Circulating mortalin in blood and activation of the alternative complement pathway as risk indicators in COVID-19 infection.

Background: Mortalin/GRP75 is a ubiquitous mitochondrial chaperone related to the cytosolic heat shock protein 70. It protects cells from various types of damages and from senescence. Our goal was to determine whether COVID-19 patients have circulating mortalin in their blood and to assess its prognostic value in anticipating disease severity. Methods: Mortalin was determined by ELISA in the sera of 83 COVID-19 patients enrolled in the study. Patients were categorized into 4 groups: critical patients who died (FATAL) or required intensive care and survived (ICU), patients of mild severity (hospitalized but not critical) who required nasal oxygen support (HOSP+O2), and patients who did not need oxygen therapy (HOSP). Results: The mortalin concentration in the serum of all COVID-19 patients in the cohort was 194-2324 pg/mL. A comparison of the mortalin levels by peak severity among the various patient groups showed a highly significant difference between the HOSP and FATAL groups and a significant difference between the HOSP and the ICU groups. COVID-19 patients who eventually failed to survive had at hospitalization a markedly higher level of mortalin in their sera. Cox regression analysis revealed a high mortality hazard (HR=3.96, p<0.01) in patients with high mortalin circulating levels (above the median, ≥651 pg/mL). This was confirmed in survival curve analysis (Kaplan-Meier; p=0.0032, log-rank test). Mortalin remained an independent predictor of mortality even after adjusting for age and sex or various complement activation products. Complement activation data collected in an earlier study in the same cohort was compared regarding the mortalin levels. Patients with higher circulating mortalin levels also had higher levels of complement C3a but reduced levels of properdin. Discussion: This is the first report on circulating mortalin in COVID-19 patients. Higher mortalin levels were associated with more severe illnesses and a higher risk of death. We claim that quantifying the blood levels of mortalin and activated complement proteins will provide important information on the prognosis of COVID-19 patients and will serve as a useful tool for guiding their clinical management and treatment.

Onset of Adverse Abdominal Events Due to Intestinal Ischemia-Reperfusion Injury after Aortic Cross-Clamping Is Associated with Elevated HSP70 Serum Levels in the Early Postoperative Phase.

Tissue injury of the viscera during open thoracoabdominal aortic (TAA) reconstructions has been reported as the aftermath of the ischemia-reperfusion mechanism following supracoeliac aortic cross-clamping. Abdominal complications after open aortic reconstructions, although rare through the intraoperative implementation of selective visceral artery blood perfusion, are associated with high rates of reinterventions and a poor prognosis. Recent animal experiments demonstrated that provoking mesenteric ischemia in rats induces the leukocyte-mediated transcription of heat-shock protein 70 (HSP70), a chaperone belonging to the danger-associated molecular pattern proteins (DAMPs). Translating these findings clinically, we investigated the serum levels of HSP70 in patients undergoing open aortic reconstructions with supracoeliac clamping. We postoperatively observed a relevant induction of HSP70, which remained significantly elevated in cases of postoperative abdominal complications (paralytic ileus, abdominal compartment syndrome, and visceral malperfusion). The receiver-operator curve analysis revealed the reliable prognostic accuracy of HSP70 as a biomarker for these complications as soon as 12 h post-operation (AUC 0.908, sensitivity 88.9%, specificity 83.3%). In conclusion, measuring HSP70 serum levels in the early postoperative phase may serve as a further adjutant in the diagnostic decision making for both the vascular surgeon and intensivist for the timely detection and management of abdominal complications following open TAA surgery.

The abnormal level of HSP70 is related to Treg/Th17 imbalance in PCOS patients.

