Investigation on the relationship between violent death, cocaine abuse and single nucleotide polymorphisms / Estudo da relação entre morte violenta, uso de cocaína e polimorfismos de nucleotídeo único

Violence is a dreadful phenomenon spread throughout the world, resulting in unfortunate events that can ultimately cause death. It is known that some countries play a much worrying role in this scenario than others. Brazil is one of them. The present study has focused on identifying the use of cocaine within 105 postmortem cases arriving at the Institute of Legal Medicine of São Paulo (IML-SP) through analytic toxicological methods and latter applying genetic testing to see whether the presence of certain single nucleotide polymorphisms (SNPs) is more predominant within users rather than non-users, which would help to better understand one's susceptibility to abuse the drug. Both blood and hair samples have been analysed through ultra-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry (UPLC-ESI-MS/MS) in order to distinguish between recent or chronic cocaine use among violent individuals whose violence has ultimately leaded to their death. Two dilute-and-shoot methods have been validated and used for this purpose, and the final residue was analysed through the UPLC-ESIMS/ MS system. From the 105 postmortem cases, a rather high proportion of cocaine and its metabolites was found. A chronic use of the drug was denoted in 53% of the cases, which were positive for cocaine and benzoylecgonine, followed by 43% for norcocaine, 40% for cocaethylene and 13% for anhydroecgonine methyl ester, in hair. As for blood, reflecting the use of cocaine prior to death, 51% of the cases have shown to be positive for benzoylecgonine, followed by 41% for cocaine, 23% for cocaethylene and 20% for norcocaine. These findings suggest a probable association between the use of the drug and risky/violent behaviours. Genetic wise, a significant difference has been observed for SNP rs4263329 from the BCHE gene in its dominant model, with higher frequencies of the genotypes A/G and G/G seen in cocaine users rather than non-users (OR=8.91; 95%CI=1.58-50.21; ρ=0.01). Likewise, also SNP rs6280 from the DRD3> gene presented a significant association in both its additive and dominant model, suggesting that the C allele may be playing a role in cocaine use as both genotypes T/C and C/C were significantly more frequent in users than non-users. This association was not lost when adjusted for covariants using logistic regression (OR=4.96; 95%CI=1.07; ρ=0.04). Finally, a statistically significant association (ρ = 0.003) was also encountered among individuals with both A/G and G/G genotypes within SNP rs4263329 and the use of cocaine HCl (f(A/G+G/G)=44.7%) versus crack-cocaine (f(A/G+G/G)=7.7%) and nonusers (f(A/G+G/G)=16.2%). In conclusion, this study has found significant associations within two SNPs related to cocaine use, however, due to several inherent limitations, these must be confirmed by further studies with a higher number of subjects and within a more controlled setting. Definite assumptions may not be made at this point and future researches are to be conducted

A violência é um fenômeno aterrador espalhado por todo o mundo, resultando em eventos que podem, em última instância, causar a morte. Sabe-se que, em alguns países esse cenário é mais preocupante que em outros. O Brasil é um deles. O presente estudo teve como objetivo identificar o uso de cocaína em 105 casos postmortem provenientes do Instituto de Medicina Legal de São Paulo (IML-SP) por meio de métodos toxicológicos analíticos e posterior aplicação de testes genéticos para verificar se a presença de determinados polimorfismos de nucleotídeo único (SNPs) é mais predominante dentro dos usuários do que dos não usuários, o que explicaria uma possível suscetibilidade de um indivíduo ao abuso da droga. Amostras de sangue e cabelo foram analisadas através de cromatografia líquida de ultra-eficiência acoplada a espectrometria de massas e ionização por electrospray (UPLC-ESI-MS/MS) para distinguir entre uso recente ou crônico de cocaína entre indivíduos violentos cuja violência levou à sua morte. Para tal, dois métodos de extração baseados na técnica de "dilute-and-shoot" foram validados e utilizados para esse fim, e o resíduo final foi analisado através de um sistema UPLC-ESI-MS/MS. Dos 105 casos postmortem, foi encontrada uma proporção significativa de cocaína e seus produtos de biotransformação. O uso crônico da droga foi denotado em 53% dos casos, sendo estes positivos para cocaína e benzoilecgonina, seguidos de 43% para norcocaína, 40% para cocaetileno e 13% para anidroecgonina metil éster, no cabelo. Quanto ao sangue, refletindo o uso de cocaína antes da morte, 51% dos casos mostraram-se positivos para benzoilecgonina, seguido de 41% para cocaína, 23% para cocaetileno e 20% para norcocaína. Esses dados corroboram a hipótese provável da relação entre o uso da droga e comportamentos de risco/violentos. Quanto à genética, uma diferença significativa foi observada para o SNP rs4263329 do gene BCHE em seu modelo dominante, com maiores frequências dos genótipos A/G e G/G vistos em usuários de cocaína ao contrário de não usuários (OR=8,91; 95%IC=1,58-50,21; ρ=0,01). Da mesma forma, também o SNP rs6280 do gene DRD3 apresentou uma associação significativa tanto no seu modelo aditivo quanto dominante, sugerindo que o alelo C pode estar desempenhando um papel no uso de cocaína, pois ambos os genótipos T/C e C/C foram significativamente mais frequentes nos usuários do que não usuários. Essa associação não foi perdida quando ajustada para co-variáveis usando regressão logística (OR=4,96; 95%IC=1,07; ρ=0,04). Finalmente, uma associação estatisticamente significativa (ρ=0,003) também foi encontrada entre indivíduos com ambos os genótipos A/G e G/G dentro do SNP rs4263329 e o uso de cocaína HCl (f(A/G + G/G)=44,7%) versus crack (f(A/G + G/G)=7,7%) e não usuários (f(A/G + G/G)=16,2%). Em conclusão, este estudo encontrou associações significativas em dois SNPs relacionados ao uso de cocaína, no entanto, devido a várias limitações inerentes, estas devem ser confirmadas por mais estudos com um maior número de indivíduos e dentro de um cenário mais controlado. Hipóteses definitivas não poderão ser feitas neste momento e futuras pesquisas devem ser conduzidas

