Search

Many recent reports suggest that the calpastatin/calpain system plays a role in cellular growth and differentiation. Defects of the calpastatin/calpain system have been linked to cellular dysfunctions, apoptosis, myocardial infarct, and dystrophies. The calpastatin/calpain system has also been implicated in post-mortem tenderization of skeletal muscle through degradation of key myofibrillar and associated proteins, a process of key importance to meat quality. In the present study we investigate the presence and activity of the calpastatin/calpain system in trout muscle samples, collected at 0, 3, 18 and 28 h post-mortem, by immunohistochemistry method. Calpastatin is a specific endogenous enzyme of cytosol, modulating the ubiquitous calpains. Calpastatin was found in samples obtainedin vivoand immediately post-mortem, but its concentration declined rapidly in samples obtained 3, 18 and 28 h post-mortem. The ubiquitous m e m-calpains, which are localized on Z line proteins and activated by intracellular Ca2+ increase, showed a rapid decline within 3 h post-mortem. By contrast p94 calpain, which is specific to skeletal muscle, showed a slow decrease post-mortem which was independent of intracellular Ca2+ increase. Our results suggest that the mechanism of activation and activity of the calpastatin/calpain system in trout is similar to that described in mammals. [ABSTRACT FROM AUTHOR]

Objectives: The Cannabinoid Receptor type 2 (CB2) is involved in inflammation and immune cell modulation. In previous studies, we demonstrated the association between the CNR2 rs35761398 polymorphism and the risk for pediatric inflammatory bowel disease (IBD). In this study, we analyzed the intestinal biopsies from Crohn disease (CD) and ulcerative colitis (UC) pediatric patients at the diagnosis to evaluate the expression of CB2 and several factors associated with IBD inflammatory pathways., Methods: We enrolled five patients with CD, five with UC, and five controls (CTR). We analyzed ileum and rectum biopsies from patients of each group evaluating the expression of CB2, Toll-like receptor 4, interleukin-6, and interleukin-1β by western blot and immunofluorescence., Results: Western blot analysis showed a significant increase of CB2 in the CD ileum and in the UC rectum biopsies and an increase of TLR4 in the UC rectum. We also observed a significant over-expression of the IL-6 in UC rectum. The immunofluorescence analysis confirmed western blot data, showing also a T-lymphocytes infiltration colocalized with CB2 expression in the CD ileum and UC rectum., Conclusions: Our results show an upregulation of CB2 in pediatric IBD, which might have implications for drug discovery., Impact: The Cannabinoid Receptor type 2 (CB2) is involved in the inflammation and modulation of the immune response in pediatric inflammatory bowel disease (IBD). CB2 receptor is more expressed in the inflamed intestine of pediatric IBD patients. CB2 could be used as a potential therapeutic target to reduce IBD-related inflammatory state in childhood., (© 2022. The Author(s), under exclusive licence to the International Pediatric Research Foundation, Inc.)

Osteoarthritis is a very disabling disease that can be treated with both non-pharmacological and pharmacological approaches. In the last years, pharmaceutical-grade chondroitin sulfate (CS) and glucosamine emerged as symptomatic slow-acting molecules, effective in pain reduction and improved function in patients affected by osteoarthritis. CS is a sulfated glycosaminoglycan that is currently produced mainly by extraction from animal tissues, and it is commercialized as a pharmaceutical-grade ingredient and/or food supplement. However, public concern on animal product derivatives has prompted the search for alternative non-extractive production routes. Thus, different approaches were established to obtain animal-free natural identical CS. On the other hand, the unsulfated chondroitin, which can be obtained via biotechnological processes, demonstrated promising anti-inflammatory properties in vitro , in chondrocytes isolated from osteoarthritic patients. Therefore, the aim of this study was to explore the potential of chondroitin, with respect to the better-known CS, in an in vivo mouse model of knee osteoarthritis. Results indicate that the treatment with biotechnological chondroitin (BC), similarly to CS, significantly reduced the severity of mechanical allodynia in an MIA-induced osteoarthritic mouse model. Decreased cartilage damage and a reduction of inflammation- and pain-related biochemical markers were also observed. Overall, our data support a beneficial activity of biotechnological unsulfated chondroitin in the osteoarthritis model tested, thus suggesting BC as a potential functional ingredient in pharmaceuticals and nutraceuticals with the advantage of avoiding animal tissue extraction., Competing Interests: DC and CS are inventors but not assignees of the patent WO/2010/136435, International Patent Application PCT/EP2010/057129. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Cimini, Boccella, Alfano, Stellavato, Paino, Schiraldi, Guida, Perrone, Donniacuo, Tirino, Desiderio and Rinaldi.)

