Protective effect of ginsenoside Rg_1 on hypoxia/reoxygenation injury and its mechanism / 中国中药杂志

This project aimed to explore the protective effect of ginsenoside Rg_1 on hypoxia/reoxygenation(H/R)-induced H9 c2 cardiomyocyte injury and its underlying signaling pathway. The H/R model of H9 c2 cardiomyocytes was established and then the cells were divided into different treatment groups. CCK-8(cell counting kit-8) was used to detect the activity of cardiomyocytes; Brdu assay was used to detect the proliferation of H9 c2 cells; the caspase-3 activity was tested, and then the protein expression was assessed by Western blot. Flow cytometry was used to evaluate the apoptosis level of cardiomyocytes. Ginsenoside Rg_1 inhibited H/R-induced cardiomyocyte apoptosis and caspase-3 activity, promoted nuclear transcription of nuclear factor erythroid-2 related factor 2(Nrf2), and enhanced the expression of the downstream heme oxygenase-1(HO-1). Ginsenoside Rg_1 could increase Nrf2 nuclear transcription and HO-1 expression with the increase of concentration(10, 20, 40, 60 μmol·L~(-1)). However, the protective effect of ginsenoside Rg_1 on cardiomyocytes was significantly weakened after the transfection of Nrf2-siRNA. Ginsenoside Rg_1 could protect cardiomyocytes by activating the Nrf2/HO-1 pathway.

Interaction between NOS3 and HMOX1 on antihypertensive drug responsiveness in preeclampsia / Interação entre os genes NOS3 e HMOX1 na resposta à terapia antihipertensiva em pré-eclâmpsia

Abstract Objective We examined the interaction of polymorphisms in the genes heme oxygenase- 1 (HMOX1) and nitric oxide synthase (NOS3) in patients with preeclampsia (PE) as well as the responsiveness to methyldopa and to total antihypertensive therapy. Methods The genes HMOX1 (rs2071746, A/T) and NOS3 (rs1799983, G/T) were genotyped using TaqMan allele discrimination assays (Applied Biosystems, Foster City, CA, USA ), and the levels of enzyme heme oxygenase-1 (HO-1) were measured using enzyme-linked immunosorbent assay (ELISA). Results We found interactions between genotypes of the HMOX-1 and NOS3 genes and responsiveness tomethyldopa and that PE genotyped as AT presents lower levels of protein HO-1 compared with AA. Conclusion We found interactions between the HMOX-1 and NOS3 genes and responsiveness to methyldopa and that the HMOX1 polymorphism affects the levels of enzyme HO-1 in responsiveness to methyldopa and to total antihypertensive therapy. These data suggest impact of the combination of these two polymorphisms on antihypertensive responsiveness in PE.

Resumo Objetivo Examinamos a interação dos polimorfismos nos genes heme oxigenase-1 (HMOX1) eóxido nítrico sintase (NOS3) empacientes compré-eclâmpsia (PE)bem como as capacidades de resposta à metildopa e à terapia anti-hipertensiva. Métodos Os polimorfismos nos genes HMOX1 (rs2071746, A/T) e NOS3 (rs1799983, G/T) foram genotipados usando TaqMan allele discrimination assays (Applied Biosystems, Foster City, CA, EUA), e os níveis da enzima heme oxigenase-1 (HO-1) foram medidos por enzyme-linked immunosorbent assay (ELISA). Resultados Foram encontradas interações entre os genótipos da HMOX-1 e NOS3 e responsividade à metildopa, e que pacientes genotipados como AT apresentam níveis mais baixos de proteína HO-1 em comparação com o genótipo AA. Conclusão Foram encontradas interações entre os genes HMOX-1 e NOS3 e responsividade à metildopa e que o polimorfismo localizado no gene HMOX1 afeta os níveis de enzima HO-1 na resposta à metildopa e à terapia anti-hipertensiva. Esses dados sugerem o impacto da combinação desses dois polimorfismos na resposta antihipertensiva na PE.

