The Effects of Rapamycin on the Intestinal Graft in a Rat Model of Cold Ischemia Perfusion and Preservation.

Attenuating the rheological and structural consequences of intestinal ischemia-reperfusion-injury (IRI) is important in transplant proceedings. Preconditioning is an often-proposed remedy. This technique uses physical or pharmacological methods to manipulate key ischemia pathways, such as oxidation, inflammation, and autophagy, prior to ischemia. This study determined the time-dependent effects of Rapamycin preconditioning on small-bowel grafts undergoing cold ischemia perfusion and preservation. Our main parameters were mucosa and cell injury and autophagy. A total of 30 male Wistar rats were divided into 5 groups: sham, preservation-control, and 3 treated groups (Rapamycin administered either 0, 30, or 60 min prior to perfusion). After perfusion, the intestines were placed in chilled IGL-1 solution for 12 h. Thereafter, they were reperfused. Histology and bioanalysis (LDH and lactate) were used to ascertain intestinal injury while immunohistochemistry was used for measuring changes in autophagy markers (Beclin-1, LC3B, and p62 proteins). The results show no significant difference amongst the groups after vascular perfusion. However, intestinal injury findings and autophagy changes demonstrate that administering Rapamycin 30 min or 60 min prior was protective against adverse cold ischemia and reperfusion of the intestinal graft. These findings show that Rapamycin is protective against cold ischemia of the small intestine, especially when administered 30 min before the onset.

Subnormothermic isolated organ perfusion with Nicorandil increased cold ischemic tolerance of liver in experimental model.

BACKGROUND: The cold ischemia -reperfusion injury may lead to microcirculatory disturbances, hepatocellular swelling, inflammation, and organ dysfunction. Nicorandil is an anti-ischemic, ATP-sensitive potassium (KATP) channel opener drug and has proved its effectiveness against hepatic Ischemia/Reperfusion (I/R) injury. OBJECTIVE: This study aimed to investigate the effect of Nicorandil on mitochondrial apoptosis, oxidative stress, inflammation, histopathological changes, and cold ischemic tolerance of the liver in an ex vivo experimental isolated-organ-perfusion model. METHODS: We used an ex vivo isolated rat liver perfusion system for this study. The grafts were retrieved from male Wistar rats (n = 5 in each), preserved in cold storage (CS) for 2 or 4 hours (group 1, 2), or perfused for 2 or 4 hours (group 3, 4) immediately after removal with Krebs Henseleit Buffer (KHB) solution or Nicorandil containing KHB solution under subnormothermic (22-25°C) conditions (group 5, 6). After 15 minutes incubation at room temperature, the livers were reperfused with acellular, oxygenated solution under normothermic condition for 60 minutes. RESULTS: In the Nicorandil perfused groups, significantly decreased liver enzymes, GLDH, TNF-alpha, and IL-1ß were measured from the perfusate. Antioxidant enzymactivity was higher in the perfused groups. Histopathological examination showed ameliorated tissue deterioration, preserved parenchymal structure, decreased apoptosis, and increased Bcl-2 activity in the Nicorandil perfused groups. CONCLUSIONS: Perfusion with Nicorandil containing KHB solution may increase cold ischemic tolerance of the liver via mitochondrial protection which can be a potential therapeutic target to improve graft survival during transplantation.

Characterizing Autophagy in the Cold Ischemic Injury of Small Bowel Grafts: Evidence from Rat Jejunum.

Cold ischemic injury to the intestine during preservation remains an unresolved issue in transplantation medicine. Autophagy, a cytoplasmic protein degradation pathway, is essential for metabolic adaptation to starvation, hypoxia, and ischemia. It has been implicated in the cold ischemia (CI) of other transplantable organs. This study determines the changes in intestinal autophagy evoked by cold storage and explores the effects of autophagy on ischemic grafts. Cold preservation was simulated by placing the small intestines of Wistar rats in an IGL-1 (Institute George Lopez) solution at 4 °C for varying periods (3, 6, 9, and 12 h). The extent of graft preservation injury (mucosal and cellular injury) and changes in autophagy were measured after each CI time. Subsequently, we determined the differences in apoptosis and preservation injury after activating autophagy with rapamycin or inhibiting it with 3-methyladenine. The results revealed that ischemic injury and autophagy were induced by cold storage. Autophagy peaked at 3 h and subsequently declined. After 12 h of storage, autophagic expression was reduced significantly. Additionally, enhanced intestinal autophagy by rapamycin was associated with less tissue, cellular, and apoptotic damage during and after the 12-h long preservation. After reperfusion, grafts with enhanced autophagy still presented with less injury. Inhibiting autophagy exhibited the opposite trend. These findings demonstrate intestinal autophagy changes in cold preservation. Furthermore, enhanced autophagy was protective against cold ischemia-reperfusion damage of the small bowels.

