Overexpression Of Hsp70 Research Articles

Small Molecule Screening Identifies HSP90 as a Modifier of RNA Foci in Myotonic Dystrophy Type 1.

Myotonic dystrophy type 1 (DM1) is a multisystemic disorder caused by a CTG triplet repeat expansion within the 3' untranslated region of the DMPK gene. Expression of the expanded allele generates RNA containing long tracts of CUG repeats (CUGexp RNA) that form hairpin structures and accumulate in nuclear RNA foci; however, the factors that control DMPK expression and the formation of CUGexp RNA foci remain largely unknown. We performed an unbiased small molecule screen in an immortalized human DM1 skeletal muscle myoblast cell line and identified HSP90 as a modifier of endogenous RNA foci. Small molecule inhibition of HSP90 leads to enhancement of RNA foci and upregulation of DMPK mRNA levels. Knockdown and overexpression of HSP90 in undifferentiated DM1 myoblasts validated the impact of HSP90 with upregulation and downregulation of DMPK mRNA, respectively. Furthermore, we identified p-STAT3 as a downstream mediator of HSP90 impacting levels of DMPK mRNA and RNA foci. Interestingly, differentiated cells exhibited an opposite effect of HSP90 inhibition displaying downregulation of DMPK mRNA through a mechanism independent of p-STAT3 involvement. This study has revealed a novel mediator for DMPK mRNA and foci regulation in DM1 cells with the potential to identify targets for future therapeutic intervention.

Treatment of Refractory Mucosal Leishmaniasis Is Associated with Parasite Overexpression of HSP70 and ATPase and Reduced Host Hydrogen Peroxide Production (Brief Report).

Mucosal leishmaniasis (ML) is a deforming type of American Tegumentary Leishmaniasis caused by Leishmania (Viannia) braziliensis that frequently does not respond to treatment. Despite its relapsing clinical course, few parasites are usually found in mucosal lesions. Host and parasite factors may be responsible for this paradox in the pathogenesis of the disease, allowing for both a low parasite burden and the inability of the host to clear and eliminate the disease. In this work, we present a clinical case of relapsing ML that was treated for 25 years without success with SbV, N-methyl glucamine, sodium stibogluconate, amphotericin B deoxycholate, gabromycin, antimonial plus thalidomide, liposomal amphotericin B, Leishvacin (a vaccine made in Brazil) and miltefosine. In a comparative analysis using nanoscale liquid chromatography coupled with tandem mass spectrometry of protein extracts of L. (V.) braziliensis promastigotes isolated from the patient and from the reference strain (MHOM/BR/94/M15176), we observed increases in ATPase and HSP70 protein levels in the parasite. We also observed an impairment in the production of hydrogen peroxide by peripheral mononuclear blood monocytes (PBMCs), as assessed by the horseradish peroxidase-dependent oxidation of phenol red. We hypothesise that these parasite molecules may be linked to the impairment of host parasiticidal responses, resulting in Leishmania persistence in ML patients.

Hsp70 Negatively Regulates Autophagy via Governing AMPK Activation, and Dual Hsp70-Autophagy Inhibition Induces Synergetic Cell Death in NSCLC Cells.

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.

HSP70 promotes pancreatic cancer cell epithelial-mesenchymal transformation and growth via the NF-κB signaling pathway.

To study the effects of HSP70 on proliferation, migration, invasion, and epithelial-mesenchymal transformation (EMT) of pancreatic cancer cells and explore its underlying mechanisms. Pancreatic cancer cell models with either reduced HSP70 or increased HSP70 expression were established. RT-qPCR and Western blot assays were used to determine mRNA and protein levels of HSP70, IKK/IκBa/NF-κB signaling pathway-related genes, and EMT markers. The CCK-8 and cell cloning assays were used to evaluate cell proliferation and cloning abilities. Transwell and wound healing assays were used to assess the invasive and migratory properties of the cells. Effects of NF-κB signaling modulation were explored using an IKK inhibitor (BAY11-7082) and an IKK overexpression vector (pCMV-IKK). Electrophoretic mobility shift assay (EMSA) and luciferase reporter assays were conducted to analyze NF-κB's promoter binding and transcriptional activities. HSP70 knockdown inhibited p-p65 nuclear translocation and reduced the expression of p-p65, p-IKKα/β, p-IκBα, N-cadherin, Vimentin, and Twist. It also decreased NF-κB's promoter binding and transcriptional activities, increased E-cadherin levels, and suppressed pancreatic cancer cell proliferation, cloning, migration, and invasion. In contrast, HSP70 overexpression led to increased expression of p-p65, p-IKKα/β, p-IκBα, N-cadherin, Vimentin, and Twist, decreased E-cadherin levels, and enhanced cell proliferation, cloning, migration, and invasion capabilities. NF-κB signaling pathway modulation reversed EMT changes induced by altered HSP70 expression levels. rhHSP70 also increased p-IKKα/β and p-IκBα protein levels. HSP70 promotes the EMT and enhances pancreatic cancer cell proliferation, migration, and invasion by activating the NF-κB pathway.

