Imidacloprid (IMI), the most widely used worldwide neonicotinoid biocide, produces cognitive disorders after repeated and single treatment. However, little was studied about the possible mechanisms that produce this effect. Cholinergic neurotransmission regulates cognitive function. Most cholinergic neuronal bodies are present in the basal forebrain (BF), regulating memory and learning process, and their dysfunction or loss produces cognition decline. BF SN56 cholinergic wild-type or acetylcholinesterase (AChE), β-amyloid-precursor-protein (βAPP), Tau, glycogen-synthase-kinase-3-beta (GSK3β), beta-site-amyloid-precursor-protein-cleaving enzyme 1 (BACE1), and/or nuclear-factor-erythroid-2-related-factor-2 (NRF2) silenced cells were treated for 1 and 14 days with IMI (1 μM to 800 μM) with or without recombinant heat-shock-protein-70 (rHSP70), recombinant proteasome 20S (rP20S) and with or without N-acetyl-cysteine (NAC) to determine the possible mechanisms that mediate this effect. IMI treatment for 1 and 14 days altered cholinergic transmission through AChE inhibition, and triggered cell death partially through oxidative stress generation, AChE-S overexpression, HSP70 downregulation, P20S inhibition, and Aβ and Tau peptides accumulation. IMI produced oxidative stress through reactive oxygen species production and antioxidant NRF2 pathway downregulation, and induced Aβ and Tau accumulation through BACE1, GSK3β, HSP70, and P20S dysfunction. These results may assist in determining the mechanisms that produce cognitive dysfunction observed following IMI exposure and provide new therapeutic tools.

The correction of protein folding is fundamental for cellular functionality and its failure can lead to severe diseases. In this context, molecular chaperones are crucial players involved in the tricky process of assisting in protein folding, stabilization, and degradation. Chaperones, such as heat shock proteins (HSP) 90, 70, and 60, operate within complex systems, interacting with co-chaperones both to prevent protein misfolding and direct to the correct folding. Chaperone targeting drugs could represent a challenging approach for the treatment of cystic fibrosis (CF), an autosomal recessive genetic disease caused by mutations in the CFTR gene, encoding for the CFTR chloride channel. In this review, we discuss the potential role of molecular chaperones as proteostasis modulators affecting CFTR biogenesis. In particular, we focused on HSP90 and HSP70, for their key role in CFTR folding and trafficking, as well as on HSP60 for its involvement in the inflammation process.

Proteostasis mechanisms, such as proteotoxic-stress response and autophagy, are increasingly recognized for their roles in influencing various cancer hallmarks such as tumorigenesis, drug resistance, and recurrence. However, the precise mechanisms underlying their coordination remain not fully elucidated. The aim of this study is to investigate the molecular interplay between Hsp70 and autophagy in lung adenocarcinoma cells and elucidate its impact on the outcomes of anticancer therapies in vitro. For this purpose, we utilized the human lung adenocarcinoma A549 cell line and genetically modified it by knockdown of Hsp70 or HSF1, and the H1299 cell line with knockdown or overexpression of Hsp70. In addition, several treatments were employed, including treatment with Hsp70 inhibitors (VER-155008 and JG-98), HSF1 activator ML-346, or autophagy modulators (SAR405 and Rapamycin). Using immunoblotting, we found that Hsp70 negatively regulates autophagy by directly influencing AMPK activation, uncovering a novel regulatory mechanism of autophagy by Hsp70. Genetic or chemical Hsp70 overexpression was associated with the suppression of AMPK and autophagy. Conversely, the inhibition of Hsp70, genetically or chemically, resulted in the upregulation of AMPK-mediated autophagy. We further investigated whether Hsp70 suppression-mediated autophagy exhibits pro-survival- or pro-death-inducing effects via MTT test, colony formation, CellTiter-Glo 3D-Spheroid viability assay, and Annexin/PI apoptosis assay. Our results show that combined inhibition of Hsp70 and autophagy, along with cisplatin treatment, synergistically reduces tumor cell metabolic activity, growth, and viability in 2D and 3D tumor cell models. These cytotoxic effects were exerted by substantially potentiating apoptosis, while activating autophagy via rapamycin slightly rescued tumor cells from apoptosis. Therefore, our findings demonstrate that the combined inhibition of Hsp70 and autophagy represents a novel and promising therapeutic approach that may disrupt the capacity of refractory tumor cells to withstand conventional therapies in NSCLC.

