What is the role of heat shock protein in abdominal organ transplantation?

ABSTRACT Ischemia-reperfusion injury is a pathophysiological event occuring after abdominal organ transplantation, and has a significant influence on prognosis and survival of the graft. It is involved in delaying the primary function or non-functioning of the graft. The objective of this study was to provide information on heat shock protein mechanisms in ischemia-reperfusion injuries in abdominal organ transplantations, and to indicate the possible factors involved that may influence the graft outcome. Several classes of heat shock proteins are part of the ischemia and reperfusion process, both as inflammatory agonists and in protecting the process. Studies involving heat shock proteins enhance knowledge on ischemia-reperfusion injury mitigation processes and the mechanisms involved in the survival of abdominal grafts, and open space to support therapeutic future clinical studies, minimizing ischemia and reperfusion injuries in abdominal organ transplantations. Expression of heat shock proteins is associated with inflammatory manifestations and ischemia-reperfusion injuries in abdominal organ transplantations and may influence graft outcomes.

Effects of melatonin against thioacetamide-induced testicular toxicity in rats / Efectos de la melatonina contra la toxicidad testicular inducida por tioacetamida en ratas

SUMMARY: This study aimed to investigate the changes in testis tissue of thioacetamide-induced rats and the effect of melatonin on these changes. Thirty-five male Wistar Albino rats were divided into five groups. Group I; Control (n=7), Group II; Melatonin (Mel) (10 mg/kg) a single dose (i.p)(n=7), Group III; Thioacetamide (TAA) (300 mg/kg) (i.p) 2 times with 24 hour intervals (n=7), Group IV; TAA (300 mg/kg) was administered at 24-hour intervals, afterwards of 10 mg/kg single dose of Mel (n=7), Group V; Mel was administered 10 mg/kg a single dose 24 hours before the administration of TAA (n=7). Testis was evaluated histologically, immunohistochemically (Heat Shock Proteins (HSP) 70 and 90), blood serum testosterone, total antioxidant status(TAS) and total oxidant status(TOS) in tissue. The tissue sections of Group III decreased seminiferous tubule diameters, and germinal epithelium spills were observed. HSP70 and HSP90 expressions were increased. There wasn't a statistically significant change in testosterone levels among the groups. While TAS levels decreased in Group III compared to control, TOS levels didn't change. HSP70 and HSP90 decreased in groups with Mel-treated. Mel was found to have both protective and therapeutic effects. According to our results, the therapeutic effect of Mel in thioacetamide-induced acute testicular injury is greater than its protective effect.

RESUMEN: Este estudio tuvo como objetivo investigar los cambios en el tejido testicular de ratas inducidas por tioacetamida y el efecto de la melatonina en estos cambios. Treinta y cinco ratas macho Wistar Albino se dividieron en cinco grupos. Grupo I; Control (n = 7), Grupo II; Melatonina (Mel) (10 mg / kg) una dosis única (i.p) (n = 7), Grupo III; Tioacetamida (TAA) (300 mg / kg) (i.p) 2 veces con intervalos de 24 horas (n = 7), Grupo IV; TAA (300 mg / kg) se administró a intervalos de 24 horas, luego de una dosis única de 10 mg / kg de Mel (n = 7), Grupo V; Mel recibió 10 mg / kg de una dosis única 24 horas antes de la administración de TAA (n = 7). Los testículos se evaluaron histológicamente, inmunohistoquímicamente (proteínas de choque térmico (PCT) 70 y 90), testosterona en suero sanguíneo, estado antioxidante total (EAT) y estado oxidante total (EOT) en el tejido. En secciones de tejido del Grupo III se observó disminución de los diámetros de los túbulos seminíferos y derrames en el epitelio germinal. Se aumentaron las expresiones HSP70 y HSP90. No hubo un cambio estadísticamente significativo en los niveles de testosterona entre los grupos. Mientras que los niveles de EAT disminuyeron en el Grupo III en comparación con el control, los niveles de EOT no cambiaron. HSP70 y HSP90 disminuyeron en los grupos tratados con Mel. Se descubrió que Mel tenía efectos protectores y terapéuticos. Según nuestros resultados, el efecto terapéutico de Mel en la lesión testicular aguda inducida por tioacetamida es mayor que su efecto protector.

