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During hypoxic incubation (1% O2) of PC12 cells, the PLD activity was transiently increased within 12h, followed by a gradual decrease. In the in vitro assay, the increased PLD activity was independent of GTPgammaS required for PLD1 or of oleic acid for PLD(OA), suggesting the activation of PLD2. The level of PLD2 protein showed no change up to 12h but a gradual decrease after 24 h. Pretreatment of cells with S. chromofuscus PLD resulted in inhibition of hypoxia-induced apoptotic cell death. In contrast, 1-butanol, but not 2-butanol, potentiated cell death. Moreover, the number of apoptotic cells significantly reduced in PC12 cells over-expressing PLD2. These results raise the possibility that PLD2 activation may play an anti-apoptotic role in hypoxia-induced cell death.

The present study was designed to elucidate the relationship between p53 and ceramide, both of which are involved in apoptotic signaling. Treatment of human glioma cells with etoposide caused apoptosis only in cells expressing functional p53. p53 activation was followed by the formation of reactive oxygen species (ROS), superoxide anion (O2-*) measured by hydroethidium oxidation into ethidium and hydrogen peroxide (H2O2) measured by oxidation of 2',7'-dichlorofluorescin (DCFH) into 2',7'-dichlorofluorescein (DCF), which was accompanied with ceramide generation through the activation of neutral, but not acid, sphingomyelinase. Superoxide dismutase (SOD), a selective antioxidant for O2-*, had no effects on p53 expression but inhibited ceramide generation and apoptotic cell death caused by etoposide. However, catalase, a specific antioxidant for H2O2, only weakly inhibited and sodium formate, a hydroxyl radical (* OH) scavenger, unaffected etoposide-induced apoptosis. Like etoposide-induced cell death, treatment of glioma cells with the O2-*-releasing agent, pyrogallol, induced typical apoptosis and ceramide generation even in the presence of catalase. In contrast, human glioma cells lacking functional p53, either due to mutation or the expression of E6 protein of human papillomavirus, were highly resistant to etoposide and exhibited no significant change in the ceramide level. Moreover, expression of functional p53 protein in glioma cells expressing mutant p53 using a temperature-sensitive human p53(Val138) induced ceramide accumulation by the activation of neutral sphingomyelinase which was dependent on the generation of O2-*. Taken together, these results suggest that p53 may modulate ceramide generation by activation of neutral sphingomyelinase through the formation of O2-*, but not its downstream compounds H2O2 or * OH.

Reduced glutathione (GSH) and N-acetylcysteine (NAC), but not other antioxidative or reducing agents, were found to inhibit cell death, both apoptosis and necrosis, induced by hypoxia in naive and nerve growth factor-differentiated PC12 cells. The level of intracellular total GSH decreased time-dependently during hypoxia, but exogenously added GSH prevented such a decrease in GSH. Pretreatment of cells with exogenous GSH or NAC resulted in inhibition of both neutral sphingomyelinase (SMase) activation and ceramide formation during hypoxia. In the in vitro assay system, neutral SMase activity was inhibited dose-dependently by GSH and NAC. Activation of caspase-3 induced by hypoxia was also inhibited by either GSH or NAC. NAC but not GSH inhibited caspase-3 activation induced by C2-ceramide. These results suggest that GSH protects cells from hypoxic injury by direct inhibition of neutral SMase activity and ceramide formation, resulting in inhibition of caspase-3 activation, and that NAC exerts an additional inhibitory effect(s) downstream of ceramide.

A 61-year-old female presented with a unique case of moyamoya disease associated with pulmonary sarcoidosis. She was admitted for sudden onset of left temporalgia with episode of numbness on face, tongue, and upper extremity on the right side. The next morning, she had symptoms of Gerstmann syndrome and her ability to speak was disturbed. Her medical history included radical resection of lung cancer on the right side. She had no symptoms of pulmonary sarcoidosis. Neuroimaging showed an infarction in the left occipital lobe. Angiography showed occlusions of the bilateral internal carotid arteries at the supraclinoid portions. Subsequently, a left superficial temporal artery-middle cerebral artery anastomosis with encephalo-myo-synangiosis was performed. Ninety-three days after admission, she suddenly developed dyspnea which resulted in death 3 hours later. Autopsy findings showed typical epithelioid granulomas of sarcoid type in the lymph nodes of the peribronchus, lung, and liver. Thrombotic emboli were found in the bilateral pulmonary arteries, and marked fibrous intimal thickening in the bilateral internal carotid arteries. Immunological reaction with inflammatory events may cause pathological changes in patients with moyamoya disease or sarcoidosis. The co-incidence in this case suggests that some common inflammatory events may be involved in the pathogenesis of these diseases.

Aim To evaluate the humoral immune response to HLA antigens in Brazilian renal transplant candidates. Methods The humoral immune response to HLA antigens was studied in 269 renal transplant candidates from the north/northwest of Parana state, Brazil. HLA typing was performed by the polymerase chain reaction sequence-specific oligonucleotide method (PCR-SSO), combined with Luminex technology using the LABType SSO kit (One Lambda, Inc., Canoga Park, CA, USA). Panel reactive antibodies (PRA) and HLA-specific antibodies determination were performed using the LABScreen (Luminex technology) LS1PRA and LS2PRA kits (One Lambda, Inc.). The Fisher’s exact test and Student’s t-test were used to compare the demographic characteristics and potential risk factors for anti-HLA antibody production (pregnancies, blood transfusions and previous transplants) between PRA-positive and PRA-negative groups. Results One hundred and eighty two (67.7%) patients had positive PRA. Potential risk factors for anti-HLA antibody production showed no significant differences between PRA-positive and PRA-negative groups. Only gender was statistically different between these groups, showing that female group was the most sensitized (p Conclusions The data of this study allowed the knowledge of HLA antibodies profile in renal transplant candidates from the Parana State (Brazil), showing high sensitization to HLA antigens. Further studies are needed to evaluate the influence of alloreactivity and HLA polymorphisms in the end-stage renal disease and the evolution of transplantation.

