Short-Term Exposure to Bisphenol A Does Not Impact Gonadal Cell Steroidogenesis In Vitro

: Bisphenol A (BPA) is a ubiquitous, synthetic chemical proven to induce reproductive disorders in both men and women. The available studies investigated the effects of BPA on male and female steroidogenesis following long-term exposure to the compound at relatively high environmental concentrations. However, the impact of short-term exposure to BPA on reproduction is poorly studied. We evaluated if 8 and 24 h exposure to 1 nM and 1 µM BPA perturbs luteinizing hormone/choriogonadotropin (LH/hCG)-mediated signalling in two steroidogenic cell models, i.e., the mouse tumour Leydig cell line mLTC1, and human primary granulosa lutein cells (hGLC). Cell signalling studies were performed using a homogeneous time-resolved fluorescence (HTRF) assay and Western blotting, while gene expression analysis was carried out using real-time PCR. Immunostainings and an immunoassay were used for intracellular protein expression and steroidogenesis analyses, respectively. The presence of BPA leads to no significant changes in gonadotropin-induced cAMP accumulation, alongside phosphorylation of downstream molecules, such as ERK1/2, CREB and p38 MAPK, in both the cell models. BPA did not impact STARD1, CYP11A1 and CYP19A1 gene expression in hGLC, nor Stard1 and Cyp17a1 expression in mLTC1 treated with LH/hCG. Additionally, the StAR protein expression was unchanged upon exposure to BPA. Progesterone and oestradiol levels in the culture medium, measured by hGLC, as well as the testosterone and progesterone levels in the culture medium, measured by mLTC1, did not change in the presence of BPA combined with LH/hCG. These data suggest that short-term exposure to environmental concentrations of BPA does not compromise the LH/hCG-induced steroidogenic potential of either human granulosa or mouse Leydig cells

Pathophysiological Role and Medicinal Chemistry of A2A Adenosine Receptor Antagonists in Alzheimer's Disease

The A(2A) adenosine receptor is a protein belonging to a family of four GPCR adenosine receptors. It is involved in the regulation of several pathophysiological conditions in both the central nervous system and periphery. In the brain, its localization at pre- and postsynaptic level in striatum, cortex, hippocampus and its effects on glutamate release, microglia and astrocyte activation account for a crucial role in neurodegenerative diseases, including Alzheimer's disease (AD). This ailment is considered the main form of dementia and is expected to exponentially increase in coming years. The pathological tracts of AD include amyloid peptide-beta extracellular accumulation and tau hyperphosphorylation, causing neuronal cell death, cognitive deficit, and memory loss. Interestingly, in vitro and in vivo studies have demonstrated that A(2A) adenosine receptor antagonists may counteract each of these clinical signs, representing an important new strategy to fight a disease for which unfortunately only symptomatic drugs are available. This review offers a brief overview of the biological effects mediated by A(2A) adenosine receptors in AD animal and human studies and reports the state of the art of A(2A) adenosine receptor antagonists currently in clinical trials. As an original approach, it focuses on the crucial role of pharmacokinetics and ability to pass the blood-brain barrier in the discovery of new agents for treating CNS disorders. Considering that A(2A) receptor antagonist istradefylline is already commercially available for Parkinson's disease treatment, if the proof of concept of these ligands in AD is confirmed and reinforced, it will be easier to offer a new hope for AD patients

Low oxygen tension primes aortic endothelial cells to the reparative effect of tissue-protective cytokines

Erythropoietin (EPO) has both erythropoietic and tissue-protective properties. The EPO analogues carbamylated EPO (CEPO) and pyroglutamate helix B surface peptide (pHBSP) lack the erythropoietic activity of EPO but retain the tissue-protective properties that are mediated by a heterocomplex of EPO receptor (EPOR) and the β common receptor (βCR). We studied the action of EPO and its analogues in a model of wound healing where a bovine aortic endothelial cells (BAECs) monolayer was scratched and the scratch closure was assessed over 24 h under different oxygen concentrations. We related the effects of EPO and its analogues on repair to their effect on BAECs proliferation and migration (evaluated using a micro-Boyden chamber). EPO, CEPO and pHBSP enhanced scratch closure only at lower oxygen (5%), while their effect at atmospheric oxygen (21%) was not significant. The mRNA expression of EPOR was doubled in 5% compared to 21% oxygen, and this was associated with increased EPOR assessed by immunofluorescence and Western blot. By contrast βCR mRNA levels were similar in 5% and 21% oxygen. EPO and its analogues increased both BAECs proliferation and migration, suggesting that both may be involved in the reparative process. The priming effect of low oxygen tension on the action of tissue-protective cytokines may be of relevance to vascular disease, including atherogenesis and restenosis

A2A Adenosine Receptor: A Possible Therapeutic Target for Alzheimer’s Disease by Regulating NLRP3 Inflammasome Activity?

