Presence of atrial natriuretic factor in normal and hyperplastic human prostate and its relationship with oxytocin localisation

In this work, we showed the presence of atrial natriuretic factor (ANF) in human prostate and compared its localisation in normal and hyperplastic conditions. ANF was localised in epithelial and stromal cells, being increased in hyperplasia, mainly in the stromal component. Moreover, we compared ANF and oxytocin positivity in the same glands, focusing on the possible relationship between the paracrine effects of these two hormones

Mitochondrial activity of human umbilical cord mesenchymal stem cells

Human umbilical cord mesenchymal stem cells (hUC-MSCs) serve as a potential cell-based therapy for degenerative disease. They provide immunomodulatory and anti-inflammatory properties, multipotent differentiation potential and are harvested with no ethical concern. It is unknown whether MSCs collected from different areas of the human umbilical cord elicit more favorable effects than others. Three MSC populations were harvested from various regions of the human umbilical cord: cord lining (CL-MSCs), perivascular region (PV-MSCs), and Wharton's jelly (WJ-MSCs). Mesenchymal markers (CD90 and CD73) were expressed by all three cell populations. Stemness marker (OCT4), endothelial cell adhesion molecular marker (CD146), and monocyte-macrophage marker (CD14) were expressed by WJ-MSCs, PV-MSCs, and CL-MSCs, respectively. Stroke presents with oxygen and glucose deprivation and leads to dysfunctional mitochondria and consequently cell death. Targeting the restoration of mitochondrial function in the stroke brain through mitochondrial transfer may be effective in treating stroke. In vitro exposure to ambient and OGD conditions resulted in CL-MSCs number decreasing the least post-OGD/R exposure, and PV-MSCs exhibiting the greatest mitochondrial activity. All three hUC-MSC populations presented similar metabolic activity and survival in normal and pathologic environments. These characteristics indicate hUC-MSCs potential as a potent therapeutic in regenerative medicine

The value of immunohistochemical research on PCNA, p53 and heat shock proteins in prostate cancer management: a review.

This review addresses the significance of the expression of proliferating cell nuclear antigen (PCNA), p53 and some heat shock proteins (Hsps) in prostate carcinoma (PC). In fact, PCNA and p53 are two widely discussed tools in PC diagno- sis, mainly because of the controversy regarding the signifi- cance of their expression during prostate cancer development and progression. At the same time, only few studies have shown the potential role of Hsps in carcinogenesis and their overexpression in pre-neoplastic and neoplastic lesions of the prostate. We briefly describe the physiological roles of Hsps in normal cells, and the significance of their immunohistochem- ical detection in PC as well as in pre-cancerous lesions of the prostate. We will also discuss the possible functional interac- tions of these molecules in both dysplastic and neoplastic cell

Helicobacter pylori and Epstein–Barr Virus Co-Infection in Gastric Disease: What Is the Correlation with p53 Mutation, Genes Methylation and Microsatellite Instability in a Cohort of Sicilian Population?

Genetic predisposition, environmental factors, and infectious agents interact in the development of gastric diseases. Helicobacter pylori (Hp) and Epstein–Barr virus (EBV) infection has recently been shown to be correlated with these diseases. A cross-sectional study was performed on 100 hospitalized Italian patients with and without gastric diseases. The patients were stratified into four groups. Significant methylation status differences among CDH1, DAPK, COX2, hMLH1 and CDKN2A were observed for coinfected (Hp-EBV group) patients; particularly, a significant presence of COX2 (p = 0.0179) was observed. For microsatellite instability, minor stability was described in the Hp-HBV group (69.23%, p = 0.0456). Finally, for p53 mutation in the EBV group, exon 6 was, significantly, most frequent in comparison to others (p = 0.0124), and in the Hp-EBV group exon 8 was, significantly, most frequent in comparison to others (p < 0.0001). A significant positive relationship was found between patients with infection (Hp, EBV or both) and p53 mutation (rho = 0.383, p = 0.0001), methylation status (rho = 0.432, p < 0.0001) and microsatellite instability (rho = 0.285, p = 0.004). Finally, we observed among infection and methylation status, microsatellite instability, and p53 mutation a significant positive relationship only between infection and methylation status (OR = 3.78, p = 0.0075) and infection and p53 mutation (OR = 6.21, p = 0.0082). According to our analysis, gastric disease in the Sicilian population has different pathways depending on the presence of various factors, including infectious agents such as Hp and EBV and genetic factors of the subject