Bucillamine prevents cisplatin-induced ototoxicity through induction of glutathione and antioxidant genes

Bucillamine is used for the treatment of rheumatoid arthritis. This study investigated the protective effects of bucillamine against cisplatin-induced damage in auditory cells, the organ of Corti from postnatal rats (P2) and adult Balb/C mice. Cisplatin increases the catalytic activity of caspase-3 and caspase-8 proteases and the production of free radicals, which were significantly suppressed by pretreatment with bucillamine. Bucillamine induces the intranuclear translocation of Nrf2 and thereby increases the expression of gamma-glutamylcysteine synthetase (gamma-GCS) and glutathione synthetase (GSS), which further induces intracellular antioxidant glutathione (GSH), heme oxygenase 1 (HO-1) and superoxide dismutase 2 (SOD2). However, knockdown studies of HO-1 and SOD2 suggest that the protective effect of bucillamine against cisplatin is independent of the enzymatic activity of HO-1 and SOD. Furthermore, pretreatment with bucillamine protects sensory hair cells on organ of Corti explants from cisplatin-induced cytotoxicity concomitantly with inhibition of caspase-3 activation. The auditory-brainstem-evoked response of cisplatin-injected mice shows marked increases in hearing threshold shifts, which was markedly suppressed by pretreatment with bucillamine in vivo. Taken together, bucillamine protects sensory hair cells from cisplatin through a scavenging effect on itself, as well as the induction of intracellular GSH.

HMOX1 Gene Promoter Polymorphism is Not Associated with Coronary Artery Disease in Koreans

BACKGROUND: The heme oxygenase-1 gene (HMOX1) promoter polymorphisms modulate its transcription in response to oxidative stress. This study screened for HMOX1 polymorphisms and investigated the association between HMOX1 polymorphisms and coronary artery disease (CAD) in the Korean population. METHODS: The study population consisted of patients with CAD with obstructive lesions (n=110), CAD with minimal or no lesions (n=40), and controls (n=107). Thirty-nine patients with CAD with obstructive lesions underwent follow-up coronary angiography after six months for the presence of restenosis. The 5'-flanking region containing (GT)n repeats of the HMOX1 gene was analyzed by PCR. RESULTS: The numbers of (GT)n repeats in the HMOX1 promoter showed a bimodal distribution. The alleles were divided into two subclasses, S25 and L25, depending on whether there were less than or equal to and more than 25 (GT)n repeats, respectively. The allele and genotype frequencies among groups were statistically not different. More subjects in the S25-carrier group had the low risk levels of high sensitivity C-reactive protein (hsCRP) for the CAD than those in the non-S25 carrier group (P=0.034). Multivariate logistic regression analysis revealed that the genotypes of (GT)n repeats were not related to CAD status. The restenosis group in the coronary angiography follow-up did not show any significant difference in HMOX1 genotype frequency. CONCLUSIONS: The HMOX1 genotypes were not found to be associated with CAD, but the short allele carrier group contained more individuals with hsCRP values reflecting low risk of cardiovascular disease in the Korean population.

Effect of dexamethasone on unfolded protein response genes (MTJ1, Grp78, Grp94, CHOP, HMOX-1) in HEp2 cell line.

The endoplasmic reticulum (ER) is related to the various signal routes that are activated in unfolded protein response (UPR). The Grp78, Grp94, CHOP, MTJ1 and HMOX1 genes expressions demonstrate UPR activity. In this study, we investigated the UPR gene expressions in larynx epidermoid carcinoma (HEp2) to which dexamethasone (dex) was applied. HEp2 cells were administered for 48 h with different combinations using 0.1 μM and 1 μM dex, 1 mM phenyl butyric acid (PBA) and 100 ng/ml lipopolysaccharide (LPS). The Grp78, Grp94, CHOP, MTJ1 and HMOX1 genes expression was determined using quantitative RT-PCR. The Grp78, MTJ1 and HMOX1 gene expression increased with the administration of 1 µM dex. CHOP expression, on the other hand, decreased with 0.1 µM dex. When dex was combined with LPS, nearly all gene expressions decreased. The increase in Grp78, Grp94, HMOX1 and MTJ1 gene expression was greater in groups in which dex was administered in combination with PBA than in groups in which dex was administered alone. Dex in low dose (0.1 μM) caused a decrease in CHOP expression in HEp2 cells and an increase in Grp78 expression, in particular. The changes in UPR genes expressions may lead to the extended survival of the cells.

