemia. Immunofluorescence staining, optical coherence tomography, and haematoxylin and eosin staining were utilized to assess changes in major neural cell classes and retinal structure degeneration at two reperfusion durations. Transcriptomics was employed to investigate alterations in the pathological process of UPOAO following ischemia and reperfusion, highlighting transcriptomic differences between UPOAO and other retinal ischemia-reperfusion models. The UPOAO model successfully replicated the acute interruption of retinal blood supply observed in RAO. 60 min of Ischemia led to significant loss of major retinal neural cells and visual function impairment. Notable thinning of the inner retinal layer, especially the ganglion cell layer, was evident post-UPOAO. Temporal transcriptome analysis revealed various pathophysiological processes related to immune cell migration, oxidative stress, and immune inflammation during the non-reperfusion and reperfusion periods. A pronounced increase in microglia within the retina and peripheral leukocytes accessing the retina was observed during reperfusion periods. Comparison of differentially expressed genes (DEGs) between the UPOAO and high intraocular pressure models revealed specific enrichments in lipid and steroid metabolism-related genes in the UPOAO model. The UPOAO model emerges as a novel tool for screening pathogenic genes and promoting further therapeutic research in RAO.

Transcriptional responses in a mouse model of silicone wire embolization induced acute retinal artery ischemia and reperfusion

Acute retinal ischemia and ischemia-reperfusion injury are the primary causes of retinal neural cell death and vision loss in retinal artery occlusion (RAO). The absence of an accurate mouse model for simulating the retinal ischemic process has hindered progress in developing neuroprotective agents for RAO. We developed a unilateral pterygopalatine ophthalmic artery occlusion (UPOAO) mouse model using silicone wire embolization combined with carotid artery ligation. The survival of retinal ganglion cells and visual function were evaluated to determine the duration of ischemia. Immunofluorescence staining, optical coherence tomography, and haematoxylin and eosin staining were utilized to assess changes in major neural cell classes and retinal structure degeneration at two reperfusion durations. Transcriptomics was employed to investigate alterations in the pathological process of UPOAO following ischemia and reperfusion, highlighting transcriptomic differences between UPOAO and other retinal ischemia-reperfusion models. The UPOAO model successfully replicated the acute interruption of retinal blood supply observed in RAO. 60 min of Ischemia led to significant loss of major retinal neural cells and visual function impairment. Notable thinning of the inner retinal layer, especially the ganglion cell layer, was evident post-UPOAO. Temporal transcriptome analysis revealed various pathophysiological processes related to immune cell migration, oxidative stress, and immune inflammation during the non-reperfusion and reperfusion periods. A pronounced increase in microglia within the retina and peripheral leukocytes accessing the retina was observed during reperfusion periods. Comparison of differentially expressed genes (DEGs) between the UPOAO and high intraocular pressure models revealed specific enrichments in lipid and steroid metabolism-related genes in the UPOAO model. The UPOAO model emerges as a novel tool for screening pathogenic genes and promoting further therapeutic research in RAO.

8709 FABP4+ Bone Marrow Endothelial Cells Support Hematopoietic Stem Cell Niches And Osteogenesis In Response To Vascular Insufficiency