BACKGROUND: Polycystic ovary syndrome (PCOS) is a disease with chronic nonspecific low-grade inflammation. The imbalance of immune cells exists in PCOS. Several studies have found that heat shock protein 70 (HSP70) may be involved in the immunological pathogenesis of PCOS, but the relationship between HSP70 and Regulatory T cell (Treg)/T helper cell 17(Th17) ratio remains unclear. This study aims to explore the correlation between HSP70 and Treg/Th17 ratio and to provide evidence for the role of HSP70 in the immunological etiology of PCOS. RESULTS: There was no significant difference in age and body mass index (BMI) between the two groups. The concentrations of basal estradiol (E2), basal follicle-stimulating hormone (FSH) did not show a significant difference between the two groups. The concentrations of basal luteinizing hormone (LH) (P < 0.01), testosterone (T) (P < 0.01), glucose (P < 0.001) and insulin (P < 0.001) in PCOS patients were significantly higher than those in the control group. The protein levels of HSP70 were significantly higher in serum in the PCOS group (P < 0.001). The percentage of Treg cells was significantly lower (P < 0.01), while the percentage of the Th17 cells of the PCOS group was significantly higher than that of the control group (P < 0.05). The ratio of Treg/Th17 in the PCOS group was significantly lower (P < 0.001). The concentrations of Interleukin (IL)-6, IL-17, and IL-23 were significantly higher, while the levels of IL-10 and Transforming growth factor-ß (TGF-ß) were significantly lower in the PCOS group (P < 0.001). Spearman rank correlation analysis showed a strong negative correlation of serum HSP70 levels with Treg/Th17 ratio, IL-10, and TGF-ß levels. In contrast, HSP70 levels were significantly positively correlated with IL-6, IL-17, IL-23, LH, insulin, and glucose levels. CONCLUSION: The abnormal level of HSP70 is correlated with Treg/Th17 imbalance and corresponding cytokines, which indicates that HSP70 may play an important role in PCOS immunologic pathogenesis.

HSP70 as a biomarker of the thin threshold between benefit and injury due to physical exercise when exposed to air pollution.

Physical exercise has acute and chronic effects on inflammatory balance, metabolic regulation, and redox status. Exercise-induced adaptations are mediated by enhanced 70-kDa heat shock protein (HSP70) levels and an improved heat shock response (HSR). Therefore, exercise could be useful against disease conditions [obesity, diabetes mellitus (DM), and exposure to atmospheric pollutants] marked by an impaired HSR. However, exercise performed by obese or diabetic subjects under pollution conditions might also be dangerous at certain intensities. Intensity correlates with an increase in HSP70 levels during physical exercise until a critical point at which the effort becomes harmful and impairs the HSR. Establishing a unique biomarker able to indicate the exercise intensity on metabolism and cellular fatigue is essential to ensure adequate and safe exercise recommendations for individuals with obesity or DM who require exercise to improve their metabolic status and live in polluted regions. In this review, we examined the available evidence supporting our hypothesis that HSP70 could serve as a biomarker for determining the optimal exercise intensity for subjects with obesity or diabetes when exposed to air pollution and establishing the fine threshold between anti-inflammatory and pro-inflammatory exercise effects.

PROGRESS STUDY: Progression of chronic kidney disease in children and heat shock proteins.

Various molecular and cellular processes are involved in renal fibrosis, such as oxidative stress, inflammation, endothelial cell injury, and apoptosis. Heat shock proteins (HSPs) are implicated in the progression of chronic kidney disease (CKD). Our aim was to evaluate changes in urine and serum HSP levels over time and their relationships with the clinical parameters of CKD in children. In total, 117 children with CKD and 56 healthy children were examined. The CKD group was followed up prospectively for 24 months. Serum and urine HSP27, HSP40, HSP47, HSP60, HSP70, HSP72, and HSP90 levels and serum anti-HSP60 and anti-HSP70 levels were measured by ELISA at baseline, 12 months, and 24 months. The urine levels of all HSPs and the serum levels of HSP40, HSP47, HSP60, HSP70, anti-HSP60, and anti-HSP70 were higher at baseline in the CKD group than in the control group. Over the months, serum HSP47 and HSP60 levels steadily decreased, whereas HSP90 and anti-HSP60 levels steadily increased. Urine HSP levels were elevated in children with CKD; however, with the exception of HSP90, they decreased over time. In conclusion, our study demonstrates that CKD progression is a complicated process that involves HSPs, but they do not predict CKD progression. The protective role of HSPs against CKD may weaken over time, and HSP90 may have a detrimental effect on the disease course.

