Heat Shock Protein 70 Protein Expression Research Articles

Ultrashort wave diathermy inhibits pulmonary inflammation in mice with acute lung injury in a HSP70 independent way: a pilot study.

Acute lung injury (ALI) is a clinical syndrome characterized by pulmonary inflammation. Ultrashort wave diathermy (USWD) has been shown to be effective at in inhibiting ALI inflammation, although the underlying mechanism remains unclear. Previous studies have demonstrated that USWD generates a therapeutic thermal environment that aligns with the temperature required for heat shock protein 70 (HSP70), an endogenous protective substance. In this study, we examined the correlation between HSP70 and USWD in alleviating lung inflammation in ALI. Forty-eight male C57BL/6 mice were randomly divided into control, model, USWD intervention (LU) 1, 2, and 3, and USWD preintervention (UL) 1, 2, and 3 groups (n = 6 in each group). The mice were pretreated with LPS to induce ALI. The UL1, 2, and 3 groups received USWD treatment before LPS infusion, while the LU1, 2, and 3 groups received USWD treatment after LPS infusion. Lung function and structure, inflammatory factor levels and HSP70 protein expression levels were detected. USWD effectively improved lung structure and function, and significantly reduced IL-1β, IL-10, TGF-β1, and TNF-α levels in both the USWD preintervention and intervention groups. However, HSP70 expression did not significantly differ across the experimental groups although the expression of TLR4 was significantly decreased, suggesting that USWD may have anti-inflammatory effects through multiple signaling pathways or that the experimental conditions should be restricted. Both USWD intervention and preintervention effectively reduced the inflammatory response, alleviated lung injury symptoms, and played a protective role in LPS-pretreated ALI mice. HSP70 was potentially regulated by USWD in this process, but further studies are urgently needed to elucidate the correlation and mechanism.

Exploring variations in heat shock protein 70 expression among vulnerable populations across urban and rural areas in Klang Valley

As global temperatures rise due to climate change, the occurrence and intensity of heat waves are anticipated to increase. Despite this anticipation, there remains a gap in comprehending the underlying defense mechanisms against heat stress, particularly exhibited by Heat Shock Proteins (HSP) within vulnerable populations. Hence, this study aims to assess the expression of heat shock protein 70 (HSP70) in response to heat exposure among vulnerable populations residing in both urban and rural areas. A total of 108 respondents were recruited using multistage sampling. This study consists of three (3) methods: Part I: face-to-face questionnaire; Part II: indoor heat exposure monitoring using Universal Thermal Climate Index (UTCI) classification; and Part III: blood sample collection to determine HSP70 protein expression using HSP70 High Sensitivity Enzyme-linked Immunosorbent Assay (ELISA) kit. The results show that urban areas were exposed to strong UTCI thermal stress, whereas rural areas experienced moderate UTCI thermal stress. A higher level of HSP70 protein expression was observed in urban (2.524 ± 0.26) than in rural respondents (2.095 ± 0.27). This study concludes that both urban and rural vulnerable populations demonstrated different patterns of HSP70 protein expression in response to various levels of heat exposure.

Electroacupuncture inhibits PDK1/Akt/HCN4 pathway to improve neurogenic urinary retention in rats.

To observe the therapeutic effect of electroacupuncture (EA) on neurogenic urinary retention rats, so as to explore the underlying mechanism of EA in treating neurogenic urinary retention by focusing on 3-phosphoinositide-dependent protein kinase 1 (PDK1)/protein kinase B (Akt)/hyperpolarization activated cyclic nucleotide-gated cation channel 4 (HCN4) pathway. Female SD rats were randomly divided into sham operation, model, EA, PDK1 inhibitor, HCN4 blocker and EA + HCN4 blocker groups, with 20 rats in each group. The model of sacral spinal cord injury was established by modified Hassan Shaker spinal cord transection method. EA (2 Hz/15 Hz, 0.5 mA) was applied to "Zhongji" (CV3) and "Zhongliao" (BL33) for 20 min, once daily for 10 days. Rats of the PDK1 inhibitor group received intraperitoneal injection of OSU-03012 (20 mg/kg), and rats of the HCN4 blocker group received intraperitoneal injection of ivabradine (10 mg/kg), both once every other day for 10 days. The urodynamic indexes of rats were detected by multi-channel physiological recorder；muscle strip test was used to detect detrusor excitability；the morphological changes of bladder were observed by HE staining. Immunofluorescence double staining was used to detect the co-expression of HCN4 and C-Kit, a specific marker of interstitial cells of Cajal in bladder. Western blot was used to detect the expression of PDK1/Akt/HCN4 pathway proteins in bladder tissue and heat shock protein 27 (HSP27), a protein related to bladder contraction function. Compared with the sham operation group, the rats in the model group showed urinary dysfunction, decreased leak point pressure, isolated detrusor spontaneous contraction frequency, fluorescence intensity of C-Kit positive cells, HCN4+/C-Kit+ co-expression, HCN4 and p-HSP27/HSP27 protein expression in bladder tissue (P<0.05), and increased maximum bladder capacity and comp-liance, minimum tension during contraction of isolated detrusor, PDK1 and p-Akt/Akt protein expression in bladder tissue (P<0.05). Meanwhile, the above index were all reversed after EA and PDK1 inhibitor intervention (P<0.05). In comparison with the EA group, the rats had severe urinary dysfunction, the urine leakage point pressure, spontaneous contraction frequency, fluorescence intensity of C-Kit positive cells, the co-expression of HCN4+/C-Kit+, and the protein expression of HCN4 and p-HSP27/HSP27 were decreased (P<0.05), the maximum bladder capacity and compliance, the minimum tension during contraction of isolated detrusor, and the protein expression of PDK1 and p-Akt/Akt in bladder tissue were increased (P<0.05) in both HCN4 blocker and EA+HCN4 blocker groups. HE staining showed exfoliated bladder epithelium and disordered layers, vacuolization of bladder wall cells, with infiltration of neutrophils in mucosal and muscular layers in the model group, which were relatively milder in the EA and PDK1 inhibitor groups, but worse in the HCN4 blocker and EA + HCN4 blocker groups. EA can improve the urinary dysfunction in rats with neurogenic urinary retention, which may be related to its effect in inhibiting the activation of PDK1/Akt pathway, promo-ting HCN4-mediated detrusor excitatory contraction and urinary electrical signal activation.