Pharmacomicrobiomics: a novel route towards personalized medicine?

Inter-individual heterogeneity in drug response is a serious problem that affects the patient's wellbeing and poses enormous clinical and financial burdens on a societal level. Pharmacogenomics has been at the forefront of research into the impact of individual genetic background on drug response variability or drug toxicity, and recently the gut microbiome, which has also been called the second genome, has been recognized as an important player in this respect. Moreover, the microbiome is a very attractive target for improving drug efficacy and safety due to the opportunities to manipulate its composition. Pharmacomicrobiomics is an emerging field that investigates the interplay of microbiome variation and drugs response and disposition (absorption, distribution, metabolism and excretion). In this review, we provide a historical overview and examine current state-of-the-art knowledge on the complex interactions between gut microbiome, host and drugs. We argue that combining pharmacogenomics and pharmacomicrobiomics will provide an important foundation for making major advances in personalized medicine.

De novo transcriptome assembly, functional annotation and differential gene expression analysis of juvenile and adult E. fetida, a model oligochaete used in ecotoxicological studies

BACKGROUND: Earthworms are sensitive to toxic chemicals present in the soil and so are useful indicator organisms for soil health. Eisenia fetida are commonly used in ecotoxicological studies; therefore the assembly of a baseline transcriptome is important for subsequent analyses exploring the impact of toxin exposure on genome wide gene expression. RESULTS: This paper reports on the de novo transcriptome assembly of E. fetida using Trinity, a freely available software tool. Trinotate was used to carry out functional annotation of the Trinity generated transcriptome file and the transdecoder generated peptide sequence file along with BLASTX, BLASTP and HMMER searches and were loaded into a Sqlite3 database. To identify differentially expressed transcripts; each of the original sequence files were aligned to the de novo assembled transcriptome using Bowtie and then RSEM was used to estimate expression values based on the alignment. EdgeR was used to calculate differential expression between the two conditions, with an FDR corrected P value cut off of 0.001, this returned six significantly differentially expressed genes. Initial BLASTX hits of these putative genes included hits with annelid ferritin and lysozyme proteins, as well as fungal NADH cytochrome b5 reductase and senescence associated proteins. At a cut off of P = 0.01 there were a further 26 differentially expressed genes. CONCLUSION: These data have been made publicly available, and to our knowledge represent the most comprehensive available transcriptome for E. fetida assembled from RNA sequencing data. This provides important groundwork for subsequent ecotoxicogenomic studies exploring the impact of the environment on global gene expression in E. fetida and other earthworm species.

Expression Levels of GABA-A Receptor Subunit Alpha 3, Gabra3 and Lipoprotein Lipase, Lpl Are Associated with the Susceptibility to Acetaminophen-Induced Hepatotoxicity