Coadministration of Quercetin and Indocyanine Green via PEGylated Phospholipid Micelles for Augmented Chem-Photothermal Combination Tumor Therapy.

A significant impediment persists in developing multicomponent nanomedicines designed to dismantle the heat shock protein (HSP)-based protective mechanism of malignant tumors during photothermal therapy. Herein, well-defined PEGylated phospholipid micelles were utilized to coencapsulate quercetin (QUE, a natural anticancer agent and potent HSP inhibitor) and indocyanine green (ICG, a photothermal agent) with the aim of achieving synchronized and synergistic drug effects. The subsequent investigations validated that the tailored micellar system effectively enhanced QUE's water solubility and augmented its cellular internalization efficiency. Intriguingly, the compositional PEGylated phospholipids induced extraordinary endoplasmic reticulum stress, thereby sensitizing the tumor cells to QUE. Furthermore, QUE played a crucial role in inhibiting the stress-induced overexpression of HSP70, thereby augmenting the photothermal efficacy of ICG. In systemic applications, the proposed nanotherapeutics exhibited preferential accumulation within tumors and exerted notable tumoricidal effects against 4T1 xenograft tumors under 808 nm near-infrared irradiation, facilitated by prominent near-infrared fluorescence imaging-guided chemo-photothermal therapy. Therefore, our strategy for fabricating multicomponent nanomedicines emerges as a coordinated platform for optimizing antitumor therapeutic efficacy and offers valuable insights for diverse therapeutic modalities.

Abstract Tu051: ROR2 Drives Right Ventricular Failure Via Proteostatic Imbalances

Background: No therapies exist to treat right ventricular failure (RVF), in part because RVF molecular drivers have been incompletely studied. We recently identified that the developmentally restricted noncanonical WNT receptor ROR2 is re-expressed in a severity-dependent manner in human RVF. Here we test in mice if, and how, the re-expression of Ror2 causes RVF. Methods/Results: We find in neonatal rat ventricular myocytes (NRVMs) that Ror2 overexpression (Ror2 OE ) both activates protein translation and reduces Hspa1a/b mRNA and Hsp70 protein, resulting in increased protein turnover, fragmented sarcomeres, intercalated disc disruption, reduced cell major axis, impairment of contractile and relaxation kinetics, and ectopy. Ror2 knockdown (Ror2 KD ) results in opposing phenotypes. Hsp70 overexpression or proteasome inhibition rescues many of the Ror2 OE phenotypes, demonstrating their causal involvement. NRVM proteomic analysis indicates that Ror2 OE activates an ERK/p90RSK/Eef2 pathway that is known to regulate translation extension. In vivo, cardiac Ror2 OE in mice causes RV dilation and dysfunction, intercalated disc disruption, reduced Hspa1b (encoding for Hsp70), proteasome activation, and reduced cardiomyocyte ubiquitin staining. The mouse pulmonary artery banding (PAB) model of RVF reveals similar Ror2 induction and proteasome activation. Ror2 KD by AAV9 in PAB reduces RV dilation and improves RV systolic and diastolic function. Finally, in cardiac samples from human RVF, we find evidence for a similar ROR2-responsive proteostatic imbalance with increased proteasome activity and ubiquitin as well as markers of increased ERK and p90RSK. Conclusions: ROR2 is induced in human and mouse RVF, and mechanistic studies in NRVMs and mice show that the induction of ROR2 causes RVF by disrupting the proteostatic balance of translation, folding, and turnover. Knockdown or suppression of ROR2 may serve as a novel RVF therapeutic target.

Chaperones Hsc70 and Hsp70 play distinct roles in the replication of bocaparvovirus minute virus of canines.

