HSP Group Research Articles

Circulating endothelial microparticles in children with Henoch–Schönlein purpura; preliminary results

The aim of this study was to investigate the levels of circulating endothelial microparticles (EMPs) in children with HSP and to determine whether there was a difference between patients with nephritis and those without nephritis. Twenty patients with HSP aged between 2.5 and 15 and 10 age-and sex-matched healthy controls were enrolled in the study. The HSP group was divided into two groups, including patients with nephritis (n = 9) and those without nephritis (n = 11). In all groups, circulating EMPs were enumerated by flow cytometry, after staining platelet-free plasma with PE-conjugated anti-CD144. At the same time, human umbilical vein endothelial cells (HUVEC) were incubated with the platelet-free plasma of patients with HSP and that of the control group. Then, circulating EMPs were counted in HUVEC supernatant incubated with the platelet-free plasma of patients and control groups, after staining the supernatant with PE-conjugated anti-CD146. Circulating EMPs were significantly higher in both the active and the remission period of the patient groups compared with the control subjects. In the patient group, there were no statistically significant differences in the level of circulating EMPs between patients with nephritis and those without nephritis. Both CD144 and 146+EMP in patients with HSP nephritis in the active period were substantially higher than in those remissions. CD144+EMP in the active period were substantially higher than in the remission period in patients without nephritis. We detected that circulating EMPs increased in patients with HSP in both active and remission periods. Although clinical and laboratory findings return to normal in the remission period, the increased circulating EMPs may show that the subclinical inflammatory process is continuous. We think that circulating EMPs could be used as a surrogate marker for subclinical inflammation in HSP.

Association of CCR5-59029 A/G and CCL3L1 Copy Number Polymorphism with HIV Type 1 Transmission/Progression among HIV Type 1-Seropositive and Repeatedly Sexually Exposed HIV Type 1-Seronegative North Indians

The CCR5Delta32 mutation does not account for HIV-1 resistance in the majority of persons who are repeatedly exposed to HIV-1 by high-risk activities but remain seronegative and uninfected. Therefore, we investigated the impact of CCR5 59029 A/G and CCL3L1 copy number polymorphism on HIV-1 disease susceptibility and progression among HIV-1-infected and HIV-1-exposed seronegative North Indians. HIV-1-seropositive (HSP, n = 196) patients, stratified on the basis of disease severity (Stages I, II, and III) and HIV-1-exposed seronegative (HES, n = 47) individuals were genotyped for CCR5-59029 A/G polymorphism by PCR-RFLP and CCL3L1 copy number by the real-time TaqMan PCR method. A group of ethnically matched HIV-1-seronegative (HSN, n = 315) healthy volunteers were also genotyped as controls. Statistical analysis was done by SPSS software. The CCR5-59029 AG genotype was significantly higher in the HES compared with the HSP group (57.44% vs. 37.24%, p = 0.014). The CCL3L1 mean copy number of HES was higher compared with the HSP groups (3.148 +/- 0.291 vs. 2.795 +/- 0.122, p = 0.212), but was not significant when compared with independent samples t test. Possession of CCL3L1 copies < or = 2 or >2 was not associated with enhanced or reduced risk of HIV-1 acquisition. Gene-gene interaction studies showed enrichment of the CCR5-59029AG*CCL3L1>2 genotype in the HES group when compared with the HSP group (31.91% vs. 15.81%, p = 0.021, OR = 0.401, CI = 0.194-0.826). The increased frequency of the CCR5-59029AG*CCL3L1>2 genotype among HES individuals led us to conclude that the CCR5-59029 AG genotype and CCL3L1 gene dose appeared to have synergistic or interactive effects and are expected to be involved in the host innate resistance to HIV-1 infection.