Human Wharton’s jelly-derived mesenchymal stem cells express several immunomodulatory molecules both in their naïve state and hepatocyte-like differentiated progeny: prospects for their use in liver diseases.

Wharton’s jelly (WJ), the main constituent of umbilical cord, is a reliable source of mesenchymal stem cells (MSC). WJ-MSC show unique ability in crossing lineage borders. As other extraembryonic mesenchymal populations (placenta and amnionderived cells), WJ-MSC express several immunomodulatory molecules, essential during the initial phases of human development. Indeed, our recent work pointed out the expression of non-classical HLA molecules as HLA-G in such cells, together with a favorable combination of B7 costimulators. Very few data in literature suggest that some of the immune features of the naïve cells are maintained after performing differentiation. The aim of this work was extending the knowledge on the expression of immunomodulatory molecules by naïve and differentiated WJ-MSC. To this purpose, WJMSC underwent differentiation to osteoblasts, adipocytes and hepatocyte-like cells. Differentiated cells were characterized, by both RT-PCR, ICC and histological stains for the acquisistion of the desired phenotypical features. RT-PCR and ICC were used to investigate the differential expression of immune-related molecules in control and differentiated cells. WJ-MSC resulted expressing diverse immunomodulatory molecules which spans from non-classical type I HLAs (i.e. HLA-E, -F, -G) , to further members of the B7 family, and of the CEA superfamily, for all of which in vivo immunomodulating functions are known. In addition, we demonstrated for the first time that the expression of these molecules is maintained after performing osteogenic, adipogenic or hepatogenic differentiation. Further experiments are undergoing to better evaluating the implications of these findings in the evolving field of liver regenerative medicine

Novel in vitro and in vivo data on the cellular localization of Hsp10 in smokers affected by COPD and in lung-derived cell lines exposed to cigarette smoke extract as stressor

Cigarette smoke is a potent stressor for the respiratory system, contributing to pathogenesis, for instance in chronic obstructive pulmonary disease (COPD), but its effects on the expression, function, and cellular localization of mitochondrial chaperonins are still largely unknown. We studied in vivo (airways biopsies) the localization of Hsp10 and Hsp60 in patients (smokers and non-smokers) affected by mild-moderate COPD, and characterized the effects of non-lethal doses of cigarette smoke extract (CSE) on the expression of these molecules in two human cell lines: lung fibroblasts (HFL-1) and bronchial epithelial (16HBE). We applied various in vitro methods: immunohistochemistry (IHC), subcellular fractionation analyses (SFA), Western blotting (WB), immunocytochemistry (ICC), and transmission electron microscopy (TEM) immunogold, and used bioinformatics and databases searches to gather structural in silico data for interpreting and complementing the in vitro results. IHC showed that in smokers and non-smokers COPD patients Hsp10 was localized in both, the cytoplasm and the nucleus of epithelial and lamina propria cells, while Hsp60 was present only in the cytosol. ICC, SFA, and WB on both CSE-exposed cell lines confirmed the presence of nuclear Hsp10, with an increasing trend in parallel to CSE concentration. TEM immunogold further confirmed Hsp10 in the nucleus, in addition to its presence in the cytoplasm and mitochondria, on both cell lines. Bioinformatics and in silico structural analyses indicated that Hsp10 can localize in extramitochondrial sites, such as the nucleus, even if Hsp10 lacks known DNA-binding motifs or nuclear import signals in its primary sequence. Our data suggest a link between exposure to exogenous oxidative stress and cell response, involving Hsp10, which would play roles different from its canonical functions. It is known that Hsp10 can display an array of functions depending on its location: cytoplasm, mitochondria, or extracellular. Here, we show for the first time the presence of Hsp10 in the nucleus of epithelial and stromal human-lung cell lines, paralleling the observations in vivo in COPD patients, and indicating that intranuclear Hsp10 levels are affected by oxidative stress due to an exogenous stressor like cigarette-smoke. The questions now are by what mechanism Hsp10 becomes a resident of the nucleus and what are its functions there.