Minute virus of canines (MVC) belongs to the genus Bocaparvovirus (formerly Bocavirus) within the Parvoviridae family and causes serious respiratory and gastrointestinal symptoms in neonatal canines worldwide. A productive viral infection relies on the successful recruitment of host factors for various stages of the viral life cycle. However, little is known about the MVC-host cell interactions. In this study, we identified that two cellular proteins (Hsc70 and Hsp70) interacted with NS1 and VP2 proteins of MVC, and both two domains of Hsc70/Hsp70 were mediated for their interactions. Functional studies revealed that Hsp70 was induced by MVC infection, knockdown of Hsc70 considerably suppressed MVC replication, whereas the replication was dramatically promoted by Hsp70 knockdown. It is interesting that low amounts of overexpressed Hsp70 enhanced viral protein expression and virus production, but high amounts of Hsp70 overexpression weakened them. Upon Hsp70 overexpressing, we observed that the ubiquitination of viral proteins changed with Hsp70 overexpression, and proteasome inhibitor (MG132) restored an accumulation of viral proteins. In addition, we verified that Hsp70 family inhibitors remarkably decreased MVC replication. Overall, we identified Hsc70 and Hsp70 as interactors of MVC NS1 and VP2 proteins and were involved in MVC replication, which may provide novel targets for anti-MVC approach.

HSP70 protects PC12 cells against TBHP-induced apoptosis and oxidative stress by activating the Nrf2/HO-1 signaling pathway.

HSP70 exhibits neuroprotective, antioxidant, and anti-apoptotic properties, which are crucial in preventing spinal cord injury (SCI) induced by oxidative stress and apoptosis. In this study, we assessed the potential protective effects and underlying mechanisms of HSP70 on tert-butyl hydroperoxide (TBHP)-damaged PC12 cells in an in vitro model of SCI. To establish the model, PC12 cells were subjected to oxidative damage induced by TBHP, followed by overexpression of HSP70. Cell viability was assessed using the CCK8 kit, intracellular reactive oxygen species level was evaluated using a commercial kit, cell apoptosis was detected using the Annexin V-APC/7-ADD Apoptosis Detection Kit, and the oxidative stress level was determined using SOD and MDA assay kits. Western blot analysis was used to detect the expression levels of Bax, cleaved caspase-3, and Bcl-2 proteins. Furthermore, immunofluorescence staining and Western bolt were used to detect the expression levels of proteins associated with the Nrf2/HO-1 signaling pathway. We found that HSP70 overexpression reduced apoptosis and oxidative stress in TBHP-induced PC12 cells. Furthermore, it activated the Nrf2/HO-1 signaling pathway. In addition, the Nrf2 inhibitor ML385 attenuated the protective effects of HSP70 on TBHP-induced PC12 cells. In conclusion, HSP70 can partially alleviate TBHP-induced apoptosis and oxidative stress in PC12 cells by promoting the Nrf2/HO-1 signaling pathway.

GL-V9 synergizes with oxaliplatin of colorectal cancer via Wee1 degradation mediated by HSP90 inhibition.

GL-V9 exhibited anti-tumour effects on various types of tumours. This study aimed to verify if GL-V9 synergized with oxaliplatin in suppressing colorectal cancer (CRC) and to explore the synergistic mechanism. The synergy effect was tested by MTT assays and the mechanism was examined by comet assay, western blotting and immunohistochemistry (IHC). Xenograft model was constructed to substantiated the synergy effect and its mechanism in vivo. GL-V9 was verified to enhance the DNA damage effect of oxaliplatin, so as to synergistically suppress colon cancer cells in vitro and in vivo. In HCT-116 cells, GL-V9 accelerated the degradation of Wee1 and induced the abrogation of cell cycle arrest and mis-entry into mitosis, bypassing the DNA damage response caused by oxaliplatin. Our findings suggested that GL-V9 binding to HSP90 was responsible for the degradation of Wee1 and the vulnerability of colon cancer cells to oxaliplatin. Functionally, overexpression of either HSP90 or WEE1 annulled the synergistic effect of GL-V9 and oxaliplatin. Collectively, our findings revealed that GL-V9 synergized with oxaliplatin to suppress CRC and displayed a promising strategy to improve the efficacy of oxaliplatin.