Influence of muscle length on muscle atrophy in the mouse tibialis anterior and soleus muscles

The tibialis anterior and soleus muscles were fixed at the stretched or shortened positions to examine the influence of muscle length on muscle atrophy. Mice were divided into control (C), hindlimb suspension (HS), hindlimb suspension with ankle joint fixation at the maximum dorsiflexion (HSD), and hindlimb suspension with ankle joint fixation at the maximum plantarflexion (HSP). During the hindlimb suspension, the length of these muscles in the HS and HSP groups was very similar. Fourteen days after the hindlimb suspension, the atrophy of the tibialis anterior muscle in the HS and HSP groups was evidently milder than that in the HSD group, and that in the HS and HSP groups was very similar, suggesting that atrophy of the tibialis anterior muscle might largely depend on muscle length. Atrophy of the soleus muscle in the HSD group was milder than that in the HS and HSP groups, indicating that atrophy of the soleus muscle might also depend on muscle length. But atrophy of this muscle in the HSP group was milder than that in the HS group. These results demonstrate that some factors induced by the joint immobilization might be effective in preventing atrophy of the soleus muscle.

Effect of phosphate solubility on phosphorus kinetics and ruminal fermentation activity in dairy goats

This study was carried out to determine the impact of phosphorus (P) solubility on its kinetics in liquid and bacterial ruminal phases and also on rumen fermentation activity. For the first time, the fractional disappearance rates of ruminal P were estimated and postprandial variations of bacterial P content were studied. Two groups were formed, each with four ruminal-canulated goats weighing 60 ± 1.4 kg, and were housed indoors within individual stalls. Their days in milk (DIM) ranged between 42 and 52. During adaptation and experimental periods, goats received twice a day (08:00 and 15:00 h) a ration consisting of 400, 300 and 300 g/kg of dry matter (DM) of sugar beet pulp silage, grass hay and experimental semi-purified concentrates, respectively. The concentrates supplied 0.71 g of P/g of dietary P either as soluble source (monocalcium phosphate, HSP group) or insoluble one (dicalcium phosphate, LSP group). Liquid and solid ruminal contents were obtained at time 0, 90, 180 and 360 min after morning feeding. Dry matter intake (DMI) and P intake (Pint) were recorded at each sampling time. Phosphorus in ruminal liquid, ruminal bacteria and volatile fatty acid (VFA) contents were quantified. DMI was 1.22 times higher for the HSP group. Available P concentration was 1.3 times higher for the same group at time 360. Fractional disappearance rates of P from rumen liquid content were not different between groups (−0.007 h −1). Postprandial changes (times 0–90) of P concentrations in liquid-associated bacteria (LAB) from LSP group suggested a relative P deficiency status (HSP = 1.6, LSP = 1.4, ±0.04 g/100 g of organic matter (OM)). Phosphorus content of solid-associated bacteria (P-SAB = 0.59 ± 0.02 g/100 g of SAB) presented less variation suggesting a capacity to use P from feedstuffs. The total and individual VFA kinetics were affected by P solubility. Under extreme conditions, where P supply derives mainly from inorganic sources, P solubility affects the kinetics of DMI and P concentrations in ruminal liquid, as well as fermentation activity. Phosphorus solubility significantly affects P content of LAB, whereas SAB appeared to adapt to variations of P concentrations in rumen liquid. However, further research is needed in these areas.

Association of RANTES −403 G/A, −28 C/G and In1.1 T/C polymorphism with HIV‐1 transmission and progression among North Indians

The relationships between host immune factors and HIV-1 disease progression are still in dispute. The RANTES SNPs exhibit distinct ethnic distribution and are associated with different effects on the course of HIV infection. Therefore, impact of RANTES gene polymorphism on HIV-1 transmission and progression needs to be evaluated. The RANTES genotypes were identified by PCR-RFLP method and confirmed by sequencing in HIV-1 seronegative (HSN; n=315), HIV-1 exposed seronegative (HES; n=47) and HIV-1 seropositive (HSP; n=196) patients classified into different clinical stages (i.e. Stages I, II, III). Fisher exact test was used for statistical analysis and Arlequin software for haplotype analysis. RANTES allele -403G, -28C and In1.1 T were the predominant allele in the subject studied. HSP group have higher frequency of RANTES In1.1 T allele compared with HSN (91.32% vs. 86.19%; P=0.013) and HES (91.32% vs. 78.72%; P=0.001). Higher frequency of RANTES In1.1 C allele in Stage III was observed, compared with Stage I (14.28% vs. 6.39%) and was significantly associated with high risk (P=0.047, OR=2.439, C.I.=1.061-5.609). Haplotype II (ACT) was significantly higher in HSP compared with HSN (9.69% vs. 1.58%) and associated with high risk (P<0.001, OR=6.655, C.I.=2.443-18.132). There were no significant differences in RANTES -403 A/G and -28 C/G genotype and allele distribution in all the groups compared. Our results implicate that RANTES In1.1 T allele and haplotype II (ACT) may be a risk factor for HIV-1 transmission while RANTES In1.1 C allele may be risk factor for disease progression among North Indians.