Nuclear localization and new isoforms detection give new insights on Hsp10 functions in normal and cigarette smoke-stressed lung cells

Heat-shock protein (Hsp)10 is the co-chaperone for Hsp60 inside mitochondria, but it also resides outside the organelle. Variations in its levels and intracellular dis- tribution have been documented in pathological conditions, e.g. cancer and chronic obstructive pulmonary disease (COPD). Cigarette smoke (CS) is a potent stressor for the respiratory system, but its effects on the expression, function, and cellular locali- zation of mitochondrial chaperonins are still largely unknown. We studied in vivo (airways biopsies) the localization of Hsp10 and Hsp60 in patients (smokers and non-smokers) affected by mild-moderate COPD, and charac- terized the effects of non-lethal doses of CS extract (CSE) on the expression of these molecules in two human cell lines: lung fibroblasts (HFL-1) and bronchial epithelial cells (16HBE). We applied various in vitro methods: IHC, subcellular fractionation analyses (SFA), western blotting (WB), ICC, transmission electron microscopy (TEM) immunogold, chromati protein extracts (CPE), as well as 2D-gel based proteomics analyses. Bioinformatics was used to gather structural in silico data. IHC showed that Hsp10 occurred in nuclei of epithelial and lamina propria cells of bronchial mucosa from non-smokers and smokers. ICC, SFA, and WB showed that 16HBE and HFL-1 cells featured nuclear Hsp10, before and after CSE exposure; TEM immunogold further confirmed this observation. Proteomics data showed that CSE stimulation did not increase the levels of Hsp10 but did elicit qualitative changes as indicated by molecular weight and isoelectric point shifts. Bioinformatics analyses indicated that Hsp10 can localize in extramitochondrial sites, such as the nucleus, even if Hsp10 lacks known DNA-binding motifs or nuclear import signals. Hsp10 nuclear levels increased after CSE stimulation in HFL-1, indicating cytosol to nucleus migration, and although Hsp10 did not bind DNA, it bound a DNA-associated protein as suggested by CPE/gel retardation experiments. Data reported here indicate that in human cells of the respiratory mucosa there are at least three different intracellular locales for Hsp10: mitochondrial, nuclear, and cyto- solic. Further experiments are en route for the definition of the mechanisms underlying the transfer of Hsp10 to the nucleus and other cellular/extracellular compartments. This work was supported by grants from University of Palermo (FFR 2012) to GLR

A multipronged approach to unveil the emerging role of Hsp60 in chronic obstructive pulmonary disease