Exogenous Hsp70 attenuates nitroglycerin-induced migraine-like symptoms in mice.

Although the connection between heat shock protein 70 (HSP70) and vestibular migraine is not clear, HSP70 is neuroprotective in other scenarios. This study aimed to investigate the potential of exogenous HSP70 for treating migraine-like symptoms in a mouse model of nitroglycerin (NTG)-induced migraine. HSP70 levels were assessed in patients with vestibular migraine and healthy individuals by ELISA. Migraine was induced in mice by NTG, and HSP70 expression was examined in the trigeminal nucleus caudalis (TNC) tissue of mice treated with NTG and NTG together with exogenous HSP70. The effects of exogenous HSP70 on migraine-like symptoms were assessed through behavioral assays. Finally, the impact of HSP70 on oxidative stress and NF-κB signaling in mice with migraine was investigated. Serum HSP70 in patients with vestibular migraine was significantly lower than that of healthy individuals. NTG administration significantly suppressed HSP70 expression in mouse TNC tissue, which was reversed by exogenous HSP70. HSP70 alleviated NTG-induced mechanical hypersensitivity, light aversion, and anxiety-like behavior. Finally, exogenous HSP70 suppressed NTG-induced oxidative stress and NF-κB signaling. Our study suggests that exogenous HSP70 may be a potential therapy for alleviating migraine symptoms and our promising finding warrants further investigation of HSP70 for clinical application.NEW & NOTEWORTHY The study suggests that exogenous HSP70 may be a potential therapy for alleviating migraine symptoms and our promising finding warrants further investigation of HSP70 for clinical application.

Analysis of HSPA1B (+1267A>G) Genetic Polymorphism in Patients with Vibration Disease in Combination with Metabolic Syndrome.

In 167 men with vibration syndrome (VS), allele and genotype frequencies of HSPA1B (+1267A>G) genetic polymorphism (rs1061581) were studied depending on the type of vibration exposure and on the presence or absence of metabolic syndrome (MetS). The examined subjects were divided into two groups: VS patients (n=80) and VS+MetS patients (n=87). The differences in the lipid profiles between groups were revealed against the background of the lack of distinctions in the age of patients for A/G and G/G genotypes carriers. An increase in A/A (p=0.03) and a decrease in A/G (p=0.04) genotype frequencies in VS patients caused by hand-transmitted vibration in comparison with those in whom the disease was caused by a combination of hand-transmitted and whole-body vibration were found. The shifts in the frequencies of the above genotypes (p=0.01) were similar in patients with both types of vibration exposure in the VS+MetS group in comparison with VS group; the carriage of genotypes with the G allele in VS group exceeded that in VS+MetS group (p=0.01).

Increased systemic HSP70B levels in spinal muscular atrophy infants.

Despite newly available treatments for spinal muscular atrophy (SMA), novel circulating biomarkers are still critically necessary to track SMA progression and therapeutic response. To identify potential biomarkers, we performed whole-blood RNA sequencing analysis in SMA type 1 subjects under 1 year old and age-matched healthy controls. Our analysis revealed the Heat Shock Protein Family A Member 7 (HSPA7)/heat shock 70kDa protein 7 (HSP70B) as a novel candidate biomarker to track SMA progression early in life. Changes in circulating HSP70B protein levels were associated with changes in circulating neurofilament levels in SMA newborns and infants. Future studies will determine whether HSP70B levels respond to molecular therapies.

PLASMA BIOCHEMISTRY PROFILES OF WILD WESTERN TIGER SNAKES (NOTECHIS SCUTATUS OCCIDENTALIS) BEFORE AND AFTER SIX MONTHS OF CAPTIVITY.