Abstract Disclosure: Y. Kim: None. M. Song: None. H. Sun: None. S. Cho: None. Background: Numerous non-communicable and aging-related diseases are linked to systemic vascular insufficiency. The consequences of vascular insufficiency in bone marrow extend to compromising osteogenesis and hematopoiesis, with a more pronounced impact on aging individuals compared to younger counterparts affected by these conditions. Recent advances in scRNA-seq have shed light on the cellular taxonomy in bone marrow; however, a comprehensive understanding of cellular complexity requires further elucidation, and there are limitations in delineating impaired osteogenesis and hematopoiesis resulting from vascular insufficiency. In this study, we employed a combined approach of scRNA-seq and intensive ex vivo investigations to elucidate the regeneration process of bone marrow microenvironment following vascular insufficiency. Method Female C57BL/6 mice at different ages (6-wkks and 14 weeks) were subjected to sham (sham group) or right femoral artery ligation surgery (ischemia group). Reperfusion duration was quantified by color laser doppler. Osteogenesis was assessed by H&E staining, DXA and microCT scan. Flow cytometry was employed to count the number of type H endothelial cell (THEC, Endomucinhi). Bone marrow regeneration capacity was examined in vitro by treating with conditioned medium (CM) of THEC and in vivo via THEC injection. scRNA-seq was performed on the bone marrow of both sham and ischemia group 7 days post-ischemia. Result In young mice, reperfusion occurred within 7 days and osteogenesis was not different between sham and ischemia group. However, in old mice, reperfusion extended to 6 weeks, and accompanied by a reduction in osteogenesis, which did not recover until 12 weeks post-ischemia compared to sham group. The percentage of THEC increased after ischemia in young mice from 1.01% to 3.12 % (n=10, p<0.05), while those were comparable between sham and ischemia group in old mice (from 0.66% to 1.02%, n = 10, p=0.56). in MC3T3-E1 preosteoblast cell line, treatment with THEC-CM upregulated gene expressions associated with bone formation such as Alpl, Bglap, and Col1a1. In vivo THEC injection in old mice following ischemia surgery resulted in a reduction in reperfusion duration and an improvement in osteogenesis. scRNA-seq of bone marrow identified a significant increase of bone marrow stromal cells (BMSCs) and Cxcl12-abundant reticular (CAR) cells 7-days post-ischemia, which provides favorable microenvironment for dormant hematopoietic stem cells (HSCs). The number of FABP4+ THEC synchronized with the increase in BMSCs and CARs. In addition, FABP4+ THEC expressed Col1a1 and Col1a2, associated with osteogenesis, along with Cxcl12 and Foxc1, linked to HSC niches. Conclusion: FABP4+ THECs contribute to favorable microenvironment for HSCs and play a pivotal role in angiogenesis-osteogenesis coupling following vascular insufficiency. Presentation: 6/1/2024

Time varying characteristic in somatosensory evoked potentials as a biomarker of spinal cord ischemic-reperfusion injury in rat.

Spinal cord ischemic-reperfusion injury (SCIRI) could occurs during surgical procedures without detection, presenting a complex course and an unfavorable prognosis. This may lead to postoperative sensory or motor dysfunction in areas innervated by the spinal cord, and in some cases, permanent paralysis. Timely detection of SCIRI and immediate waring can help surgeons implement remedial intervention to prevent irreversible spinal cord injury. Therefore, it is crucial to develop a precise and effective method for early detection of SCIRI. This study utilized rat models to simulate intraoperative SCIRI and employed somatosensory evoked potentials (SEP) for continuous monitoring during surgery. In this study, SEP signal changes were examined in six groups with varying severities of SCIRI and one normal control group. SEP signal changes were examined during operations in different groups and correlated with postoperative behavioral and histopathological data. The result demonstrated specific changes in SEP signals during SCIRI, termed as time-varying characteristics, which are associated with the duration of ischemia and subsequent reperfusion. Time-varying characteristics in SEP could potentially serve as a new biomarker for the intraoperative detection of SCIRI. This finding is significant for clinical surgeons to identify and guide early intervention of SCIRI timely. Additionally, this measurement is easily translatable to clinical application.

Buyang Huanwu Decoction attenuates cerebral ischemia-reperfusion injury in rats via miR-26a-5p mediated PTEN/PI3K/Akt signaling pathway

