慢性前列腺炎患者精浆中热休克蛋白的表达及临床意义

目的　探讨慢性前列腺炎患者精浆中热休克蛋白 70 (HSP 70 )的表达及临床意义。　方法　采用酶联免疫吸附测定法 (ELISA)对 10例慢性细菌性前列腺炎 (Ⅱ型 )、10例炎症型慢性非细菌性前列腺炎即炎症型慢性骨盆疼痛综合征 (ⅢA 型 )、10例非炎症型慢性非细菌性前列腺炎即非炎症型慢性骨盆疼痛综合征 (ⅢB 型 )患者及 6例健康志愿者精浆中HSP 70含量进行测定 ,并与慢性前列腺炎症状评分 (修订版 )进行相关性研究。　结果　Ⅱ型、ⅢA 型、ⅢB 型慢性前列腺炎患者及正常对照组精浆HSP 70含量分别为 (10 .74± 4 .0 2 )ng/ml、(2 .5 8± 0 .96 )ng/ml、(2 .78± 1.12 )ng/ml及 (6 .5 7±2 .72 )ng/ml。Ⅱ型患者精浆HSP 70含量显著高于正常对照组 ,P <0 .0 1,差异有显著性意义。ⅢA 型与ⅢB 型患者HSP 70含量均显著低于正常对照组 ,P <0 .0 1,差异有显著性意义。而ⅢA 型与ⅢB 型患者HSP 70含量间差异无显著性意义 ,P >0 .0 5。各组精浆HSP 70含量与慢性前列腺炎症状评分间均呈显著的负相关关系 (P <0 .0 1)。　结论　测定精浆中HSP 70的表达对各种类型慢性前列腺炎诊断有一定的临床应用价值 ,并可作为评价慢性前列腺炎病情程度的分子生物学指标。HSP 70在慢性前列腺炎的治疗方面也具有广阔的应用前景。     

Insulin potentiates expression of myocardial heat shock protein 70.

OBJECTIVE: Since insulin stimulates nitric oxide (NO) production and an increase in NO following heat shock is required for myocardial heat shock protein 70 (Hsp70) synthesis, we hypothesized that insulin would enhance myocardial Hsp70 synthesis by augmenting NO signaling. We examined whether a physiologic dose of insulin increased myocardial Hsp70 in unstressed and heat shock treated rats. METHODS: Adult male Sprague-Dawley rats were assigned to groups: (1) control, (2) insulin injected (200 microU/gm body weight), (3) heat shock treated (core body temperature 42 degrees C for 15 min), (4) heat shock and insulin treated, (5) L-nitroarginine methyl ester (L-NAME) and heat shock and insulin treated, (6) sodium nitroprusside (SNP) and heat shock and insulin treated. Six hours later, myocardial Hsp70 content and localization was analyzed. RESULTS: Hsp70 was increased in heat shock treated hearts (120.6+/-16.8 ng/mg protein, P < 0.001) vs. control (12.9+/-2.0 ng/mg protein), or insulin treated hearts (15.5+/-0.83 ng/mg protein). In addition, Hsp70 was increased in the heat shock and insulin treated hearts (164.4+/-7.53 ng/mg protein) compared to control, insulin only (P = 0.001) or heat shock only treated hearts (P = 0.01). L-NAME did not abolish the insulin induced increase in Hsp70 in heat shocked hearts (195.2+/-13.4 ng/mg protein, P = 0.21) and SNP did not further enhance Hsp70 in the insulin and heat shocked group (188.9+/-8.2 ng/mg protein, P = 0.71). Western analysis and confocal microscopy revealed a lowlevel expression of myocardial Hsp70 in response to insulin. Hsp70 was localized primarily in blood vessels after insulin or heat shock treatments. CONCLUSIONS: Insulin caused a low-level expression of myocardial Hsp70 and potentiated Hsp70 synthesis in response to heat shock. The ability of insulin to potentiate Hsp70 after heat shock is independent of NO signaling as it was not altered by either LNAME or SNP pretreatment. Blood vessels appear to be the primary site of Hsp70 after insulin or heat shock treatment.     

