Interplay between the Chaperone System and Gut Microbiota Dysbiosis in Systemic Lupus Erythematosus Pathogenesis: Is Molecular Mimicry the Missing Link between Those Two Factors?

Systemic lupus erythematosus (SLE) is a multifactorial autoimmune disease characterized by self-immune tolerance breakdown and the production of autoantibodies, causing the deposition of immune complexes and triggering inflammation and immune-mediated damage. SLE pathogenesis involves genetic predisposition and a combination of environmental factors. Clinical manifestations are variable, making an early diagnosis challenging. Heat shock proteins (Hsps), belonging to the chaperone system, interact with the immune system, acting as pro-inflammatory factors, autoantigens, as well as immune tolerance promoters. Increased levels of some Hsps and the production of autoantibodies against them are correlated with SLE onset and progression. The production of these autoantibodies has been attributed to molecular mimicry, occurring upon viral and bacterial infections, since they are evolutionary highly conserved. Gut microbiota dysbiosis has been associated with the occurrence and severity of SLE. Numerous findings suggest that proteins and metabolites of commensal bacteria can mimic autoantigens, inducing autoimmunity, because of molecular mimicry. Here, we propose that shared epitopes between human Hsps and those of gut commensal bacteria cause the production of anti-Hsp autoantibodies that cross-react with human molecules, contributing to SLE pathogenesis. Thus, the involvement of the chaperone system, gut microbiota dysbiosis, and molecular mimicry in SLE ought to be coordinately studied.

Nanovesicular Mediation of the Gut-Brain Axis by Probiotics: Insights into Irritable Bowel Syndrome.

BACKGROUND: Dysbiosis, influenced by poor diet or stress, is associated with various systemic diseases. Probiotic supplements are recognized for stabilizing gut microbiota and alleviating gastrointestinal issues, like irritable bowel syndrome (IBS). This study focused on the tryptophan pathways, which are important for the regulation of serotonin levels, and on host physiology and behavior regulation. METHODS: Nanovesicles were isolated from the plasma of subjects with chronic diarrhea, both before and after 60 days of consuming a probiotic mix (Acronelle®, Bromatech S.r.l., Milan, Italy). These nanovesicles were assessed for the presence of Tryptophan 2,3-dioxygenase 2 (TDO 2). Furthermore, the probiotics mix, in combination with H2O2, was used to treat HT29 cells to explore its cytoprotective and anti-stress effect. RESULTS: In vivo, levels of TDO 2 in nanovesicles were enhanced in the blood after probiotic treatment, suggesting a role in the gut-brain axis. In the in vitro model, a typical H2O2-induced stress effect occurred, which the probiotics mix was able to recover, showing a cytoprotective effect. The probiotics mix treatment significantly reduced the heat shock protein 60 kDa levels and was able to preserve intestinal integrity and barrier function by restoring the expression and redistribution of tight junction proteins. Moreover, the probiotics mix increased the expression of TDO 2 and serotonin receptors. CONCLUSIONS: This study provides evidence for the gut-brain axis mediation by nanovesicles, influencing central nervous system function.

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.

NF-kB Regulation and the Chaperone System Mediate Restorative Effects of the Probiotic Lactobacillus fermentum LF31 in the Small Intestine and Cerebellum of Mice with Ethanol-Induced Damage.

Probiotics are live microorganisms that yield health benefits when consumed, generally by improving or restoring the intestinal flora (microbiota) as part of the muco-microbiotic layer of the bowel. In this work, mice were fed with ethanol alone or in combination with the probiotic Lactobacillus fermentum (L. fermentum) for 12 weeks. The modulation of the NF-κB signaling pathway with the induction of Hsp60, Hsp90, and IkB-α by the probiotic occurred in the jejunum. L. fermentum inhibited IL-6 expression and downregulated TNF-α transcription. NF-κB inactivation concurred with the restoration of the intestinal barrier, which had been damaged by ethanol, via the production of tight junction proteins, ameliorating the ethanol-induced intestinal permeability. The beneficial effect of the probiotic on the intestine was repeated for the cerebellum, in which downregulation of glial inflammation-related markers was observed in the probiotic-fed mice. The data show that L. fermentum exerted anti-inflammatory and cytoprotective effects in both the small intestine and the cerebellum, by suppressing ethanol-induced increased intestinal permeability and curbing neuroinflammation. The results also suggest that L. fermentum could be advantageous, along with the other available means, for treating intestinal diseases caused by stressors associated with inflammation and dysbiosis.