This study aims to investigate the mechanism of Buyang Huanwu Decoction in treatment of cerebral ischemia-reperfusion injury in rats. A total of 180 SD rats were randomly divided into 5 different groups: sham group, model group, Buyang Huanwu Decoction group, Buyang Huanwu Decoction + miR-26a-5p agomir(agomir) group, Buyang Huanwu Decoction + miR-26a-5p agomir negative control(agomir NC) group. There were 36 rats in each group. Each group was then subdivided into three subgroups for the duration of reperfusion(3, 7, 14 d). A ligature-induced middle cerebral artery occlusion(MCAO) model was carried out on all groups other than sham group. Reperfusion was performed following ischemia for 90 min. Buyang Huanwu Decoction group, agomir group, and agomir NC group were given Buyang Huanwu Decoction twice daily by gavage 24 h after the formation of the model. Sham group and model group were given an equal amount of physiological saline by gavage until the day before sacrifice. At 24 h after ischemia induction, miR-26a-5p agomir was injected into the lateral ventricle in agomir group, miR-26a-5p NC in agomir NC group, and equal amounts of physiological saline in the other groups. 24 h after ischemia induction, BrdU was intraperitoneally injected once daily until the day before sacrifice. Modified neurological severity score(mNSS) was used to evaluate neurological deficits, 2,3,5-triphenyltetrazolium chloride(TTC) staining was used to determine the cerebral infarct volume, TUNEL staining was used to assess the apoptosis of parenchymal ischemic brain tissue, and double immunofluorescence staining was used to examine BrdU/NeuN double positive neurons in the parenchymal ischemic brain tissue to evaluate the neuronal regeneration. We employed a luciferase reporter assay to identify and validate that the target gene of miR-26a-5p is PTEN. Real-time quantitative polymerase chain reaction(RT-qPCR) was used to assess gene expression levels of PTEN and miR-26a-5p and Western blot to assess the protein levels of PTEN, PI3K, p-PI3K, Akt, and p-Akt. The results revealed that compared with model group, Buyang Huanwu Decoction treatment promoted neural function recovery, reduced the cerebral infarct volume, increased the number of BrdU~+/NeuN~+ neurons, upregulated the expression of miR-26a-5p, regulated the PTEN/PI3K/Akt signaling pathway, and promoted neuronal regeneration in the cerebral ischemia-reperfusion rats. These effects were significantly enhanced after lateral ventricle injection of miR-26a-5p agomir. The findings prove that Buyang Huanwu Decoction treatment can promote neural function recovery, reduce the cerebral infarct volume, and promote neuronal regeneration in a cerebral ischemia-reperfusion rat model, which is likely to be achieved via miR-26a-5p mediated PTEN/PI3K/Akt signaling pathway.

S-Nitroso Human Serum Albumin Enhances Left Ventricle Hemodynamic Performance and Reduces Myocardial Damage after Local Ischemia-Reperfusion Injury.

Endothelial nitric oxide (NO) production is crucial in maintaining vascular homeostasis. However, in the context of ischemia-reperfusion (I/R) injury, uncoupled endothelial nitric oxide synthase (eNOS) can exacerbate reactive oxygen species (ROS) generation. Supplementation with S-nitroso human serum albumin (S-NO-HSA) offers a potential solution by mitigating eNOS uncoupling, thereby enhancing NO bioavailability. In a study conducted at the University of Verona, male rats underwent thoracotomy followed by 30 min left anterior descendant coronary (LAD) occlusion and subsequent reperfusion. Hemodynamic parameters were meticulously assessed using a conductance catheter inserted via the carotid artery. The rats were stratified into two main groups based on reperfusion duration and the timing of drug infusion, with the effects of S-NO-HSA evaluated after 2 or 24 h. Remarkably, intravenous administration of S-NO-HSA, initiated before or during ischemia, exhibited notable benefits. It significantly improved left ventricular function, safeguarded energetic substrates such as phosphocreatine and ATP, and sustained glutathione levels akin to basal conditions, indicative of diminished oxidative stress. The data from this study strongly suggest a protective role for S-NO-HSA in mitigating I/R injury induced by LAD artery occlusion, a phenomenon observed at both 2 and 24 h post-reperfusion. These findings underscore the promising therapeutic potential of NO supplementation in alleviating myocardial damage subsequent to ischemic insult.

Isoliquiritigenin alleviates myocardial ischemia-reperfusion injury by regulating the Nrf2/HO-1/SLC7a11/GPX4 axis in mice