Binding of Caspase-3 Prodomain to Heat Shock Protein 27 Regulates Monocyte Apoptosis by Inhibiting Caspase-3 Proteolytic Activation

Caspase-3 is an essential executioner of apoptosis responsible for regulating many important cellular processes, among them the number of circulating monocytes, central players in the innate immune response. The activation of caspase-3 requires its processing from an inactive precursor. Here we show that the small heat shock protein 27 (Hsp27) associates with caspase-3 and protein-protein interaction experiments in vivo and with purified proteins demonstrate a direct interaction between Hsp27 and the amino-terminal prodomain of caspase-3. Using an in vitro caspase-3 activation assay, our results further establish that the interaction of Hsp27 with the caspase-3 prodomain inhibits the second proteolytic cleavage necessary for caspase-3 activation, revealing a novel mechanism for the regulation of this effector caspase. Hsp27 expression in monocytes is constitutive. Consistent with a central role of Hsp27 in blocking caspase-3 activation, Hsp27 down-regulation by double-stranded RNA interference induces apoptosis of macrophages, whereas Hsp27 overexpression increases the life span of monocytes by inhibiting apoptosis. Highlighting the importance of cell partitioning in the regulation of apoptosis, immunofluorescence, and subcellular fractionation studies revealed that whereas both caspase-3 and Hsp27 are cytoplasmic in fresh monocytes (i.e. not undergoing apoptosis), Hsp27 moves to the nucleus during apoptosis, a relocalization that can be blocked by promoting the differentiation of monocytes to macrophages or by inhibiting cell death. These results reveal a novel mechanism of caspase-3 regulation and underscore a novel and fundamental role of Hsp27 in the regulation of monocyte life span.

Uterus Preservation in Surgical Correction of Urogenital Prolapse

Objective: This study aimed to evaluate the efficacy of colposacropexy with uterine preservation as therapy for uterovaginal prolapse. Surgical techniques, efficacy and overall results are described. Methods: In this prospective, controlled study, 34 of the 72 consecutive patients with symptomatic uterovaginal prolapse were treated with colposacropexy with uterus conservation (hysterocolposacropexy, HSP) and the other 38 with hysterectomy followed by sacropexy (CSP). Anchorage was achieved with two rectangular meshes in CSP and with one posterior rectangular and one anterior Y-shaped mesh in HSP. Check-ups were scheduled at 3, 6 and 12 months and then yearly. Pre-operative patient characteristics, operative and post-operative events and follow-up results were recorded. Mean follow-up was 51 months (range 12–115). Results: No significant differences emerged in demographic and clinical characteristics between the HSP and CSP groups. Mean operating times, intra-operative blood loss and hospital stay were significantly less after HSP ( p < 0.001). At follow-up success rates were similar in the two groups in terms of uterine and upper vaginal support (100%). Recurrent low-grade cystoceles developed in 1/38 (2.6%) in the CSP group and in 5/34 (14.7%) in the HSP group ( p = NS), recurrent low-grade rectocele developed in 6/38 (15.8%) and in 3/34 (8.8%) patients respectively ( p = NS). No patient required surgery for recurrent vault or uterus prolapse. Urodynamic results showed that pressure/flow parameters improved significantly ( p < 0.001) in both groups. Thirty-one of the 34 patients (91%) in the HSP group and 33/38 (86.8%) in the CSP group were satisfied and would repeat surgery again. Conclusions: Colposacropexy provides a secure anchorage, restoring an anatomical vaginal axis and a good vaginal length. HSP can be safely offered to women who request uterine preservation. Whether the uterus was preserved or not, patients had similar results in terms of prolapse resolution, urodynamic outcomes, improvements in voiding and sexual dysfunctions. HSP has shorter operating times and less blood loss.