O objetivo deste estudo foi analisar a prevalencia de dialise em mulheres no estado do Parana, municipio de Maringa e regiao metropolitana. Estudo quantitativo, descritivo, de corte transversal. Foi estudada a prevalencia de dialise em mulheres no estado do Parana, municipio de Maringa e regiao metropolitana, no periodo de 1999 a 2007, atraves do Sistema de Informacoes Ambulatoriais do Sistema Unico de Saude (SIA/SUS), no programa do Departamento de Informatica do Sistema Unico de Saude (DATASUS). Houve aumento da taxa de prevalencia de dialise em mulheres no estado do Parana, municipio de Maringa e regiao metropolitana no periodo de 1999 a 2007. No estado do Parana, houve um crescimento de 1,71 mulheres em dialise por 100.000 habitantes do sexo feminino anualmente (p?0,05), as faixas etarias 30 a 59 anos e 60 anos e mais se revelaram como fatores de risco para realizar dialise no sexo feminino. No municipio de Maringa e regiao metropolitana, houve um crescimento de 0,88 de mulheres em dialise por 100.000 habitantes do sexo feminino anualmente (p?0,05), a faixa etaria 60 anos e mais se revelou como fator de risco para realizar dialise no sexo feminino.

Aim To analyse the distribution of HLA class I (A and B) and HLA class II (DRB1) alleles of chronic renal patients and the association with negative and positive PRA, based in sensitization events. Methods One hundred fifty chronic kidney patients from southern region of Brazil, were subdivided into 4 groups, according to the patients’ PRA results, regardless of the occurrence or not of one of the sensitiveness events, such as pregnancy (P), transfusion (S) or transplantation (T), characteristics grouped and identified as PST: Group 1, PRA = 0 (n = 79); Group 2, PRA > 0 (n = 71); Group 3, PRA between 0 and 50% (n = 43); Group 4, PRA > 50% (n = 28). Control group comprised 71 nonsensitized chronic kidney patients (PRA = 0), classified as positive for any of the PST events. HLA genotyping was undertaken by the RSSO-LABType methodology. PRA were obtained by LABScreen methodology. Fisher’s exact test and Odds Ratio (OR) with 95% confidence interval (CI) were used for statistical analysis. Further, p rates were significant when lower than 0.05. Results No significant differences with regard to loci HLA-A, HLA-B and HLA-DRB1 were found in Group 1 when compared to control group. No significant differences with regard to loci HLA-A and HLA-DRB1 were found in Groups 2 and 3. However, allele HLA-B ∗ 08 had a significant negative association in Group 2 (p = 0.0258; OR = 0.27; 95%IC = 0.09-0.83) and in Group 3 (p = 0.0334; OR = 0.22; 95%IC = 0.05-0.98) when compared to control group. Significant positive association in Group 4 was found for alleles HLA-A ∗ 26 (p= 0.0101; OR = 4.57; 95%IC = 1.42-14.64), HLA-B ∗ 58 (p = 0.0074; OR = 13.82; 95%IC = 1.58- 121.17) and HLA-B ∗ 13 (p = 0.0218; OR = 17.97; 95%IC= 0.91-353.64). No significant differences were found within the HLA-DRB1 alleles when compared to control group. Conclusions There are HLA alleles that influence chronic kidney patients to sensitization of HLA molecules. HLA Class I alleles seem to be more involved to susceptibility and/or resistance to the development of anti-HLA antibodies.

Aim The aim of this study was to evaluate the frequencies of the erythrocyte phenotypes ABO, Rh and Duffy in CKD patients in the southern region of Brazil. Methods Samples from CKD patients underwent direct hemaglutination methods to define ABO and Rh phenotypes and agglutination in gel columns for Duffy phenotypes. Pearson’s χ 2 test was employed for statistical analysis and Odds Ratio (OR), confidence interval (CI) at 95% and error α equal to 5% were calculated. Data from other studies with voluntary healthy blood and bone marrow donors were used for the comparison of analysis. Results Representatives of all erythrocyte phenotypes ABO were found among the 231 patients under analysis. No statistically significant difference was reported in the general frequencies ( χ 2 = 3.504; p = 0.320) and among the four phenotypes when they were compared with healthy volunteer blood donors. With regard to the Rh phenotype, 89.2% of patients were Rh positive. No statistically significant difference was reported in the frequency of Rh erythrocyte phenotypes in chronic renal patients and in bone marrow volunteer donors. Phenotypes Fy(a+b+) and Fy(a+b−) were found respectively in 39.0% and 25.1% of kidney patients under analysis. When data on bone marrow volunteer donors of the same region were compared, rates were statistically significant, or rather, the phenotype Fy(a+b+) with OR = 0.691 (IC 0.4975-0.9612) and p = 0.0343 and phenotype Fy(a+b−) with OR = 2.347 (IC 1.543-3.571) and p = 0.0001. Conclusions We conclude that Fy(a+b−) phenotype is associated with increased risk for chronic kidney disease.