热休克蛋白70抑制大鼠颅脑损伤诱生型一氧化氮合酶mRNA的表达

目的 探讨热休克蛋白７０（ＨＳＰ７０）对大鼠感染性脑损伤（感脑）诱生型一氧化氮合酶（ｉＮＯＳ）的表达及一氧化氮（ＮＯ）合成的影响。方法 将大鼠７２只随机分为正常对照组、感染性脑损伤组和热休克处理组,每组又ｈ共３个时间点。采用百日咳菌液通过左颈内动脉注入制成大鼠感染性脑损伤模型,用Ｗｅｓｔｅｒｎ印迹杂交技术检测各组各时间点的ＨＳＰ７０的表达,同时用原位杂交方法检测各组ｉＮＯＳｍＲＮＡ的表达及Ｇｒｉｅｓｓ法测定各组的ＮＯ含量的变化。结果 Ｗｅｓｔｅｒｎ印迹杂交分析结果表明,大鼠感脑各组及正常组有一定量的ＨＳＰ７０表达,而热休克处理组的ＨＳＰ７０的量明显高于感脑组（Ｐ〈０．０１）。原位杂交结果提示ｉＮＯＳ在感脑的大脑皮质神经细胞４、８、２４ｈ开始表达,可见明显的杂交信号,而休克处理组仅有少量的阳性颗粒。ＮＯ含量在感脑     

Heat shock protein response in a prostate tumor model to interstitial thermotherapy: Implications for clinical treatment

Hyperthermia is being utilized individually and in conjunction with other therapies in treating malignant and benign tumors, though few studies have examined cellular effects of elevated temperatures in the prostate model. Highly conserved proteins of the 70 kDa heat shock protein family (HSP 70) are produced in response to environmental stresses, including heat, and are found in all organisms. HSPs are an indicator of cell damage, are associated with thermotolerance, and provide cells with transient resistance to subsequent thermal challenges. Transient thermotolerance is important in the determination of temperature, duration, and sequencing for treatments. This preliminary study analyzes the HSP 70 response of the Dunning R3327 adenocarcinoma model to a single 50°C 1 hr treatment. Elevated HSP levels were found between 10 and 16 hr, returning to baseline by 24 hr. As some fractions of the cells are able to produce HSP 70 following treatment, the data suggest that currently utilized clinical temperatures (42–46°C) administered for 1 hr are inadequate. HSP levels in response to hyperthermia, radiation, and chemotherapy may be useful in finding optimal treatment regimens for prostate cancer. © 1993 wiley-Liss, Inc.     

Correlation of Hsp70-1 and Hsp70-2 gene expression with the degree of graft-versus-host reaction in a rat skin explant model

BACKGROUND: Graft-versus-host disease (GVHD) is the most serious complication after allogeneic hematopoietic stem-cell transplantation. A human skin explant assay has been used to predict the risk of GVHD in patients by histological grading of graft-versus-host reactions (GVHR). New molecular markers of GVHR might help to further increase the predictive value of the assay. METHODS: A rat skin explant assay has been developed to further aid in identifying potential novel molecular markers. RESULTS: In inbred rat strains GVHR were observed in skin explants co-cultured with allogeneic lymphocytes stimulated against minor or major histocompatibility antigens. The histological signs of GVHR were similar to those observed in human skin explant assays and acute GVHD lesions occurring in rats after experimental bone marrow transplantation. Heat shock protein (HSP) 70 has been shown to be expressed during GVHR. We therefore investigated the expression of the three major histocompatibility complex (MHC)-linked HSP70 genes in rat skin explants. The two major stress-inducible genes Hsp70-1 and Hsp70-2 were found to be upregulated in the allogeneic rat skin explant assays. The increase in mRNA correlated with the GVHR grade (I-IV). Interestingly, the expression of the third MHC-linked Hsp70 gene Hsp70-3 was not found to be augmented during GVHR. CONCLUSION: The observed induction of the MHC-encoded Hsp70-1 and Hsp70-2 genes might serve as new markers of GVHR and as potentially novel diagnostic tools for GVHD.     