Effect of Pioglitazone on endoplasmic reticulum stress regarding in situ perfusion rat model.

BACKGROUND: Ischemia-reperfusion injury (IRI) can cause insufficient microcirculation of the transplanted organ and results in a diminished and inferior graft survival rate. OBJECTIVE: This study aimed to investigate the effect of different doses of an anti-diabetic drug, Pioglitazone (Pio), on endoplasmic reticulum stress and histopathological changes, using an in situ perfusion rat model. METHODS: Sixty male Wistar rats were used and were divided into six groups, consisting of the control group, vehicle-treated group and four Pio-treated groups (10, 20, 30 and 40 mg/kg Pio was administered). The rats were perfused through vena cava and an outflow on the abdominal aorta occurred. Following the experiment, kidneys and livers were collected. The level of the endoplasmic reticulum stress markers (XBP1 and Caspase 12) was analyzed using Western blot and histopathological changes were evaluated. RESULTS: Histopathological findings were correlated with the Western blot results and depict a protective effect corresponding to the elevated dosage of Pioglitazone regarding in situ perfusion rat model. CONCLUSIONS: In our study, Pioglitazone can reduce the endoplasmic reticulum stress, and the most effective dosage proved to be the 40 mg/kg Pio referencing the kidney and liver samples.

Histological and Mechanical Assessment of Decellularized Porcine Biografts, and Its Biological Evaluation following Aortic Implantation during Mid-Term Follow-Up.

AIMS: Prosthetic graft infection frequently requires graft replacement. Among other options, a biological graft could serve as an alternative choice. Decellularization reduces tissue immunogenicity. Our aim was to determine an efficient decellularization method and to evaluate the decellularized porcine biografts' adaptability. METHODS: Four different protocols were implemented to decellularize porcine aortic segments (n = 4). Cell removal effectiveness and matrix structure preservation were histologically examined. Mechanical tests were performed. Decellularized porcine grafts were interpositioned in a porcine aorta. After a 6-month period, implanted samples were removed and evaluated using light and electron microscopy. RESULTS: Histological results showed complete removal of cells and preserved connective tissue fiber structure following decellularization, using sodium dodecyl sulfate and sodium azide. Pressure tests demonstrated similar compliance to fresh vessels. In 9 out of 10 cases, pigs survived the follow-up period. Graft rejection, intimal hyperplasia, reocclusion and/or aneurysm formation were not observed. Presence of host cells and neoendothelialization were microscopically confirmed. CONCLUSIONS: This decellularization protocol enables a cost-effective preparation of biological grafts featuring reduced immunogenicity. The implanted grafts did not degenerate during the 6-month follow-up period, the lack of graft rejection suggests acceptable immunological tolerance, while recipient cells migrate into, proliferate and differentiate, thus creating the possibility for further use as an optional vascular graft.

The effect of trimetazidine in reducing the ischemia-reperfusion injury in rat epigastric skin flaps.

BACKGROUND: Ischemia-reperfusion injury may lead to insufficient microcirculation and results in partial flap loss during the free flap surgeries. OBJECTIVE: This study aimed to investigate the effect of trimetazidine (TMZ) on oxidative stress, inflammation and histopathological changes, using the epigastric skin flap model in rats. METHODS: 40 male Wistar rats were used, that were divided into four groups. Control group, non-treated ischemic (I/R)-group and two trimetazidine treated groups (preischemically, postischemically) were established. To create ischemia in the skin flap, the superficial epigastric vessels were clamped for six hours, followed by twenty-four hours of reperfusion. Blood samples and biopsies from skin flaps were collected at the end of the reperfusion period. The inflammatory response, the degree of oxidative stress (by measuring the plasma level of malondialdehyde (MDA), reduced glutathione (GSH); sulfhydryl (-SH) groups) and histopathological changes were evaluated. RESULTS: Inflammatory response, and oxidative stress were significantly attenuated in the trimetazidine treated groups, compared to the non-treated ischemic group. Histopathological findings were also correlated with the biochemical results. CONCLUSION: In our study trimetazidine could reduce the ischaemia-reperfusion injury, even after an unexpected ischemic period, so it is a promising drug during free tissue transfer, replantation or during revascularization procedures in the future.