The A2A adenosine receptor, a member of the P1 purinergic receptor family, plays a crucial role in the pathophysiology of different neurodegenerative illnesses, including Alzheimer’s disease (AD). It regulates both neurons and glial cells, thus modulating synaptic transmission and neuroinflammation. AD is a complex, progressive neurological condition that is the leading cause of dementia in the world’s old population (>65 years of age). Amyloid peptide-β extracellular accumulation and neurofibrillary tangles constitute the principal etiologic tracts, resulting in apoptosis, brain shrinkage, and neuroinflammation. Interestingly, a growing body of evidence suggests a role of NLRP3 inflammasome as a target to treat neurodegenerative diseases. It represents a tripartite multiprotein complex including NLRP3, ASC, and procaspase-1. Its activation requires two steps that lead with IL-1β and IL-18 release through caspase-1 activation. NLRP3 inhibition provides neuroprotection, and in recent years adenosine, through the A2A receptor, has been reported to modulate NLRP3 functions to reduce organ damage. In this review, we describe the role of NLRP3 in AD pathogenesis, both alone and in connection to A2A receptor regulation, in order to highlight a novel approach to address treatment of AD

Calcium to phosphorous ratio (Ca/P) as helpful index to recognize primary hyperparathyroidism, but not primary hypoparathyroidism: a big-data approach.

Background Primary hyperparathyroidism (HyperPT) and primary hypoparathyroidism (HypoPT) are often underdiagnosed. Several strategies have been investigated in the past in order to identify diagnostic parameters, although the diagnosis of both HyperPT and HypoPT remains challenging so far, especially in asymptomatic patients. Calcium (Ca) and phosphorus (P) are inversely related together, thus the Ca/P ratio could be an useful tool to define these conditions. Recently, we proposed for the first time a cut-off of 3.5 for Ca/P ratio for the diagnosis of HyperPT. Aim To evaluate the diagnostic value of the Ca/P ratio for HyperPT and HypoPT through a big-data approach. Methodology A retrospective, observational, case-control study on big-data was carried out. All examinations of parathyroid hormone (PTH), Ca and P performed at the laboratory of Modena Hospital from 2010 to 2016 were consecutively included. We considered only patients between 18 and 90 years of age. Laboratory ranges of normality for both PTH and Ca were used to divide records in HyperPT, HypoPT and controls. Statistical analysis The diagnostic accuracy of Ca/P ratio was investigated using receiver operator characteristics (ROC) curves in order to define cut-off points, which show higher sensitivity and specificity for the identification of affected patients. Results 46 597 records were considered. 576 HyperPT (1.2%), 323 HypoPT (0.7%) and 45 698 controls (98.1%) were found. Ca/P ratio was significantly different among groups (P!0.001). In particular, Ca/P ratio was significantly higher in HyperPT than controls (P!0.001). For the diagnosis of HyperPT, the threshold of 3.17 for Ca/P ratio was obtained by means of the ROC curve analysis, with 85% of both sensitivity and specificity. HypoPT showed lower Ca/P ratio compared to controls (P!0.001), although no useful threshold for the diagnosis was found at ROC curve because of the low sensitivity. Conclusions We confirm the high sensitivity and specificity of Ca/P ratio for the diagnosis of HyperPT using the largest cohort of patients available so far in the literature. On the contrary, Ca/P ratio does not contribute to identify patients with HypoPT and further researches are needed to better describe this condition. In conclusion, Ca/P ratio is a simple and inexpensive diagnostic tool to recognize HyperPT

Calcium to phosphorous ratio (Ca/P) as helpful index to recognize primary hyperparathyroidism, but not primary hypoparathyroidism: a big-data approach.