Ischemia-reperfusion (I/R) injury, a multifaceted pathological process, occurs when the prolongation of reperfusion duration triggers ferroptosis-mediated myocardial damage. Isoliquiritigenin (ISL), a single flavonoid from licorice, exhibits a wide range of pharmacological impacts, but its function in ferroptosis caused by myocardial I/R injury remains unclear. This study delved into the protective effect of ISL on myocardial I/R injury-induced ferroptosis and its mechanism. Neonatal mouse cardiomyocytes (NMCM) underwent hypoxia/reoxygenation (H/R) to simulate the pathological process of myocardial I/R. ISL significantly attenuated H/R-triggered production of reactive oxygen species in NMCM, reduced the expression of malondialdehyde and the activity of lactate dehydrogenase, enhanced superoxide dismutase and catalase activity, and increased the expression of nuclear factor E2-related factor 2 (Nrf2) and its downstream heme oxygenase 1 (HO-1), thereby mitigating oxidative stress damage. CCK8 experiment revealed that the ferroptosis inhibitor Ferrostatin-1 significantly improved myocardial cell viability after 24 h of reoxygenation, and ISL treatment showed a similar effect. ISL reduced intracellular free iron accumulation, up-regulated glutathione peroxidase 4 (GPX4) and solute carrier family 7 member 11 (SLC7A11) expression, and inhibited lipid peroxidation accumulation, thereby alleviating ferroptosis. The Nrf2-specific inhibitor ML385 counteracted ISL's defensive role against H/R-triggered oxidative stress damage and ferroptosis. In vivo experiments further confirmed that by regulating the translocation of Nrf2 into the nucleus, ISL treatment increased the levels of HO-1, GPX4, and SLC7A11, inhibited the expression of ACSL4, Drp1 to exert the antioxidant role, alleviated mitochondrial damage, and ferroptosis, ultimately reducing myocardial infarction area and injury induced by I/R. ML385 nearly abolished ISL's protective impact on the I/R model by inhibiting Nrf2 function. In summary, ISL is capable of mitigating oxidative stress, mitochondrial damage, and cardiomyocyte ferroptosis caused by I/R, thereby reducing myocardial injury. A key mechanism includes triggering the Nrf2/HO-1/SLC7A11/GPX4 pathway to prevent oxidative stress damage and cardiomyocyte ferroptosis caused by I/R.

Determination of Maximum Tolerable Cold Ischemia Time in a Mouse Model of Cervical Heterotopic Uterus Transplantation.

Uterus transplantation (UTx) is an emerging treatment for uterine factor infertility. Determining the maximum tolerable cold ischemia time is crucial for successful UTx. However, the limit for cold ischemia in the uterus is unclear. This study aimed to examine cold ischemia's effects on mouse uteri and identify the maximum cold ischemia duration that uteri can endure. We systematically assessed the tolerance of mouse uteri to extended cold ischemia, 24 h, 36 h, and 48 h, using the cervical heterotopic UTx model. Multiple indicators were used to evaluate ischemia-reperfusion injury, including reperfusion duration, macroscopic examination, oxidative stress, inflammation, and histopathology. The function of transplants was evaluated through estrous cycle monitoring and embryo transfer. Mouse uteri subjected to 48 h of cold ischemia exhibited significant delays and insufficiencies in reperfusion, substantial tissue necrosis, and loss of the estrous cycle. Conversely, uteri that underwent cold ischemia within 36 h showed long survival, regular estrous cycles, and fertility. Our study demonstrated that mouse uteri can endure at least 36 h of cold ischemia, extending the known limits for cold ischemia and providing a pivotal reference for research on the prevention and treatment of cold ischemic injury in UTx.

Cerebral ischemia-induced gene expression changes in diabetic mice from acute to subacute stage

Diabetes is a risk factor for stroke; however, its impact on stroke progression at the genomic level is not well understood. To address this gap, we used transcriptome sequencing to explore the relationship between lncRNA and mRNA expression patterns and the reperfusion duration in the cortex of diabetically induced stroke mice. First, focal ischemia was induced in adult male ob/ob mice, which were then subjected to reperfusion periods of one, three, or seven days. Total RNA was extracted from the ischemic cortical tissue for RNA sequencing, and the resulting reads were aligned to the GRCm38 murine reference genome. A total of 672 mRNAs and 301 lncRNAs were identified as differentially expressed one day post-stroke, 1195 mRNAs and 66 lncRNAs at three days post-stroke, and 1069 mRNAs and 75 lncRNAs at seven days post-stroke. Stage-specific differentially expressed mRNAs were bioinformatically analyzed and found significantly enriched in processes such as apoptosis, angiogenesis, and lipid metabolism at one, three, and seven days post-stroke, respectively. Stage-specific DElncRNA-mRNA cis-regulatory relationships were constructed using these biological processes as examples, revealing the potential roles of four pairs of lncRNA-mRNAs (Gm39787-Lcn2, Gm46111-Drd2, D3300500i16Rik-Fosl1, and Gm41689-Egr1) in apoptosis. Additionally, Gm40237-Tie1 and Gm52352-Pdgfrb are associated with angiogenesis and lipid metabolism, respectively. In conclusion, our study demonstrated that lncRNA and mRNA expression in the cortex of transient focal ischemia-induced diabetic mice undergo extensive alterations, providing insights into complex molecular interactions underlying diabetic stroke.

Evaluation of time-dependent phenotypes of myocardial ischemia-reperfusion in mice.