HSP70 attenuates neuronal necroptosis through the HSP90α-RIPK3 pathway following neuronal trauma.

Necroptosis, a newly defined regulatable necrosis with membrane disruption, has been demonstrated to participate in trauma brain injury (TBI) related neuronal cell death. Heat shock protein 70 (HSP70) is a stress protein with neuroprotective activity, but the potential protective mechanisms are not fully understood. Here, we investigated the effects of HSP70 regulators in a cellular TBI model induced by traumatic neuronal injury (TNI) and glutamate treatment. We found that necroptosis occurred in cortical neurons after TNI and glutamate treatment. Neuronal trauma markedly upregulated HSP70 protein expression within 24h. The results of immunostaining and lactate dehydrogenase release assay showed that necroptosis following neuronal trauma was inhibited by HSP70 activator TRC051384 (TRC), but promoted by the HSP70 inhibitor 2-phenylethyenesulfonamide (PES). In congruent, the expression and phosphorylation of receptor interacting protein kinase 3 (RIPK3) and mixed lineage kinase domain-like protein (MLKL) were differently regulated by HSP70. Furthermore, the expression of HSP90α induced by neuronal trauma was further promoted by PES but decreased by TRC. The data obtained from western blot showed that the phosphorylation of RIPK3 and MLKL induced by HSP70 inhibition were reduced by RIPK3 inhibitor GSK-872 and HSP90α inhibitor geldanamycin (GA). Similarly, inhibition of HSP90α with GA could partially prevented the increased necroptosis induced by PES. Taken together, HSP70 activation exerted protective effects against neuronal trauma via inhibition of necroptosis. Mechanistically, the HSP90α-mediated activation of RIPK3 and MLKL is involved in these effects.

Expression of HSP27 and BAX/BCL-2 apoptosis factor in silicosis rat model of fibrosis

To study the expression of heat shock protein 27(HSP27), BAX and BCL-2 apoptosis in silicosis rat model, and to explore the correlation between HSP27 and BAX and BCL-2 apoptosis. Silicosis model was established by the oropharyngeal and endotracheal intubation. Forty SPF healthy adult Wistar male rats were randomly divided into 4 groups, with 10 rats in each group. Silicosis group for 6 weeks(feeding for 6 weeks), silicosis group for 8 weeks(feeding for 8 weeks): oropharyngeal and tracheal perfusion of 50 mg/mL SiO_2 suspension 1.0 mL/mouse; Model control group for 6 weeks and model control group for 8 weeks: 1.0 mL saline was infused into the oropharynx and trachea. Immunohistochemical staining was used to detect the expression of HSP27, BAX and BCL-2 in the right lower lung of silicosis model group at 6 and 8 weeks and model control group at 6 and 8 weeks. Western blot was used to detect the protein expression of HSP27, BAX and BCL-2 in the left lower lobe lung tissue of silicosis model group at 6 and 8 weeks and model control group at 6 and 8 weeks, respectively. Immunofluorescence staining was used to detect the colocalization of HSP27 with pro-apoptotic factor BAX and HSP27 with anti-apoptotic factor BCL-2. Compared with the model control group at 6 weeks and 8 weeks, the expression of HSP27 and pro-apoptotic factor BAX in fibrotic region increased, and the expression of anti-apoptotic factor BCL-2 decreased in silicosis model group at 6 weeks and 8 weeks(P&lt;0.05). Immunofluorescence staining showed that there was colocalization of HSP27 and pro-apoptotic factor BAX in the fibrotic region. Correlation analysis showed that the correlation coefficient between HSP27 and pro-apoptotic factor BAX was r=0.94, indicating a positive correlation between them, while the correlation coefficient between HSP27 and anti-apoptotic factor BCL-2 was r=-0.81, indicating a negative correlation between them. High expression of HSP27 and pro-apoptotic factor BAX and low expression of anti-apoptotic factor BCL-2 exist in silicosis rats, and their expression is correlated.