Heat shock protein expression in the transplanted human kidney

Heat shock proteins (HSPs) have been shown to represent potential target molecules for T-cell-mediated allograft rejection in heart and kidney transplants. In the present study, we therefore investigated the expression of HSP subtypes 60, 72, and 73 in normal kidneys and qualitative and/or quantitative changes in rejected renal allografts. Six normal kidney tissue specimens, three biopsies from patients with minimal change nephritis, as well as 37 biopsies and eight transplant nephrectomy specimens of patients with renal allograft rejection were studied. Type and severity of rejection were assessed according to the Banff classification. Immunohistochemical demonstration of HSP expression was performed using specific monoclonal antibodies after wet autoclave antigen retrieval on sections from either Carnoy-fixed (biopsies) or formalin-fixed (transplant nephrectomies) and paraffin-embedded tissue. The expression was scored in a semiquantitative manner. All three subtypes were found to be constitutively expressed in normal kidney tissue and in noninflammatory minimal change nephritis, albeit with a characteristic compartmental and cellular distribution. Rejection resulted in a higher immunohistochemical scoring for all three HSP subtypes in compartments in which they were normally present; in addition, a de novo expression of HSP60 was found in the vascular compartment and, moreover, infiltrating mononuclear cells were strongly immunoreactive for HSP60 and HSP73. Only quantitative differences were observed for HSP72 immunoreactivity. These results indicate that rejection episodes are paralleled by an increased but differential expression of HSPs in the glomerular, tubular, and vascular compartments of the kidney. This enhancement as well as the de novo appearance of HSP60 on vascular endothelial cells might explain the presence of HSP-reactive T lymphocytes in rejected allografts.     

Heat shock protein expression and injury optimization for laser therapy design

BACKGROUND AND OBJECTIVES: Hyperthermia can induce heat shock protein (HSP) expression in tumor regions where non-lethal temperature elevation occurs, enhancing cell viability and resistance to chemotherapy and radiation treatments typically employed in conjunction with thermal therapy. However, HSP expression control has not been incorporated into current thermal therapy design. Treatment planning models based on achieving the desired post-therapy HSP expression and injury distribution in the tumor and healthy surrounding tissue can enable design of more effective thermal therapies that maximize tumor destruction and minimize healthy tissue injury. STUDY DESIGN/MATERIALS AND METHODS: An optimization algorithm for prostate cancer laser therapy design was integrated into a previously developed treatment planning model, permitting prediction and optimization of the spatial and temporal temperature, HSP expression, and injury distributions in the prostate. This optimization method is based on dosimetry guidelines developed from measured HSP expression kinetics and injury data for normal and cancerous prostate cells and tumors exposed to hyperthermia. RESULTS: The optimization model determines laser parameters (wavelength, power, pulse duration, fiber position, and number of fibers) necessary to satisfy prescribed HSP expression and injury distributions in tumor and healthy tissue. Optimization based on achieving desired injury and HSP expression distributions within the tumor and normal tissue permits more effective tumor destruction and diminished injury to healthy tissue compared to temperature driven optimization strategies. CONCLUSIONS: Utilization of the treatment planning optimization model can permit more effective tumor destruction by mitigating tumor recurrence and resistance to chemotherapy and radiation arising from HSP expression and insufficient injury.     