Effects of ACE-I or ARB on the prolonging of survival in hypertensive rats with systolic heart failure: Relationship between the survival and the amelioration of cardiac dysfunction or LV fibrosis

The aim of the present study was to assess the effects of angiotensin coverting enzyme inhibitor (ACE-I) perindopril and angiotensin receptor blocker (ARB) candesartan on the survival of Dahl salt-sensitive (DS) hypertensive rats. In addition, we also studied the effects of ACE-I and ARB on LV contractile dysfunction and LV fibrosis for exploring to the mechanism contributing to the improvement of the survival. At the age of 6 weeks, the DS rats were divided into the following four groups: DS rats fed a low-salt diet (group LS); DS rats fed a high-salt diet (group HS); DS rats fed a high-salt diet and treated with perindopril [1mg/kg/day] (group HS-P), and DS rats fed a high-salt diet and treated with candesartan [1mg/kg/day] (group HS-C). At the age of 16 weeks, studies of systemic and cardiac hemodynamics, LV remodeling and LV fibrosis were then evaluated. Compared with the HS group, systolic blood pressure in both the HS-P group and the HS-C group did not significantly decrease and although the transition to systolic heart failure was inhibited, LV hypertrophy did develop in both groups. LV contractile function in both groups improved to almost the same degree as that in the LS group. The effect of perindopril was a fraction greater than that of candesartan (%FS: 49.9 ± 6.8%, 45.7 ± 5.4%, respectively) as opposed to the HS group figures of 33.9 ± 7.0%. In contrast, the 100-day survival rate in the HS-C group significantly improved compared to that in the HS-P group (100% to 40%, respectively). Histological examinations using Azan-Mallory stain demonstrated that that candesartan had more success than perindopril in decreasing LV fibrosis. The study showed that candesartan significantly improved survival rate compared to perindopril in DS hypertensive rats. Candesartan was far more successful than perindopril in decreasing LV fibrosis, which is probably a key factor in improving the survival rate for hypertensive heart failure.

Heat Shock Protein 70 Gene Transfection Protects Mitochondrial and Ventricular Function Against Ischemia-Reperfusion Injury

Background Upregulation of heat shock protein 70 (HSP70) is beneficial in cardioprotection against ischemia-reperfusion injury, but the mechanism of action is unclear. We studied the role of HSP70 overexpression through gene therapy on mitochondrial function and ventricular recovery in a protocol that mimics clinical donor heart preservation. Methods and Results Hemagglutinating virus of Japan (HVJ)-liposome technique was used to transfect isolated rat hearts via intracoronary infusion of either the HSP70 gene (HSP group, n=16) or no gene (CON group, n=16), which was heterotopically transplanted into recipient rats. Four days after surgery, hearts were either perfused on a Langendorff apparatus for 30 minutes at 37°C (preischemia studies [n=8/group]) or perfused for 30 minutes at 37°C, cardioplegically arrested for 4 hours at 4°C, and reperfused for 30 minutes at 37°C (postischemia studies [n=8/group]). Western blotting and immunohistochemistry confirmed HSP70 upregulation in the HSP group. Postischemic mitochondrial respiratory control indices (RCIs) were significantly better preserved in HSP than in CON hearts: NAD + -linked RCI values were 9.54±1.1 versus 10.62±0.46 before ischemia (NS) but 7.98±0.69 versus 1.28±0.15 after ischemia ( P &lt;0.05), and FAD-linked RCI values were 6.87±0.88 versus 6.73±0.93 before ischemia (NS) but 4.26±0.41 versus 1.34±0.13 after ischemia ( P &lt;0.05). Postischemic recovery of mechanical function was greater in HSP than in CON hearts: left ventricular developed pressure recovery was 72.4±6.4% versus 59.7±5.3% ( P &lt;0.05), maximum dP/dt recovery was 77.9±6.6% versus 52.3±5.2% ( P &lt;0.05), and minimum dP/dt recovery was 72.4±7.2% versus 54.8±6.9% ( P &lt;0.05). Creatine kinase release in coronary effluent after reperfusion was 0.20±0.04 versus 0.34±0.06 IU · min −1 · g wet wt −1 ( P &lt;0.05) in HSP versus in CON hearts. Conclusions HSP70 upregulation protects mitochondrial function after ischemia-reperfusion injury; this was associated with improved preservation of ventricular function. Protection of mitochondrial function may be important in the development of future cardioprotective strategies.

