Walter Klimecki

PMID: 12573264;Abstract:

TLR9 is a mammalian Toll-like receptor homologue that appears to function as an innate immune pattern recognition protein for motifs that are far more common in bacterial than in mammalian DNA. The gene was sequenced in 71 subjects from three self-identified U.S. ethnic groups to identify single-nucleotide polymorphisms (SNPs). A total of 20 SNPs were found of which only 20% were in the public dbSNP database. Four SNPs were relatively common in all three ethnic samples. Using these four SNPs, seven distinct haplotypes were statistically inferred, of which four accounted for 75% or more chromosomes. These four haplotypes could be distinguished from each other by the alleles of two SNPs (-1237 and 2848). Five exploratory nested case-control disease-association studies (asthma, DVT, MI, and COPD in European Americans and asthma in African Americans) were performed by genotyping DNA collected from four ongoing cohort studies. There was evidence suggesting increased risk for asthma with a C allele at -1237 (odds ratio 1.85, 95%CI 1.05 to 3.25) among European Americans (genotypes available from 67 cases and 152 controls). No other significant disease associations were detected. Replication of this finding in other, larger samples is needed. This study suggests that there is substantial diversity in human TLR9, possibly associated with asthma in Europeans but not African Americans. No association was detected with three other diseases potentially related to innate immunity. © 2003 Elsevier Science (USA). All rights reserved.

Autophagy is a critical cellular process orchestrating the lysosomal degradation of cellular components in order to maintain cellular homeostasis and respond to cellular stress. A growing research effort over the last decade has proven autophagy to be essential for constitutive protein and organelle turnover, for embryonic/neonatal survival and for cell survival during conditions of environmental stress. Emphasizing its biological importance, dysfunctional autophagy contributes to a diverse set of human diseases. Cellular stress induced by xenobiotic exposure typifies environmental stress, and can result in the induction of autophagy as a cytoprotective mechanism. An increasing number of xenobiotics are notable for their ability to modulate the induction or the rate of autophagy. The role of autophagy in normal cellular homeostasis, the intricate relationship between cellular stress and the induction of autophagy, and the identification of specific xenobiotics capable of modulating autophagy, point to the importance of the autophagic process in toxicology. This review will summarize the importance of autophagy and its role in cellular response to stress, including examples in which consideration of autophagy has contributed to a more complete understanding of toxicant-perturbed systems.

PMID: 12406828;Abstract:

Toll-like receptor 4 (TLR4) is the principal receptor for bacterial endotoxin recognition, and functional variants in the gene confer endotoxin-hyporesponsiveness in humans. Furthermore, there is evidence that endotoxin exposure during early life is protective against the development of atopy and asthma, although this relationship remains poorly understood. It is therefore possible that genetic variation in the TLR4 locus contributes to asthma susceptibility. In this study we characterize the genetic diversity in the TLR4 locus and test for association between the common genetic variants and asthma-related phenotypes. In a cohort of 90 ethnically diverse subjects, we resequenced the TLR4 locus and identified a total of 29 single nucleotide polymorphisms. We assessed five common polymorphisms for evidence of association with asthma in two large family-based cohorts: a heterogeneous North American cohort (589 families), and a more homogenous population from northeastern Quebec, Canada (167 families). Using the transmission-disequilibrium test, we found no evidence of association for any of the polymorphisms tested, including two functional variants. Furthermore, we found no evidence for association between the TLR4 variants and four quantitative intermediate asthma- and atopy-related phenotypes. Based on these results, we found no evidence that genetic variation in TLR4 contributes to asthma susceptibility.

SOCS-1 is a critical regulator of multiple signaling pathways, including those activated by cytokines that regulate Ig H chain class switching to IgE. Analysis of mice with mutations in the SOCS-1 gene demonstrated that IgE levels increase with loss of SOCS-1 alleles. This suggested that overall SOCS-1 acts as an inhibitor of IgE expression in vivo. A genetic association study was performed in 474 children enrolled in the Tucson Children's Respiratory Study to determine if genetic variation in the SOCS-1 locus correlates with altered levels of IgE. Carriers of the C-allele for a novel, 3' genomic single nucleotide polymorphism (SNP) in the SOCS-1 gene (SOCS1+1125G > C; rs33932899) were found to have significantly lower levels of serum IgE compared with those of homozygotes for the G-allele. Analysis demonstrated that the SOCS1+1125G > C SNP was in complete linkage disequilibrium with an SNP at position SOCS1-820G > T (rs33977706) of the SOCS-1 promoter. Carriers of the T-allele at the SOCS1-820G > T were also found to be associated with the decreased IgE. The promoter SNP increased transcriptional activity of the SOCS-1 promoter in reporter assays and human B cells. Consistent with this observation, the presence of this polymorphism within the promoter abolished binding of yin yang-1, which is identified as a negative regulator of SOCS-1 transcriptional activity. These data suggest that genetic variation in the SOCS-1 promoter may affect IgE production.

Arsenic is a widespread environmental toxicant with a diverse array of molecular targets and associated diseases, making the identification of the critical mechanisms and pathways of arsenic-induced cytotoxicity a challenge. In a variety of experimental models, over a range of arsenic exposure levels, apoptosis is a commonly identified arsenic-induced cytotoxic pathway. Human lymphoblastoid cell lines (LCL) have been used as a model system in arsenic toxicology for many years, but the exact mechanism of arsenic-induced cytotoxicity in LCL is still unknown. We investigated the cytotoxicity of sodium arsenite in LCL 18564 using a set of complementary markers for cell death pathways. Markers indicative of apoptosis (phosphatidylserine externalization, PARP cleavage, and sensitivity to caspase inhibition) were uniformly negative in arsenite exposed cells. Interestingly, electron microscopy, acidic vesicle fluorescence, and expression of LC3 in LCL 18564 identified autophagy as an arsenite-induced process that was associated with cytotoxicity. Autophagy, a cellular programmed response that is associated with both cellular stress adaptation as well as cell death appears to be the predominant process in LCL cytotoxicity induced by arsenite. It is unclear, however, whether LCL autophagy is an effector mechanism of arsenite cytotoxicity or alternatively a cellular compensatory mechanism. The ability of arsenite to induce autophagy in lymphoblastoid cell lines introduces a potentially novel mechanistic explanation of the well-characterized in vitro and in vivo toxicity of arsenic to lymphoid cells.