The Major Heat Shock Proteins, Hsp70 and Hsp90, in 2-Methoxyestradiol-Mediated Osteosarcoma Cell Death Model

2-Methoxyestradiol is one of the natural 17β-estradiol derivatives and a potential novel anticancer agent currently being under evaluation in advanced phases of clinical trials. However, the mechanism of anticancer action of 2-methoxyestradiol has not been yet fully established. In our previous studies we have demonstrated that 2-methoxyestradiol selectively induces the expression and nuclear translocation of neuronal nitric oxide synthase in osteosarcoma 143B cells. Heat shock proteins (Hsps) are factors involved in the regulation of expression and activity of nitric oxide synthases. Herein, we chose osteosarcoma cell lines differed in metastatic potential, metastatic 143B and highly metastatic MG63.2 cells, in order to further investigate the anticancer mechanism of 2-methoxyestradiol. The current study aimed to determine the role of major heat shock proteins, Hsp90 and Hsp70 in 2-methoxyestradiol-induced osteosarcoma cell death. We focused on the implication of Hsp90 and Hsp70 in control under expression of neuronal nitric oxide synthase, localization of the enzyme, and further generation of nitro-oxidative stress. To give the insight into the role of Hsp90 in regulation of anticancer efficacy of 2-methoxyestradiol, we used geldanamycin as a potent Hsp90 inhibitor. Herein, we evidenced that inhibition of Hsp90 controls the protein expression of 2-methoxyestradiol-induced neuronal nitric oxide synthase and inhibits enzyme nuclear translocation. We propose that decreased level of neuronal nitric oxide synthase protein after a combined treatment with 2-methoxyestradiol and geldanamycin is directly associated with the accompanying upregulation of Hsp70 and downregulation of Hsp90. This interaction resulted in abrogation of anticancer efficacy of 2-methoxyestradiol by geldanamycin

Does sars-cov-2 trigger stress-induced autoimmunity by molecular mimicry? A hypothesis

Viruses can generate molecular mimicry phenomena within their hosts. Why should severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) not be considered one of these? Information in this short review suggests that it might be so and, thus, encourages research aiming at testing this possibility. We propose, as a working hypothesis, that the virus induces antibodies and that some of them crossreact with host’s antigens, thus eliciting autoimmune phenomena with devasting consequences in various tissues and organs. If confirmed, by in vitro and in vivo tests, this could drive researchers to find effective treatments against the virus

Molecular mimicry in the post-COVID-19 signs and symptoms of neurovegetative disorders?

Many individuals who have severe forms of COVID-19 experience a suite of neurovegetative signs and symptoms (eg, tachycardia) after their recovery, suggesting that the imbalance of the sympathetic-parasympathetic activity of the autonomic nervous system1 could continue for many weeks or months after respiratory symptoms stop. Moreover, a reduction of the parasympathetic tone could have a role in restricting the cholinergic anti-inflammatory pathway, thus favouring hyperinflammation and cytokine storm in the most severe phases of the disease. As reported by Guglielmo Lucchese in The Lancet Microbe,2 SARS-CoV-2 can damage the nervous system via an indirect mechanism, resulting in a high prevalence of autoantibodies, mainly against unknown autoantigens in the brain, in cerebrospinal fluid from patients with neurological complications.2 The cause of low vagal tone and SARS-CoV-2 has not yet been investigated sufficiently and here we would like to share some original data supporting the putative role of molecular mimicry as the culprit of COVID-19 pathogenesis, including the post-COVID-19 neurovegetative syndrome.2, 3, 4, 5 Using methods that have been previously described,3 we looked specifically at the human proteins expressed in vagal nuclei and ganglia. As shown in the appendix (pp 1–2), we found that 22 of these proteins share peptides that could putatively generate a T-cell or B-cell driven autoimmune response. The location and function of these proteins are described in the appendix (pp 3–24). Fibres of the vagal nerve originate from four nuclei located in the medulla oblongata—ie, the dorsal motor nucleus, the nucleus ambiguus, the solitary nucleus, and, to a lesser extent, the spinal trigeminal nucleus. These fibres contribute to the somatic and visceral motricity, somatic and visceral sensibility, and the sense of taste. The visceral motor inputs originate specifically from the dorsal motor nucleus and nucleus ambiguus and are directed towards the heart, the airways, and the gastrointestinal system. Moreover, the vagal visceral innervation includes two sensory ganglia of the peripheral nervous system—the nodose ganglion and the jugular ganglion. In particular, peripheral fibres of the neurons of the nodose ganglion not only innervate the taste buds on the epiglottis, the chemoreceptors of the aortic bodies, and baroreceptors in the aortic arch, but they also provide sensory innervation to the circulatory, respiratory, and gastrointestinal systems. An impairment of the vagal innervation of the heart can lead to tachycardia at rest, which is often seen by clinicians during physical examination of patients who have recovered from a severe form of COVID-19.1 We found that the dorsal motor nucleus, nucleus ambiguus, nodose ganglion, and jugular ganglion can all host neurons presenting proteins with epitopes in common with SARS-CoV-2 proteins, and the peptide TGRLQSL is embedded in one immunoreactive linear epitope that has already been experimentally validated in the human host (Immune Epitope Database and Analysis Resource identification number 36724) to be able to generate an autoimmune response. We share our findings to prompt further studies assessing whether severe forms of COVID-19 could produce transitory or permanent damage in some vagal structure and whether this can, in turn, be responsible for the low vagal tone and the related clinical signs and symptoms