Bioaccumulation and biological effects of hydrogenated cement particles in the marine bivalve, Mytilusgalloprovincialis

The decommissioning and normal functioning of nuclear facilities can result in the production and release of airborne particles in the environment. Aquatic biota are expected to be exposed to these particles considering that nuclear facilities are often located near water bodies. Aerosols, such as cement dust, can interact with radionuclides as well as with heavy metals, and therefore elicit not only radiological impacts but also chemical toxicity. In the present study, we aimed to determine the effects of hydrogenated cement particles (HCPs) as a first step before evaluating any radiotoxicity of tritiated cement particles in the marine mussels, Mytilus galloprovincialis. Responses at different levels of biological organisation were assessed, including clearance rate (CR), tissue specific accumulation, DNA damage and transcriptional expression of key stress related genes. Acute (5 h) and medium-term, chronic (11 d) exposures to 1000 μg L−1 HCPs showed that bioaccumulation, assessed using Cu as a proxy and determined by inductively coupled plasma mass spectrometry, was time and tissue dependent. The highest levels of Cu were found in the digestive gland (DG) after 11 d. HCP exposure caused changes in the expression of oxidative and other stress-related genes, including mt20 in DG and gst and sod in the gill after 5 h exposure, while an overexpression of hsp70 in the gill was observed after 11 d. Genotoxic effects in haemocytes were observed after 11 d of HCP exposure. Multivariate analysis indicated that oxidative stress is the most probable factor contributing to overall physiological dysfunction. Our results provide a baseline to perform further studies employing tritiated cement particles. Specifically, future work should focus on the DG since only this tissue showed significant bioaccumulation when compared to the negative control.

Capsaicin combined with stem cells improved mitochondrial dysfunction in PIG3V cells, an immortalized human vitiligo melanocyte cell line, by inhibiting the HSP70/TLR4/mTOR/FAK signaling axis.

Vitiligo is a common autoimmune skin disease. Capsaicin has been found to exert a positive effect on vitiligo treatment, and mesenchymal stem cells (MSCs) are also confirmed to be an ideal cell type. This study aimed to explore the influence of capsaicin combined with stem cells on the treatment of vitiligo and to confirm the molecular mechanism of capsaicin combined with stem cells in treating vitiligo. PIG3V cell proliferation and apoptosis were detected using CCK-8 and TUNEL assays, MitoSOX Red fluorescence staining was used to measure the mitochondrial ROS level, and JC-1 staining was used to detect the mitochondrial membrane potential. The expression of related genes and proteins was detected using RT‒qPCR and Western blotting. Coimmunoprecipitation was used to analyze the protein interactions between HSP70 and TLR4 or between TLR4 and mTOR. The results showed higher expression of HSP70 in PIG3V cells than in PIG1 cells. The overexpression of HSP70 reduced the proliferation of PIG3V cells, promoted apoptosis, and aggravated mitochondrial dysfunction and autophagy abnormalities. The expression of HSP70 could be inhibited by capsaicin combined with MSCs, which increased the levels of Tyr, Tyrp1 and DCT, promoted the proliferation of PIG3V cells, inhibited apoptosis, activated autophagy, and improved mitochondrial dysfunction. In addition, capsaicin combined with MSCs regulated the expression of TLR4 through HSP70 and subsequently affected the mTOR/FAK signaling pathway CONCLUSIONS: Capsaicin combined with MSCs inhibits TLR4 through HSP70, and the mTOR/FAK signaling pathway is inhibited to alleviate mitochondrial dysfunction and autophagy abnormalities in PIG3V cells.

Phytomediated stress modulates antioxidant status, induces overexpression of CYP6M2, Hsp70, α-esterase, and suppresses the ABC transporter in Anopheles gambiae (sensu stricto) exposed to Ocimum tenuiflorum extracts.