Central post-stroke pain (CPSP) and associated depression remain poorly understood and pharmacological treatments are unsatisfactory. Recently, microglia activation was suggested to be involved in CPSP pathophysiology. The goal of this study was to investigate the effectiveness of a co-ultramicronized combination of N-palmitoylethanolamide and luteolin (PEALut) in a mouse model of thalamic hemorrhage (TH)-induced CPSP. TH was established through the collagenase-IV injection in thalamic ventral-posterolateral-nucleus. PEALut effects in CPSP-associated behaviors were evaluated during a 28-days observation period. We found that repeated administrations of co-ultra PEALut significantly reduced mechanical hypersensitivity after TH, as compared to vehicle, by reducing the early microglial activation in the perilesional site. Moreover, PEALut prevented the development of depressive-like behavior (21 days post-TH). These effects were associated with the restoration of synaptic plasticity in LEC-DG pathway and monoamines levels found impaired in TH mice. Hippocampal MED1 and TrkB expressions were significantly increased in TH compared to sham mice 21 days post-TH, whereas BDNF levels were decreased. PEALut restored MED1/TrkB/BDNF expression in mice. Remarkably, we found significant overexpression of MED1 in the human autoptic brain specimens after stroke, indicating a translational potential of our findings. These results pave the way for better-investigating depression in TH- induced CPSP, together with the involvement of MED1/TrkB/BDNF pathway, proposing PEALut as an adjuvant treatment., (Copyright © 2022. Published by Elsevier Inc.)

The bioactive form of vitamin D, 1,25-dihydroxyvitamin D (1,25D3), exerts immunomodulatory actions resulting in neuroprotective effects potentially useful against neurodegenerative and autoimmune diseases. In fact, vitamin D deficiency status has been correlated with painful manifestations associated with different pathological conditions. In this study, we have investigated the effects of vitamin D deficiency on microglia cells, as they represent the main immune cells responsible for early defense at central nervous system (CNS), including chronic pain states. For this purpose, we have employed a model of low vitamin D intake during gestation to evaluate possible changes in primary microglia cells obtained from postnatal day(P)2-3 pups. Afterwards, pain measurement and microglia morphological analysis in the spinal cord level and in brain regions involved in the integration of pain perception were performed in the parents subjected to vitamin D restriction. In cultured microglia, we detected a reactive-activated and proliferative-phenotype associated with intracellular reactive oxygen species (ROS) generation. Oxidative stress was closely correlated with the extent of DNA damage and increased β-galactosidase (B-gal) activity. Interestingly, the incubation with 25D3 or 1,25D3 or palmitoylethanolamide, an endogenous ligand of peroxisome proliferator-activated-receptor-alpha (PPAR-α), reduced most of these effects. Morphological analysis of ex-vivo microglia obtained from vitamin-D-deficient adult mice revealed an increased number of activated microglia in the spinal cord, while in the brain microglia appeared in a dystrophic phenotype. Remarkably, activated (spinal) or dystrophic (brain) microglia were detected in a prominent manner in females. Our data indicate that vitamin D deficiency produces profound modifications in microglia, suggesting a possible role of these cells in the sensorial dysfunctions associated with hypovitaminosis D.