A mouse model of myocardial ischemia-reperfusion (I/R) is widely used to study myocardial ischemia-reperfusion injury (I/RI). However, few studies focus on the direct comparison of the extent of pathological events resulting from variant durations of ischemia and reperfusion process. A mouse model of I/RI was established by ligation and perfusion of the left anterior descending coronary artery (LAD), and the dynamic changes were recorded by electrocardiogram at different stages of I/R. Subsequently, reperfusion duration was used as a variable to directly compare the phenotypes of different myocardial injury degrees induced by 3 h, 6 h and 24 h reperfusion from myocardial infarct size, myocardial apoptosis, myocardial enzyme, and inflammatory cytokine levels. All mice subjected to myocardial I/R surgery showed obvious myocardial infarction, extensive myocardial apoptosis, dynamic changes in serum myocardial enzyme and inflammatory cytokines, at least for the first 24 h of reperfusion. The infarct size and apoptosis rates gradually increased with the extension of reperfusion time. The peaks of serum myocardial enzyme and inflammatory cytokines occurred at 6 h and 3 h of reperfusion, respectively. We also established I/R mice models with 30 and 60 mins of ischemia. After 21 days of remodeling, longer periods of ischemia increased the degree of fibrosis and reduced cardiac function. In summary, we conclude that reperfusion durations of 3 h, 6 h, and 24 h induces different injury phenotypes in ischemia-reperfusion mouse model. At the same time, the ischemia duration before reperfusion also affects the degree of cardiac remodeling.

CCA repair or ECA ligation-Which middle cerebral artery occlusion is better in the reperfusion mouse model?

A reliable animal model is essential for ischemic stroke research. The implications of the external carotid artery (ECA) transection or common carotid artery (CCA) ligation have been described. Thus, a modified animal model, the CCA-repair model, has been established, and studies have shown that the CCA-repair model has potential advantages over the CCA-ligation model. However, whether the CCA-repair model is superior to the ECA-ligation model remains unclear. Sixty male C57BL/6 mice were randomly assigned to establish the CCA-repair (n = 34) or ECA-ligation (n = 26) models. Cerebral blood flow before middle cerebral artery occlusion (MCAO), immediately after MCAO and reperfusion were monitored and the operation duration, postoperative body weight, and food intake within 7 days, and the number of intraoperative and postoperative deaths within 7 days were recorded in the two models. Modified neurological severity scores and Bederson (0-5) scores were used to evaluate postoperative neurological function deficits on Days 1/3/5/7. 2,3,5-Triphenyltetrazolium chloride staining was used to quantify lesion volume on Day 7 after the operation. We found the establishment of the CCA-repair model required a longer total operation duration (p = 0.0175), especially the operation duration of reperfusion (p < 0.0001). However, there was no significant difference in body weight and food intake development, lesion volume and intragroup variability, neurological function deficits, mortality, and survival probability between the two groups. The CCA-repair model has no significant advantage over the ECA-ligation model. The ECA-ligation model is still a better choice for focal cerebral ischemia.

Safety and Tolerability of Direct Ischemic Postconditioning Following Thrombectomy for Acute Ischemic Stroke.

Experimental studies have demonstrated the neuroprotection of ischemic postconditioning (IPostC) in acute ischemic stroke by attenuating ischemia-reperfusion injury. This study aimed to investigate the safety and tolerability of direct IPostC in both a dog model and patients with acute ischemic stroke treated with thrombectomy. The study involved 2 parts. First, IPostC was induced by repeated balloon inflation and deflation in dogs, where a low-pressure balloon was navigated to the anterior spinal artery, and 4 cycles of 5-minute ischemia followed by 5-minute reperfusion were performed. Vascular injuries were assessed using angiography and vascular tissue specimens. Then, a 3+3 dose-escalation trial was conducted in patients with acute ischemic stroke following successful thrombectomy recanalization. Patients received direct IPostC with ischemia and reperfusion durations in progressive increments of 0, 1, 2, 3, 4, and 5 minutes ×4 cycles. Major adverse responses were defined as vessel perforation, rupture, dissection, reocclusion, severe vasospasm, thrombotic events, and rupture of the balloon. IPostC was investigated in 4 dogs. No vessel perforation or rupture, dissection, or vasospasm was observed under the angiography. Only 1 vessel experienced mild injury between the intima and the internal elastic membrane detected on a histopathologic slide. Then, 18 patients were recruited. The duration of IPostC was progressively escalated with no major response happened. No patient experienced agitation, discomfort, or other tolerability issues. Five patients (27.8%) experienced any intracranial hemorrhage after thrombectomy, and 1 (5.6%) was symptomatic. At 3-month follow-up, no patient died, and 9 patients (50%) achieved functional independence. Direct IPostC inducing by 4 cycles of 5-minute ischemia followed by 5-minute reperfusion is safe, feasible, and tolerable in patients with acute ischemic stroke treated with thrombectomy. Further investigations are needed to determine the safety and preliminary efficacy of direct IPostC. URL: https://www. gov; Unique identifier: NCT05153655.