Enhancement of parthenolide-induced apoptosis by a PKC-alpha inhibition through heme oxygenase-1 blockage in cholangiocarcinoma cells

Cholangiocarcinoma (CC) is a chemoresistant intrahepatic bile duct carcinoma with a poor prognosis. The aims of this study were to identify molecular pathways that enhance sesquiterpene lactone parthenolide (PTL)-induced anticancer effects on CC cells. The effects of PTL on apoptosis and hemoxygenase-1 (HO-1) induction were examined in CC cell lines. The enhancement of PTL-mediated apoptosis by modulation of HO-1 expression and the mechanisms involved were also examined in an in vitro cell system. Low PTL concentrations (5 to 10 micrometer) led to Nrf2-dependent HO-1 induction, which attenuated the apoptogenic effect of PTL in Choi-CK and SCK cells. PTL-mediated apoptosis was enhanced by the protein kinase C-alpha inhibitor Ro317549 (Ro) through inhibition of expression and nuclear translocation of Nrf2, resulting in blockage of HO-1 expression. Finally, HO-1 silencing resulted in enhancement of apoptotic cell death in CC cells. The combination of PTL and Ro efficiently improved tumor growth inhibition compared to treatment with either agent alone in an in vivo subcutaneous tumor model. In conclusion, the modulation of HO-1 expression substantially improved the anticancer effect of PTL. The combination of PTL and Ro could prove to be a valuable chemotherapeutic strategy for CC.

The Heme Oxygenase-1 Genotype is a Risk Factor to Renal Impairment of IgA Nephropathy at Diagnosis, Which is a Strong Predictor of Mortality

The induction of heme oxygenase-1 (HO-1) ameliorates oxidative stress and inflammatory process, which play important roles in IgA nephropathy. We hypothesized length polymorphism in the promoter region of the HO-1 gene, which is related to the level of gene transcription, is associated with disease severity of IgA nephropathy. The subjects comprised 916 patients with IgA nephropathy and gene data. Renal impairment was defined as an estimated glomerular filtration rate less than 60 mL/min/1.73 m(2) at diagnosis. The short (S: 28) (GT) repeats in the HO-1 gene was determined. The frequencies of S/S, S/M, M/M, S/L, L/M, and L/L genotypes were 7.2%, 6.9%, 3.1%, 30.8%, 22.7%, and 29.4%, respectively. The baseline characteristics were not different. In the S/S genotypic group, the renal impairment rate was 18.2%, which was lower than 32.2% in the group with M/M, L/M, or L/L genotype. The odds ratio of renal impairment in S/S genotype, compared to that in M/M, L/M, or L/L genotype, was 0.216 (95% confidence interval, 0.060-0.774, p=0.019). The HO-1 gene promoter length polymorphism was related to the renal impairment of IgA nephropathy at diagnosis, which is an important risk factor for mortality in IgA nephropathy patients.

Curcumin reduces cold storage-induced damage in human cardiac myoblasts

Curcumin is a polyphenolic compound possessing interesting anti-inflammatory and antioxidant properties and has the ability to induce the defensive protein heme oxygenase-1 (HO-1). The objective of this study was to investigate whether curcumin protects against cold storage-mediated damage of human adult atrial myoblast cells (Girardi cells) and to assess the potential involvement of HO-1 in this process. Girardi cells were exposed to either normothermic or hypothermic conditions in Celsior preservation solution in the presence or absence of curcumin. HO-1 protein expression and heme oxygenase activity as well as cellular damage were assessed after cold storage or cold storage followed by re-warming. In additional experiments, an inhibitor of heme oxygenase activity (tin protoporphyrin IX, micrometer) or siRNA for HO-1 were used to investigate the participation of HO-1 as a mediator of curcumin- induced effects. Treatment with curcumin produced a marked induction of cardiac HO-1 in normothermic condition but cells were less responsive to the polyphenolic compound at low temperature. Cold storage-induced damage was markedly reduced in the presence of curcumin and HO-1 contributed to some extent to this effect. Thus, curcumin added to Celsior preservation solution effectively prevents the damage caused by cold- storage; this effect involves the protective enzyme HO-1 but also other not yet identified mechanisms.

Heme oxygenase 1 overexpression increases iron fluxes in Caco-2 cells

Heme oxygenase-1 is a microsomal enzyme that, when induced by stress, protects the cells from oxidative injury. Heme oxygenase-1 participates in the cleavage of the heme ring producing biliverdin, CO and ferrous Fe. The released Fe becomes part of intracellular Fe pool and can be stored in ferritin or released by an iron exporter. The mechanism by which heme enters cells is not completely understood, although it had been suggested that it might be internalized by an endocytosis process. In this study, we expressed a full-length Heme oxygenase-1 cDNA in Caco-2 cells and measured intracellular iron content, heme-iron uptake and transport and immunolocalization of heme oxygenase-1 in these cells. We found that heme oxygenase-1 expressing cells showed increased apical heme iron uptake and transepithelial transport when compared to control cells. These results suggested that heme oxygenase-1 mediates heme iron influx and efflux in intestinal cells.