[Examination of cytoprotective and anti-inflammatory effect of PACAP-38 on small bowel autotransplantation]. / A PACAP-38 citoprotektív és antiinflammatórikus hatásának vizsgálata vékonybél-autotranszplantációs modellben.

INTRODUCTION: The small intestine is one of the most sensitive organs to ischemia-reperfusion injury during transplantation. Cytoprotective effect of pituitary adenylate cyclase-activating polypeptide (PACAP) is well known. The aim of our study was to measure changes of PACAP-38-like immunoreactivities and cytokine levels in intestinal grafts stored PACAP-38 containing preservation solution. MATERIAL AND METHODS: Small-bowel autotransplantation was performed on male Wistar rats (n = 56). Grafts were stored in University of Wisconsin (UW) solution at 4 °C for 1 (GI), 3 (GII), and 6 hours (GIII); and in PACAP-38 containing UW solution for 1 (GIV), 3 (GV), and 6 hours (GVI). Reperfusion lasted 3 hours in each group. Intestinal PACAP-38 immunoreactivities were measured by radioimmunoassay. To measure cytokine from tissue homogenates we used rat cytokine array and Luminex Multiplex Immunoassay. RESULTS: Levels of PACAP-38-like and PACAP-27-like immunoreactivities decreased by preservation time compared to control. This decrease was significant following 6 hours cold storage (p < 0.05). Values remained significantly higher in grafts stored in PACAP-38 containing UW. Expressions of sICAM-1, L-selectin, tissue inhibitor of metalloproteinase-1 were increased in GIII and were decreased in GVI. CONCLUSION: PACAP-38 increased tissue levels of PACAP-38 and PACAP-27, and decreased cytokine expression. This indicates that PACAP-38 has anti-inflammatory and cytoprotective effects in intestinal autotransplantation model.

[Examination of warm and cold ischemic injury of small intestine by differential scanning calorimetry]. / A vékonybél meleg és hideg ischaemiás károsodásának kimutatása differenciál pásztázó kalorimetriás vizsgálattal.

INTRODUCTION/AIM: Small bowel is extremely sensitive to ischemia/reperfusion injury. Therefore, we compared the effect of warm and cold ischemia on the intestinal structural changes by differential scanning calorimetry (DSC) method. MATERIALS AND METHODS: Warm and cold ischemia groups were established on Wistar rats with 1, 3 and 6 h ischemic times (n = 30). Intestinal biopsies were collected after laparotomy and at the end of the ischemia periods. DSC measurement was performed on the mucosa, muscular layer and total intestinal wall. RESULTS: Our DSC data confirmed that longer warm ischemia period caused more severe damage in the structure of the mucosa and muscular layers. The results of transition temperature and calorimetric enthalpy suggest that these changes can be reduced using cold ischemic procedure in University of Wisconsin solution. However, the extent of thermal destruction of each layers following cold preservation injury was significantly different compared to normal bowel structure. CONCLUSION: In this study we quantitatively measured structural changes in the gut following ischemia using DSC. This thermodynamic method may provide basis for further investigations in different bowel stress models.

[Effect of ischemic postconditioning on oxidative stress and structural tissue changes in intestinal warm ischemic and autotransplantation models]. / Az ischaemiás posztkondicionálás hatása az oxidatív stresszre és a szöveti struktúrára vékonybél meleg ischaemiás és autotranszplantációs modellekben.

INTRODUCTION/AIM: Our study investigated the effect of ischemic postconditioning (IPO) in intestinal warm ischemia/reperfusion (I/R) and autotransplantation models. MATERIALS AND METHODS: Warm ischemia was performed by occlusion of superior mesenteric artery for 1, 3 and 6 hours in white domestic pigs (n = 15). Prior to 3 hours reperfusion the intestine was postconditioned by 3 cycles of 30-seconds ischemia and 30-seconds reperfusion (IPO protocol). In the cold ischemia group (n = 15) the bowel was preserved in University of Wisconsin solution for 1, 3, and 6 hours. Prior to 3 hours reperfusion IPO protocol was applied, too. Tissue samples were collected after laparotomy (control) and at the end of the reperfusion periods. As far as oxidative stress markers, malondialdehyde and reduced glutathione (GSH) levels and superoxide dismutase (SOD) activity were determined. Tissue damage was evaluated by qualitative (Park-classification) and quantitative (Scion Image) methods. RESULTS: As regards oxidative stress parameters, lipidperoxidation decreased and the protective effect of endogenous antioxidants (GSH, SOD) retained significantly by IPO procedure at the end of reperfusion. Tissue injury correlated significantly by the duration of warm ischemia and cold preservation. Quantitative analysis demonstrated that IPO ameliorated tissue injury in each group (p < 0.05). CONCLUSION: IPO significantly attenuated intestinal oxidative stress and morphological damages in warm and cold I/R models.

