E. Laszlo

PACAP is an endogenous protective factor-insights from PACAP-deficient mice.

Pituitary adenylate cyclase-activating polypeptide (PACAP) is a widespread neuropeptide with a diverse array of biological functions. Not surprisingly, the lack of endogenous PACAP therefore results in a variety of abnormalities. One of the important effects of PACAP is its neuroprotective and general cytoprotective role. PACAP protects neurons and other tissues against ischemic, toxic, and traumatic lesions. Data obtained from PACAP-deficient mice provide evidence that endogenous PACAP also has protective functions. Mice lacking PACAP are more vulnerable to different in vitro and in vivo insults. The present review summarizes data on the increased sensitivity of PACAP-deficient mice against harmful stimuli. Mice lacking PACAP respond with a higher degree of injury in cerebral ischemia, autoimmune encephalomyelitis, and axonal lesion. Retinal ischemic and excitotoxic injuries also produce increased cell loss in PACAP-deficient mice. In peripheral organs, kidney cell cultures from PACAP-deficient mice are more sensitive to oxidative stress and in vitro hypoxia. In vivo, PACAP-deficient mice have a negative histological outcome and altered cytokine response in kidney and small intestine ischemia/reperfusion injury. Large intestinal inflammation, toxic lesion of the pancreas, and doxorubicin-induced cardiomyopathy are also more severe with a lack of endogenous PACAP. Finally, an increased inflammatory response has been described in subacute endotoxin-induced airway inflammation and in an oxazolone-induced allergic contact dermatitis model. In summary, lack of endogenous PACAP leads to higher vulnerability in a number of injuries in the nervous system and peripheral organs, supporting the hypothesis that PACAP is part of the endogenous cytoprotective machinery.

Mice deficient in pituitary adenylate cyclase activating polypeptide (PACAP) show increased susceptibility to in vivo renal ischemia/reperfusion injury

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide with well-known cytoprotective effects. We have reported earlier that PACAP decreases mortality and the degree of tubular atrophy in a rat model of renal ischemia/reperfusion injury. Recently, we have shown that kidney cultures isolated from PACAP deficient mice show increased susceptibility to renal oxidative stress. Based on these previous studies, we raised the question whether PACAP deficient mice display increased sensitivity to in vivo kidney ischemia/reperfusion. PACAP⁻/⁻ mice underwent 45 or 60 min of renal ischemia followed by 2 weeks reperfusion. Kidneys were processed for histological analysis. Sections stained with PAS-haematoxylin were graded for the following parameters: degree of tubular dilation, Bowmanns capsule dilation, lymphocyte and macrophage infiltration, thyroidization and the disappearance of the PAS-positive glycocalyx from under the brush border. In other sets of experiments, tissue cytokine expression and the level of the endogenous antioxidant superoxide dismutase (SOD) were also determined after 60 min ischemia/reperfusion. Our results show that while intact kidneys were not different between wild-type and PACAP deficient mice, marked differences were observed in the histological structures in groups that underwent ischemia/reperfusion. PACAP deficient mice had a worse histological outcome, with significantly higher histological scores for all tested parameters. Cytokine expression was also markedly different between wild-type and PACAP deficient mice. In addition, the level of SOD was significantly lower in PACAP⁻/⁻ animals after ischemia/reperfusion. In conclusion, the lack of endogenous PACAP leads to higher susceptibility to in vivo renal ischemia/reperfusion, suggesting that PACAP has an endogenous renoprotective effect.

Effects of PACAP on Oxidative Stress-Induced Cell Death in Rat Kidney and Human Hepatocyte Cells

