Wren JD, Sonntag WE, Ungvari Z. Caloric restriction confers persistent anti-oxidative, pro-angiogenic, and anti-inflammatory effects and promotes anti-aging miRNA expression profile in cerebromicrovascular endothelial cells of aged rats.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a neuropeptide with highly potent neurotrophic and neuroprotective effects. PACAP and its receptors occur in the retina and PACAP has been applied in animal models of metabolic retinal disorders to reduce structural and functional damage. Furthermore, PACAP has been implicated as a potential anti-diabetic peptide. Our aim has been to investigate, by using a complex morphological, immunochemical and molecular biological approach, whether PACAP attenuates diabetic retinopathy. Diabetes was induced in rats with a single streptozotocin injection. PACAP was injected intravitreally into one eye (100 pmol) three times during the last week of a 3-week survival period. Retinas were processed for the following procedures: routine histology, immunohistochemistry (single and double labeling, whole-mount), quantitative reverse transcription with the polymerase chain reaction and Western blotting. Cone photoreceptors and dopaminergic amacrine and ganglion cells degenerated in diabetic retinas and glial fibrillary acidic protein were upregulated in Müller glial cells. The number of cones, the length of their outer segments and the cell number in the ganglion cell layer were decreased. PACAP ameliorated these structural changes. Moreover, PACAP increased the levels of PAC1-receptor and tyrosine-hydroxylase as detected by molecular biological methods. Thus, PACAP has significant protective effects in the diabetic retina. PACAP treatment attenuates neuronal cell loss in diabetic retinopathy, the protective effects of PACAP probably being mediated through the activation of PAC1-receptor. These results suggest that PACAP has a therapeutic potential in diabetic retinopathy.

Pituitary adenylate cyclase activating polypeptide (PACAP) is a pleiotropic and multifunctional neuropeptide having important roles in various physiological processes. Recent trends in PACAP research point to the clinical introduction of PACAP or its analogs/fragments possibly in the near future. Recently, we have shown the presence of PACAP in human plasma, milk, placenta, and follicular fluid samples. However, relatively few data are available on PACAP in human tissues from patients with different disorders. The aim of the present study was to determine, by radioimmunoassay, the tissue level of PACAP38-like immunoreactivity (LI) and PACAP27-LI in different primary non-small cell lung cancer, colon tumor samples, and in cardiac muscle samples from patients suffering from ischemic heart disease and valvular disorders. We also labeled the PAC1 receptors in human cardiac cells. All samples showed significantly higher PACAP38-LI compared with PACAP27-LI. We found significantly lower levels of PACAP38-LI and PACAP27-LI in tumoral and peripheral samples compared with normal healthy tissue in both lung and colon cancers. Further investigations are necessary to describe the exact function of PACAP in oncogenesis. We showed that PACAP38-LI and PACAP27-LI are significantly higher in ischemic heart diseases compared with valvular abnormalities, suggesting that PACAP might play a role in ischemic heart disorders.

Aging impairs angiogenic capacity of cerebromicrovascular endothelial cells (CMVECs) promoting microvascular rarefaction, but the underlying mechanisms remain elusive. PACAP is an evolutionarily conserved neuropeptide secreted by endothelial cells and neurons, which confers important antiaging effects. To test the hypothesis that age-related changes in autocrine PACAP signaling contributes to dysregulation of endothelial angiogenic capacity, primary CMVECs were isolated from 3-month-old (young) and 24-month-old (aged) Fischer 344 x Brown Norway rats. In aged CMVECs, expression of PACAP was decreased, which was associated with impaired capacity to form capillary-like structures, impaired adhesiveness to collagen (assessed using electric cell-substrate impedance sensing [ECIS] technology), and increased apoptosis (caspase3 activity) when compared with young cells. Overexpression of PACAP in aged CMVECs resulted in increased formation of capillary-like structures, whereas it did not affect cell adhesion. Treatment with recombinant PACAP also significantly increased endothelial tube formation and inhibited apoptosis in aged CMVECs. In young CMVECs shRNA knockdown of autocrine PACAP expression significantly impaired tube formation capacity, mimicking the aging phenotype. Cellular and mitochondrial reactive oxygen species production (dihydroethidium and MitoSox fluorescence, respectively) were increased in aged CMVECs and were unaffected by PACAP. Collectively, PACAP exerts proangiogenic effects and age-related dysregulation of autocrine PACAP signaling may contribute to impaired angiogenic capacity of CMVECs in aging.

Objective: Pituitary adenylate cyclase activating polypeptide (PACAP) is a pleiotropic and multifunctional neuropeptide widely distributed throughout the body. It is involved in the regulation of various physiological and pathophysiological processes, such as reproduction, thermoregulation, motor activity, brain development, neuronal survival, inflammation and pain. Since little is known about its distribution in humans, our aim was to examine PACAP-38 in human plasma. Furthermore, based on the presence of vasoactive intestinal peptide, structurally the closest to PACAP, in milk and PACAP and its receptors in the mammary gland, our aim was to study PACAP-38 in human milk. Design and methods: The presence of PACAP-38 was determined by mass spectrometry in plasma samples from healthy male and female volunteers (age: 20-40), as well as in plasma and milk samples from lactating women (age: 20-35). PACAP concentration was measured with a specific and sensitive RIA. Results: Our results revealed that PACAP-38 is present in human plasma, its concentration is relatively stable in healthy volunteers and it is not significantly altered by gender, age, food intake or hormonal cycle in females. However, PACAP-38 plasma levels significantly increased in lactating women having 1-6 month-old babies. Moreover, this study is the first which provides evidence for the presence of PACAP-38 in the human milk with levels 5-20-fold greater in the milk whey than in the respective plasma samples. Conclusions: We found PACAP-38 in human plasma and its increase during the first 6 months of the lactation period. A prominent, nearly 10-fold higher concentration of this peptide was detected in human milk. Based on the literature, several important actions of milk-derived PACAP-38 can be suggested such as mammary gland proliferation, nutrient transfer as well as regulation of growth/differentiation of certain tissues of the neonates. The novelty of the present descriptive data provides a basis for further investigations on the mechanism of PACAP-38 secretion in human milk and its functional significance.

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Pituitary adenylate cyclase-activating polypeptide (PACAP) is a neuropeptide with widespread occurrence in the nervous system and peripheral organs, including the mammary gland. Previously, we have shown that PACAP38 is present in the human milk at higher levels than in respective blood samples. However, it is not known how PACAP levels and the expression of PAC1 receptor change during lactation. Therefore, the aim of our study was to investigate PACAP38-like immunoreactivity (PACAP38-LI) in human colostrums and transitional and mature milk during lactation and to compare the expression of PAC1 receptors in lactating and non-lactating mammary glands. We found that PACAP38-LI was significantly higher in human colostrum samples than in the transitional and mature milk. PACAP38-LI did not show any significant changes within the first 10-month period of lactation, but a significant increase was observed thereafter, up to the examined 17th month. Weak expression of PAC1 receptors was detected in non-lactating sheep and human mammary glands, but a significant increase was observed in the lactating sheep samples. In summary, the present study is the first to show changes of PACAP levels in human milk during lactation. The presence of PACAP in the milk suggests a potential role in the development of newborn, while the increased expressions of PAC1 receptors on lactating breast may indicate a PACAP38/PAC1 interaction in the mammary gland during lactation.