Influence of PACAP on oxidative stress and tissue injury following small-bowel autotransplantation.

Tissue injury caused by cold preservation and reperfusion remains an unsolved problem during small-bowel transplantation. Pituitary adenylate cyclase-activating polypeptide (PACAP) is present and plays a central role in the intestinal physiology. This study investigated effect of PACAP-38 on the oxidative stress and tissue damage in autotransplanted intestine. Sham-operated, ischemia/reperfusion, and autotransplanted groups were established in Wistar rats. In ischemia/reperfusion groups, 1 h (group A), 2 h (group B), and 3 h (group C) ischemia followed by 3 h of reperfusion was applied. In autotransplanted groups, total orthotopic intestinal autotransplantation was performed. Grafts were preserved in University of Wisconsin (UW) solution and in UW containing 30 microg PACAP-38 for 1, 2, 3, and 6 h. Reperfusion lasted 3 h in all groups. Endogenous PACAP-38 concentration was measured by radioimmunoassay. To determine oxidative stress parameters, malondialdehyde, reduced glutathione, and superoxide dismutase were measured in tissue samples. Tissue damage was analyzed by qualitative and quantitative methods on hematoxylin/eosin-stained sections. Concentration of endogenous PACAP-38 significantly decreased in groups B and C compared to sham-operated group. Preservation solution containing PACAP-38 ameliorated bowel tissue oxidative injury induced by cold ischemia and reperfusion. Histological results showed that preservation caused destruction of the mucous, submucous, and muscular layers, which were further deteriorated by the end of reperfusion. In contrast, PACAP-38 significantly protected the intestinal structure. Ischemia/reperfusion decreased the endogenous PACAP-38 concentration in the intestinal tissue. Administration of PACAP-38 mitigated the oxidative injury and histological lesions in small-bowel autotransplantation model.

Changes of PACAP immunoreactivities and cytokine levels after PACAP-38 containing intestinal preservation and autotransplantation.

Small bowel is one of the most sensitive organs to ischemia-reperfusion injury, which is a significant problem during transplantation. Pituitary adenylate cyclase-activating polypeptide (PACAP) has cytoprotective effect in ischemic injuries of various tissues. The aim of our study was to measure changes of PACAP-38 and PACAP-27 immunoreactivities and cytokine levels in intestinal grafts stored in PACAP-38-containing preservation solution. Small bowel autotransplantation was performed on male Wistar rats. Grafts were stored in University of Wisconsin (UW) solution at 4 °C for 1 h (group (G)I), for 3 h (GII), and for 6 h (GIII) and in PACAP-38-containing UW solution for 1 h (GIV), for 3 h (GV), and for 6 h (GVI). After preservation, performing vessel anastomosis reperfusion began, which lasted 3 h in each group. Tissue biopsies were collected after laparotomy (control) and at the end of the reperfusion periods. Intestinal PACAP-38 and PACAP-27 immunoreactivities were measured by radioimmunoassay. To measure cytokines from tissue homogenates, we used rat cytokine array and Luminex Multiplex Immunoassay. Levels of PACAP-38 and PACAP-27 immunoreactivity decreased after 1 and 3 h preservation compared to control levels. This decrease was significant following 6 h cold storage (p < 0.05). Values remained significantly higher in grafts stored in PACAP-38-containing UW. Cytokine array revealed that expression of the soluble intercellular adhesion molecule-1 (CD54) and L-selectin (CD62L/LECAM-1) was increased in GIII. Both 6 h cold storage in PACAP-38-containing UW solution and 3 h reperfusion caused strong reduction in these cytokines activation in GVI. RANTES (CCL5) levels were increased in all groups. Strong activation of the tissue inhibitor of metalloproteinase-1 was in GIII. However, PACAP-38-containing cold storage could decrease its activation in GVI. Furthermore, strong activation of the tissue inhibitor of metalloproteinase-1 was detected in 6 h preserved grafts without PACAP-38 (GIII). PACAP-38-containing cold storage could decrease its activation in GVI. Our present study showed that PACAP-38 and PACAP-27 immunoreactivities decreased in a time-dependent manner during intestinal cold preservation, which could be ameliorated by administration of exogenous PACAP-38 to the preservation solution. Moreover, PACAP-38 could attenuate tissue cold ischemic injury-induced changes in cytokine expression.