Oxidative stress plays an important role in various renal and hepatic pathologies, and reduction of oxidative stress-induced processes is an important protective strategy in tissues of diverse origins against harmful stimuli. Pituitary adenylate cyclase activating polypeptide (PACAP) is a well-known cytotrophic and cytoprotective peptide. PACAP promotes cell survival in numerous cells and tissues exposed to various stimuli. Protective effects of PACAP have been shown in the kidney, but it is not known whether PACAP is protective against oxidative stress in renal cells. Little is known about the effects of PACAP in the liver. The aim of the present study was to investigate whether PACAP is protective against oxidative stress in primary rat kidney cell culture and whether PACAP has any effect on cell survival in human WRL-68 hepatocytes and HEP-G2 hepatocellular carcinoma cells subjected to oxidative stress. Cells were exposed to various concentrations of H2O2 with or without PACAP co-treatment and cell viability was evaluated with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide test (MTT). We found that oxidative stress induced a significant decrease in cell viability in both cell lines. PACAP could dose-dependently increase the percentage of living cells in kidney cells, but it failed to do so in hepatocytes. Given the survival-promoting effects of PACAP against oxidative stress in rat kidney, we conducted a further experiment to determine whether PACAP influences the markers of oxidative stress in vivo. We have proven earlier that PACAP was effective in kidney ischemia/reperfusion injury in vivo. In the present study, we determined the levels of the oxidative stress marker malondialdehyde and the activity of the scavenger molecules glutathione (GSH) and superoxide dismutase (SOD) following kidney ischemia/reperfusion in rats. We found that PACAP significantly increased the level of GSH and counteracted the marked reduction of SOD activity after ischemia/reperfusion in vivo. In summary, the present study showed that while PACAP was able to significantly increase the cell survival in primary kidney cell cultures exposed to oxidative stress, possibly involving interaction with the endogenous scavenger system, it failed to influence the viability of normal or cancerous hepatocytes.

Effects of pituitary adenylate cyclase activating polypeptide in the urinary system, with special emphasis on its protective effects in the kidney

Pituitary adenylate cyclase activating polypeptide (PACAP) is a widespread neuropeptide with diverse effects in the nervous system and peripheral organs. One of the most well-studied effects of PACAP is its cytoprotective action, against different harmful stimuli in a wide variety of cells and tissues. PACAP occurs in the urinary system, from the kidney to the lower urinary tract. The present review focuses on the nephroprotective effects of PACAP and summarizes data obtained regarding the protective effects of PACAP in different models of kidney pathologies. In vitro data show that PACAP protects tubular cells against oxidative stress, myeloma light chain, cisplatin, cyclosporine-A and hypoxia. In vivo data provide evidence for its protective effects in ischemia/reperfusion, cisplatin, cyclosporine-A, myeloma kidney injury, diabetic nephropathy and gentamicin-induced kidney damage. Results accumulated on the renoprotective effects of PACAP suggest that PACAP is an emerging candidate for treatment of human kidney pathologies.

Presence of Pituitary Adenylate Cyclase Activating Polypeptide and Its Type I Receptor in the Rat Kidney

Pituitary adenylate cyclase activating polypeptide (PACAP), a multifunctional neuropeptide, has 2 active forms, PACAP38 and PACAP27. It is now well-established that PACAP has several actions also in peripheral organs, including renoprotective effects. The peptide itself has not been previously identified in the rat kidney. The first aim of our study was to identify PACAP in the rat kidney using mass spectrometry and radioimmunoassay (RIA). Receptor mRNA and binding studies revealed the existence of all 3 PACAP receptors (PAC1, VPAC1, and VPAC2) in the kidney, but their exact localization in histologic sections was not evident. Because most of the cytoprotective effects of PACAP relate to its specific PAC1 receptor, our second aim was to identify the cell types wherein the PAC1 receptor is expressed in the rat kidney. Mass spectrometry revealed the presence of PACAP38 in the kidney. RIA measurements showed both PACAP38- and PACAP27-like immunoreactivities in kidney homogenates, with PACAP38 being dominant. Immunohistochemistry revealed PAC1 receptor-like immunoreactivity in kidney sections, mainly expressed in cortical tubular epithelial cells. These results showed PACAP to be endogenously present in the kidney. The tubular localization of the PAC1 receptor provides the basis for the renal effects of the peptide under physiologic and pathologic conditions.

Mice deficient in neuropeptide pacap demonstrate increased sensitivity to in vitro kidney hypoxia

One of the well-known effects of pituitary adenylate cyclase activating polypeptide (PACAP) is its neuroprotective and cytoprotective actions including renoprotective effects. Mice deficient in endogenous PACAP exhibit several behavioral, metabolic, and developmental alterations. Furthermore, PACAP-deficient mice have larger infarct volume in a model of cerebral ischemia, delayed axonal regeneration, and increased cell death in cerebellar oxidative stress. We have previously demonstrated that PACAP-deficient mice have increased susceptibility to in vitro oxidative stress, which can be counteracted by exogenous PACAP treatment. These results demonstrate that endogenous PACAP has a protective role against various stressors. The objective of the present study was to investigate whether endogenous PACAP has a protective effect in the kidney against in vitro hypoxia. Kidney cell cultures were isolated from wild-type and PACAP-deficient mice, and cell viability was assessed after in vitro hypoxia induced using CoCl(2). The sensitivity of cells from PACAP-deficient mice was increased to hypoxia: both after 24 and 48 hours of exposure, cell viability was significantly reduced compared with that in control wild-type mice. These results show that endogenous PACAP protects against noxious stimuli in the kidney and that PACAP may act as a stress sensor in renal cells.