DNA strand breaks induced by nuclear hijacking of neuronal NOS as an anti-cancer effect of 2-methoxyestradiol

2-Methoxyestradiol (2-ME) is a physiological metabolite of 17β-estradiol. At pharmacological concentrations, 2-ME inhibits colon, breast and lung cancer in tumor models. Here we investigated the effect of physiologically relevant concentrations of 2-ME in osteosarcoma cell model. We demonstrated that 2-ME increased nuclear localization of neuronal nitric oxide synthase, resulting in nitro-oxidative DNA damage. This in turn caused cell cycle arrest and apoptosis in osteosarcoma cells. We suggest that 2-ME is a naturally occurring hormone with potential anti-cancer properties

Plausible role of estrogens in pathogenesis, progression and therapy of lung cancer

Malignant neoplasms are among the most common diseases and are responsible for the majority of deaths in the developed world. In contrast to men, available data show a clear upward trend in the incidence of lung cancer in women, making it almost as prevalent as breast cancer. Women might be more susceptible to the carcinogenic effect of tobacco smoke than men. Furthermore, available data indicate a much more frequent mutation of the tumor suppressor genep53 in non-small cell lung cancer (NSCLC) female patients compared to males. Another important factor, however, might lie in the female sex hormones, whose mitogenic or carcinogenic effect is well known. Epidemiologic data show a correlation between hormone replacement therapy (HRT) or oral contraceptives (OCs), and increased mortality rates due to the increased incidence of malignant tumors, including lung cancer. Interestingly, two types of estrogen receptors have been detected in lung cancer cells: ERα and ERβ. The presence of ERα has been detected in tissues and non-small-cell lung carcinoma (NSCLC) cell lines. In contrast, overexpression of ERβ is a prognostic marker in NSCLC. Herein, we summarize the current knowledge on the role of estrogens in the etiopathogenesis of lung cancer, as well as biological, hormonal and genetic sex-related differences in this neoplasm

Preferential uptake of polyunsaturated fatty acids by colorectal cancer cells

Although a growing body of evidence suggests that colorectal cancer (CRC) is associated with alterations of fatty acid (FA) profiles in serum and tumor tissues, available data about polyunsaturated fatty acid (PUFA) content in CRC patients are inconclusive. Our study showed that CRC tissues contained more PUFAs than normal large intestinal mucosa. However, serum levels of PUFAs in CRC patients were lower than in healthy controls. To explain the mechanism of PUFA alterations in CRC, we measured FA uptake by the colon cancer cells and normal colon cells. The levels of PUFAs in colon cancer cell culture medium decreased significantly with incubation time, while no changes were observed in the medium in which normal colon cells were incubated. Our findings suggest that the alterations in tumor and serum PUFA profiles result from preferential uptake of these FAs by cancer cells; indeed, PUFAs are essential for formation of cell membrane phospholipids during rapid proliferation of cancer cells. This observation puts into question potential benefits of PUFA supplementation in CRC patients

Morphological alterations and stress protein variations in lung biopsies obtained from autopsies of covid-19 subjects

Molecular chaperones, many of which are heat shock proteins, play a role in cell stress response and regulate the immune system in various ways, such as in inflammatory/autoimmune reactions. It would be interesting to study the involvement of these molecules in the damage done to COVID-19-infected lungs. In our study, we performed a histological analysis and an immunomorphological evaluation on lung samples from subjects who succumbed to COVID-19 and subjects who died from other causes. We also assessed Hsp60 and Hsp90 distribution in lung samples to determine their location and post-translational modifications. We found histological alterations that could be considered pathognomonic for COVID-19-related lung disease. Hsp60 and Hsp90 immunopositivity was significantly higher in the COVID-19 group compared to the controls, and immunolocalization was in the plasma membrane of the endothelial cells in COVID-19 subjects. The colocalization ratios for Hsp60/3-nitrotyrosine and Hsp60/acetylate-lisine were significantly increased in the COVID-19 group compared to the control group, similar to the colocalization ratio for Hsp90/acetylate-lisine. The histological and immunohistochemical findings led us to hypothesize that Hsp60 and Hsp90 might have a role in the onset of the thromboembolic phenomena that lead to death in a limited number of subjects affected by COVID-19. Further studies on a larger number of samples obtained from autopsies would allow to confirm these data as well as discover new biomarkers useful in the battle against this disease