Immunohistochemical study of amelogenin binding proteins in an amelogenin point mutation mouse / Estudio inmunohistoquímico de proteínas de unión de la amelogenina en un ratón con mutación puntual de amelogenina

Amelogenin is one of the enamel matrices secreted by ameloblasts. A mutation of the amelogenin gene can cause hereditary dental enamel defects known as amelogenesis imperfecta (AI). Since lysosome-associated membrane protein-1 (LAMP-1), -3 (LAMP-3), and 78kDa glucose-related protein (Grp78) were identified as binding proteins of amelogenin, several studies have suggested the involvement of these binding proteins with the cell kinetics of ameloblasts in normal or abnormal conditions. The purpose of this study is to investigate the distribution of these amelogenin binding proteins in the ameloblast cell differentiation of mice with a point mutation of the amelogenin gene (Amelx*). The incisors of Amelx* mice had a white opaque color and the tooth surface was observed to be rough under a scanning electron microscope. Among the sequential ameloblast cell differentiation in the Amelx* mice, the shape of ameloblasts at the transition stage was irregular in comparison to those in wild-type (WT) mice. Immunostaining of Grp78 revealed that the whole cytoplasm of the transition stage ameloblasts was immunopositive for Grp78 antibody, while only the distal part of cell was positive in the WT mice. Furthermore, in the Amelx* mice, the cytoplasm of the transition stage ameloblasts was immunopositive for LAMP-1 and LAMP-3. These results suggest that Amelx* may cause the abnormal distribution of amelogenin binding proteins in the cytoplasm of ameloblasts.

La amelogenina es una de las matrices de esmalte secretadas por los ameloblastos. Una mutación del gen de amelogenina puede causar defectos hereditarios del esmalte dental conocidos como amelogénesis imperfecta (AI). Dado que la proteína de membrana asociada a lisosoma-1 (LAMP-1), -3 (LAMP-3) y la proteína relacionada con la glucosa de 78 kDa (Grp78) se identificaron como proteína de unión a amelogenina, varios estudios han sugerido la participación de estas proteínas con la cinética celular de los ameloblastos en condiciones normales o anormales. El objetivo del estudio fue investigar la distribución de LAMP-1, LAM-3 y Grp78 durante la diferenciación celular de ameloblastos de ratones con una mutación puntual del gen de amelogenina (Amelx*). Los incisivos de los ratones Amelx* presentaron un color blanco opaco y se observó en microscopio electrónico de barrido que la superficie del diente era áspera. La diferenciación celular secuencial y la forma de los ameloblastos en la etapa de transición en los ratones Amelx* fue irregular en comparación con los ratones silvestres (RS). La inmunotinción de Grp78 reveló que todo el citoplasma de los ameloblastos en etapa de transición fue inmunopositivo para el anticuerpo Grp78, mientras que solo la parte distal de la célula fue positiva en los ratones RS. Además, en ratones Amelx*, el citoplasma de los ameloblastos en etapa de transición fue inmunopositivo para LAMP-1 y LAMP-3. Estos resultados sugieren que Amelx* puede causar distribución anormal de proteínas de unión a amelogenina en el citoplasma de los ameloblastos.

Cardiac protein expression patterns are associated with distinct inborn exercise capacity in non-selectively bred rats

In the present study, we successfully demonstrated for the first time the existence of cardiac proteomic differences between non-selectively bred rats with distinct intrinsic exercise capacities. A proteomic approach based on two-dimensional gel electrophoresis coupled to mass spectrometry was used to study the left ventricle (LV) tissue proteome of rats with distinct intrinsic exercise capacity. Low running performance (LRP) and high running performance (HRP) rats were categorized by a treadmill exercise test, according to distance run to exhaustion. The running capacity of HRPs was 3.5-fold greater than LRPs. Protein profiling revealed 29 differences between HRP and LRP rats (15 proteins were identified). We detected alterations in components involved in metabolism, antioxidant and stress response, microfibrillar and cytoskeletal proteins. Contractile proteins were upregulated in the LVs of HRP rats (α-myosin heavy chain-6, myosin light chain-1 and creatine kinase), whereas the LVs of LRP rats exhibited upregulation in proteins associated with stress response (aldehyde dehydrogenase 2, α-crystallin B chain and HSPβ-2). In addition, the cytoskeletal proteins desmin and α-actin were upregulated in LRPs. Taken together, our results suggest that the increased contractile protein levels in HRP rats partly accounted for their improved exercise capacity, and that proteins considered risk factors to the development of cardiovascular disease were expressed in higher amounts in LRP animals.