BACKGROUND: Primary hyperparathyroidism (HyperPT) and primary hypoparathyroidism (HypoPT) are often underdiagnosed. Several strategies have been investigated in the past in order to identify diagnostic parameters, although the diagnosis of both HyperPT and HypoPT remains challenging so far, especially in asymptomatic patients. Calcium (Ca) and phosphorus (P) are inversely related together, thus the Ca/P ratio could be an useful tool to define these conditions. Recently, we proposed for the first time a cut-off of 3.5 for Ca/P ratio for the diagnosis of HyperPT. AIM: to evaluate the diagnostic value of the Ca/P ratio for HyperPT and HypoPT through a big-data approach. METHODOLOGY: a retrospective, observational, case-control study on big-data was carried out. All examinations of parathyroid hormone (PTH), Ca and P performed at the laboratory of Modena Hospital from 2010 to 2016 were consecutively included. We considered only patients between 18 and 90 years of age. Laboratory ranges of normality for both PTH and Ca were used to divide records in HyperPT, HypoPT and controls. Statistical analysis: The diagnostic accuracy of Ca/P ratio was investigated using receiver operator characteristics (ROC) curves in order to define cut-off points, which show higher sensitivity and specificity for the identification of affected patients. RESULTS: 46597 records were considered. 576 HyperPT (1.2%), 323 HypoPT (0.7%) and 45698 controls (98.1%) were found. Ca/P ratio was significantly different among groups (p<0.001). In particular, Ca/P ratio was significantly higher in HyperPT than controls (p<0.001). For the diagnosis of HyperPT, the threshold of 3.17 for Ca/P ratio was obtained by means of the ROC curve analysis, with 85% of both sensitivity and specificity. HypoPT showed lower Ca/P ratio compared to controls (p<0.001), although no useful threshold for the diagnosis was found at ROC curve because of the low sensitivity. CONCLUSIONS: We confirm the high sensitivity and specificity of Ca/P ratio for the diagnosis of HyperPT using the largest cohort of patients available so far in the literature. On the contrary, Ca/P ratio does not contribute to identify patients with HypoPT and further researches are needed to better describe this condition. In conclusion, Ca/P ratio is a simple and inexpensive diagnostic tool to recognize HyperPT

Pharmacology of Adenosine Receptors: Recent Advancements

Adenosine receptors (ARs) are widely acknowledged pharmacological targets yet are still underutilized in clinical practice. Their ubiquitous distribution in almost all cells and tissues of the body makes them, on the one hand, excellent candidates for numerous diseases, and on the other hand, intrinsically challenging to exploit selectively and in a site-specific manner. This review endeavors to comprehensively depict the substantial advancements witnessed in recent years concerning the development of drugs that modulate ARs. Through preclinical and clinical research, it has become evident that the modulation of ARs holds promise for the treatment of numerous diseases, including central nervous system disorders, cardiovascular and metabolic conditions, inflammatory and autoimmune diseases, and cancer. The latest studies discussed herein shed light on novel mechanisms through which ARs exert control over pathophysiological states. They also introduce new ligands and innovative strategies for receptor activation, presenting compelling evidence of efficacy along with the implicated signaling pathways. Collectively, these emerging insights underscore a promising trajectory toward harnessing the therapeutic potential of these multifaceted targets

Two human menopausal gonadotrophin (hMG) preparations display different early signaling in vitro

Commercial hMG drugs are marketed for the treatment of infertility and consist of highly purified hormones acting on receptors expressed in target gonadal cells. Menopur\uae and Meriofert\uae are combined preparation of FSH and hCG and are compared in vitro herein. To this purpose, the molecular composition of the two drugs was analyzed by immunoassay. The formation of FSH receptor and LH/hCG receptor (FSHR; LHCGR) heteromer, intracellular Ca2+ and cAMP activation, \u3b2-arrestin 2 recruitment and the synthesis of progesterone and estradiol were evaluated in transfected HEK293 and human primary granulosa lutein cells treated by drugs administered within the pg-mg/ml concentration range. Molecular characterization revealed that Meriofert\uae has a higher FSH:hCG ratio than Menopur\uae which, in turn, displays the presence of LH molecules. While both drugs induced similar FSHR-LHCGR heteromeric formations and intracellular Ca2+ increase, Meriofert\uae had a higher potency than Menopur\uae in inducing a cAMP increase. Moreover, Meriofert\uae revealed a higher potency than Menopur\uae in recruiting \u3b2-arrestin 2, likely due to different FSH content modulating the tridimensional structure of FSHR-LHCGR-\u3b2-arrestin 2 complexes, as evidenced by a decrease in bioluminescence resonance energy transfer signal. This drug-specific activation of intracellular signaling pathways is consistent with the molecular composition of these preparations and impacts downstream progesterone and estradiol production, with Menopur\uae more potent than Meriofert\uae in inducing the synthesis of both the steroids. These findings are suggestive of distinct in-vivo activities of these preparations, but require cautious interpretation and further validation from clinical studies