This study aimed to analyze the prospective associations during adolescence between depressive symptoms and response styles to positive affect and to examine gender differences. A longitudinal study was conducted with three waves separated by 1 year each to assess a non-clinical sample of 622 Spanish adolescents who were 13 and 14 years old (50.2% boys, 49.8% girls). The participants completed self-report measures of depressive symptoms and responses to positive affect (emotion-focused positive rumination, self-focused positive rumination and dampening of positive emotion). The results showed that the increase in depressive symptoms was associated with an increase in dampening and decreases in emotion-focused and self-focused positive rumination. Furthermore, girls presented more depressive symptoms, as well as higher dampening and lower self-focused positive rumination, than boys. The conclusions highlight the need to consider responses to positive affect in explaining gender differences in depressive symptoms during mid-adolescence, as well as in designing prevention programs., (Copyright © 2017 The Foundation for Professionals in Services for Adolescents. Published by Elsevier Ltd. All rights reserved.)

The peptides orexin A (OXA) and orexin B, deriving from the cleavage of the precursor molecule prepro-orexin, bind two G-coupled transmembrane receptors, named as receptor 1 (OX1R) and receptor 2 for orexin, showing different affinity-binding properties. First discovered in the rat hypothalamus, orexins and their receptors have been also found in many peripheral tissues where they exert neuroendocrine, autocrine and paracrine functions. Because inconclusive data on their localization in the mammalian prostate are reported, the aim of this study was to investigate the presence of prepro-orexin, OXA and OX1R in the human normal and hyperplastic gland. Immunohistochemistry revealed the localization of both OXA and OX1R in the cytoplasm of the follicular exocrine epithelium of all tested normal and hyperplastic prostates. Positive immunostaining was mainly observed in the basal cells of the stratified epithelium, and only rarely in the apical cells. The expression of mRNAs coding for prepro-orexin and OX1R and of proteins in the tissues was also ascertained by polymerase chain reaction and Western blotting analysis, respectively. In order to gain insights into the functional activity of OXA in the prostate, we administered different concentrations of OXA to cultured prostatic epithelial cells PNT1A. We first demonstrated that PNT1A cells express OX1R. The addition of OXA did not affect PNT1A cell proliferation, while it enhanced cAMP synthesis and Ca(2+) release from intracellular storage. Overall, our results definitely demonstrate the expression of OXA and OX1R in the human prostate, and suggest an active role for them in the metabolism of the gland., (© 2013 The Authors Journal of Anatomy © 2013 Anatomical Society.)

The orexins A (oxA) and B are peptides discovered in the rat hypothalamus and successively found in some peripheral organs of the mammalian body. They binds two protein G-coupled receptors defined receptor 1 (ox1r) and 2 for orexins, the first of which is highly specific for oxA while the second binds both the peptides with equal affinity. This work aimed to detect the presence of oxA and ox1r in the testis of the South American camelid alpaca (Vicugna pacos) and investigate the role played by them on Leydig cell steroidogenesis. The species alpaca acquired, in the last years, increasing zootechnical interest for the quality of the wool produced and its breeding spread from the country of origin to USA, Australia and Europe. Immunohistochemistry allowed us to detect oxA in Leydig and Sertoli cells, spermatogonia, resting spermatocytes, round and oval spermatids. Ox1r-immunoreactivity was found in Leydig cells and round, oval and elongated spermatids. The expression of the two peptides in tissue extracts was established by using Western blotting technique. Such results demonstrated that in the alpaca testis exists in a cellular complex able to produce and/or internalize oxA. Finally, the effect of oxA on steroidogenesis was investigated by means of in vitro cultured thin testis slices which were added with oxA or/and Müllerian Inhibiting Substance (MIS), a steroidolitic agent basally produced by the Sertoli cell. OxA evoked increase of testosterone production while MIS a decrease. The consecutive addition of oxA and MIS, or vice versa, highlighted an antagonistic interplay between the two substances which has been thought to be the main molecular event at the basis of the oxA-stimulated steroidogenesis mechanism., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Urocortin (UCN) is a 40-amino-acid peptide and a member of the corticotropin-releasing hormone (CRH) family, which includes CRH, urotensin I, sauvagine, UCN2 and UCN3. The biological actions of CRH family peptides are mediated via two types of G-protein-coupled receptors, namely CRH type 1 receptor (CRHR1) and CRH type 2 receptor (CRHR2). The biological effects of these peptides are mediated and modulated not only by CRH receptors but also via a highly conserved CRH-binding protein (CRHBP). Our aim was to investigate the expression of UCN, CRHR1, CRHR2 and CRHBP by immunohistochemistry, Western blot and reverse transcription with the polymerase chain reaction (RT-PCR) in the horse thyroid gland. The results showed that UCN, CRHR1 and CRHR2 were expressed in the thyroid gland, whereas CRHBP was not expressed. Specifically, UCN immunoreactivity (-IR) was found in the thyroid follicular cells, CRHR2-IR in the C-cells and CRHR1-IR in blood vessels. Western blot analysis and RT-PCR experiments confirmed the immunohistochemical data. These results suggest that a regulatory system exists in the mammalian thyroid gland based on UCN, CRHR1 and CRHR2 and that UCN plays a role in the regulation of thyroid physiological functions through a paracrine mechanism.