Heat shock protein 70 gene transfection protects mitochondrial and ventricular function against ischemia-reperfusion injury.

Upregulation of heat shock protein 70 (HSP70) is beneficial in cardioprotection against ischemia-reperfusion injury, but the mechanism of action is unclear. We studied the role of HSP70 overexpression through gene therapy on mitochondrial function and ventricular recovery in a protocol that mimics clinical donor heart preservation. Hemagglutinating virus of Japan (HVJ)-liposome technique was used to transfect isolated rat hearts via intracoronary infusion of either the HSP70 gene (HSP group, n=16) or no gene (CON group, n=16), which was heterotopically transplanted into recipient rats. Four days after surgery, hearts were either perfused on a Langendorff apparatus for 30 minutes at 37 degrees C (preischemia studies [n=8/group]) or perfused for 30 minutes at 37 degrees C, cardioplegically arrested for 4 hours at 4 degrees C, and reperfused for 30 minutes at 37 degrees C (postischemia studies [n=8/group]). Western blotting and immunohistochemistry confirmed HSP70 upregulation in the HSP group. Postischemic mitochondrial respiratory control indices (RCIs) were significantly better preserved in HSP than in CON hearts: NAD(+)-linked RCI values were 9.54+/-1.1 versus 10.62+/-0.46 before ischemia (NS) but 7.98+/-0.69 versus 1.28+/-0.15 after ischemia (P<0.05), and FAD-linked RCI values were 6.87+/-0.88 versus 6.73+/-0.93 before ischemia (NS) but 4.26+/-0.41 versus 1.34+/-0.13 after ischemia (P<0.05). Postischemic recovery of mechanical function was greater in HSP than in CON hearts: left ventricular developed pressure recovery was 72.4+/-6.4% versus 59.7+/-5.3% (P<0.05), maximum dP/dt recovery was 77.9+/-6.6% versus 52.3+/-5.2% (P<0.05), and minimum dP/dt recovery was 72.4+/-7.2% versus 54.8+/-6.9% (P<0.05). Creatine kinase release in coronary effluent after reperfusion was 0.20+/-0.04 versus 0.34+/-0.06 IU. min(-1). g wet wt(-1) (P<0.05) in HSP versus in CON hearts. HSP70 upregulation protects mitochondrial function after ischemia-reperfusion injury; this was associated with improved preservation of ventricular function. Protection of mitochondrial function may be important in the development of future cardioprotective strategies.

Myocardial self-preservative effect of heat shock protein 70 on an immature lamb heart

Background. Heat shock proteins have been shown to enhance myocardial tolerance of ischemia-reperfusion injury and are induced in the myocardium of many animals by various stressors.Methods. To assess the effects and time course of the inducible form of heat shock protein 70, we raised the rectal temperature of 15 neonatal lambs to 43°C for 15 minutes. At 15, 30, 60, and 120 minutes and 24 hours after heat shock, hearts were subjected to immunoblot analysis for heat shock protein (hsp 72/73). Twenty-four hours after heat shock, neonatal lamb hearts (n = 8) were subjected to 2 hours of cold cardioplegic ischemia (HSP group). Eight neonatal lamb hearts without heat shock served as control. After 60 minutes of reperfusion, left ventricular systolic and diastolic function, coronary blood flow (CBF), myocardial oxygen consumption (MVO2), and lactate levels were measured. Endothelial function was assessed by measuring in situ coronary vascular resistance response to acetylcholine and trinitroglycerine.Results. The HSP group showed a significantly higher recovery of systolic function as well as MVO2, and a lower lactate level compared to the control group at 60 minutes after reperfusion. Recovery of coronary endothelial function was also significantly better in the HSP group than in the control group. Inducible form of HSP 70 was expressed 15 minutes after heat shock and continued to be observed at 24 hours after the stress.Conclusions. Heat shock stress associated with the production of inducible heat shock proteins improved the recovery of ventricular function as well as endothelial function and aerobic metabolism after hypothermic cardioplegic ischemia. Induction of heat shock proteins by any means prior to planned hypothermic ischemia may lead to a new approach for myocardial protection.