Pro-inflammatory cytokines/chemokines, TNF-α, IL-6 and MCP-1, as biomarkers for the nephro- and pneumoprotective effect of silibinin after hepatic ischemia/reperfusion: Confirmation by immunohistochemistry and qRT-PCR.

The present study investigated the potential nephro- and pneumoprotective effect of silibinin (Si) after hepatic ischemia-reperfusion (I/R) injury, by measuring pro-inflammatory factors. Sixty-three rats were randomly assigned into three groups, as follows: (a) the sham group (n = 7 rats), subjected to opening and closing the abdomen; (b) the control group (n = 28 rats), subjected to 45-min hepatic ischemia followed by reperfusion; and (c) the silibinin group (n = 28), subjected to 45-min hepatic ischemia followed by intravenous administration of lyophilised SLB-HP-β-CD before reperfusion. Control and silibinin groups were further subdivided into time-point groups, according to the duration of reperfusion. TNF-α, IL-6 and MCP-1 expressions were determined immunohistochemically and by qrT-PCR at each time-point. Kidney TNF-α expression was significantly lower at 180 and 240 min, while lung TNF-α expression was significantly lower at 240 min. Comparison between the control and Si group at the same time-points showed very strong evidence of difference at 240 min, with the levels of IL-6 shifting towards lower values in the Si group. Finally, we found a high MCP-1 expression after 120 min. We conclude that hepatic I/R injury remotely increases pro-inflammatory mediators in the kidney and lung, whereas silibinin shows a time-dependent nephro- and pneumoprotective effect.

Contribution of Myocyte Apoptosis to Myocardial Injury in an &amp;lt;i&amp;gt;in Vivo&amp;lt;/i&amp;gt; Rabbit Preparation of Ischemia-Reperfusion

Objective: Exposure of the heart to repeated, brief episodes of coronary occlusion/reperfusion prevents lethal myocyte injury. Necrosis and apoptosis, two seemingly distinct mechanisms of cell death caused by ischemia could contribute independently to progressive loss of myocardium. Studies suggest that ischemic conditioning (IC) lessens myocyte injury by decreasing apoptosis. The goal of this study was to examine cell death in rabbit hearts subject to ischemia-reperfusion injury without (nIC) or with pretreatment by IC. Methods: In the control study, anesthetized, male rabbits (n = 4/group) underwent 30-min regional coronary occlusion (CO) and either 3, 6 or 24h reperfusion (REP). In the IC study, rabbits were pretreated by IC (2 cycles of 5-min CO and 10-min REP) before 30-min CO and subsequent REP. Additional groups were evaluated with 60, or 120-min CO followed by up to 96 h REP. Agarose electrophoresis was used to detect DNA laddering and poly (ADP-ribose) polymerase (PARP; chromatin bound nuclear DNA repair enzyme) was assessed in myocardial biopsies. Results: Genomic DNA from nIC and IC hearts showed no oligonucleosomal fragmentation. In addition, we did not detect any cleavage of PARP; however, myocardial PARP levels decreased when CO and REP durations were prolonged. Conclusion:Absence of genomic DNA fragmentation or PARP cleavage in an in vivo preparation of ischemia-reperfusion injury does not support the view that apoptosis contributes markedly to post-ischemic tissue necrosis.

Autophagy-lysosome dysfunction is involved in gastric ischemia-reperfusion injury by promoting microglial activation in the paraventricular nucleus.