Up-regulation of androgen receptor by heat shock protein 27 and miR-1 induces pathogenesis of androgenic alopecia.

The pathogenesis of androgenetic alopecia (AGA) is related to the level of androgen and its metabolic pathways. The binding of androgen and androgen receptor (AR) depends on the assistance of heat shock protein 27 (HSP27). HSP27 combined with microRNAs (miR)-1 can regulate AR levels. However, it is not clear whether HSP27 and miR-1 jointly participate in the pathogenesis of AGA. This study aims to investigate the role of AR up-regulation in the pathogenesis of AGA and underlying mechanisms. A total of 46 male AGA patients (AGA group), who admitted to the First Affiliated Hospital of Guangzhou Medical University from September 2019 to February 2020, and 52 healthy controls admitted to the same period were enrolled in this study. Serum levels of dihydrotestosterone (DHT) and HSP27 in patients and healthy controls were measured by ELISA. Western blotting was used to detect the protein expression of HSP27 and AR in scalp tissues of patients and the healthy controls. The levels of HSP27, AR, and miR-1 were analyzed using real-time PCR. Human dermal papilla cells were transfected with HSP27 siRNA to inhibit the expression of HSP27. MiR-1 and miR-1 inhibitors were transfected simultaneously or separately into cells and then the changes in AR protein expression were detected. The levels of DHT and HSP27 in the AGA group were (361.4±187.7) pg/mL and (89.4±21.8) ng/mL, respectively, which were higher than those in the control group [(281.8±176.6) pg/mL and (41.2±13.7) ng/mL, both P<0.05]. However, there was no significant difference in serum HSP27 and AR levels among AGA patients with different degrees of hair loss (P>0.05). Correlation analysis showed that there was a positive correlation between HSP27 level and DHT level in the AGA patients (P<0.05). The level of HSP27 mRNA in scalp tissue was negatively correlated with that of miR-1 mRNA (P<0.05). Compared with the control group, the levels of HSP27 protein, AR protein, HSP27 mRNA, and AR mRNA in scalp tissues of AGA group were significantly increased (P<0.05). The up-regulation of HSP27 in scalp tissues of AGA patients was closely related to the increased levels of AR. However, the level of miR-1 in scalp tissues of AGA patients was significantly down-regulated, contrary to the expression of AR (P<0.05). Further in cell studies showed that inhibition of HSP27 or miR-1 expression in human dermal papilla cells could inhibit the expression of AR, and inhibition of both HSP27 and miR-1 expression was found to have an accumulative effect on AR, with statistically significant differences (all P<0.05). HSP27 could combine with miR-1 to up-regulate AR levels, which is closely related to the development of AGA.

Changes In Activity of HSPA1A and HSPA1L is Associated with Early Acute Pancreatitis Severity

Introduction: Genome assessment shows promise as a potential method for predicting acute pancreatitis (AP) severity. We hypothesized that Heat Shock Protein 70 (HSP70) single nucleotide polymorphisms (SNPs) and expression changes may play a role in early detection of acute pancreatitis severity. Methods: A total of 57 AP patients and 52 age- and sex-matched healthy controls were studied. Peripheral blood samples from pancreatitis patients were collected upon admission. Two SNPs of the HSP70-gene family were selected. RNA was extracted parallel to genomic DNA from AP patients (N=12) and healthy controls (N=21).Gene expression of two HSP70 family members HSPA1A and HSPA1L was measured using TaqMan gene expression assays via reverse transcription quantitative polymerase chain reaction (RT-qPCR). Results: Major allele frequency in HSPA1A gene was 1 (A>G) for AP patients and 1 (A>G) for controls. For HSPA1A, the gene major allele frequency was 0.672 (A>C) for patients and 0.7 (A>C) for controls, the major allele frequencies were 0.638 (A>G) and 0.672 (G>C) for patients and 0.85 (A>G) and 0.65 (G>C) for controls, respectively. The allele frequencies differed between AP patients with organ failure and those with mild disease. Following gene expression of HSPA1L and SNP relationship analysis, AP patients with heterozygous genotype had higher expression levels as compared to non-AP patients (p=0.014). Conclusions: Our data suggest that polymorphism of the HSP70 promoter region may be a risk factor for developing severe acute pancreatitis. Further study to determine the urine levels of HSP70 protein expression is needed to confirm the protective mechanism of HSP70.

Cyclophosphamide inhibits the progression of Meniere's disease by reducing the generation of circulating immune complex.