The incorporation of phytoactive compounds in the management of malarial vectors holds promise for the development of innovative and efficient alternatives. Nevertheless, the molecular and physiological responses that these bioactive substances induce remain underexplored. This present study investigated the toxicity of different concentrations of aqueous and methanol extracts of Ocimum tenuiflorum against larvae of Anopheles gambiae (sensu stricto) and unraveled the possible underlying molecular pathways responsible for the observed physiological effects. FTIR and GCMS analyses of phytoactive compounds in aqueous and methanol crude extracts of O. tenuiflorum showed the presence of OH stretching vibration, C = C stretching modes of aromatics and methylene rocking vibration; ring deformation mode with high levels of trans-β-ocimene, 3,7-dimethyl-1,3,6-octatriene in aqueous extract and 4-methoxy-benzaldehyde, 1,3,5-trimethyl-cyclohexane and o-cymene in methanol extract. The percentage mortality upon exposure to methanol and aqueous extracts of O. tenuiflorum were 21.1% and 26.1% at 24 h, 27.8% and 36.1% at 48 h and 36.1% and 45% at 72 h respectively. Using reverse transcription quantitative polymerase chain reaction (RT-qPCR), down-regulation of ABC transporter, overexpression of CYP6M2, Hsp70, and α-esterase, coupled with significantly increased levels of SOD, CAT, and GSH, were observed in An. gambiae (s.s.) exposed to aqueous and methanol extracts of O. tenuiflorum as compared to the control. Findings from this study have significant implications for our understanding of how An. gambiae (s.s.) larvae detoxify phytoactive compounds.

Sexual Dimorphism in Mitochondrial Metabolism and Insulin Sensitivity Is Controlled by HSP72-ERa Crosstalk

HSP72 is highly induced during exercise, and muscle expression is reduced in obese and diabetic men. Muscle-specific HSP72 overexpression protects male mice against diet-induced obesity and insulin resistance. The current study objective was to determine the mechanisms by which HSP72 controls metabolic function and to ascertain whether these mechanisms are impacted by biological sex. We interrogated the impact of biological sex and estradiol action in the context of HSP72 loss and gain of expression in mice. HSP72 is a binding partner of the mitochondrial quality control protein Parkin. HSP72-KO led to metabolic dysfunction via impaired Parkin action, by promoting the retention of enlarged, fission incompetent mitochondria with reduced respiratory capacity. In contrast, female KOs remained lean and insulin sensitive with small spherical muscle mitochondria. Muscle estrogen receptor alpha transcript and protein (Esr1/ERa) was induced in female HSP72-KO vs. wildtype (WT), and this enhanced Parkin-DRP1-induced fatty acid oxidation and underpinned improvements in exercise capacity. The phenotype of female HSP72-KO mice recapitulates that of muscle-specific ESR1 overexpression animals. Because muscle ERa expression was identical between male genotypes, KO vs. WT, we treated male KO mice with 17β-estradiol and the treatment restored mitochondrial form/function as well as insulin sensitivity. In summary, biological sex is critical for understanding the impact of HSP72 in governing metabolic health and insulin sensitivity. National Institutes of Health DK060484 and P30DK063491. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Mechanistic study of heat shock protein 60-mediated apoptosis in DF-1 cells

Heat shock proteins (HSP) are a group of highly conserved molecular chaperones found in various organisms and have been associated with tumorigenesis, tumor progression, and metastasis. However, the relationship between HSP60 and apoptosis remains elusive. The aim of this study was to explore the role and regulatory mechanisms of apoptosis in response to altered HSP60 expression. We generated DF-1 cell lines of both HSP60 overexpression and knockdown and assessed their impact on apoptosis levels using ELISA and flow cytometry analyses. Additionally, we examined the transcription and protein expression levels of apoptosis-related signaling factors using fluorescence quantitative PCR (qPCR) and Western blotting analyses. Heat shock proteins 60 overexpression led to a significant decrease in apoptosis levels in DF-1 cells, which could be attributed to the downregulation of BAX and BAK expression, the upregulation of Bcl-2, and the decreased expression of Caspase 3. Conversely, HSP60 knockdown led to a substantial increase in apoptosis levels in DF-1 cells, facilitated by the downregulation of BAX and Bcl-2 expression, and the upregulation of BAK expression, which increased Caspase 3 levels, thereby promoting apoptosis. The findings of our study provide the first evidence of the inhibitory effect of HSP60 on apoptosis in DF-1 cells. These observations have significant implications for disease progression and cancer research, with potential medical applications.

Lead induced genotoxicity and hepatotoxicity in zebrafish (Danio rerio) at environmentally relevant concentration: Nrf2-Keap1 regulated stress response and expression of biomarker genes

Genotoxic and hepatotoxic potentials of Pb at an environmentally relevant concentration (5 ppm) in zebrafish were investigated in the present study. Erythrocytic nuclear abnormality tests revealed the increased frequencies of abnormal erythrocytes after Pb exposure, indicating a strong genotoxic potential of Pb. Multiple stress-related parameters were further evaluated in liver, the major detoxifying organ. Pb caused increased production of ROS, which in turn caused severe oxidative stress. As a result, lipid peroxidation was increased, whereas reduced glutathione level and catalase activity was decreased. Alterations in liver histoarchitecture also served as evidence of Pb-induced hepatotoxicity. Pb-induced ROS stress triggered upregulation of Nrf2, Nqo1, Ho1; downregulation of Keap1, and altered mRNA expressions of Mn-sod, Cu/Zn-sod, gpx1, cyp1a, ucp2 suggesting involvement of Nrf2-Keap1-ARE signaling in cellular defence. Nrf2-keap1 is a sensitive biomarker of Pb-induced ROS stress. Overexpression of Hsp70 and other genes in hepatocytes might help cell survival under oxidative stress generation.