Urban wildlife often suffer poorer health than their counterparts living in more pristine environments due to exposure to anthropogenic stressors such as habitat degradation and environmental contamination. As a result, the health of urban versus nonurban snakes might be assessed by differences in their plasma biochemistries. We compared the plasma profiles of western tiger snakes (Notechis scutatus occidentalis) from a heavily urbanized wetland and a natural, nonurbanized wetland. Despite the urbanized snakes having lower body mass index, we found no significant difference between the plasma profiles of the two populations. We collected snakes from each population and kept them in captivity for 6 mo, providing them with stable conditions, uncontaminated (exempt from heavy metals and pesticides) food and water, and lowered parasite intensity in an attempt to promote better health through depuration. After captivity, snakes experienced a significant improvement in body mass index and significant changes in their plasma profiles. Snakes from the natural wetland initially had more variation of DNA damage; mean concentration of DNA damage in all snakes slightly decreased, but not significantly, after captivity. We present the plasma biochemistry profiles from western tiger snakes both before and after captivity and suggest a period of removal from natural stressors via captivity may offer a more reliable result of how plasma profiles of healthy animals might appear.

Prognosis and predictive value of heat-shock proteins expression in oral cancer: A PRISMA-compliant meta-analysis.

BACKGROUND: Heat-shock proteins (HSP) is a key chaperone protein which maintains intracellular proteostasis and is expressed on the surface of solid and hematological malignancies. Several studies have reported paradoxical evidence of the association between HSP expression and prognosis of oral cancer. To address the discrepancy, we carried out the meta-analysis to assess the role of HSP such as: HSP70, HSP90, HSP27, HSP60, and HSP105 in susceptibility, progression, and prognosis of oral cancer. MATERIALS AND METHODS: We retrieved the PubMed, Embase, Web of science, China National Knowledge Infrastructure (CNKI), and Wanfang databases to acquire the eligible studies which were associated with HSP70, HSP90, HSP27, HSP60, and HSP105 protein expression and oral cancer. We applied hazard ratio (HR) and its 95% confidence interval (95% CI) to assess the value of HSP protein expression in overall survival of oral cancer; odds ratio (OR) and its 95% CI were used to evaluate the association of risk and clinical features of oral cancer. Funnel plot, Begg test, and Egger line regression test were utilized to observe publication bias among studies. All statistical analysis was performed with Stata 14.0 software (Stata Corporation, College Station, TX). RESULTS: A total of 26 studies were included in the present meta-analysis. On based of the results, HSP70 and HSP27 had no significant association with progression of oral cancer. However, the pooled HR and 95% CI revealed a significant well effects of HSP70 and HSP27 expression on survival of oral cancer. Moreover, the susceptibility of oral cancer was significantly associated with HSP70 and HSP60 overexpression. CONCLUSION: HSP70 and HSP27 protein overexpression might be valuable biomarkers for the prognosis of oral cancer. And HSP70 and HSP60 might have potential predictive effects on the risk of oral cancer.

Different responses of apoptotic, inflammatory and heat shock protein gene expression to a single bout of high-intensity interval exercise between physically active and inactive men.

High mechanical load of muscles may induce muscular apoptosis on the one hand and adaptation to exercise on the other. This study aimed to explore whether changes of circulatory levels of inflammation, apoptosis and heat shock proteins (HSPs) messenger RNA (mRNA) following single bout of high-intensity interval exercise (HIIE) differs between physically active (PA) and inactive (PI) men. Nine PA and 9 PI (peak oxygen consumption: 2.6 ± 0.4 vs. 2.0 ± 0.2 L·min-1) healthy men (age: 28.7 ± 6.3 vs. 30.2 ± 4.5 years and body mass index: 2.6 ± 2.1 vs. 23.3 ± 2.8 kg·m-2) performed HIIE, comprising 4 repeats of a Wingate test (load: 0.050 kg·kg-1 body weight). Blood samples were collected before exercise, 5 min after HIIE, and 24 h after HIIE for measuring mRNA of inflammation markers interleukin-6 (IL-6) and tumor necrosis factor alpha (TNFα), apoptosis markers including Bcl-2, Bax, and HSP27, HSP60, HSP70, HSP90 using quantitative real-time PCR analysis. Post-HIIE IL-6, TNFα and HSP60 were higher in the PI than the PA group 5 min after exercise (p = 0.003, effect size (ES) = 1.59; p = 0.007, ES = 1.59 and p = 0.027, ES = 1.10 respectively). HSP70 acutely increased only in the PA group (p = 0.024, ES = 1.20). The increase in Bcl-2 (p = 0.047, ES = 1.08) and Bax (p = 0.024, ES = 1.20) levels were higher in the PI group 5 min after HIIE. The present study indicated that the response of inflammatory, apoptosis and HSP gene expressions to HIIE in blood of healthy male volunteers strongly depends on their level of regular physical activity. Novelty: Blood IL-6 and HSP60 mRNA levels following high intensity exercise may indicate metabolic stress. Increased blood HSP70 mRNA in physically active men may show an alternative apoptosis suppression pathway.