Data mining-based statistical analysis of biological data uncovers hidden significance: clustering Hashimoto’s thyroiditis patients based on the response of their PBMC with IL-2 and IFN-γ secretion to stimulation with Hsp60

The pathogenesis of Hashimoto’s thyroiditis includes autoimmunity involving thyroid antigens, autoantibodies, and possibly cytokines. It is unclear what role plays Hsp60, but our recent data indicate that it may contribute to pathogenesis as an autoantigen. Its role in the induction of cytokine production, pro- or anti-inflammatory, was not elucidated, except that we found that peripheral blood mononucleated cells (PBMC) from patients or from healthy controls did not respond with cytokine production upon stimulation by Hsp60 in vitro with patterns that would differentiate patients from controls with statistical significance. This “negative” outcome appeared when the data were pooled and analyzed with conventional statistical methods. We re-analyzed our data with non-conventional statistical methods based on data mining using the classification and regression tree learning algorithm and clustering methodology. The results indicate that by focusing on IFN-γ and IL-2 levels before and after Hsp60 stimulation of PBMC in each patient, it is possible to differentiate patients from controls. A major general conclusion is that when trying to identify disease markers such as levels of cytokines and Hsp60, reference to standards obtained from pooled data from many patients may be misleading. The chosen biomarker, e.g., production of IFN-γ and IL-2 by PBMC upon stimulation with Hsp60, must be assessed before and after stimulation and the results compared within each patient and analyzed with conventional and data mining statistical methods

Probiotics as Potential Therapeutic Agents: Safeguarding Skeletal Muscle against Alcohol-Induced Damage through the Gut–Liver–Muscle Axis

Probiotics have shown the potential to counteract the loss of muscle mass, reduce physical fatigue, and mitigate inflammatory response following intense exercise, although the mechanisms by which they work are not very clear. The objective of this review is to describe the main harmful effects of alcohol on skeletal muscle and to provide important strategies based on the use of probiotics. The excessive consumption of alcohol is a worldwide problem and has been shown to be crucial in the progression of alcoholic liver disease (ALD), for which, to date, the only therapy available is lifestyle modification, including cessation of drinking. In ALD, alcohol contributes significantly to the loss of skeletal muscle, and also to changes in the intestinal microbiota, which are the basis for a series of problems related to the onset of sarcopenia. Some of the main effects of alcohol on the skeletal muscle are described in this review, with particular emphasis on the “gut-liver-muscle axis”, which seems to be the primary cause of a series of muscle dysfunctions related to the onset of ALD. The modulation of the intestinal microbiota through probiotics utilization has appeared to be crucial in mitigating the muscle damage induced by the high amounts of alcohol consumed

Hsp60 Post-translational Modifications: Functional and Pathological Consequences

Hsp60 is a chaperone belonging to the Chaperonins of Group I and typically functions inside mitochondria in which, together with the co-chaperonin Hsp10, maintains protein homeostasis. In addition to this canonical role, Hsp60 plays many others beyond the mitochondria, for instance in the cytosol, plasma-cell membrane, extracellular space, and body fluids. These non-canonical functions include participation in inflammation, autoimmunity, carcinogenesis, cell replication, and other cellular events in health and disease. Thus, Hsp60 is a multifaceted molecule with a wide range of cellular and tissue locations and functions, which is noteworthy because there is only one hsp60 gene. The question is by what mechanism this protein can become multifaceted. Likely, one factor contributing to this diversity is post-translational modification (PTM). The amino acid sequence of Hsp60 contains many potential phosphorylation sites, and other PTMs are possible such as O-GlcNAcylation, nitration, acetylation, S-nitrosylation, citrullination, oxidation, and ubiquitination. The effect of some of these PTMs on Hsp60 functions have been examined, for instance phosphorylation has been implicated in sperm capacitation, docking of H2B and microtubule-associated proteins, mitochondrial dysfunction, tumor invasiveness, and delay or facilitation of apoptosis. Nitration was found to affect the stability of the mitochondrial permeability transition pore, to inhibit folding ability, and to perturb insulin secretion. Hyperacetylation was associated with mitochondrial failure; S-nitrosylation has an impact on mitochondrial stability and endothelial integrity; citrullination can be pro-apoptotic; oxidation has a role in the response to cellular injury and in cell migration; and ubiquitination regulates interaction with the ubiquitin-proteasome system. Future research ought to determine which PTM causes which variations in the Hsp60 molecular properties and functions, and which of them are pathogenic, causing chaperonopathies. This is an important topic considering the number of acquired Hsp60 chaperonopathies already cataloged, many of which are serious diseases without efficacious treatment