Endolymphatic hydrops is a characteristic pathological manifestation of Meniere's disease (MD) that has been previously associated with autoimmunity. Interest in the circulating immune complex (CIC) has increased due to its reported role in the occurrence of MD. The present study aimed to investigate the potential value of serum CIC concentration in the diagnosis of MD and the therapeutic potential of cyclophosphamide (CTX) for the treatment of MD. In the present study, guinea pigs were immunized with isologous crude inner ear antigens to establish an autoimmune MD model. Pure tone audiometry, Vestibular-evoked myogenic potential test, electrocochleography test and auditory brainstem response was applied in this study for assessing the severity of MD in guinea pigs. ELISA was applied to measure CIC, tumor necrosis factor α (TNF-α) and heat shock protein 70 (HSP70) expression levels in the serum samples of different groups of patients. Western blotting was applied to detect the protein expression of HSP70 in inner ear tissues in guinea pigs. Hematoxylin and eosin staining was applied to visualize the spiral ganglions in spiral ganglions models. CIC expression in the inner ear was detected by immunohistochemistry. In vivo experiments were performed to confirm the therapeutic effects of CTX in MD. Serum concentrations of CIC, TNF-α and HSP70 were found to be significantly higher in patients with MD, which were also associated with increases in hearing classification and the severity of endolymphatic hydrops. Using a guinea pig MD model, ELISA results revealed significantly increased serum CIC, TNF-α and HSP70 concentrations compared with those in the control group. ABR results showed that the thresholds in the CTX group were notably decreased compared with that in the dexamethasone group, whereas CIC concentrations in the serum were reduced following dexamethasone and CTX treatments compared with those after saline treatment. In the inner ear tissues, the CIC concentration in CTX group was lower than that in the dexamethasone group. Similarly, reductions in HSP70 and TNF-α concentrations was also observed in a similar manner. Immunohistochemistry staining found notably lower CIC deposition in the inner ear tissues following CTX treatment than that in dexamethasone group. Taken together, higher CIC expression can be used as a biomarker for MD diagnosis. The efficacy of CTX in MD was found to be higher compared with that in dexamethasone, which may be associated with the effective inhibition of CIC, HSP70 and TNF-α generation.

Effect of aerobic training on heat shock protein 27 and brain-derived neurotrophic factor in hippocampus and prefrontal cortex of type II diabetic rats

Background: According to the experimental studies, the levels of heat shock protein 27 (HSP27) and brain-derived neurotrophic factor (BDNF) are decreased in diabetes which is associated with impaired cognitive function in the brain. Therefore, the increased expression of these two proteins in some brain regions associated with learning and memory can be one of the positive effects of aerobic training on cognition improvement at diabetes. The aim of the present study was to evaluate the effects of aerobic treadmill exercise training (ATET) on HSP27 and BDNF proteins in the prefrontal cortex and hippocampus of streptozotocin (STZ)-induced diabetes in rats. Methods: Twenty healthy adult male Wistar rats approximately 3-month old, weighing 250 ± 25gr were used in this study. Experimental diabetes was induced by a high-fat diet (HFD) for 4 weeks and a low dose of STZ (35 mg/kg, intraperitoneally). Then diabetic animals were divided into two groups (n=10 per each group): control group (C) and aerobic training group (E). The exercise program was treadmill running at 13 m/min, 25 min/day, for 5 days/week at 0˚ slope for the first week, with a gradual increase to 27 m/min for 60 min/day in week 8. Un-paired t-test was used to compare two groups with the considered significant at p&lt;0.05. Results: The results showed that ATET significantly increased protein expression of HSP27 and BDNF in the hippocampus and prefrontal cortex as compared to C group. Conclusion: Our findings showed that aerobic treadmill exercise program can increase the protein expression of HSP27 and BDNF associated with cognition and synaptic plas­ticity and thus counteract the neurodegenerative diseases of the nervous system.

Effects of mancozeb on heat Shock protein 70 (HSP70) and its relationship with the thermal physiology of Physalaemus henselii (Peters, 1872) tadpoles (Anura: Leptodactylidae).

Negative impacts on amphibians have been reported due to contamination by agrochemicals. However, until now, no study has tested the effect of the fungicide mancozeb (MZ) on thermal tolerance and its relationship with the expression of heat shock proteins (HSPs). MZ is the best-selling broad-spectrum fungicide in the world, which negatively affects non-target organisms. Here, we tested for the first time the effects of MZ on critical thermal maximum (CTmax) and its relationship to the expression of heat shock protein 70 (HSP70) in tadpoles of Physalameus henselii, a colder-adapted species in southernmost of the Neotropical region. A sublethal concentration of 2mg/L was used. We found that the CTmax of the MZ-treated group was lower than that of the control group. In addition, there was an increase in HSP70 expression in tadpoles exposed to MZ and in tadpoles that underwent heat treatment. However, tadpoles subjected to MZ and heat treatment showed no induced HSP70 protein expression. Our results demonstrated that sublethal doses of the fungicide MZ negatively affected the thermal physiology and heat shock protein expression in tadpoles of P. henselii by inducing an increase in HSP70 concentration and by reducing the critical CTmax supported by tadpoles. It is important to understand the relationship between environmental contamination and physiological thermal limits in our current scenario of high rates of habitat conversion associated with unrestricted use of agrochemicals, as well as the challenging environmental changes induced by global warming.