Inflammation is a major component of chronic obstructive pulmonary disease (COPD) and its cause and mechanisms are still incompletely understood. For example, the role of heat shock proteins (Hsps), many of which are molecular chaperones, has not been explored in detail in COPD, despite the fact that these molecules are known to participate in inflammation in other diseases. It has been shown that extracellular Hsps can signal certain types of T cells, macrophages, dendritic cells, and neutrophils and, thereby, elicit inflammation and immunity. However, these phenomena have not been investigated in COPD despite: a) the increasing awareness of Hsp participation in inflammation and immunity; and b) the fact that this disease is waiting for new knowledge to benefit from effective treatment and continues to be one of the commonest and most serious illnesses in the Western countries. We developed a strategy to study Hsps in COPD involving a multipronged approach, using in vivo and in vitro methods, which would, at least in part, compensate for the limitations inherent to the analysis of human diseases. We determined the levels of six Hsps in bronchial mucosa biopsies, as well as several inflammatory markers, from patients at various stages compared to smoker and non-smoker controls by immunohistochemistry, and found significant increase of Hsp60, Hsp10, and Hsp40 in COPD but no changes for Hsp27, Hsp70 and Hsp90. We also found that the increase in Hsp60 positively correlated with number of neutrophils, and it localized in them. Hsp60 has been implicated in human inflammatory pathology; hence it was pertinent to investigate whether the chaperonin originated only in the neutrophils or also in other cells. In vitro experiments showed that in bronchial epithelial cells submitted to oxidative stress, a characteristic of COPD mucosa, Hsp60 was overexpressed and was released into the extracellular medium. Other measurements indicated that NFkB-p65 was involved in the hsp60-gene upregulation whereas HSF-1 apparently was not. All the data we obtained using a battery of complementary in vivo and in vitro methods coincided to indicate that Hsp60 plays an active role in inflammation in COPD. Hence, one can infer that the chaperonin does contribute to the etiology and/or pathogenesis of COPD and that it is pertinent to investigate this aspect of Hsp60 biology-COPD pathology with renewed intensity. The results could have a significant impact on the developing of strategies for diagnosis, determining prognosis, and treatment that should be centered on Hsp60

Anisotropy and chemical composition of ultra-high energy cosmic rays using arrival directions measured by the Pierre Auger Observatory

The Pierre Auger Collaboration has reported evidence for anisotropy in the distribution of arrival directions of the cosmic rays with energies $E>E_{th}=5.5\times 10^{19}$ eV. These show a correlation with the distribution of nearby extragalactic objects, including an apparent excess around the direction of Centaurus A. If the particles responsible for these excesses at $E>E_{th}$ are heavy nuclei with charge $Z$, the proton component of the sources should lead to excesses in the same regions at energies $E/Z$. We here report the lack of anisotropies in these directions at energies above $E_{th}/Z$ (for illustrative values of $Z=6,\ 13,\ 26$). If the anisotropies above $E_{th}$ are due to nuclei with charge $Z$, and under reasonable assumptions about the acceleration process, these observations imply stringent constraints on the allowed proton fraction at the lower energies

Convergent Sets of Data from In Vivo and In Vitro Methods Point to an Active Role of Hsp60 in Chronic Obstructive Pulmonary Disease Pathogenesis

BACKGROUND: It is increasingly clear that some heat shock proteins (Hsps) play a role in inflammation. Here, we report results showing participation of Hsp60 in the pathogenesis of chronic obstructive pulmonary diseases (COPD), as indicated by data from both in vivo and in vitro analyses. METHODS AND RESULTS: Bronchial biopsies from patients with stable COPD, smoker controls with normal lung function, and non-smoker controls were studied. We quantified by immunohistochemistry levels of Hsp10, Hsp27, Hsp40, Hsp60, Hsp70, Hsp90, and HSF-1, along with levels of inflammatory markers. Hsp10, Hsp40, and Hsp60 were increased during progression of disease. We found also a positive correlation between the number of neutrophils and Hsp60 levels. Double-immunostaining showed that Hsp60-positive neutrophils were significantly increased in COPD patients. We then investigated in vitro the effect on Hsp60 expression in bronchial epithelial cells (16HBE) caused by oxidative stress, a hallmark of COPD mucosa, which we induced with H\u2082O\u2082. This stressor determined increased levels of Hsp60 through a gene up-regulation mechanism involving NFkB-p65. Release of Hsp60 in the extracellular medium by the bronchial epithelial cells was also increased after H\u2082O\u2082 treatment in the absence of cell death. CONCLUSIONS: This is the first report clearly pointing to participation of Hsps, particularly Hsp60, in COPD pathogenesis. Hsp60 induction by NFkB-p65 and its release by epithelial cells after oxidative stress can have a role in maintaining inflammation, e.g., by stimulating neutrophils activity. The data open new scenarios that might help in designing efficacious anti-inflammatory therapies centered on Hsp60 and applicable to COP