Heat shock protein 60 expression in patients undergoing cardiac operations

AIM: Cardiomyocytes respond to stress with the expression of different heat shock proteins (HSP). The mitochondrial HSP60 is known to be expressed by various stress factors, including ischemia and reperfusion. The aim of this study was to investigate if HSP60 is increased in human myocardium after cardiac surgery. METHODS: To determine if heat shock protein 60 accumulated in the myocardium of patients undergoing car-diac operations, right atrial samples before and after extracorporeal circulation were excised and immediately frozen in liquid nitrogen. Patients: we obtained 10 sequential right atrial specimens from 5 male patients in sinus rhythm undergoing elective cardiac surgery. Measures: the HSP60 protein level was determined by SDS-PAGE, Western blot and quantified by optical densitometry according to the immunoreactive bands of actin. RESULTS: The HSP60 concentration was unchanged in hearts after a single episode of hypothermic ischemia and reperfusion. Immunoblot analysis demonstrated HSP60 expression in all hearts. There was no correlation with the endurance of cardiopulmonary bypass or reperfusion time. CONCLUSION: These findings indicate that myocardial HSP60 of patients undergoing cardiac operations is not increased after an obligatory period of ischemia, cardioplegic arrest and reperfusion. This might reflect an effective cardioprotection during ECC.     

Heat shock protein expression in cerebral vessels after subarachnoid hemorrhage

OBJECTIVE: The mechanisms of cerebral vasospasm after subarachnoid hemorrhage (SAH) remain controversial. Recent data have implicated two small heat shock proteins (HSPs), namely HSP20 and HSP27, in the regulation of vascular tone. Increases in the phosphorylation of HSP20 are associated with vasorelaxation, and increases in the phosphorylation of HSP27 are associated with impaired vasorelaxation. Therefore, we hypothesized that alterations in the expression and/or phosphorylation of these two small HSPs might play a role in cerebral vasospasm after SAH. METHODS: A rat model of endovascular perforation was used to induce SAH. Middle cerebral arteries were harvested from control animals, sham-treated animals, and animals with SAH, 48 hours after SAH induction. Dose-response curves for endothelium-independent (sodium nitroprusside, 10(-8) to 10(-4) mol/L) and endothelium-dependent (bradykinin, 10(-10) to 10(-5) mol/L) relaxing agents were recorded ex vivo. Physiological responses were correlated with the expression and phosphorylation of HSP20 and HSP27 by using one- and two-dimensional immunoblots. RESULTS: There was impaired endothelium-independent and endothelium-dependent relaxation in cerebral vessels after SAH. These changes were associated with decreased expression of both total and phosphorylated HSP20 and increases in the amount of phosphorylated HSP27. CONCLUSION: In this model, impaired relaxation of cerebral vessels after SAH was associated with increases in the amount of phosphorylated HSP27 and decreases in the expression and phosphorylation of HSP20. These data are consistent with alterations in the expression and phosphorylation of these small HSPs in other models of vasospasm.     

Heat shock protein 27 in chronic allograft nephropathy: a local stress response

BACKGROUND: Heat shock protein (HSP) 27 plays a cytoprotective role through its antioxidant, antiapoptotic, and actin-stabilizing properties during cell stress. The authors hypothesized that HSP27 is involved in chronic allograft nephropathy (CAN), a chronic state of inflammation and stress. METHODS: The authors used the Fisher 344-to-Lewis model of CAN. Transplants were performed in 3-month-old recipient rats. HSP27 mRNA and protein levels were determined using semiquantitative polymerase chain reaction, microarray (stress-toxicity, GEArray) analyses, gene sequencing, immunoblotting, and immunohistochemical analyses at 10 days and 6 months posttransplant. P38 mitogen-activated protein kinase (MAPK), manganese (Mn) superoxide dismutase (SOD), copper-zinc (CuZn) SOD, FasL, Bax, hypoxia-inducible factor (HIF)-1alpha, and CD3 lymphocytes were studied in parallel as selective biomarkers of oxidative stress (OS), apoptosis, hypoxia, and graft-infiltrating immune cells. RESULTS: Six months after transplantation, kidney allografts displayed histologic and functional features of CAN, including tubular atrophy, interstitial fibrosis, glomerulosclerosis, and increased proteinuria and serum creatinine levels. HSP27 mRNA and protein levels in CAN were reduced by 50% and 85%, respectively (P=0.04). Immunohistochemical analyses revealed a "shift" in HSP27 from the medulla to the cortex in allografts with CAN. Bax, phosphorylated p38-MAPK, HIF-1alpha, and MnSOD followed a parallel relocation pattern. CD3 lymphocyte density and tubular FasL expression were also greater in the cortex of allografts with CAN. Time-course analyses revealed that most of these changes were present as early as 10 days posttransplant. CONCLUSIONS: The shift of HSP27 from the medulla to the cortex, combined with greater CD3, p38-MAPK, Bax, FasL, HIF-1alpha, and MnSOD immunoreactivity in this area of the kidney, likely represents an allograft-level response to CAN-related OS-hypoxia.     