Photobiomodulation therapy on expression of HSP70 protein and tissue repair in experimental acute Achilles tendinitis.

The aim of the present study was to investigate the effects of photobiomodulation (PBM) therapy on the expression of heat shock protein 70 (HSP70) and tissue repair in an experimental model of collagenase-induced Achilles tendinitis. Thirty Wistar rats (aged 12weeks) were randomly distributed among control group (n = 8), tendinitis group (n = 11), and LED group (n = 11). Tendinitis was induced in the tendinitis and LED groups through a peritendinous injection of collagenase (100μl). The LED group animals received the first irradiation 1h after injury. A 630 ± 20nm, 300-mW continuous wave light-emitting diode (LED), spot size 1cm2, was placed in contact with the skin. One point over the tendon was irradiated for 30s, delivering 9J (9J/cm2). LED irradiation was performed once daily for 7days, with the total energy delivered being 63J. The tendons were surgically removed and expression of the HSP70 protein was calculated using semi-quantitative analyses of immunohistochemistry (HSCORE). Number of fibroblasts and amount of collagen were measured using histological and histochemical analyses. An increase in the mean HSCORE for HSP70, in the number of fibroblasts, and in the amount of collagen were found in the LED group compared with those in the tendinitis and control group (P≤ 0.05). PBM therapy increased the expression of the HSP70, number of fibroblasts, and amount of collagen in the acute Achilles tendinitis in rats.

Effects of transport stress on pathological injury and expression of main heat shock proteins in the caprine stomach

BackgroundTransportation is necessary to introduce new breeds of goats to the farm and move the adult meat goat from the farm to the slaughterhouse. However, these actions may give rise to transport stress. Heat shock proteins (HSPs) are playing some important regulate roles during transport stress. The aim of this study was to evaluate the effects of transport stress on the pathological injury and HSPs expression in the stomach of goats. A total of three batches of Ganxi goats from western Jiangxi province were enrolled in this study. For each batch, twelve healthy adult male goats were randomly divided into three groups (four goats per batch and per group): Control group, stress group transported during 2 h and stress group transported during 6 h.ResultsOur results showed that the different degrees of stomach walls damage, with the change of expression levels of heat shock protein 27 (HSP27), heat shock protein 70 (HSP70) and heat shock protein 90 (HSP90), occurred after goats transportation. In rumen, the mRNA and protein expressions of HSP27 and HSP70 were increased after transport stress, but not HSP90. In reticulum, all three HSPs mRNA and protein levels were upregulated after 2 h transport, but decreased after 6 h transport. In omasum, HSP27 and HSP70 mRNA and protein were increased after transport stress, however, HSP90 mRNA level only had a slightly enhancement after transport stress. In abomasum, HSP70 and HSP90 mRNA and protein levels were increased after transport stress, but HSP27 was decreased after transport stress.ConclusionsTaken together, these results revealed that the pathological changes in the gastric tissues and the stomach HSPs expression in goats are related to transport stress and duration. Moreover, this study also provides some new data to advocate reducing transport stress of goats and improving animal welfare.

Effects of Combine Exercise on HSP70 and SOD1 Expression of Aorta, Skeletal Muscle and Myocardium in High Fat Diet induced Obese Aging Rats

PURPOSE: Exercise improve myocardial cell protection and vascular function through cell repair and suppression of oxidative stress in cardiovascular diseases caused by aging. This study aimed to investigate the effect of combine exercise on HSP70 and SOD1 protein expression of aorta, skeletal muscle and myocardium in high fat diet induced obese aging rats.METHODS: Male 50-week-old Sprague Dawley rats (n=40) were divided into normal diet (ND, n=10), normal diet+exercise (NDEx, n=10), high fat diet (HFD, n=10), and high fat diet+exercise (HFDEx, n=10) groups. After six weeks on a high fat diet to induce obesity, a 12-week combine exercise program was implemented, which combine exercise (treadmill running+ladder climbing) three times a week for 45 minutes per session.RESULTS: Body weight was significantly decreased after 12 weeks combine exercise program compared to the ND group (p&lt;.05) and HFDEx group compared to the HFD group (p&lt;.05), respectively. After completing the 12-week exercise program, heat shock protein 70 (HSP70) and superoxide dismutase 1 (SOD1) expressions were significantly (p&lt;.05) higher in the NDEx group compared to the ND group in the myocardium. Also, SOD1 protein expression was significantly (p&lt;.05) higher in the NDEx group compared to the ND group and HFDEx group compared to the HFD group in the skeletal muscle.CONCLUSIONS: In conclusion, combine exercise intervention of high fat diet-induced obesity resulted in decreased cell repair protein and antioxidant enzyme protein in the myocardium. Therefore, it is thought that combine exercise intervention for obese induced rats improved the cell repair protein and antioxidant enzyme activity of the myocardium.