Role of heat shock protein 70 in silibinin-induced apoptosis in bladder cancer.

Hsp70 (heat shock protein 70) plays critical roles in cancer cell proliferation and apoptosis. Recently, accumulating evidences have demonstrated the cancer promoting effects of Hsp70 in bladder cancer. The development of novel therapeutic approaches targeting Hsp70 thus received great attention from researchers. In this study, we demonstrated that silibinin, a natural polyphenolic flavonoid isolated from the milk thistle, targeted Hsp70 by inhibiting its transcription in bladder cancer cells. We also demonstrated that knockdown of endogenous Hsp70 enhanced silibinin-induced apoptosis, while overexpression of exogenous Hsp70 could partially reverse the effects of silibinin-induced cell apoptosis. Furthermore, we found that silibinin could activate HSF1/Hsp70-regulated mitochondrial apoptotic pathway. Mechanically, silibinin inhibited the interaction between Apaf-1 and Hsp70, thus increasing the recruitment of pro caspase-9. Results from in vivo study demonstrated that silibinin suppressed the growth of bladder cancer xenografts, which was accompanied with the activation of caspase-3 and downregulation of HSF1 and Hsp70. Taken together, our data indicates that silibinin induces mitochondrial apoptosis via inhibiting HSF1/Hsp70 pathway and also suggests the therapeutic potential of silibinin in the treatment of bladder cancer.

The effect of HSP70 overexpression on the cerebral cortex development

Heat shock proteins (HSPs) make up a large family of molecular chaperones, recognized for their role in protein maturation, refolding, and degradation. HSP70 was shown to promote cell survival during several pathological processes in the brain, such as stroke, neurodegenerative diseases, epilepsy, and trauma [1]. In addition, HSPs serve to promote the proper embryonic and postnatal development of many organ systems, such as the central nervous system [2]. Heat shock proteins demonstrate specific expression patterns throughout the development of the nervous system, notably during crucial embryonic and postnatal moments [1]. During embryonic development, neural and glial progenitors must survive within a hypoxic microenvironment while performing energetically demanding actions, such as cell migration and neurite outgrowth. HSPs can activate or inhibit development pathways in the nervous system that modulate cell differentiation, neurite outgrowth, cell migration, or angiogenesis [1]. Indeed, recent studies demonstrated that HSP70 directly regulates the development of the nervous system by modulating signaling cascades involved in cell growth and migration [3]. Additionally, research demonstrated [4] that introducing HSP70 from an external source significantly augments the populations of proliferating cells and differentiated neuroblasts within the mouse hippocampus. Nevertheless, some researchers [1] contend that overexpression of HSPs may negatively impact cell survival. Therefore, the precise role of these chaperones remains largely unexplored. In our study, we used in utero electroporation to introduce plasmids that controlled HSP70 overexpression into neuron progenitors of mouse embryos on the 14th day of gestation. Additionally, plasmid DNA encoding GFP was used to facilitate subsequent visualization of transformed cells. Brain samples were collected on the 18th day of gestation for immunohistochemical analysis of the sections. Confocal microscopy was used to compare the characteristics of neuronal migration in both control and HSP70 overexpression conditions. Cells that received plasmids inducing HSP70 were discovered to migrate at a lower pace in comparison to the control. Additionally, it is hypothesized that the induction of HSP70 expression could lead to neuronal malformations and impact the development of neurites. In the future, the study of cytoarchitectonics in the cortex will continue, examining the identification of electroporated cells in individual neuron populations using Satb2 and Ctip2 markers. Additionally, the differentiation of these cells will be assessed by counting those that have not exited the mitotic cycle, and the hypothesis of apoptosis induction in cells electroporated with HSP70 will be tested.