Ischemia/reperfusion-induced Kidney Injury in Heterozygous PACAP-deficient Mice

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide with very diverse distribution and functions. Among others, PACAP is a potent cytoprotective peptide due to its antiapoptotic, anti-inflammatory, and antioxidant actions. This also has been shown in different kidney pathologies, including ischemia/reperfusion-induced kidney injury. Similar protective effects of the endogenous PACAP are confirmed by the increased vulnerability of PACAP-deficient mice to different harmful stimuli. Kidneys of homozygous PACAP-deficient mice have more severe damages in renal ischemia/reperfusion and kidney cell cultures isolated from these mice show increased sensitivity to renal oxidative stress. In our present study we raised the question of whether the partial lack of the PACAP gene is also deleterious, i.e. whether heterozygous PACAP-deficient mice also display more severe damage after renal ischemia/reperfusion. Mice underwent 45 or 60 minutes of ischemia followed by 2 weeks reperfusion. Histological evaluation of the kidneys was performed and individual histopathological parameters were graded. Furthermore, we investigated apoptotic markers, cytokine expression, and the activity of superoxide dismutase (SOD) enzyme 24 hours after 60 minutes of renal ischemia/reperfusion. We found no difference between the intact kidneys of wild-type and heterozygous mice, but marked differences could be observed following ischemia/reperfusion. Heterozygous PACAP-deficient mice had more severe histological alterations, with significantly higher histopathological scores for most of the tested parameters. Higher level of the proapoptotic pp38 MAPK and of some proinflammatory cytokines, as well as lower activity of the antioxidant SOD could be found in these mice. In conclusion, the partial lack of the PACAP gene results in worse outcomes in cases of renal ischemia/reperfusion, confirming that PACAP functions as an endogenous protective factor in the kidney.

The effects of pituitary adenylate cyclase activating polypeptide in renal ischemia/reperfusion

Pituitary adenylate cyclase activating polypeptide (PACAP ) is a multifunctional neuropeptide occurring in the nervous system as well as in the peripheral organs. Beneficial action of PACAP has been shown in different pathological processes. The strong protective effects of the peptide are probably due to its complex modulatory actions in antiapoptotic, anti-inflammatory and antioxidant pathways. In the kidney, PACAP is protective in models of diabetic nephropathy, myeloma kidney injury, cisplatin-, gentamycin- and cyclosporin-induced damages. Numerous studies have been published describing the protective effect of this peptide in renal ischemia/reperfusion. The present review focuses on the ischemia/reperfusion-induced kidney injury and gives a brief summary about the results published in this area.

Changes in Pituitary Adenylate Cyclase-Activating Polypeptide Following Renal Ischemia-Reperfusion in Rats

PACAP (pituitary adenylate cyclase-activating polypeptide) occurs in two biologically active forms, with 38 and 37 amino acid residues (PACAP38 and PACAP27). In mammalian tissues, PACAP38 is the dominant form. Diverse effects have been described in the cardiovascular, respiratory, gastrointestinal, and urogenital systems. PACAP is known for its strong cytoprotective effects, which are present endogenously as well, as proven by knockout studies and results showing that PACAP is up-regulated following diverse injuries. Little is known about such effects in the kidney. We have previously shown that PACAP is protective in renal ischemia-reperfusion injury. Therefore, the aim of the present study was to investigate the changes of endogenous PACAP following 60-minute renal ischemia using radioimmunoassay. Changes were observed within 24 hours following renal vessel clamping. In the cortex, an acute decrease was followed by an increase on the intact side, and levels returned to original ones on the operated side. In the medulla, changes were only observed on the clamped side: a marked up-regulation was detected in PACAP38-like immunoreactivity within the first 24 hours. The present study showed that PACAP38- and PACAP27-like immunoreactivities sensitively react to renal ischemia-reperfusion, the physiological importance of which awaits further investigation.