Effect Of Aerobic Exercise On Cardiac Diastolic Dysfunction: Role Of Heat Shock Protein 27

PURPOSE: To evaluate the role of heat shock protein 27 (HSP27) in the effect of aerobic exercise on cardiac diastolic dysfunction in type 2 diabetic rats. METHODS: Forty male Sprague-Dawley rats were randomly divided into four groups: control (C), aerobic exercise (A), diabetes (D), and diabetes plus aerobic exercise (DA). Type 2 diabetes was induced by feeding with a high-fat high-sugar diet for 7 weeks followed by a single intraperitoneal injection of streptozotocin (30mg/kg) in the rats. Aerobic exercise was performed on a rodent treadmill at 21m/min for 60 min, 5 days per week for 8 weeks. Metabolic factors, such as fasting blood glucose (FBG), triglycerides (TG), cholesterol (CHOL) and lipoproteins, were determined by a standard procedure. Cardiac structure and function, such as left ventricular diastolic upper diameter (LVIDd), left ventricular diastolic terminal volume (EDV) and ejection fraction (EF), were measured using echocardiography. Myocardial HSP27 protein expression and phosphorylation were determined using western blot, and myocardial HSP27-titin colocalization were determined using double immunofluorescence. Two-way ANOVAs with post-hoc tests were used to assess differences between groups. RESULTS: No pathological changes were observed in the myocardial structure in all four groups. Compared to the C group, the D group had significantly higher levels of FBG, TG, and CHOL (all p<0.01), and significantly lower LVIDd and EDV (both p<0.01). Compared to the D group, the DA group had significantly lower levels of FBG, TG, and CHOL (all p<0.01) and significantly higher LVIDd and EDV (both p<0.05). Interestingly, compared to the C group, the D group showed significantly lower myocardial HSP27 phosphorylation and HSP27-titin colocalization (both p<0.05), while the DA group showed significantly higher myocardial HSP27 protein expression and phosphorylation, and HSP27-titin colocalization than the D group (all p<0.01). CONCLUSION: Decreases in myocardial HSP27 phosphorylation and HSP27-titin colocalization are likely involved in early diastolic dysfunction in diabetic rats. Increased HSP27 phosphorylation and titin colocalization may be an important mechanism by which aerobic exercise restores diastolic function in diabetic rats.

Expression and clinical significance of HSP27 and its phosphorylation in lower extremity arteriosclerosis obliterans

ObjectivesThis study set out to analyze the difference of heat shock protein 27 (HSP27) and its phosphorylation in patients with lower extremity arteriosclerosis obliterans (LEASO) at different stages. This research also examined their clinical significance in this disease.MethodsBlood samples from 60 patients with LEASO were collected and divided into two groups according to ankle-brachial index (ABI): group A (ABI ≤ 0.43) and group B (ABI > 0.43). The expression of HSP27 in each stage of Fontaine was measured by ELISA, and the difference of HSP27 concentration and ABI between the two groups was analyzed. Meanwhile, three normal femoral artery specimens (normal group) and three atherosclerotic femoral artery specimens (lesion group) were collected, and HSP27 and its Phospho-HSP27 (Ser15), Phospho-HSP27 (Ser78) and Phospho-HSP27 (Ser82) were detected by western blotting. The data of the protein level between the normal group and the lesion group was made a statistical analysis.ResultsHSP27 concentration in group A was (40.73 ± 15.99) ng/ml, and ABI was 0.26 ± 0.20. HSP27 concentration in group B was (66.30 ± 24.70) ng/ml, and ABI was 0.64 ± 0.20. The protein expression of HSP27 and its phosphorylation in the normal group was 0.82 ± 0.13, 0.66 ± 0.12, 0.91 ± 0.24 and 0.90 ± 0.16, respectively; the protein expression of the lesion group was 0.45 ± 0.08, 0.42 ± 0.09, 0.39 ± 0.12 and 0.58 ± 0.11.ConclusionPatients with higher LEASO Fontaine stage and lower ABI had a lower HSP27 concentration. Serum HSP27 concentration was negatively correlated with the severity of LEASO, while HSP27 concentration was positively correlated with ABI value. The content of HSP27 and its phosphorylation of lesion group is significantly lower than that of normal group, which may be closely related to the occurrence and development of atherosclerosis.

Sevoflurane inhibits neuronal apoptosis and expressions of HIF-1 and HSP70 in brain tissues of rats with cerebral ischemia/reperfusion injury.