HSP70 as a Diagnostic and Prognostic Marker in Egyptian Women With Breast Cancer.

BACKGROUND: Heat shock protein 70 (HSP70) is a significant cellular stress response protein that has intrinsic and extrinsic pathways to protect cells against apoptosis. It is one of the most induced proteins in cancer cells. The aim of the present study is to investigate the significant role of the HSP70 expression in Egyptian patients with breast cancer (BC) and its potential to be as a diagnostic and prognostic marker. MATERIALS AND METHODS: HSP70 was examined in 155 cases in this prospective study; patients were subdivided into 3 groups: 60 patients with malignant metastatic disease, 60 patients with malignant non-metastatic disease, and 35 patients with benign lesions as control. HSP70 expression was detected using enzyme-linked immunosorbent assay (ELISA) and immunohistochemistry (IHC). RESULTS: Most cases of breast cancer expressed HSP70 in both serum (98.3%) and tumor tissue (90%). A strong positive correlation was found between HSP70 IHC and ELISA (r = 0.811). The mean HSP70 levels, as detected in both patients' serum by ELISA and tumor tissue by IHC, was significantly higher in patients with BC than in benign cases (P = .001). HSP70 was significantly higher in patients with metastatic BC than in those with non-metastatic BC (P = .001). HSP70 showed positive correlation with tumor size (pT stage) and number of lymph node metastases (P ≤ .001). CONCLUSION: HSP70 is over-expressed in patients with metastatic and non-metastatic BC than in benign cases. A high level of HSP70 either in patient's serum or in tumor tissue correlated significantly with advanced disease in patients with BC. This present study suggests that HSP70 can serve as a BC biomarker for early screening, diagnosis, and follow-up.

Heat Shock Protein 70 Is Associated With Cardioversion Outcome and Recurrence of Symptomatic Recent Onset Atrial Fibrillation in Hypertensive Patients.

ABSTRACT: Accumulating evidence indicates that heat shock proteins (HSPs) may represent a suitable biomarker to predict atrial fibrillation (AF). We investigated the relation of circulating serum HSP70 (sHSP70) with inflammatory cytokines and recurrence of symptomatic recent onset AF (ROAF). We enrolled 90 patients with ROAF (the duration from onset of symptoms ≤24 hours) and 30 controls. Patients received amiodarone for cardioversion and rhythm control. The association of serum HSP70, serum interleukin-2 (sIL-2), and serum interleukin-4 (sIL-4) with the presence of cardioversion and AF recurrence within a year was investigated. Toll-like receptor 4 (TLR4) signaling dependence for IL-2 and IL-4 induction in response to stimulation with HSP70 was tested in rat aortic vascular smooth muscle cell cultures. Patients had higher sHSP70 and sIL-2 and lower sIL-4 compared with controls. Serum HSP70 was independently associated with ROAF (P = 0.005) and correlated with sIL-2 (r = 0.494, P < 0.001) and sIL-4 (r = -0.550, P < 0.001). By 48 hours, 71 of the 90 patients were cardioverted, with noncardioverted patients having higher sHSP70 and sIL-2 and lower sIL-4, which were the only independent factors associated with cardioversion. AF recurred in 38 of the 71 cardioverted patients in 1 year. A cutoff value of sHSP70 ≥0.65 ng/mL and sIL-2 ≥0.21 pg/mL was the only independent factor associated with AF recurrence (hazard ratio: 3.311, 95% confidence interval: 1.503-7.293, P = 0.003 and hazard ratio: 3.144, 95% confidence interval: 1.341-7.374, P = 0.008, respectively). The exposure of smooth muscle cell to HSP70 in vitro increased the expression of IL-2 (5×) and IL-4 (1.5×) through TLR4-dependent and receptor-independent mechanisms. In conclusion, sHSP70 and sIL-2 might constitute a prognostic tool for determining the cardioversion and recurrence likelihood in ROAF.