Orexins (OxA and OxB) and their receptors (Ox1R and Ox2R), originally detected in the hypothalamus, have also been localized in multiple cerebral areas and peripheral organs. Thus, in addition to their central function in the regulation of food intake, arterial blood pressure, heart rate, sleep/wake cycle, sexual behaviour, arousal, and hypothalamic/hypophyseal axis, these neuropeptides may exert a local action in various peripheral organs and tissues. Emerging evidence suggests a main role of OxA and its highly specific receptor Ox1R in the male genital tract of mammals. We previously demonstrated OxA localization in Sertoli cells and spermatids of rat testis. Here, we show positive stainings of Ox1R in developing spermatocytes, and spermatids of rat testis by immunohistochemistry. The expression of Ox1R mRNA and the protein in the tissue was also established by reverse-transcription polymerase chain reaction and Western blotting respectively. The addition of OxA to fresh testis slices significantly increased testosterone (T) secretion which, conversely, was inhibited by Mullerian inhibiting substance (MIS). The sequential treatment of testis samples with the two substances highlighted an antagonizing activity of OxA versus MIS in regulating T levels. Furthermore, the stimulating effect on T production by OxA was prevented by the addition of the selective Ox1R inhibitor SB-408124. Overall, our findings suggest that locally secreted OxA interacting with Ox1R activates signals which antagonize MIS action in the control of T levels in mammalian testis.

Urocortin (UCN), a 40 amino acid peptide, is a corticotrophin-releasing hormone (CRH)-related peptide. The biological actions of CRH family peptides are mediated via two types of G protein-coupled receptors, CRH type 1 receptor (CRHR1) and CRH type 2 receptor (CRHR2). The aim of the present study was to investigate the expression of UCN, CRHR1 and CRHR2 by immunohistochemistry, western blot and real-time RT-PCR in the bovine adrenal gland to clarify the mechanisms of the intra-adrenal CRH-based regulatory system. The results showed that UCN, CRHR1 and CRHR2 were expressed in both the cortex and medulla. Specifically, UCN-immunoreactivity (IR) was distributed in the outer part of the zona fasciculata and in the zona reticularis of the cortex and in the medulla. UCN and CRHR2 mRNA expression levels were higher in the cortex than in the medulla, while CRHR1 mRNA levels were undetectable in the cortex. These results suggest that UCN, CRHR1 and CRHR2 are expressed in the bovine adrenal gland and that UCN might play a role in the intra-adrenal CRH-based regulatory system through an autocrine mechanism., (Copyright © 2011 Elsevier Inc. All rights reserved.)

We measured Cd, Cr, Hg, Mn, and Pb levels in samples of fodder, milk, dairy products, and tissues collected from 12 ovine farms in the regions of Campania and Calabria (Southern Italy). The areas in which the farms are located show different levels of anthropogenic pressure. The main purpose of this study is the identification and the analysis of relationships among metal concentrations observed in samples representative of different links in the food chain. Particularly, we apply univariate, bivariate, and multivariate statistical techniques to identify the correlation structure of our data set and to evaluate the influence of anthropogenic activity. We discuss the results, focusing the analysis on the spatial and the temporal patterns of metal concentrations.