To observe the influences of sevoflurane on neuronal apoptosis and expressions of hypoxia-inducible factor 1 (HIF-1), and heat-shock protein 70 (HSP70) in brain tissues of rats with cerebral ischemia/reperfusion (I/R) injury. A total of 60 Sprague-Dawley rats were selected and divided into sham-operation group (Sham group, n=20), cerebral I/R model group (Model group, n=20), and 3% sevoflurane treatment group (Sevoflurane group, n=20). The rats in each group received neurological scoring, and the blood and brain tissues were collected to detect the concentrations of serum K+, Na+ and glucose (Glu). Enzyme-linked immunosorbent assay (ELISA) was adopted to measure the levels of inflammatory factors [tumor necrosis factor-β (TNF-β) and interleukin-6 (IL-6)] and oxidative stress [catalase (CAT), malondialdehyde (MDA) and superoxide dismutase (SOD)]. Terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay was performed to determine the nerve cell apoptosis in the brain tissues. The gene and protein expressions of Caspase-3, HIF-1, and HSP70 in the brain tissues were measured via quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR) and Western blotting. In Sevoflurane group, the content of serum Glu and Na+ was decreased markedly, that of K+ was increased notably, and the levels of TNF-β, IL-1 and IL-6 were lowered remarkably compared with those in Model group (p<0.05). Moreover, the neurological score was reduced evidently (p<0.05). Model group had significantly strengthened the activity of MDA and CAT and decreased SOD content, while Sevoflurane group exhibited the opposite results. TUNEL staining showed that there were distinctly more apoptotic cells that were dominated by glial cells in Model group and fewer apoptotic cells in Sevoflurane group. It was indicated in gene assay that the messenger ribonucleic acid (mRNA) expression levels of HIF-1, HSP70, and Caspase-3 in Model group were remarkably higher than those in Sham group and Sevoflurane group (p<0.05). According to the results of Western blotting, the protein expressions of HIF-1 and HSP70 in Sevoflurane group were markedly lower than those in Model group. Sevoflurane can reduce the content of inflammatory factors, inhibit apoptosis, and reduce the expressions of HIF-1 and HSP70 in the case of cerebral I/R injury, thus exerting protective effects on rats with cerebral I/R injury.

Resveratrol Plays Protective Roles on Kidney of Uremic Rats via Activating HSP70 Expression.

Objective To investigate the protective effects of resveratrol on kidney of uremic rats and to explore whether the mechanism is associated with heat shock protein 70 (HSP70) expression. Methods Sixty male Sprague Dawley rats were randomly separated into 5 groups, including sham group, uremic model group, and different doses of resveratrol group (5 mg/kg, 10 mg/kg, and 20 mg/kg). The serum creatinine (Cr) and urea nitrogen (BUN) levels were detected by Automatic Biochemical Analyzer (ABA). The pathological changes of renal tissues and the renal interstitial fibrosis were analyzed by hematoxylin-eosin (HE) and Masson, respectively. The expression of HSP70 protein in renal tissues was detected by immunohistochemistry. The expression of HSP70 and NF-κB pathway-related proteins were detected by Western blot. To further validate the protective role of resveratrol through activating HSP70 in uremic rats, HSP70 activator (17-AAG) and HSP70 inhibitor group (MKT-077) were used. Results In the model group, the levels of Cr and BUN in serum were significantly increased, and the renal interstitial collagen deposition was also obviously increased (p < 0.05). Compared with the model group, the levels of Cr and BUN in different doses of resveratrol groups were remarkably declined, and the renal interstitial collagen deposition was declined (p < 0.05). Resveratrol also significantly improved the renal tissue lesions when compared with the model group. In renal tissues, different doses of resveratrol treatment remarkably raised HSP70 and p-IκBα expression and also remarkably declined the level of p-P65 protein (p < 0.05). Meanwhile, the effect of 17-AAG was similar to 20 mg/kg resveratrol on NF-κB pathway-related proteins expression. After the added MKT-077 in the resveratrol treatment group, the levels of HSP70 and p-IκBα in the renal tissue were remarkably declined; however, the levels of p-P65 protein was remarkably raised (p < 0.05). Conclusion Resveratrol played a protective role on the kidney of uremic rats through activating HSP70 expression.

Ovary Proteome Analysis Reveals RH36 Regulates Reproduction via Vitellin Uptake Mediated by HSP70 Protein in Hard Ticks.

Ticks are blood-sucking vector arthropods, which play an important role in transmitting pathogens between humans and animals. RH36 is an immunomodulatory protein expressed in the salivary glands, but not other organs, of partially fed Rhipicephalus haemaphysaloides ticks, and it reaches its peak on the day of tick engorgement. RH36 gene silencing inhibited tick blood feeding and induced a significant decrease in tick oviposition, indicating that another function of immunosuppressor RH36 was regulating tick reproduction. Why did RH36 protein expressed uniquely in the salivary gland regulate tick reproduction? RH36 regulated positively the expression of vitellogenin in ovary, which indicated RH36 protein played an important role in the integration of nutrition and reproduction. According to proteomic analysis, heat shock protein 70 (HSP70) was significantly down-regulated in the immature ovary of post-engorged ticks. In addition, gene silencing of HSP70 not only inhibited tick blood-sucking and the expression of vitellogenin, but also increased tick death rate. These results suggested RH36 affected tick vitellogenin uptake and then regulated ovary cell maturation by modulating the expression of HSP70 protein, and finally controlled tick oviposition.