The hypothalamic paraventricular nucleus (PVN) is a specific center in the brain that regulates gastric mucosal injury following gastric ischemia-reperfusion (GI-R) injury. This study aimed to investigate whether autophagy-lysosome dysfunction in the PVN tissues of GI-R rats is involved in the gastric injury, and the underlying molecular mechanisms. The rat model of GI-R was established by clamping the celiac artery for 30 min and reperfusion for different hours (1, 3, and 6 h). The gastric injury was evaluated by hematoxylin and eosin staining of the stomachand the gastric mucosal index. The autophagy-lysosome dysfunction in the PVN was evaluated by the protein levels of LC3 II and Beclin-1 (markers for autophagosome activity) and the activity of acid phosphatase (a representative lysosomal enzyme). Immunohistochemical staining of ionized calcium-binding adaptor molecule 1 in the PVN was performed to evaluate microglial activation. Reactive oxygen species (ROS) content and phosphorylated γ-aminobutyric acid B receptor (p-GABAB R) expression in the PVN were also examined. The results revealed that, in GI-R rats, the shorter the reperfusion duration, the more severe the gastric mucosal damage. The autophagy-lysosome dysfunction exhibited by GI-R rats further enhanced microglial activation, ROS production, p-GABAB R expression, and gastric injury. In addition, activating microglial cells increased ROS production, p-GABAB R expression, and gastric injury in GI-R rats,while inhibiting microglial activation resulted in the opposite results. Taken together, autophagy-lysosome dysfunction induced by GI-R aggravated the gastric injury by inducing microglia activation.

Assessment of retinal oxygen metabolism, visual function, thickness and degeneration markers after variable ischemia/reperfusion in rats

After total retinal ischemia induced experimentally by ophthalmic vessel occlusion followed by reperfusion, studies have reported alterations in retinal oxygen metabolism (MO2), delivery (DO2), and extraction fraction (OEF), as well as visual dysfunction and cell loss. In the current study, under variable durations of ischemia/reperfusion, changes in these oxygen metrics, visual function, retinal thickness, and degeneration markers (gliosis and apoptosis) were assessed and related. Additionally, the prognostic value of MO2 for predicting visual function and retinal thickness outcomes was reported. Sixty-one rats were divided into 5 groups of ischemia duration (0 [sham], 60, 90, 120, or 180 min) and 2 reperfusion durations (1 h, 7 days). Phosphorescence lifetime and blood flow imaging, electroretinography, and optical coherence tomography were performed. MO2 reduction was related to visual dysfunction, retinal thinning, increased gliosis and apoptosis after 7-days reperfusion. Impairment in MO2 after 1-h reperfusion predicted visual function and retinal thickness outcomes after 7-days reperfusion. Since MO2 can be measured in humans, findings from analogous studies may find value in the clinical setting.

Late thrombectomy for ischaemic stroke

Patients with ischaemic stroke tend to delay seeking professional help compared with patients with acute myocardial infarction with ST elevation (STEMI). Moreover, for patients with STEMI, a diagnosis of ongoing ischaemia can be made with an electrocardiogram in the ambulance. The situation is vastly different for a patient who has had a stroke. Those with cortical infarcts—the case for most patients—are often not aware of their acute deficit, or they cannot communicate because of language problems. Moreover, patients with cerebral ischaemia do not have pain, as patients with STEMI do. They also typically do not wake up when a stroke occurs during sleep. Notification of the stroke to emergency services often depends on the alertness of partners and bystanders, and it should not be surprising that patients have delays in presenting to the emergency room, typically 3–4 h later than patients with a STEMI. 1 Al'Aref SJ Wong SC Swaminathan RV et al. Analysis of reperfusion time trends in patients with ST-elevation myocardial infarction across New York State from 2004 to 2012. Int J Cardiol. 2017; 232: 140-146 Summary Full Text Full Text PDF PubMed Scopus (5) Google Scholar , 2 de Havenon A Alexander MD Nogueira RG et al. Duration of symptomatic stroke and successful reperfusion with endovascular thrombectomy for anterior circulation large vessel occlusive stroke. J Neurointerv Surg. 2021; 01 (neurintsurg-2020-016961.) Google Scholar In patients arriving after more than 6 h from onset, diagnosis of ongoing ischaemia caused by intracranial large vessel occlusion has to be made with CT angiography and perfusion imaging, which further adds to the delay. Thrombectomy for anterior circulation stroke beyond 6 h from time last known well (AURORA): a systematic review and individual patient data meta-analysisThese findings strengthen the evidence for benefit of endovascular thrombectomy in patients with evidence of reversible cerebral ischaemia across the 6–24 h time window and are relevant to clinical practice. Our findings suggest that in these patients, thrombectomy should not be withheld on the basis of mode of presentation or of the point in time of presentation within the 6–24 h time window. Full-Text PDF