Ocrelizumab (OCR) is a humanized anti-CD20 monoclonal antibody approved for both Relapsing and Primary Progressive forms of Multiple Sclerosis (MS) treatment. OCR is postulated to act via rapid B cell depletion; however, by analogy with other anti-CD20 agents, additional effects can be envisaged, such as on Protein Kinase C (PKC). Hence, this work aims to explore novel potential mechanisms of action of OCR in peripheral blood mononuclear cells from MS patients before and after 12 months of OCR treatment. We first assessed, up-stream, PKCβII and subsequently explored two down-stream pathways: hypoxia-inducible factor 1 alpha (HIF-1α)/vascular endothelial growth factor (VEGF), and human antigen R (HuR)/manganese-dependent superoxide dismutase (MnSOD) and heat shock proteins 70 (HSP70). At baseline, higher levels of PKCβII, HIF-1α, and VEGF were found in MS patients compared to healthy controls (HC); interestingly, the overexpression of this inflammatory cascade was counteracted by OCR treatment. Conversely, at baseline, the content of HuR, MnSOD, and HSP70 was significantly lower in MS patients compared to HC, while OCR administration induced the up-regulation of these neuroprotective pathways. These results enable us to disclose the dual positive action of OCR: anti-inflammatory and neuroprotective. Therefore, in addition to B cell depletion, the effect of OCR on these molecular cascades can contribute to counteracting disease progression.

Mastitis typically arises from bacterial invasion, where host cell apoptosis significantly contributes to the inflammatory response. Gram-positive bacteria predominantly utilize the virulence factor lipoteichoic acid (LTA), which frequently leads to chronic breast infections, thereby impacting dairy production and animal husbandry adversely. This study employed LTA to develop models of mastitis in cow mammary gland cells and mice. Transcriptomic analysis identified 120 mRNAs associated with endocytosis and apoptosis pathways that were enriched in the LTA-induced inflammation of the Mammary Alveolar Cells-large T antigen (MAC-T), with numerous differential proteins also concentrated in the endocytosis pathway. Notably, actin-related protein 2/3 complex subunit 3 (ARPC3), actin-related protein 2/3 complex subunit 4 (ARPC4), and the heat shock protein 70 (HSP70) are closely related. STRING analysis revealed interactions among ARPC3, ARPC4, and HSP70 with components of the apoptosis pathway. Histological and molecular biological assessments confirmed that ARPC3, ARPC4, and HSP70 were mainly localized to the cell membrane of mammary epithelial cells. ARPC3 and ARPC4 are implicated in the mechanisms of bacterial invasion and the initiation of inflammation. Compared to the control group, the expression levels of these proteins were markedly increased, alongside the significant upregulation of apoptosis-related factors. While HSP70 appears to inhibit apoptosis and alleviate inflammation, its upregulation presents novel research opportunities. In conclusion, we deduced the development mechanism of ARPC3, ARPC4, and HSP70 in breast inflammation, laying the foundation for further exploring the interaction mechanism between the actin-related protein 2/3 (ARP2/3) complex and HSP70.

One of the fundamental mechanisms developed by the host to contain the highly infectious and rapidly proliferating SARS-coronavirus is elevation of body temperature, a natural fallout of which is heat shock proteins over-expression. Here, for the first time, we demonstrate that the SARS-CoV-2 exploits the host Heat shock protein 70 (Hsp70) chaperone for its entry and propagation, and blocking it can combat the infection. SARS-CoV-2 infection as well as febrile temperature enhanced Hsp70 expression in host Vero E6 cells. Furthermore, heat shock or viral infection elevated the host cell autophagic response which is a prerequisite for viral propagation. In addition, Hsp70 protein demonstrated strong interaction with host Angiotensin-converting enzyme 2 (ACE2) as well as the receptor binding domain (RBD) of the SARS-CoV-2 Spike protein, indicating that interaction of Hsp70 with ACE2 and Spike protein may serve to protect them during febrile conditions. Suppressive and prophylactic treatment of Vero E6 cells with Hsp70 inhibitor PES, 2-(3-chlorophenyl) ethynesulfonamide (PES-Cl), abrogated viral infection more potently than the currently used drug Remdesivir. In conclusion, our study not only provides a fundamental insight into the role of host Hsp70 in SARS-CoV-2 pathogenesis, it paves the way for development of potent and irresistible anti-viral therapeutics.