Effect of moxibustion on expressions of HSP70 mRNA and protein in gastric cancer-bearing rats

Objective: To observe the effect of moxibustion on the mRNA and protein expressions of heat-shock protein 70 (HSP70) in gastric cancer-bearing rats. Methods: A total of 40 healthy Sprague-Dawley (SD) rats were adaptively fed for one week. The gastric cancer model was prepared by Walker-256 cancer tissue transplantation. After 7 d, 10 rats were randomly selected to verify the successful modeling, and the remaining 30 rats were divided into a model group, a moxibustion group and an infrared group by the random number table method, with 10 rats in each group. After enrollment, the moxibustion group received suspended moxibustion at Zhongwan (CV 12), Guanyuan (CV 4) and bilateral Zusanli (ST 36), (the first group of acupoints) on the 1st day, and suspended moxibustion at bilateral Pishu (BL 20) and Weishu (BL 21), (the second group of acupoints) on the 2nd day, 20 min each time, once a day. Moxibustion was alternately performed every other day at the two groups of acupoints for 21 d. From the day of enrollment, rats in the infrared group were irradiated with the infrared radiation at the stomach area on the 1st day, and at the T12-T13 interspinous region on the 2nd day, 20 min each time, once a day, and the two locations were alternately irradiated every other day for 21 d. During the treatment, rats in the model group were intervened by grasping and fixation without treatment. At the end of the treatment, blood was collected from the inner eye orbit, and the HSP70 expression in peripheral blood was determined by enzyme linked immunosorbent assay (ELISA). Rats were sacrificed, the tumor volume and growth inhibition rate were measured. The position and changes of HSP70 in gastric cancer were observed by streptavidin-perosidase (SP); HSP70 protein expression was determined by ELISA; HSP70 mRNA expression in cancer tissues was determined by reverse transcription-polymerase chain reaction (RT-PCR) assay. Results: In comparison of the model group, the volume growth of the gastric cancer in the moxibustion group was significantly restricted (P<0.01); the volume growth inhibition rate in the moxibustion group was 37.93%; the HSP70 expression in peripheral blood and the cancer tissues was significantly increased (both P<0.01); the expression of HSP70 mRNA and HSP70 content in gastric tumor were both obviously increased in the moxibustion group (P<0.01); and a large amount of HSP70 was released to the outside of cancer cells in the moxibustion group. In comparison of the model group, the volume growth of the gastric cancer in the infrared group was slightly restricted (P<0.05) with a volume growth inhibition rate of 15.89%; the HSP70 expression in the infrared group was increased significantly in peripheral blood (P<0.01) and in the gastric cancer tissues (P<0.05); more HSP70 was released outside of the cancer cells in the infrared group. In comparison of the infrared group, the volume growth of gastric cancer was more restricted in the moxibustion group (P<0.05), and the HSP70 expression in the gastric cancer tissues was also higher (P<0.05); more HSP70 was released outside of the cancer cells in the moxibustion group. Conclusion: Moxibustion and infrared treatment inhibit the gastric cancer growth in the gastric cancer-bearing rats, up-regulate the HSP70 expression in gastric cancer tissues, and promote the production and extracellular release of HSP70, and the effect of moxibustion is more obvious. Conclusion: Moxibustion and infrared treatment inhibit the gastric cancer growth in the gastric cancer-bearing rats, up-regulate the HSP70 expression in gastric cancer tissues, and promote the production and extracellular release of HSP70, and the effect of moxibustion is more obvious.

Inhibition of BAG-1 induced SH-SY5Y cell apoptosis without affecting Hsp70 expression.

To further explore the role of BCL-2 associated anthanogen-1 (BAG-1) in neuronal apoptosis and whether the effect of BAG-1 depends on heat shock protein 70 (HSP70). RNA interference (RNAi) technology was used to inhibit the expression of BAG-1 in SH-SY5Y cells. Hypoxia-reoxygenation injury model in the SH-SY5Y cells was established. Cell Counting Kit-8 (CCK-8) was performed for cell viability. Annexin V-APC/7-AAD double-staining followed by flow cytometry was used to measure cell apoptosis. Quantitative reverse-transcription polymerase chain reaction and Western blot analysis were used to detect the messenger RNA (mRNA) and protein expression of genes, respectively. BAG-1 gene silencing decreased SH-SY5Y cell viability and promoted SH-SY5Y cell apoptosis after hypoxia-reoxygenation. However, the down-regulation of BAG-1 had no effect on the mRNA and protein expression of HSP70. BAG-1 could protect SH-SY5Y cells from the hypoxia-reoxygenation injury without affecting HSP70 expression.