Proteolytic activity of recombinant DegP from Chromohalobacter salexigens BKL5

Background: DegP is a serine protease that specifically cleaves and refolds unfolding proteins in the periplasmic space of the cells. To date, there is no information regarding DegP from halophilic bacteria. Chromohalobacter salexigens BKL5 is a moderately halophilic bacterium that has the ability to grow in a media containing more than 15% salt. Therefore, the objectives of this work were to clone and overexpress DegP-encoding gene from C. salexigens BKL5 and characterize its biochemical properties. Results: DegP-encoding gene was overexpressed in Escherichia coli BL21(DE3) CodonPlus in an active form. SDS-PAGE analysis showed that the molecular weight of the recombinant DegP was 45 kDa. Size-exclusion chromatography analysis suggested that recombinant DegP was present in two multimeric states, hexameric and dodecameric, with molecular weights of 297.9 and 579.12 kDa, respectively. Both conformations were enzymatically active when casein was used as substrate for enzymatic assay. Circular dichroism analysis showed that recombinant DegP was composed of 0.21­0.29 helical content, which was comparable to the helical content in the crystal structure of E. coli DegP. The basic/acidic residue ratio of recombinant DegP was 0.56, which was slightly higher than that of DegP from extreme halophiles (average, 0.45) but significantly lower than that of DegP from nonhalophiles (average, 0.94). Conclusions: Recombinant DegP from C. salexigens BKL5 showed proteolytic activity when ß-casein was used as a substrate. In silico analysis indicated that recombinant DegP had characteristics similar to those of halophilic proteins depending on its amino acid composition.

Endoplasmic reticulum stress and renal lesion in mice with combination of high-fat diet and streptozotocin-induced diabetes

ABSTRACT PURPOSE : To investigate in the kidney the pathologic changes and expression of GRP78 and CHOP in the Kunming (KM) mice with combination of high-fat diet and streptozotocin-induced diabetes. METHODS : Sixty two male KM mice were randomly divided into a normal control (NC) group (n=20) and a high-fat diet (HFD) group (n=42). After a four-week dietary manipulation, the KM mice in the HFD group were injected intraperitoneally with streptozotocin to induce diabetes. After diabetic models were successfully established, the kidneys were excised and conserved for further test. RESULTS : No significant difference in the body weight was observed after the dietary manipulation (p=0.554). After the streptozotocin was injected, fasting blood glucose levels in the diabetes group (DM) were significantly higher than that in the NC group (p＜0.0001). Glomerular atrophy observed under light microscope in the DM group was more serious compared with the NC group. The expression of GRP78 and CHOP in the kidneys of the mice in the DM group were higher compared with the NC group. CONCLUSION : Renal lesion occurs in the diabetic Kunming mice induced by combination of high-fat diet and low-dose streptozotocin, and endoplasmic reticulum stress and CHOP may contribute to the injury process.

Ellagic acid inhibits proliferation and induces apoptosis in human glioblastoma cells

PURPOSE: To investigate the anticancer activity of ellagic acid (EA) in U251 human glioblastoma cells and its possible molecular mechanism. METHODS: The cells were treated with EA at various concentrations for different time periods. Cell viability and cell proliferation were detected by cell counting kit-8(CCK-8) assay and live/dead assay respectively. Cell apoptosis were measured with Annexin V-FITC/PI double staining method by flow cytometry and Mitochondrial membrane potential assay separately. Cell cycle was measured with PI staining method by flow cytometry. The expressions of Bcl-2, Survivin, XIAP, Caspase-3, Bax, JNK, p-JNK, ERK1/2, p-ERK1/2, p38, p-p38, DR4, DR5, CHOP and GRP78-related proteins were detected by western blot after EA treatment. RESULTS: Cell viability and proliferation of glioblastoma cells treated with EA were significantly lower than the control group. EA caused robust apoptosis of the glioblastoma cells compared to the control group. EA significantly decreased the proportion at G0/G1 phases of cell cycling accompanied by increased populations at S phase in U251 cell lines. And the expressions of anti-apoptotic proteins were dramatically down-regulated. CONCLUSION: Ellagic acid potentially up-regulated DR4, DR5 and MAP kinases (JNK, ERK1/2 and p38). EA also caused significant increase in the expressions of CHOP and GRP78. Our findings suggest that EA would be beneficial for the treatment of glioblastoma.