The molecular chaperone heat shock proteins 70 (Hsp70) play a pivotal role in preserving cellular integrity and managing stress. This study extensively examined two Hsp70 proteins, On-Hsp70cBi, inducible, and On-Hsp70cBc, constitutively expressed, in Nile tilapia (Oreochromis niloticus) utilizing in silico analysis, homology modeling, and functional annotation. Employing the SWISS-MODEL program for homology modeling, the proposed models underwent thorough reliability assessment via ProSA, Verify 3D, PROVE, ERRAT, and Ramachandran plot analyses. Key features of On-Hsp70cBi and On-Hsp70cBc included amino acid lengths (640 and 645) and molecular weights (70,233.48 and 70,773.17 Da). Moreover, theoretical isoelectric points (pI = 5.63 and 5.28), indicated their acidic nature. Counts of negatively and positively charged residues (95 and 86; 95 and 81) revealed neutrality, while instability index (II) values of 35.27 (On-Hsp70cBi) and 38.85 (On-Hsp70cBc) suggested stability. Aliphatic index (AI) values were notably high for both proteins (84.58 and 82.85), indicating stability across a broad temperature range. Domain architecture analysis showed both proteins to contain an MreB/Mbl domain. Protein-protein interaction analysis identified the co-chaperone Stip1 as a primary functional partner. Comparative modeling yielded highly reliable 3D models, showcasing structural similarity to known proteins and predicted binding sites. Additionally, both proteins are primarily localized in the cytoplasm. Functional analysis predicted an AMP-PNP binding site for On-Hsp70cBi and an ATP binding site for On-Hsp70cBc. These findings deepened our understanding of Hsp70cBc and Hsp70cBi in Nile tilapia, underscoring their significance in fish physiology and warranting further investigation, thus advancing our knowledge of these proteins\' roles in cellular processes and stress responses, potentially impacting fish health and resilience.

OBJECTIVE: Tendinopathies are diseases that often entail long-term treatment consisting of analgesics, physiotherapy, orthotics, and sparing. The aim of this study was to investigate the effect of a single application of a high-energy PEMF (pulsed electromagnetic field) on pain perception and blood born inflammation parameters.
METHODS: 34 patients were randomly assigned to a verum group (10 min PEMF, 0,78 T) or a placebo group (10 min sham condition). Prior to and up to one week after the patient blinded treatment (t1-t5), local pain state was assessed by means of algometry as pain pressure threshold (PPT). Accordingly, heat-shock protein 70 (HSP70) levels were analysed. Statistical analyses included 2‑way ANOVA (2 × 5). The clinical trial was registered (DRKS00031321).
RESULTS: After randomization and drop-out (verum n = 17, placebo n = 13) baseline-analyses did not reveal significant between-group differences for PPT (p = 0,096), for HSP70 (p = 0,524), or any other sample characteristics (p > 0,05). Pain reduction during one week of observation showed to be significantly higher (p = 0,045, η2 = 0,013) for the PEMF group (PPT: +83 bis +139%) compared to the placebo group (PPT: +10 bis +36%). There were no HSP70 associated effects.
CONCLUSIONS: A single bout of high energy PEMF led to an immediate pain relief in tendinopathy patients lasting at least for one week, but the hypothesized underlying HSP70 associated inflammatory pathway could not be confirmed.
UNASSIGNED: ZIELSETZUNG: Tendinopathien sind Erkrankungen, die häufig eine langfristige Behandlung mit Analgetika, Physiotherapie, Orthesen und Schonung erfordern. Ziel dieser Studie war es, die Wirkung einer einmaligen Anwendung von hochenergetischem PEMF (pulsed electromagnetic field) auf das Schmerzempfinden und unspezifische Entzündungsparameter zu untersuchen.
METHODS: 34 Patienten wurden nach dem Zufallsprinzip einer Verumgruppe (10 min PEMF; 0,78 T) oder einer Placebogruppe (10 min Scheinbehandlung) zugeteilt. Vor und bis zu einer Woche nach der patientenverblindeten Behandlung (t1–t5) wurde der lokale Schmerzzustand (Algometrie) als Druckschmerzschwelle (PPT, pain pressure threshold) ermittelt. Gleichzeitig wurden Hitzeschockprotein 70 (HSP70) Blutkonzentrationen analysiert. Das zweifaktorielle Datenmodell wurde varianzanalytisch ausgewertet (2-way ANOVA). Die Studie wurde als „clinical trial“ registriert (DRKS00031321).
UNASSIGNED: Nach Randomisierung und Drop-out (Verum n = 17, Placebo n = 13) ergaben Baseline-Analysen keine signifikanten Gruppenunterschiede für PPT (p = 0,096) oder HSP70 (p = 0,524), oder in Stichprobenmerkmalen (p > 0,05). Für die PEMF Gruppe zeigte sich ein signifikant stärkerer Anstieg (p = 0,045, η2 = 0,013) der Druckschmerzschwelle (PPT: +83 bis + 139 %) als für die Placebo Gruppe (PPT: +10 bis + 36 %). Für HSP70 zeigten sich keine assoziierten Effekte.
UNASSIGNED: Eine einmalige Anwendung von hochenergetischem PEMF führte zu einer sofortigen Placebo-Effekt bereinigten Schmerzlinderung über eine Woche bei Tendinopathie-Patienten, aber der vermutete zugrunde liegende HSP70-assoziierte Entzündungsweg konnte nicht bestätigt werden.