Immunomorphological Patterns of Chaperone System Components in Rare Thyroid Tumors with Promise as Biomarkers for Differential Diagnosis and Providing Clues on Molecular Mechanisms of Carcinogenesis.

Hurthle cell (HC), anaplastic (AC), and medullary (MC) carcinomas are low frequency thyroid tumors that pose several challenges for physicians and pathologists due to the scarcity of cases, information, and histopathological images, especially in the many areas around the world in which sophisticated molecular and genetic diagnostic facilities are unavailable. It is, therefore, cogent to provide tools for microscopists to achieve accurate diagnosis, such as histopathological images with reliable biomarkers, which can help them to reach a differential diagnosis. We are investigating whether components of the chaperone system (CS), such as the molecular chaperones, can be considered dependable biomarkers, whose levels and distribution inside and outside cells in the tumor tissue could present a distinctive histopathological pattern for each tumor type. Here, we report data on the chaperones Hsp27, Hsp60, and Hsp90. They presented quantitative levels and distribution patterns that were different for each tumor and differed from those of a benign thyroid pathology, goiter (BG). Therefore, the reported methodology can be beneficial when the microscopist must differentiate between HC, AC, MC, and BG.

Structural and Dynamic Disturbances Revealed by Molecular Dynamics Simulations Predict the Impact on Function of CCT5 Chaperonin Mutations Associated with Rare Severe Distal Neuropathies.

Mutations in genes encoding molecular chaperones, for instance the genes encoding the subunits of the chaperonin CCT (chaperonin containing TCP-1, also known as TRiC), are associated with rare neurodegenerative disorders. Using a classical molecular dynamics approach, we investigated the occurrence of conformational changes and differences in physicochemical properties of the CCT5 mutations His147Arg and Leu224Val associated with a sensory and a motor distal neuropathy, respectively. The apical domain of both variants was substantially but differently affected by the mutations, although these were in other domains. The distribution of hydrogen bonds and electrostatic potentials on the surface of the mutant subunits differed from the wild-type molecule. Structural and dynamic analyses, together with our previous experimental data, suggest that genetic mutations may cause different changes in the protein-binding capacity of CCT5 variants, presumably within both hetero- and/or homo-oligomeric complexes. Further investigations are necessary to elucidate the molecular pathogenic pathways of the two variants that produce the two distinct phenotypes. The data and clinical observations by us and others indicate that CCT chaperonopathies are more frequent than currently believed and should be investigated in patients with neuropathies.

From Dysbiosis to Neurodegenerative Diseases through Different Communication Pathways: An Overview.

The microbiome research field has rapidly evolved over the last few decades, becoming a major topic of scientific and public interest. The gut microbiota (GM) is the microbial population living in the gut. The GM has many functions, such as maintaining gut homeostasis and host health, providing defense against enteric pathogens, and involvement in immune system development. Several studies have shown that GM is implicated in dysbiosis and is presumed to contribute to neurodegeneration. This review focuses mainly on describing the connection between the intestinal microbiome alterations (dysbiosis) and the onset of neurodegenerative diseases to explore the mechanisms that link the GM to nervous system health, such as the gut-brain axis, as well as the mitochondrial, the adaptive humoral immunity, and the microvesicular pathways. The gut-brain communication depends on a continuous bidirectional flow of molecular signals exchanged through the neural and the systemic circulation. These pathways represent a possible new therapeutic target against neuroinflammation and neurodegeneration. Progress in this context is desperately needed, considering the severity of most neurodegenerative diseases and the current lack of effective treatments.

The Chaperone System in Breast Cancer: Roles and Therapeutic Prospects of the Molecular Chaperones Hsp27, Hsp60, Hsp70, and Hsp90.

Breast cancer (BC) is a major public health problem, with key pieces of information needed for developing preventive and curative measures still missing. For example, the participation of the chaperone system (CS) in carcinogenesis and anti-cancer responses is poorly understood, although it can be predicted to be a crucial factor in these mechanisms. The chief components of the CS are the molecular chaperones, and here we discuss four of them, Hsp27, Hsp60, Hsp70, and Hsp90, focusing on their pro-carcinogenic roles in BC and potential for developing anti-BC therapies. These chaperones can be targets of negative chaperonotherapy, namely the elimination/blocking/inhibition of the chaperone(s) functioning in favor of BC, using, for instance, Hsp inhibitors. The chaperones can also be employed in immunotherapy against BC as adjuvants, together with BC antigens. Extracellular vesicles (EVs) in BC diagnosis and management are also briefly discussed, considering their potential as easily accessible carriers of biomarkers and as shippers of anti-cancer agents amenable to manipulation and controlled delivery. The data surveyed from many laboratories reveal that, to enhance the understanding of the role of the CS in BS pathogenesis, one must consider the CS as a physiological system, encompassing diverse members throughout the body and interacting with the ubiquitin-proteasome system, the chaperone-mediated autophagy machinery, and the immune system (IS). An integrated view of the CS, including its functional partners and considering its highly dynamic nature with EVs transporting CS components to reach all the cell compartments in which they are needed, opens as yet unexplored pathways leading to carcinogenesis that are amenable to interference by anti-cancer treatments centered on CS components, such as the molecular chaperones.

