Ruolo delle heat shock proteins nella patogenesi della miastenia gravis

Myasthenia Gravis (MG) is an autoimmune disorder of the neuromuscolar junction mediated by specific antibodies to the nicotinic acetylcholine receptors. Several clinical and experimental studies have confirmed that the HSPs are involved in the process of regulation of some autoimmune disorders such as arthritis, diabetes, atherosclerosis and multiple sclerosis. The involvement of HSPs within different autoimmune disorders allows to hypothesize their presence in the pathogenesis of the MG, i.e. as suggested by the existence of a cross-reactivity among the antigenic epitopes of Hsp60 and AChR. Moreover, since a cross-reactivity between human and bacterial epitopes of Hsp60 has also been described, we would like to stress the role played by chronic and/or persistent infections, such as those from Chlamydia Trachomatis. © Capsula Eburnea, 2010

Myelin pathology: Involvement of molecular chaperones and the promise of chaperonotherapy

The process of axon myelination involves various proteins including molecular chaperones. Myelin alteration is a common feature in neurological diseases due to structural and functional abnormalities of one or more myelin proteins. Genetic proteinopathies may occur either in the presence of a normal chaperoning system, which is unable to assist the defective myelin protein in its folding and migration, or due to mutations in chaperone genes, leading to functional defects in assisting myelin maturation/migration. The latter are a subgroup of genetic chaperonopathies causing demyelination. In this brief review, we describe some paradigmatic examples pertaining to the chaperonins Hsp60 (HSPD1, or HSP60, or Cpn60) and CCT (chaperonin-containing TCP-1). Our aim is to make scientists and physicians aware of the possibility and advantages of classifying patients depending on the presence or absence of a chaperonopathy. In turn, this subclassification will allow the development of novel therapeutic strategies (chaperonotherapy) by using molecular chaperones as agents or targets for treatment

Alzheimer’s disease and molecular chaperones: Current knowledge and the future of chaperonotherapy

Background: Alzheimer’s disease (AD) is a dementia, a neurodegenerative condition, and a protein-misfolding disease or proteinopathy, characterized by protein deposits, extracellular plaques and intracellular neurofibrillary tangles, which contain the AD’s typical pathological proteins, abnormal [1]-amyloid and hyperphosphorylated tau, respectively, and are located predominantly in the cortex of the frontal, parietal, and temporal brain lobes. What is the role of molecular chaperones in AD? Data indicate that molecular chaperones, also known as Hsp, are involved in AD, probably displaying protective roles and/or acting as pathogenic factors as it occurs in chaperonopathies in which case AD would be suitable to chaperonotherapy. Hsp60, Hsp70, and Hsp90 can be augmented and overexpressed or diminished and downregulated in various situations in AD affected tissues and cells, indicating they are active during disease development and progression. Question: What is the role of molecular chaperones in AD? Data indicate that molecular chaperones, also known as Hsp, are involved in AD, probably displaying protective roles and/or acting as pathogenic factors as it occurs in chaperonopathies in which case AD would be suitable to chaperonotherapy. Objective: Investigate the role of Hsp in AD, focusing on Hsp60, Hsp70, and Hsp90. Method: Critical examination of published data. Results: Hsp60, Hsp70, and Hsp90 can be augmented and overexpressed or diminished and downregulated in various situations in AD affected tissues and cells, indicating they are active during disease development and progression. Conclusion and Perspectives: Notwithstanding that the roles and mechanisms of action of chaperones in AD are still incompletely understood, there is already enough evidence to encourage the development of therapeutic strategies targeting them, either to block their activity in case they promote disease progression or to boost their performance when they are protective. The latter is an example of positive chaperonotherapy, which also includes chaperone replacement via gene or protein administration. On the contrary, if a chaperone is found to help the disease, it has to be blocked or eliminated, which constitute modalities of negative chaperonotherapy

Curcumin affects HSP60 folding activity and levels in neuroblastoma cells

The fundamental challenge in fighting cancer is the development of protective agents able to interfere with the classical pathways of malignant transformation, such as extracellular matrix remodeling, epithelial\u2013mesenchymal transition and, alteration of protein homeostasis. In the tumors of the brain, proteotoxic stress represents one of the main triggering agents for cell transformation. Curcumin is a natural compound with anti-inflammatory and anti-cancer properties with promising potential for the development of therapeutic drugs for the treatment of cancer as well as neurodegenerative diseases. Among the mediators of cancer development, HSP60 is a key factor for the maintenance of protein homeostasis and cell survival. High HSP60 levels were correlated, in particular, with cancer development and progression, and for this reason, we investigated the ability of curcumin to affect HSP60 expression, localization, and post-translational modifications using a neuroblastoma cell line. We have also looked at the ability of curcumin to interfere with the HSP60/HSP10 folding machinery. The cells were treated with 6, 12.5, and 25 \ub5M of curcumin for 24 h, and the flow cytometry analysis showed that the compound induced apoptosis in a dose-dependent manner with a higher percentage of apoptotic cells at 25 \ub5M. This dose of curcumin-induced a decrease in HSP60 protein levels and an upregulation of HSP60 mRNA expression. Moreover, 25 \ub5M of curcumin reduced HSP60 ubiquitination and nitration, and the chaperonin levels were higher in the culture media compared with the untreated cells. Furthermore, curcumin at the same dose was able to favor HSP60 folding activity. The reduction of HSP60 levels, together with the increase in its folding activity and the secretion in the media led to the supposition that curcumin might interfere with cancer progression with a protective mechanism involving the chaperonin

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

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

EXOSOMES: CAN DOCTORS STILL IGNORE THEIR EXISTENCE?

With this invited commentary we want to draw the attention of young medical doctors, the main readers of this journal, towards the existence and importance of a group of nanovesicles released by human cells: the exosomes. These vesicles are incontinently se-creted as a mean of cell-to-cell communication. They are involved in a number of physiol-ogic processes as well as in the pathogenesis of, virtually, all human diseases. They can be isolated from all biological fluids, like blood, urine, sweat, sperm, crevicular fluid, bile, etc., and their composition in terms of proteins, RNA and lipids is different in pathology that in physiologic conditions. It is therefore possible to predict that they will become an important diagnostic and therapeutic tool in medicine

The Molecular Anatomy of Human Hsp60 and its Similarity with that of Bacterial Orthologs and Acetylcholine Receptor Reveal a Potential Pathogenetic Role of Anti-Chaperonin Immunity in Myasthenia Gravis.

Heat-shock protein 60 (Hsp60) is ubiquitous and highly conserved being present in eukaryotes and prokaryotes, including pathogens. This chaperonin, although typically a mitochondrial protein, can also be found in other intracellular sites, extracellularly, and in circulation. Thus, it can signal the immune system and participate in the development of inflammation and immune reactions. Both phenomena can be elicited by human and foreign Hsp60 (e.g., bacterial GroEL), when released into the blood by infectious agents. Consequently, all these Hsp60 proteins become part of a complex autoimmune response characterized by multiple cross reactions because of their structural similarities. In this study, we demonstrate that Hsp60 proteins from humans and two common pathogens, Chlamydia trachomatis and Chlamydia pneumoniae, share various sequence segments of potentially highly immunogenic epitopes with acetylcholine receptor α1 subunit (AChRα1). The structural data indicate that AChRα1 antibodies, implicated in the pathogenesis of myasthenia gravis, could very well be elicited and/or maintained by self- and/or bacterial Hsp60

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