The role of GST polymorphism in reperfusion induced oxidative stress, inflammatory responses and clinical complications after surgical and percutaneous coronary intervention.

BACKGROUND: Patients having coronary artery disease treated by coronary bypass or PCI procedure are exposed to tissue damage because of the phenomenon called reperfusion injury. Reperfusion injury can be characterized/monitored by oxidative stress parameters, inflammatory markers and by post-operative complication rate. OBJECTIVE: Beyond the obvious factors determining its severity (affected myocardial mass, ischaemic time, collateral circulation etc.) we examined the GST enzyme group's most cardio selective member, GSTP1 and its genetic polymorphism if there is any genetically determined preventive effect on the above-mentioned parameters. MATERIALS AND METHODES: We have performed randomized prospective study in the Heart Institute of Pecs with 862 patients, treated by coronary bypass or PCI procedure. Blood samples were taken a day before, one hour, one day, one week after the operation. Leucocyte count (WBC), myeloperoxidase (MPO), thiol group (SH); Superoxide dismutase (SOD), malondialdehyde (MDA), reduced Glutathione (GSH) level was checked in different periods of time as a comparison. The onset of myocardial damage and the corresponding necro enzyme level changes were registered in the perioperative period. Our patient's GSTP1 allele pair combinations (A, B, or C) were determined by real time PCR method. RESULTS: In patients with GSTP1 AA genotype we have found significance level reaching plasma concentration rise in SOD and MDA, and drop in GSH, SH. The CKMB concentration rise in the post-operative 24 hours was significantly higher in the GSTP1 AA group. CONCLUSIONS: According to our results the AA allele combination can be considered as a risk factor. GSTP1-AA allele pair has negative effect on ischemia-reperfusion tolerance of the heart. In case of cardiovascular interventions, the study of GST enzyme polymorphisms can be an independent risk stratification factor in determining the perioperative risk in the future.

Pentoxifylline attenuates the local and systemic inflammatory response after infrarenal abdominal aortic ischemia-reperfusion.

AIMS: We studied the new anti-inflammatory effects of non-specific phosphodiesterase (PDE) inhibitor pentoxifylline (PTX) on ischaemia-reperfusion injury and postconditioning of the lower extremities. We aimed to examine the oxidative stress parameters (OSP), the inflammatory response and the changes in structure of skeletal muscle after revascularization surgery. METHODS: 50 Wistar rats in five groups underwent a 60 min infrarenal aortic cross clamping. After the ischaemia in IR+PC group ischemic postconditioning was performed, intermittent 15 seconds reperfusion, 15 seconds ischaemic periods were applied four times. The ischemic phase was followed by a 120 min of reperfusion. In IR+PTX group the animals were treated with PTX. In IR+PC+PTX group both ischemic postconditioning and PTX treatment were performed. Blood samples and biopsy from quadriceps muscle were collected. Plasma malondialdehyde, reduced glutathione, -SH-groups, TNF-alpha, IL-6 concentrations and superoxide dismutase enzyme activity were measured. RESULTS: The levels of OSP and the inflammatory proteins were significantly higher in the IR group. PTX treatment and PC could significantly decrease the levels of OSP and inflammatory proteins. When the animals were co-treated with PTX and PC the results were even better. CONCLUSIONS: Inhibition of PDE by PTX could markedly decrease the inflammatory response and moderate the ischaemia-reperfusion damages after lower limb ischemia and reperfusion. Administration of PTX could potentiate the beneficial effects of PC.

Sodium Pentosan Polysulfate Reduced Renal Ischemia-Reperfusion-Induced Oxidative Stress and Inflammatory Responses in an Experimental Animal Model.

Acute kidney injury (AKI) remains an independent risk factor for mortality and morbidity after vascular surgery (affecting the renal arteries) or aortic surgery (requiring suprarenal aortic clamping). These types of vascular surgery produce renal ischemia/reperfusion (I/R) injury, a common cause of AKI. The present studies aimed at monitoring the course of renal I/R injury at the cellular level and investigating the efficacy of long-term preoperative and single-shot intraoperative administration of sodium pentosan polysulfate (PPS) to protect renal tissue from acute I/R injury both in native and diabetic kidneys in rats. Western blot analyses of the proapoptotic (bax) and antiapoptotic (bcl-2) signaling pathways, as well as the extent of DNA damage (phospho-p53), were performed. Oxidative stress followed upon the termination of malondialdehyde, reduced glutathione, thiol group, and superoxide dismutase plasma levels. Inflammatory changes were measured by the determination of serum tumor necrosis factor-α and interleukin-1 levels. Morphological changes were detected by histological examinations. Our results showed that the long-term administration of PPS has an advantage in reducing I/R kidney injury in diabetic rats, while high-dose, single-shot parenteral administration of PPS prior to revascularization might be useful in nondiabetic rats.