Heat shock protein 70 induction and its urinary excretion in a model of acetaminophen nephrotoxicity

Acetaminophen (APAP) is an analgesic–antipyretic drug widely used in children. In the present study, we used an in vivo model of APAP-induced nephrotoxicity in male Wistar rats. We analyzed whether toxic doses of APAP could induce heat shock protein 70 (HSP70) in the kidney and whether HSP70 could be detected in urine. Renal function and histological evaluation of the kidneys were performed at different times after APAP administration (1,000 mg/kg body weight i.p.). Cellular injury was assessed by Triton X-100 solubilization of Na⁺/K⁺ ATPase. Renal and hepatic glutathione levels were also measured. Urinary N-acetyl-β-D glucosaminidase (NAG) excretion increased 4 h after intoxication. At this time, urea and creatinine were at control levels and a slight degree of histological alteration was detected. Kidney microscopic evaluation, Na⁺/K⁺ ATPase solubility, creatinine, and urea levels and NAG excretion did not differ from those of controls 48 h after APAP administration. HSP70 was detected in urine obtained from 4 to 24 h after APAP administration. HSP70 abundance in renal cortex was increased at early time points and 48 h after APAP administration. Urinary HSP70 excretion would be a marker of its renal induction combined with the loss of tubule integrity. NAG would be a suitable early biomarker of APAP-induced nephrotoxicity.     

谷氨酰胺诱导热休克蛋白70表达对高氧肺损伤的干预效果

目的观察谷氨酰胺诱导大鼠肺组织表达热休克蛋白70(HSP70)的作用及HSP70对高氧肺损伤的保护作用。方法健康清洁级雄性Wistar大鼠18只(体重252~286g),随机均分为谷氨酰胺组(G组)和对照组(C组):G组以0.75g/kg谷氨酰胺行腹腔注射,每天注射1次,连续3d;C组大鼠每天腹腔注射同等容积的生理盐水。于注射后第4天每组各取3只大鼠的肺组织,用免疫印迹法(Western blotting)检测HSP70表达,其余12只大鼠全部放入高氧环境(氧浓度95%)中继续喂养,观察在高氧环境下第6天两组大鼠肺组织形态学改变。结果注射谷氨酰胺后第4天,G组大鼠肺组织HSP70蛋白表达水平明显升高,其灰度值为20.34±2.26;C组HSP70蛋白表达水平较低,其灰度值为1.82±0.67,G组明显高于C组(P<0.05)。G组第6天肺部炎症表现为肺泡内极少量红细胞渗出;而C组病理改变为肺泡大小不等,肺泡腔变大,肺泡壁变薄,有肺大泡形成,肺泡内有出血和炎症细胞浸润。结论大鼠腹腔注射谷氨酰胺可明显增加肺组织HSP70表达,HSP70可减轻高氧肺损伤时肺组织炎性改变。     

热休克大鼠缺血肾脏诱导型热休克蛋白的表达

目的研究在热休克预处理大鼠整体后,大鼠肾脏在冷缺血环境下诱导型HSP70的表达.方法用热休克方法和药物川芎嗪分别对动物整体进行预处理后,通过组织切片和免疫组化方法观察诱导型HSP70的表达情况,利用计算机图像灰度分析法测定不同缺血时间下大鼠肾脏组织的灰度值,以间接量化反映诱导型HSP70的表达.结果在非缺血状态下处理组的灰度值较对照组有明显增强(P<0.001);在不同的缺血时间下,肾脏诱导型HSP70的表达有一定规律性对照组在缺血30min时表达最高;热休克两组表达更高,时间持续到60min;川芎嗪组较热休克组次之,但高于对照组.结论热休克、缺血和川芎嗪均可以诱导肾脏组织诱导型HSP70的表达,并且在时间和量上有一定规律性.     