The chaperone system in glioblastoma multiforme and derived cell lines: diagnostic and mechanistic implications.

BACKGROUND: Glioblastoma multiforme (GBM) is the most common and malignant primary brain tumor in adults. Novel treatments are needed to counteract the molecular mechanisms of GBM growth and drug resistance. The chaperone system (CS) members are typically cytoprotective but some, termed Hsp, can become pathogenic and participate in carcinogenesis, along with the vascular endothelial growth factor (VEGF), and we investigated them in GBM biopsies and derived cell lines. The objectives were to identify diagnostic-prognostic biomarkers and gather information for developing chaperonotherapy. METHODS: Cell lines from GBMs were established, characterized (morphology, growth characteristics, and specific markers), and stored. Chaperones and angiogenic factors [Hsp10, Hsp27, Hsp60, Hsp70, Hsp90, FLT-1 (VEGFR-1), FLK1 (KDR, VEGFR-2), and FLT-4 (VEGFR-3)] were observed in cells by immunofluorescence while the chaperones were measured in tumor tissue by immunohistochemistry. RESULTS: Four cell lines were derived from four different GBMs; the cells were spindle shaped or polygonal and grew at high rates as adherent monolayers or clusters without evidence of contact inhibition. The astrocyte-specific glial fibrillary acidic protein (GFAP); and the neuronal NSE, malignancy VIM, and proliferation PCNA, markers were determined. The cells expressed GFAP but no NSE, indicating that they were primary glioblastoma cell lines, with high levels of Hsp10, Hsp27, Hsp60, Hsp90, and Flk1; and low levels of Hsp70, Flt1, and Flt4. CONCLUSIONS: Four cell lines were established derived from four out of ten GBM tumors studied. The cell lines showed intense positivity for chaperones studied and factors connected to malignancy and the tumors showed increased levels of chaperones, making them potential diagnostic-prognostic biomarkers and targets for anti-cancer compounds.

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.

Molecular Chaperones and Thyroid Cancer.

Thyroid cancers are the most common of the endocrine system malignancies and progress must be made in the areas of differential diagnosis and treatment to improve patient management. Advances in the understanding of carcinogenic mechanisms have occurred in various fronts, including studies of the chaperone system (CS). Components of the CS are found to be quantitatively increased or decreased, and some correlations have been established between the quantitative changes and tumor type, prognosis, and response to treatment. These correlations provide the basis for identifying distinctive patterns useful in differential diagnosis and for planning experiments aiming at elucidating the role of the CS in tumorigenesis. Here, we discuss studies of the CS components in various thyroid cancers (TC). The chaperones belonging to the families of the small heat-shock proteins Hsp70 and Hsp90 and the chaperonin of Group I, Hsp60, have been quantified mostly by immunohistochemistry and Western blot in tumor and normal control tissues and in extracellular vesicles. Distinctive differences were revealed between the various thyroid tumor types. The most frequent finding was an increase in the chaperones, which can be attributed to the augmented need for chaperones the tumor cells have because of their accelerated metabolism, growth, and division rate. Thus, chaperones help the tumor cell rather than protect the patient, exemplifying chaperonopathies by mistake or collaborationism. This highlights the need for research on chaperonotherapy, namely the development of means to eliminate/inhibit pathogenic chaperones.

JNK pathway and heat shock response mediate the survival of C26 colon carcinoma bearing mice fed with the mushroom Pleurotus eryngii var. eryngii without affecting tumor growth or cachexia.