[Cerebral hyperperfusion syndrome and blood pressure control]. / Cerebralis hyperperfusiós szindróma és vérnyomáskontroll.

INTRODUCTION: Cerebral hyperperfusion syndrome is a rare, hardly known condition, which can result in serious complications either after surgical or endovascular revascularization. Recognition of the typical triad (headache, seizure, focal neurological deficit) and the prompt radiological diagnosis (sonography, computed tomography) are crucial to achieve a favourable outcome. AIM: The aim of the authors was to select the endangered group and set up an effective therapeutic protocol based their own experience in combination with relevant literature data. METHOD: From the beginning of 2010 up to now three cases with these symptoms pursuant to the criteria of cerebral hyperperfusion syndrome have been recognized by the authors. RESULTS: Each of the three patients were treated by similar principles on intensive care unit, but the applied therapy resulted in complete remission in one patient only. CONCLUSIONS: At present there is no efficient diagnostic way to screen the endangered group, hence the only opportunity for prevention is the appropriate perioperative blood pressure control. If symptoms have developed already, urgent treatment is required.

Inhibition of Glutathione S-Transferase by Ethacrynic Acid Augments Ischemia-Reperfusion Damage and Apoptosis and Attenuates the Positive Effect of Ischemic Postconditioning in a Bilateral Acute Hindlimb Ischemia Rat Model.

AIMS: We studied the effects of the inhibition of the endogene antioxidant glutathione-S-transferase (GST) by ethacrynic acid (EA) on ischemia-reperfusion (IR) injury and postconditioning (PC) in the lower extremities. We aimed to examine the oxidative stress parameters (OSP), inflammatory response and activation of proapoptotic signaling proteins (PSP) after revascularization surgery. METHODS: Sixty Wistar rats were divided into 6 groups: control, IR, PC, EA-control, IR and administration of EA (IR/EA) and PC and administration of EA (PC/EA). The IR, PC, IR/EA and PC/EA groups underwent 60 min of infrarenal aortic cross-clamping. After that, PC was performed in the PC and PC/EA groups. In 3 of the groups, the animals were treated with EA (EA-control, IR/EA and PC/EA groups) as well. The ischemia was followed by 120 min of reperfusion. Blood samples and biopsy specimens were collected from the quadriceps muscle. Plasma malondialdehyde, reduced glutathione, thiol/sulfhydryl group levels, TNF-α and IL-6 concentrations and superoxide-dismutase enzyme activity were measured. RESULTS: The levels of the OSP and the inflammatory proteins were higher in the EA-administered groups. The ratio of phosphorylated PSP was higher in the EA-administered groups and the protective effect of PC did not develop. CONCLUSIONS: Inhibition of GST by EA augmented the IR damage. GST inhibition was associated with a different activation of the mitogen-activated protein kinases and the PSP, regulating these pathways in the process of apoptosis and PC.

[Dilemmas of the reconstruction of the major pelvic artery due to infectious aortic graft complication]. / A medencei veroér helyreállításának dilemmái szeptikus aortagraft esete kapcsán.

INTRODUCTION: In the pelvic region thrombendarterectomy and bypass procedures are the most commonly performed procedures to treat peripheral artery occlusive diseases with chronic, severe circulation failure caused by atherosclerosis. Biologic and synthetic grafts can also be used in bypass surgeries. Application of synthetic grafts can acutely increase the development of the infectious graft complication and its mortality is still between 70 and 75% in pelvic processes. We describe the difficulties and dilemmas of an infectious aortobifemoral graft. CASE PRESENTATION: 58-year-old female patient with right lower limb trophic ulcer underwent a DSA examination showing a bilateral iliac occlusion and aortobifemoral bypass surgery with Dacron graft implantation was performed. Re-occlusion and infection of the graft led to an in situ silver Dacron graft replacement. Due to the one-sided re-occlusion, a femoro-femoral crossover bypass surgery applying silver graft was performed. Despite the previously described procedures the infectious process got worse and autologous deep vein reconstruction was required beside the removal of the infectious synthetic grafts at the same time. DISCUSSION: There are local and extraanatomical solutions to reduce infectious graft complication. In pelvic infections bypass surgeries using autologous deep vein can show the best results. This procedure is the trustworthiest but also the most straining technique due to the extension of surgical time and increased blood loss. The proper surgical strategy should be selected on individual bases including cardiopulmonary load ability, patient age and technical/infrastructural possibilities.