Reaction Kinetics Modeling of eHsp70 Induced by Norepinephrine in Response to Exercise Stress.

Exercise elicits a systemic adaptation reaction, involving both neuroendocrine and cellular/paracrine stress responses, exemplified by the sympathoadrenergic activity and the release of cellular Hsp70 into the circulation. Regular sports training is known to result in increased fitness. In this study, we characterized the plasma norepinephrine and Hsp70 levels and modeled their relationship in response to exercise stress by bicycle ergometer in 12 trained judoka athletes and in 10 healthy controls. Resting norepinephrine was similar in both groups, whereas Hsp70 was significantly higher in controls compared to athletes. Intense exercise load induced both norepinephrine and Hsp70 elevation. However, both norepinephrine and Hsp70 were significantly lower in athletes compared to the control group. A reaction kinetic model was developed that provided a quantitative description of norepinephrine-facilitated extracellular Hsp70 release, congruent with the experimental data. Our study indicates that exercise-induced norepinephrine and extracellular Hsp70 may be coordinated responses to physiological stress, which are robustly affected by regular sports activity.

A study on stress response and fertility parameters in phenotypically thermotolerant and thermosensitive dairy cows during summer heat stress.

It is well documented that heat stress (HS) causes subfertility in dairy cows. However, during the last ten years we have been observing that, under high temperature-humidity index (THI ≥ 75), despite the overall reduced fertility, some cows conceive at the first artificial insemination (AI). Here, we examined distinctive features of cows with conserved fertility under severe HS. From the databases of three herds, 167 lactating Holstein cows were selected; group TT cows (n = 57) conceived in the previous summer (THI ≥ 75) at the 1st AI, and group TS (n = 110) failed to conceive at the same period after at least 2 consecutive AIs. The animals calved in spring, and in August, blood samples were collected during a hot day (THI ≥ 81) for the determination of cortisol and HSP70 concentrations. In one farm, the validity of fertility data of the previous year was re-examined. In 28 cows from group TT and in 39 cows from group TS, the conception rate was examined during July and August. In 6 cows from each group (TT and TS) the oestrous cycles were synchronized, ovulation was induced with GnRH (THI = 80), and the concentration of the pre-ovulatory LH surge was determined in 9 blood samples. The progesterone concentration in the ensuing cycle was determined in blood samples collected every other day. Overall, cortisol and HSP70 were significantly lower in TT group compared to TS. More (p < .05) animals from group TT conceived at the first AI compared with those from group TS. The induced pre-ovulatory LH surge peaked at higher level (p < .002) in group TT than in group TS, while no difference was recorded among groups either in mean progesterone concentrations or in the duration of the ensuing oestrous cycle. These results are highly suggestive that thermotolerance in some dairy cows is an inherent characteristic, warranting further genetic investigation.

Phase 1 study of the HSP90 inhibitor onalespib in combination with AT7519, a pan-CDK inhibitor, in patients with advanced solid tumors.