In the last few years, there has been emerging interest in developing treatments against human diseases using natural bioactive content. Here, the powder of the edible mushroom Pleurotus eryngii var. eryngii was mixed with the normal diet of mice bearing C26 colon carcinoma. Interestingly, it was evidenced by a significant increase in the survival rate of C26 tumor-bearing mice accompanied by a significant increase in Hsp90 and Hsp27 protein levels in the tumors. These data were paralleled by a decrease in Hsp60 levels. The mushroom introduced in the diet induced the inhibition of the transcription of the pro-inflammatory cytokines IL-6 and IL-1 exerting an anti-inflammatory action. The effects of the mushroom were mediated by the activation of c-Jun NH2-terminal kinases as a result of metabolic stress induced by the micronutrients introduced in the diet. In the tumors of C26 bearing mice fed with Pleurotus eryngii there was also a decreased expression of the mitotic regulator survivin and the anti-apoptotic factor Bcl-xL as well as an increase in the expression levels of Atg7, a protein that drives autophagy. In our hypothesis the interplay of these molecules favored the survival of the mice fed with the mushroom. These data are promising for the introduction of Pleurotus eryngii as a dietary supplement or as an adjuvant in anti-cancer therapy.

Sex-based differences after a single bout of exercise on PGC1α isoforms in skeletal muscle: A pilot study.

To date, there are limited and incomplete data on possible sex-based differences in fiber-types of skeletal muscle and their response to physical exercise. Adult healthy male and female mice completed a single bout of endurance exercise to examine the sex-based differences of the peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC1α), heat shock protein 60 (Hsp60), interleukin 6 (IL-6) expression, as well as the Myosin Heavy Chain (MHC) fiber-type distribution in soleus and extensor digitorum longus (EDL) muscles. Our results showed for the first time that in male soleus, a muscle rich of type IIa fibers, endurance exercise activates specifically genes involved in mitochondrial biogenesis such as PGC1 α1 isoform, Hsp60 and IL-6, whereas the expression of PGC1 α2 and α3 was significantly upregulated in EDL muscle, a fast-twitch skeletal muscle, independently from the gender. Moreover, we found that the acute response of different PGC1α isoforms was muscle and gender dependent. These findings add a new piece to the huge puzzle of muscle response to physical exercise. Given the importance of these genes in the physiological response of the muscle to exercise, we strongly believe that our data could support future research studies to personalize a specific and sex-based exercise training protocol.

The Role of Molecular Chaperones in Virus Infection and Implications for Understanding and Treating COVID-19.

The COVID-19 pandemic made imperative the search for means to end it, which requires a knowledge of the mechanisms underpinning the multiplication and spread of its cause, the coronavirus SARS-CoV-2. Many viruses use members of the hosts' chaperoning system to infect the target cells, replicate, and spread, and here we present illustrative examples. Unfortunately, the role of chaperones in the SARS-CoV-2 cycle is still poorly understood. In this review, we examine the interactions of various coronaviruses during their infectious cycle with chaperones in search of information useful for future research on SARS-CoV-2. We also call attention to the possible role of molecular mimicry in the development of autoimmunity and its widespread pathogenic impact in COVID-19 patients. Viral proteins share highly antigenic epitopes with human chaperones, eliciting anti-viral antibodies that crossreact with the chaperones. Both, the critical functions of chaperones in the infectious cycle of viruses and the possible role of these molecules in COVID-19 autoimmune phenomena, make clear that molecular chaperones are promising candidates for the development of antiviral strategies. These could consist of inhibiting-blocking those chaperones that are necessary for the infectious viral cycle, or those that act as autoantigens in the autoimmune reactions causing generalized destructive effects on human tissues.

Lipid chaperones and associated diseases: a group of chaperonopathies defining a new nosological entity with implications for medical research and practice.

Fatty acid-binding proteins (FABPs) are lipid chaperones assisting in the trafficking of long-chain fatty acids with functions in various cell compartments, including oxidation, signaling, gene-transcription regulation, and storage. The various known FABP isoforms display distinctive tissue distribution, but some are active in more than one tissue. Quantitative and/or qualitative changes of FABPs are associated with pathological conditions. Increased circulating levels of FABPs are biomarkers of disorders such as obesity, insulin resistance, cardiovascular disease, and cancer. Deregulated expression and malfunction of FABPs can result from genetic alterations or posttranslational modifications and can be pathogenic. We have assembled the disorders with abnormal FABPs as chaperonopathies in a distinct nosological entity. This entity is similar but separate from that encompassing the chaperonopathies pertaining to protein chaperones. In this review, we discuss the role of FABPs in the pathogenesis of metabolic syndrome, cancer, and neurological diseases. We highlight the opportunities for improving diagnosis and treatment that open by encompassing all these pathological conditions within of a coherent nosological group, focusing on abnormal lipid chaperones as biomarkers of disease and etiological-pathogenic factors.

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.