Histological Assessment of Rat Retinas with Ischemia-Reperfusion Injury

Introduction: Retinal ischemia-reperfusion (IR) injury occurs in pathological situations that interrupt the blood flow to the retina, such as is the case during central retinal artery occlusion (CRAO). The animal models described in the literature are based on the pressure produced by the weight of a given quantity of saline elevated to a certain height; however, to establish these parameters it is necessary to perform mathematical calculations that cannot be easily redone in the case of punctual variations of intraocular pressure (IOP). The aim of this study was to present a new system that allows us to reproduce the conditions of retinal IR and thereby properly assess the level of injury in retinal histological samples. Methods: We developed a retinal IR model in WAG/RijHsd rats based on CRAO through increasing IOP. To develop this model, we produced ischemia for 1 h using a hydrostatic pressure system that maintained a constant high IOP and then allowed reperfusion for 1 h. The injury attributable to IR was assessed by histological examination of retinal samples, determining whether there was histological damage and/or dendritic swelling and counting the outer nuclear layer cells showing cytoplasmic swelling. Results: The increase in IOP to 150 mm Hg produced CRAO, in turn causing observable histological damage and dendritic swelling in all retinas subjected to IR. Counting the number of cells showing cytoplasmic swelling yielded a mean of 102.5 ± 35 cells/field. The contralateral retinas were healthy, showing no significant changes. Conclusion: The retinal IR model proposed is simple, reproducible, and allows variable durations of ischemia and reperfusion, and most importantly, it allows easy correction by adjusting the pressure of the sphygmomanometer, of any change in IOP to keep the ischemia stable, without having to recalculate the elevation height of the ischemia induction system. Moreover, the damage caused by IR can be effectively assessed by the type of histopathological assessment performed. For these reasons, it can be considered a reliable method for studying drugs that may prevent retinal IR injury.

Abstract P524: Impact of Intra-Procedural Workflow and Time Metrics of Establishing Fast Reperfusion on Clinical Outcomes in the ESCAPE-NA1 Trial

Background: Time from imaging to establishing reperfusion is a major influencer of clinical outcomes and over the years thrombectomy techniques have evolved rapidly. This has led to improvements in achieving fast and complete reperfusion. We analyzed the impact of various intra-procedural techniques and tools on the speed of reperfusion and correlated procedural duration with probability of achieving good clinical outcomes. Methods: We analyzed intra-procedural time metrics and examined factors leading to delays during EVT. The relationship between outcome (mRS Scale) and procedural time from arterial puncture to time of achieving mTICI 2b-3 First Reperfusion (FRE) was modeled using logistic regression. Results: The various procedural time metrics are summarized in Figure 1. Every 10-minute increase in FRE time reduced the probability of achieving functional independence(90-day modified Rankin Scale 0-2) by 6.7% (P=0.021, adjusted). The medianFRE timewas 25min (IQR 17-39) and was significantly longer in patients with tandem occlusions(median 34min, p 0.0005). General anesthesia vs procedural sedation vs no sedation use did not significantly alter the FRE time (p = 0.1453). The use of BGC (54.2%) was nominally longer FRE (median 26min “IQR 18-38” vs 23ming, “IQR 16-38”; p 0.095)while the use of contact aspiration (n=213) vs retrievable stents (n=676) as the first approach was associated with a shorter FRE time (21min “IQR 14-35” vs 26 min “IQR18-40”, p =0.001). Conclusions: Puncture to first reperfusion time is a significant predictor of clinical outcome in theESCAPE-NA1 trial. Various procedural and anatomical factors influence this timemetric. Figure: Intra-Procedural workflow time metrics expressed in medians and 90th percentiles. The cumulative times are calculated for each major milestone in the procedure for upto three attempts. First reperfusion duration where TICI 2b was achieved is shown in comparison to other procedural time metrics.

Duration of symptomatic stroke and successful reperfusion with endovascular thrombectomy for anterior circulation large vessel occlusive stroke

BackgroundIt has been reported that longer time intervals from stroke onset to endovascular therapy are associated with lower rates of successful reperfusion in acute ischemic stroke patients with large vessel...