Cells exposed to stressors of various origin activate protective mechanisms that include the expression of heat shock proteins (Hsps)/molecular chaperones belonging to several families. Well-characterized inducible Hsp70 is present in all human cell-types and biological fluids, including blood, urine, and saliva. The presence of anti-Hsp70 autoantibodies in the serum of healthy individuals has already been confirmed, and their elevated titers positively correlated with the severity of several pathological conditions, including coeliac disease and dermatitis herpetiformis - a cutaneous manifestation of coeliac disease. Here, using an indirect enzyme-linked immunosorbent assay, we demonstrate, for the first time, that anti-Hsp70 autoantibodies are present in the saliva and urine of healthy individuals. Although the occurrence of anti-Hsp70 autoantibodies in the biological fluids of healthy individuals is intriguing, their physiological role is currently unknown. It is believed that antibodies reacting with self-molecules present in the serum of healthy individuals are part of natural autoantibody pool with multiple regulatory functions. On the other hand, some autoantibodies (e.g., typical of autoimmune bullous skin diseases or systemic lupus erythematosus) may be present before the onset of the disease and serve as specific predictive biomarkers. Therefore, we would like to initiate a discussion or future research direction on the use of anti-Hsp70 autoantibodies as a potential \"biomarker\" in the diagnosis or prediction of autoimmune diseases. Our findings can be considered in biomedical research to develop noninvasive, inexpensive and easy-to-use tests. Nevertheless, large-scale comparative studies should be initiated, involving the collection and analysis of biological samples such as saliva or urine from patients suffering from autoimmune diseases or other inflammatory or neoplastic diseases, to determine whether the levels of anti-Hsp70 autoantibodies are indeed elevated and whether they correlate with the clinical picture of any disease or established biomarkers.

The association of clinical, pathological, and immunohistochemical characteristics of papillary thyroid cancer with cause-specific mortality was analyzed in a case-control study within a cohort of patients from the Altai Regional Oncology Center. According to multivariate analysis, the independent predictors of fatal outcome within 10 years after surgery in patients living in Altai region are nuclear pattern of Hsp70 expression, thyroid capsular invasion, Ki-67 expression index >7%, and patient\'s age >45 years for men and >50 years for women. The prognostic model based on these features contributes to a significant improvement in the individual prognostic performance for papillary thyroid cancer in the modeling sample. The model has high statistical significance (χ2=64.73; p<0.001) and discriminative power (AUC=0.950, prediction accuracy 88.5%).