Evaluation of heat shock protein 70 expression in renal parenchyma subjected to shockwave lithotripsy

PURPOSE: In this experimental study in a rabbit model, renal parenchymal heat shock protein 70 (hsp70) levels were assessed in an attempt to evaluate the traumatic effects of high-energy shockwaves (HESW), which have been found to induce transient ischemia during the procedure. MATERIALS AND METHODS: Eighteen white New Zealand rabbits, each weighing 3 to 5 kg, were included in the study. The animals were divided into three groups, and various numbers of shockwaves (1000, 1500, or 2000) were applied to the same kidney of all animals under fluoroscopic localization with a Stonelith V5 lithotripter. Untreated contralateral kidneys were evaluated as controls. Following HESW application, the treated and untreated kidneys were removed surgically after 24 hours or 7 days. Tissue hsp70 levels were assessed by an immunohistochemistry method. RESULTS: During early follow-up (24 hours), both treated and untreated kidneys demonstrated moderate to severe hsp70 positivity. The number of positive tubules increased as the number of shockwaves increased, and positivity became more evident, possibly because of a higher degree of tissue damage. Contralateral kidneys demonstrated a limited degree of hsp70 positivity, although it was not as evident as in the treated kidneys. Assessment of tissue hsp70 levels during late follow-up (7 days) demonstrated moderate or limited degrees of positivity in the treated kidneys. Limited or no positivity could be demonstrated in the untreated kidneys during this period. CONCLUSIONS: Taking the known traumatic effects of HESW and the results of this study into account, the increasing positivity of hsp70 in parallel with the increasing number of shockwaves led us to think about a possible limited degree of ischemia induced by this procedure, as the traumatic effects of HESW were pronounced, as judged by tissue hsp70 positivity.     

丙酮酸乙酯调节脓毒症小鼠肝细胞HSP70表达的研究

目的研究丙酮酸乙酯（EP）对脓毒症小鼠肝细胞热休克蛋白70（HSPT0）的调节作用。方法制作小鼠盲肠结扎穿孔模型（CLP），应用丙酮酸乙酯林格氏液（REPS）与乳酸钠林格氏液（RLS）对小鼠进行液体复苏，60只小鼠分3组，各20只：假手术组、CLP模型＋REPS复苏组、CLP模型＋RLS复苏组，检测肝组织丙二醛（MDA）及肝细胞HSP70的表达。结果脓毒症小鼠较假手术组MDA浓度增高，P〈0．01。EP显著提高脓毒症小鼠肝组织的抗氧化能力，REPS组肝组织MDA浓度低于RLS组【（48．18±598）μmol／g．prot vs（78．34±11．16）μmol／gprot，P〈0．01]；REPS组小鼠肝细胞HSPT0表达较RLS组增高[（28．76±5．69）vs（20．04±4．93），P〈0．051。HSP70表达与MDA值呈负相关（r=-0．733，P〈0．01）。结论EP具有的抗氧化作用能提高脓毒症小鼠肝细胞的HSPT0表达。     

Absence of correlation between cytolytic T lymphocytes and lethal murine graft-versus-host disease in response to minor histocompatibility antigens

Lethally irradiated mice were transplanted with bone marrow plus spleen cells from H-2 identical donor mice. Five of the six recipient strains developed lethal GVHD, but the sixth strain did not develop any signs of GVHD. Spleen cells from all six transplanted strains were cytotoxic to recipient strain target cells in a short-term chromium release assay. The cytotoxic spleen cells were antigen-specific for recipient strain target cells, Thy-1+ and Lyt-2+--and some were also Lyt-1+. These data demonstrate for the first time that the presence of antirecipient cytotoxic T lymphocytes (CTL) does not correlate with lethal GVHD. Although CTL may contribute to the pathogenesis of GVHD in response to minor histocompatibility antigens, they do not appear to be the primary effector mechanism.