Vascular permeability increase and plasma volume loss induced by endotoxin was attenuated by hypertonic saline with or without dextran.

Endotoxin given intravenously is known to cause plasma leakage and subsequent loss of circulating plasma volume. Hypertonic saline resuscitation has been successfully applied in hemorrhagic and traumatic shock, but its application for the treatment or prevention of septic or endotoxin shock is less well studied. Our aim was to investigate the effects of endotoxin on plasma leakage in hamsters when administered in two different ways: applied locally to the hamster cheek pouch microcirculation or systemically by i.v. injection. The cheek pouch was studied by intravital microscopy using FITC-labeled dextran as a tracer of plasma leakage. Escherichia coli lipopolysaccharide (LPS) was continuously added into the superfusion buffer of the cheek pouch preparation during 120 min in two control groups (each n = 6) and two further groups (each n = 6) treated with either hypertonic saline (HS) or hypertonic saline and dextran (HSD). Treatment was given as an i.v. injection 0.35 mL NaCl 7.5%/100 g b.w. during 4 min starting 15 min prior to the start of endotoxin application. Endotoxin caused a reversible increase in the number of postcapillary venular leaks with a maximal response at 70 min after start of endotoxin application. The maximal responses were reduced to 36% in the HS-treated and to 37% in the HSD-treated group in comparison to what was seen in the control groups. In the second part of the study endotoxin was given i.v. 0.3 mg/kg to anesthetized hamsters (n = 41) and arterial blood samples were collected at 0, 60, 120, and 180 min after endotoxin injection for measurement of hematocrit and plasma FITC-dextran concentration. Hamsters were divided into seven groups: untreated control group (n = 6); HSC control group given only an i.v. injection of hypertonic saline (n = 6); LPS group given endotoxin 0.3 mg/kg during 1 min (n = 9); HSp group given hypertonic saline (NaCl 7.5%) 10 min prior to i.v. endotoxin (n = 6); HSa group given hypertonic saline 10 min after i.v. endotoxin (n = 6); HSD group given hypertonic saline with dextran 40, 10 min prior to i.v. endotoxin (n = 6); HSD control group given only i.v. hypertonic saline + dextran and no endotoxin (n = 2). Injection of endotoxin caused a significant increase in hematocrit, which was counteracted by hypertonic saline treatment, with or without dextran, probably due to reduced extravasation of plasma in postcapillary venules.

Intracellular translocation of cellular and heat shock induced proteins upon heat shock in Drosophila Kc cells.

The intracellular distribution of cellular and heat shock induced proteins (hsp) was studied during a heat shock to Drosophila Kc cells. The different hsp's are unequally distributed between the intracellular organelles; while hsp 84 shows an exclusive cytoplasmic location, hsp 70 is equally distributed between the nucleus and the cytoplasm. The low molecular weight (MW) hsp's 22--26 are mainly concentrated in the nuclear fraction. Hsp 34 also shows a preferential nuclear localization but differs from the 22-26 hsp group with regard to its solubility in salt. During the heat shock (HS) treatment, a major preexisting microsomal polypeptide of relative mass (Mr) 45 000 with some properties similar to cytoskeletal proteins is translocated to the nuclear pellet. This translocation does not occur during arsenite treatment which also induces the synthesis of some hsp's. Furthermore, the hsp's synthesized in response to this arsenite treatment do not become associated with the nucleus as during the HS. Consequently the nuclear translocation of the 45 000 Mr protein does not seem to be involved in the induction of the high MW group of hsp's. Furthermore, it suggests that the hsp's which are found in the nuclear pellet upon HS do not have a nuclear localization per se. It rather reflects a temperature-dependent translocation of certain cellular proteins and hsp to the nucleus during a HS.