Substance misuse prevention as corporate social responsibility.

All sectors of society should be involved in reducing substance misuse, including businesses. However, the business sector is typically involved only to the extent that their products compel them to be (e.g., alcohol producers promoting responsible alcohol consumption). This article examines why business participation has been limited and how embedding prevention within a framework of health promotion could increase participation. It reviews both Hungarian and international cases, concluding that although corporate social responsibility (CSR) offers a framework to approach substance misuse reduction, a different perception of the role of the business sector is necessary to make it viable.

[Endovascular treatment of subclavian artery pseudoaneurysm as a delayed complication after surgery for aorto-bifemoral graft infection]. / Aortobifemoralis graft gennyedés megoldásának kései szövodményeként jelentkezo subclavia álaneurysma endovascularis kezelése.

CASE REPORT: In this article we present a relatively rare vascular surgical complication and an uncommon treatment of it. In this case we used an aorto-bifemoral bypass on a patient with Leriche syndrome. The implanted Y-graft got infected and we were forced to remove it. Having inserted the abdominal aortic graft, an axillobifemoral bypass was also applied to secure the circulation of the lower limbs. However, the graft occluded later on, and 37 months after the inital surgery a rather large pseudoaneurysm developed at the origin of the graft in the right subclavian artery. Another surgical intervention was indicated to prevent embolisation, rupture and compression. Instead of the conventional surgical method (resection, interposition) we did an endovascular procedure. We removed the false aneurysm by inserting a covered stent, using catheter technique, into the right brachial artery and therefore prevented the previously mentioned complications. DISCUSSION: This minimal invasive method is very useful for high risk patients to prevent the injury of neighbouring anatomical structures in the region as well as minimize blood loss and potential complications of long term anaesthesia when open surgery is done.

[Evaluation of calf muscle pump function after subfascial endoscopic perforator dissection]. / Lábszgári izompumpa-funkció vizsgálata endoszkópos perforans dissectio után.

INTRODUCTION: Perforating vein incompetence can be demonstrated in many cases of chronic venous insufficiency. Subfascial endoscopic perforating vein surgery has become an accepted method in the treatment of perforator insufficiency over the past decade. However, what the hemodynamic consequences of perforating vein interruption are, is not clear. OBJECTIVE: To analyse the results of endoscopic perforating vein surgery performed in the authors' department over the past four years. To compare the outcome of the operation in patients with deep venous reflux with those who only had superficial and perforating vein reflux. To analyze the changes in calf muscle pump function with photoplethysmography carried out before and after the operation. METHODS: Subfascial endoscopic perforator dissection was performed on 53 patients who suffered from severe chronic venous insufficiency mainly with active ulcer. With the follow-up of 51 patients, a prospective clinical trial was carried out which consisted of physical examination and Doppler ultrasound, six weeks after the operation and every three months thereafter. The data were analyzed with standard statistical methods. RESULTS: Clinical symptoms improved and proved to be durable in 33 (64%) of 51 patients during the follow-up. The healing rate of 40 patients with venous ulcer was 82% after the operation. 62% of the patients were ulcer-free on a long term. While of the 33 patients with deep venous reflux 17 (52%) showed durable improvement, of the 18 patient who had only superficial and perforating vein insufficiency only 2 patients did not experience improvement. Average venous refilling time of the 33 patients who underwent photoplethysmography increased from 12.5 sec to 14.6 sec after endoscopic perforating vein surgery although significant improvement was demonstrated only on patients who had only superficial venous reflux together with perforating vein insufficiency. CONCLUSIONS: Subfascial endoscopic perforating vein surgery proved to be a successful alternative in the authors' clinical trial in treating patients with severe chronic venous insufficiency. Mid-term evaluation showed excellent results with patients without deep venous reflux and acceptably good outcome with patients with deep venous insufficiency. Improvement of clinical symptoms after endoscopic perforator dissection was followed by significant improvement of calf muscle pump function only on patients without deep venous reflux.