PURPOSE: We conducted a phase 1 trial of the HSP90 inhibitor onalespib in combination with the CDK inhibitor AT7519, in patients with advanced solid tumors to determine the safety profile and maximally tolerated dose, pharmacokinetics, preliminary antitumor activity, and to assess the pharmacodynamic (PD) effects on HSP70 expression in patient-derived PBMCs and plasma. METHODS: This study followed a 3 + 3 trial design with 1 week of intravenous (IV) onalespib alone, followed by onalespib/AT7519 (IV) on days 1, 4, 8, and 11 of a 21-days cycle. PK and PD samples were collected at baseline, after onalespib alone, and following combination therapy. RESULTS: Twenty-eight patients were treated with the demonstration of downstream target engagement of HSP70 expression in plasma and PBMCs. The maximally tolerated dose was onalespib 80 mg/m2 IV + AT7519 21 mg/m2 IV. Most common drug-related adverse events included Grade 1/2 diarrhea (79%), fatigue (54%), mucositis (57%), nausea (46%), and vomiting (50%). Partial responses were seen in a palate adenocarcinoma and Sertoli-Leydig tumor; a colorectal and an endometrial cancer patient both remained on study for ten cycles with stable disease as the best response. There were no clinically relevant PK interactions for either drug. CONCLUSIONS: Combined onalespib and AT7519 is tolerable, though below monotherapy RP2D. Promising preliminary clinical activity was seen. Further benefit may be seen with the incorporation of molecular signature pre-selection. Further biomarker development will require the assessment of the on-target impact on relevant client proteins in tumor tissue.

Evaluating Serum Heat Shock Protein Levels as Novel Biomarkers for Atrial Fibrillation.

Background: Staging of atrial fibrillation (AF) is essential to understanding disease progression and the accompanied increase in therapy failure. Blood-based heat shock protein (HSP) levels may enable staging of AF and the identification of patients with higher risk for AF recurrence after treatment. Objective: This study evaluates the relationship between serum HSP levels, presence of AF, AF stage and AF recurrence following electrocardioversion (ECV) or pulmonary vein isolation (PVI). Methods: To determine HSP27, HSP70, cardiovascular (cv)HSP and HSP60 levels, serum samples were collected from control patients without AF and patients with paroxysmal atrial fibrillation (PAF), persistent (PeAF) and longstanding persistent (LSPeAF) AF, presenting for ECV or PVI, prior to intervention and at 3-, 6- and 12-months post-PVI. Results: The study population (n = 297) consisted of 98 control and 199 AF patients admitted for ECV (n = 98) or PVI (n = 101). HSP27, HSP70, cvHSP and HSP60 serum levels did not differ between patients without or with PAF, PeAF or LSPeAF. Additionally, baseline HSP levels did not correlate with AF recurrence after ECV or PVI. However, in AF patients with AF recurrence, HSP27 levels were significantly elevated post-PVI relative to baseline, compared to patients without recurrence. Conclusions: No association was observed between baseline HSP levels and the presence of AF, AF stage or AF recurrence. However, HSP27 levels were increased in serum samples of patients with AF recurrence within one year after PVI, suggesting that HSP27 levels may predict recurrence of AF after ablative therapy.

Targeted Natural Killer Cell-Based Adoptive Immunotherapy for the Treatment of Patients with NSCLC after Radiochemotherapy: A Randomized Phase II Clinical Trial.

PURPOSE: Non-small cell lung cancer (NSCLC) is a fatal disease with poor prognosis. A membrane-bound form of Hsp70 (mHsp70) which is selectively expressed on high-risk tumors serves as a target for mHsp70-targeting natural killer (NK) cells. Patients with advanced mHsp70-positive NSCLC may therefore benefit from a therapeutic intervention involving mHsp70-targeting NK cells. The randomized phase II clinical trial (EudraCT2008-002130-30) explores tolerability and efficacy of ex vivo-activated NK cells in patients with NSCLC after radiochemotherapy (RCT). PATIENTS AND METHODS: Patients with unresectable, mHsp70-positive NSCLC (stage IIIa/b) received 4 cycles of autologous NK cells activated ex vivo with TKD/IL2 [interventional arm (INT)] after RCT (60-70 Gy, platinum-based chemotherapy) or RCT alone [control arm (CTRL)]. The primary objective was progression-free survival (PFS), and secondary objectives were the assessment of quality of life (QoL, QLQ-LC13), toxicity, and immunobiological responses. RESULTS: The NK-cell therapy after RCT was well tolerated, and no differences in QoL parameters between the two study arms were detected. Estimated 1-year probabilities for PFS were 67% [95% confidence interval (CI), 19%-90%] for the INT arm and 33% (95% CI, 5%-68%) for the CTRL arm (P = 0.36, 1-sided log-rank test). Clinical responses in the INT group were associated with an increase in the prevalence of activated NK cells in their peripheral blood. CONCLUSIONS: Ex vivo TKD/IL2-activated, autologous NK cells are well tolerated and deliver positive clinical responses in patients with advanced NSCLC after RCT.