Mechanisms Involved in Exercise-Induced Cardioprotection: A Systematic Review / Mecanismos Envolvidos na Cardioproteção Induzida pelo Exercício Físico: Uma Revisão Sistemática

Acute myocardial infarction is the leading cause of morbidity and mortality worldwide. Furthermore, research has shown that exercise, in addition to reducing cardiovascular risk factors, can also protect the heart against injury due to ischemia and reperfusion through a direct effect on the myocardium. However, the specific mechanism involved in exerciseinduced cardiac preconditioning is still under debate.

O infarto agudo do miocárdio é a principal causa de mortalidade e de morbidade na população mundial. Por outro lado, pesquisas já demonstraram que o exercício físico, além de reduzir os fatores de risco cardiovascular, também é capaz de promover cardioproteção contra lesões por isquemia e reperfusão, por meio de um efeito direto no miocárdio. No entanto, o mecanismo específico envolvido no pré-condicionamento cardíaco induzido pelo exercício ainda é alvo de discussão.

Knockdown of interleukin-10 induces the redistribution of sigma1-receptor and increases the glutamate-dependent NADPH-oxidase activity in mouse brain neurons

BACKGROUND: In the central nervous system, interleukin-10 (IL-10) provides trophic and survival effects directly on neurons, modulates neurite plasticity, and has a pivotal importance in the neuronal regeneration in neurodegenerative and neuroinflammatory conditions. This cytokine is primarily produced by glial cells and has beneficial effects on the neuronal viability. However, the mechanisms of IL-10-elicited neuroprotection are not clear. RESULTS: Membrane preparations, isolated from wild-type (Wt) and IL-10 knockout (KO) mice brain were used in this study. It has been shown that compared to wild-type mice, in IL-10 KO mice brain, the amount of immunoglobulin binding protein (BiP) is greatly increased, whereas the content of sigma receptor-1 (SigR1) is not changed significantly. Co-immunoprecipitation experiments have shown that the association of SigR1 with small GTPase Rac1 (Ras-related C3 botulinum toxin substrate 1), NR2B subunit of NMDA-receptor (NMDAR) and inositol-3-phosphate receptor (IP3R) is higher in the IL-10 KO mice brain than in the Wt mice brain. Besides, we have found that either glutamate or sigma ligands, separately or together, do not change glutamate-induced NADPH-oxidase (NOX) activity in Wt-type mice brain membrane preparations, whereas in IL-10 KO mice high concentration of glutamate markedly increases the NOX-dependent production of reactive oxygen species (ROS). Glutamate-dependent ROS production was decreased to the normal levels by the action of sigma-agonists. CONCLUSIONS: It has been concluded that IL-10 deprivation, at least in part, can lead to the induction of ER-stress, which causes BiP expression and SigR1 redistribution between components of endoplasmic reticulum (ER) and plasma membrane. Moreover, IL-10 deficiency can change the specific organization of NMDAR, increasing the surface expression of SigR1-sensitive NR2B-containing NMDAR. In these conditions, glutamate-dependent ROS production is greatly increased leading to the initiation of apoptosis. In this circumstances, sigma-ligands could play a preventive role against NMDA receptor-mediated excitotoxicity.

Detección de Helicobacter Pylori en placas de ateroma / Elicobacter Pilori detected in atheroma plague

Introducción: la infección y la inflamación crónica han sido implicadas como agentes etiológicos para la ateraesclerasis (ATE). Varios estudios han relacionado a la infección por H. Pylori (HP) con la EC, especialmente con los linajes mas virulentos (linaje Cag A). Objetivo: demostrar la presencia del HP en placas de endarterectomías, utilizando una técnica Inmunohistoquimica (lHQ) específica que revela una reacción Ag-Ac mediante un cromógeno. Material y Métodos: se estudiaron 34 placas ATE de distintos territorios vasculares. Se fijaron en formol descalcificándolas en ácido fórmico según necesidad. Fueron incluidos en parafina, cortados y coloreados con H-E y técnicas de IHQ específicas para HP. Luego fueron desparafinados y tratados térmicamente con una solución de recuperación antigénica (lnmuno DNA Retriever with Citrate) utilizando olla a presión. La IHQ se efectuó con un sistema de alta sensibilidad Biotina-Estreptavidina-Peroxidasa-DAB). La observación morfológica evaluó células inflamatorias mononucleares y la identificación de la bacteria en la pared o la luz vascular. Resultados: de los 34 casos estudiados, en 14 se pudo identificar el bacilo en sus diferentes formas (41,17%), asociado a signos de inflamación crónica. Conclusión: el HP estuvo presente en un número sustancial de lesiones ATE y se asoció con inflamación. Estudios recientes sugieren que la presencia de Hp, demostrada por técnicas de IHQ, potenciaría los FR para ATE, induciendo una respuesta celular inflamatoria crónica por irritación persistente de la pared arterial.