Akt Is Controlled by Bag5 through a Monoubiquitination to Polyubiquitination Switch.

The serine-threonine kinase Akt plays a fundamental role in cell survival, metabolism, proliferation, and migration. To keep these essential processes under control, Akt activity and stability must be tightly regulated; otherwise, life-threatening conditions might prevail. Although it is well understood that phosphorylation regulates Akt activity, much remains to be known about how its stability is maintained. Here, we characterize BAG5, a chaperone regulator, as a novel Akt-interactor and substrate that attenuates Akt stability together with Hsp70. BAG5 switches monoubiquitination to polyubiquitination of Akt and increases its degradation caused by Hsp90 inhibition and Hsp70 overexpression. Akt interacts with BAG5 at the linker region that joins the first and second BAG domains and phosphorylates the first BAG domain. The Akt-BAG5 complex is formed in serum-starved conditions and dissociates in response to HGF, coincident with BAG5 phosphorylation. BAG5 knockdown attenuated Akt degradation and facilitated its activation, whereas the opposite effect was caused by BAG5 overexpression. Altogether, our results indicate that Akt stability and signaling are dynamically regulated by BAG5, depending on growth factor availability.

Unveiling the role of hypoxic macrophage-derived exosomes in driving colorectal cancer progression.

The crosstalk between tumor cells and macrophages under hypoxic conditions has been acknowledged as a pivotal determinant in the progression of colorectal cancer (CRC). Previous research has underscored the significance of exosomes derived from hypoxic tumor cells in facilitating tumor progression through inducing the polarization of macrophages towards the M2-like phenotype. The precise influence of hypoxic macrophage-derived exosomes (HMDEs) on the progression of CRC has not yet been fully elucidated. The objective of this study was to investigate the role of HMDEs in the progression of CRC. We discovered that there was an elevated release of exosomes derived from macrophages in hypoxic conditions. Additionally, the hypoxia-induced macrophage-derived exosomes played a crucial role in promoting the progression of CRC. We have also demonstrated that HMDEs have the ability to induce cell cycle transition and inhibit cell apoptosis, thereby promoting the growth of CRC cells. Furthermore, the underlying molecular mechanisms of these effects have been identified. The overexpression of Hif-1α results in its direct interaction with distinct regions (-521- -516 bp and -401- -391 bp) of the Hsp90 promoter during hypoxic circumstances. This binding event led to the overexpression of Hsp90 and the subsequent elevation of Hsp90 protein levels within HMDEs. Importantly, the crucial interaction between Hsp90 and Lats1 resulted in the deactivation of Lats1 and the inhibition of Yap phosphorylation. Ultimately, this series of events lead to the deactivation of the Hippo signaling pathway. Our in vivo and in vitro studies presented compelling evidence for the crucial role of hypoxic macrophage-derived exosomal Hsp90 in promoting CRC progression through the inhibition of the Hippo signaling pathway. These findings represented a significant advancement in our comprehension of the complex interplay between macrophages and CRC cells under hypoxic conditions.

Phosphorylated Hsp27 promotes adriamycin resistance in breast cancer cells through regulating dual phosphorylation of c-Myc

Chemotherapy resistance of breast cancer cells is one of the major factors affecting patient survival rate. Heat shock protein 27 (Hsp27) is a member of the small heat shock protein family that has been reported to be associated with chemotherapy resistance in tumor cells, but the exact mechanism is not fully understood. Here, we explored the regulation of Hsp27 in adriamycin-resistant pathological conditions of breast cancer in vitro and in vivo. We found that overexpression of Hsp27 in MCF-7 breast cancer cells reversed DNA damage induced by adriamycin, and thereby reduced subsequent cell apoptosis. Non-phosphorylated Hsp27 accelerated ubiquitin-mediated degradation of c-Myc under normal physiological conditions. After stimulation with adriamycin, Hsp27 was phosphorylated and translocated from the cytoplasm into the nucleus, where phosphorylated Hsp27 upregulated c-Myc and Nijmegen breakage syndrome 1 (NBS1) protein levels thus leading to ATM activation. We further showed that phosphorylated Hsp27 promoted c-Myc nuclear import and stabilization by regulating T58/S62 phosphorylation of c-Myc through a protein phosphatase 2A (PP2A)-dependent mechanism. Collectively, the data presented in this study demonstrate that Hsp27, in its phosphorylation state, plays a critical role in adriamycin-resistant pathological conditions of breast cancer cells.