Hspa13 Promotes Plasma Cell Production and Antibody Secretion.

The generation of large numbers of plasma cells (PCs) is a main factor in systemic lupus erythematosus (SLE). We hypothesize that Hspa13, a member of the heat shock protein family, plays a critical role in the control of PC differentiation. To test the hypothesis, we used lipopolysaccharide (LPS)-activated B cells and a newly established mouse line with a CD19cre-mediated, B cell-specific deletion of Hspa13: Hspa13 cKO mice. We found that Hspa13 mRNA was increased in PCs from atacicept-treated lupus-prone mice and in LPS-stimulated plasmablasts (PBs) and PCs. A critical finding was that PBs and PCs [but not naïve B cells and germinal center (GC) B cells] expressed high levels of Hspa13. In contrast, the Hspa13 cKO mice had a reduction in BPs, PCs, and antibodies induced in vitro by LPS and in vivo by sheep red blood cells (SRCs)- or 4-hydroxy-3-nitrophenylacetyl (NP)-immunization. Accordingly, the Hspa13 cKO mice had reduced class-switched and somatically hypermutated antibodies with defective affinity maturation. Our work also showed that Hspa13 interacts with proteins (e.g., Bcap31) in the endoplasmic reticulum (ER) to positively regulate protein transport from the ER to the cytosol. Importantly, Hspa13 mRNA was increased in B220+ cells from patients with multiple myeloma (MM) or SLE, whereas Hspa13 cKO led to reduced autoantibodies and proteinuria in both pristane-induced lupus and lupus-prone MRL/lpr mouse models. Collectively, our data suggest that Hspa13 is critical for PC development and may be a new target for eliminating pathologic PCs.

Autologous extracellular Hsp70 exerts a dual role in rheumatoid arthritis.

Extracellular heat shock proteins (Hsp) influence the adaptive immune response and may ameliorate pathogenesis of autoimmune diseases. While some preclinical observations suggest that highly conserved bacterial and/or murine Hsp70 peptides have potential utility in treatment of rheumatoid arthritis (RA) via induction of T regulatory cells (Treg), the role of extracellular inducible human Hsp70 in adaptive immune processes requires further investigation. The present study evaluated Hsp70 influence on inflammatory cytokine-mediated modulation of T cell immunophenotype in ways that influence RA onset and severity. Initial experiments in the present investigation revealed that serum levels of Hsp70 are approximately 2-fold higher in RA patients versus healthy control subjects. To explore the effect of extracellular Hsp70 on key processes underlying the adaptive immune system, the effects of a highly pure, substrate-, and endotoxin-free human Hsp70 on polarization of the T helper cell subpopulations, including CD4+IL-17+ (Th17), CD4+FoxP3+ (Treg), CD4+IFN-γ+ (Th1), and CD4+IL-4+ (Th2), were studied in naïve human peripheral blood mononuclear cell (PBMC) cultures stimulated with anti-CD3/28 mAb. Major findings included an observation that while Hsp70 treatment increased Th17 frequencies and Th17/Treg ratio, the frequency of Th1 cells and the Th1/Th2 ratio were significantly decreased in the Hsp70-treated PBMC cultures. Moreover, data shown here provides preliminary suggestion that major contributing Hsp70-mediated immunomodulation includes interleukin 6 (IL-6) influence on Th17/Treg and Th1/Th2, since expression of this inflammatory cytokine is enhanced by in vitro Hsp70 treatment. These results are nevertheless preliminary and require further investigation to validate the above model.