Introduction: In general, infection and chronic inflammation have be en implied as etiologic agents for atherosclerosis and in particular coronary illness (CI). Several studies have correlated the infection of Helicobacter pylori with CI, especially with virulent strains (lineage Cag A). Objective: Demonstrate the immunohistochemical presence of H. Pylori in atherosclerotic plaques obtained from endarterectomy of different vascular regions. Material and methods: 34 atherosclerotic plaques of different vascular areas were studied, (25 men and 9 women). The tissues were fixed with 10% neutral buffered-formalin and decalcifying in formic acid 5% was used when necessary. The tissue sections were included in paraffin, cut and colored with H&E and subjected to Immunohistochemistry (lHC) of H.Pylori. Briefly, tissues were deparaffinized and thermally treated with a citrate-based solution of antigenic retrieval (lmmunoDNA Retriever with Citrate, BlO SB, Santa Barbara, CA) using a water bath at 95°C for 1 hour. The IHC was conducted using a high sensitivity BiotinStreptavidin-HRP-DAB IHC system (lmmunoDetector HRPIDAB, BlO SB). The microscopic observation evaluated the' presence of mononuclear inflammatory cells and the identification of the bacteria in the wall or the vascular lumen. Results: Of the 34 cases studied 14 were positive, where one could identify the bacillus in their different forms (41, 17%) associated with chronic inflammation.

Expresión de proteínas del choque térmico HSP72 y HSP73 en casos colombianos de Linfoma de Hodgkin positivos y negativos para virus de Epstein Barr / Expression of heat shock proteins HSP72 and HSP73 in Epstein-Barr positive or negative Hodgkin lymphoma

Background: The expression of heat shock proteins (HSP70) in tumor cells or virus infected cells is important for the induction of specific cellular immune response. They are implicated in transport of immunodominants peptides in the endoplasmic reticulum, activation of antigen presenting cells and cross priming of CD8 T cells. Aim: To analyze the expression of HSP70 protein in its constitutive (HSP73) and inducible forms (HSP72) in Hodgkin's lymphoma (HL), infected or not by Epstein Barr virus (EBV) and to assess its relationship with pathological subtype, clinical stages and treatment response. Material and methods: The analysis of HSP73 and HSP72 was done by immunoperoxidase on routinely processed paraffin sections with prior antigen retrieval. Results: Sixty seven cases were studied. The expression of HSP73 and HSP72 was detected in 19.4 and 17.9% of samples respectively. The infiltrating lymphocytes expressed HSP72 in 58% of cases. The pathological subtypes with the higher expression in lymphocytes were mixed cellularity and nodular sclerosis. No differences in HSP70 expression were observed, according to clinical stage, treatment response or the presence of EBV. Conclusions: The expression of HSP72 on lymphocytes suggests that this protein plays an important role in the induction and amplification of anti-tumor immune response (Rev Méd Chile 2003; 131: 1375-81).

Translocation of heat shock proteins into isolated nuclei of Aedes albopictus cells

Labelled heat shock proteins of 82 kDa,70 and a group of low molecular weight are present in the post-mitochondrial supernatant obtained from Aedes albopictus cells.These proteins in the post-mitochondrial extract are transported specifically to the nuclear compartment when incubated with isolated unlabelled nuclei.The relative amounts of each protein transported were 4.7,24.1 and 3,1% for the 82-k-Daand 70-kDa and for the low molecular weght proteins, respectively.Incubation of radiolabelled post-mitochondrial extract with unlabelled nuclei at 30 C or 37 C did not modify the perecentage of proteins translocated