Drug-induced liver injury (DILI) is the serious and fatal drug-associated adverse effect, but its incidence is very low and individual variation in severity is substantial. Acetaminophen (APAP)-induced liver injury accounts for >50% of reported DILI cases but little is known for the cause of individual variations in the severity. Intrinsic genetic variation is considered a key element but the identity of the genes was not well-established. Here, pre-biopsy method and microarray technique was applied to uncover the key genes for APAP-induced liver injury in mice, and a cause and effect experiment employing quantitative real-time PCR was conducted to confirm the correlation between the uncovered genes and APAP-induced hepatotoxicity. We identified the innately and differentially expressed genes of mice susceptible to APAP-induced hepatotoxicity in the pre-biopsied liver tissue before APAP treatment through microarray analysis of the global gene expression profiles (Affymetrix GeneChip® Mouse Gene 1.0 ST for 28,853 genes). Expression of 16 genes including Gdap10, Lpl, Gabra3 and Ccrn4l were significantly different (t-test: FDR <10%) more than 1.5 fold in the susceptible animals than resistant. To confirm the association with the susceptibility to APAP-induced hepatotoxicity, another set of animals were measured for the expression level of selected 4 genes (higher two and lower two genes) in the liver pre-biopsy and their sensitivity to APAP-induced hepatotoxicity was evaluated by post hoc. Notably, the expressions of Gabra3 and Lpl were significantly correlated with the severity of liver injury (p<0.05) demonstrating that these genes may be linked to the susceptibility to APAP-induced hepatotoxicity.

Predictive and Prognostic Biomarkers of Respiratory Diseases due to Particulate Matter Exposure

Air pollution is getting severe and concerns about its toxicity effects on airway and lung disease are also increasing. Particulate matter (PM) is major component of air pollutant. It causes respiratory diseases, such as asthma, chronic obstructive pulmonary disease, lung cancer, and so on. PM particles enter the airway and lung by inhalation, causing damages to them. Especially, PM2.5 can penetrate into the alveolus and pass to the systemic circulation. It can affect the cardiopulmonary system and cause cardiopulmonary disorders. In this review, we focused on PM-inducing toxicity mechanisms in the framework of oxidative stress, inflammation, and epigenetic changes. We also reviewed its correlation with respiratory diseases. In addition, we reviewed biomarkers related to PM-induced respiratory diseases. These biomarkers might be used for disease prediction and early diagnosis. With recent trend of using genomic analysis tools in the field of toxicogenomics, respiratory disease biomarkers associated with PM will be continuously investigated. Effective biomarkers derived from earlier studies and further studies might be utilized to reduce respiratory diseases.

Toxicogenomics and its application in safety evaluation of traditional Chinese medicine / 中国中药杂志

Toxicogenomics (TGx) refers to a set of technologies that assess genome-wide responses after toxic agent exposure. Altered gene expression patterns that are caused by specific exposures reveal how toxicants may disrupt cellular processes and lead to side effects. Development and application of " omics" technology facilitate the toxicogenomic research which sharing and interpretation of the enormous amount of biological information generated in toxicologic field. In recent years TGx has been widely valued and successfully applied as an effective research tool to evaluate the toxic effects of traditional Chinese medicine (TCM). Here we reviewed current progress in the field of TGx and focused on its application in traditional Chinese medicine safety evaluation, especially in revealing the mechanism, finding potential toxic biomarkers and studying compatibility detoxification of TCM.

Etanol e isopropanol: genotoxicidad y teratogénesis evaluada aplicando el modelo de Drosophila melanogaster / Ethanol and isopropanol: genotoxicity and teratogenicity evaluated using Drosophila melanogaster as a model system

El etanol y el isopropanol son, de los alcoholes alifáticos de cadena corta, los más frecuentemente asociados a la actividad humana tanto a nivel industrial como en el entorno doméstico. En este trabajo se presentan los principales hallazgos reportados en la literatura para ensayos de genotoxicidad y teratogénesis en modelos experimentales de distinto nivel de complejidad, con especial énfasis en Drosophila melanogaster. El metabolismo de estos alcoholes es semejante en Drosophila y en humanos por lo cual la mosca es un buen modelo in vivo para la evaluación de sus potenciales efectos tóxicos, genotóxicos y teratogénicos.

Ethanol and isopropanol are two of the short chain aliphatic alcohols more frequently associated to the human environment, both in the industrial and domestic conditions. The aim of this work was to present the main findings reported in the literature about their genotoxicity and teratogenicity in experimental models of different level of complexity, with special emphasis in Drosophila melanogaster. Taking into account that the metabolism of both alcohols in Drosophila and humans is similar, the fly is a good model for the evaluation of their potentially toxic, genotoxic and teratogenic effects.

Natural occurrence of nivalenol and mycotoxigenic potential of Fusarium graminearum strains in wheat affected by head blight in Argentina

The principal agents of Fusarium head blight in the main cropping area of Argentina were investigated in heavily infected samples. The ability of the isolates to produce trichothecenes was determined by GC and HPLC. Fusarium graminearum was the predominant species and of 33 isolates, 10 produced deoxinivalenol (DON) (0.1- 29 mg kg-1), 13 produced both deoxinivalenol (1.0- 708 mg kg-1) and nivalenol (0.1- 6.2mg kg-1), 12 produced 3-acetyldeoxinivalenol (0.1- 14 mg kg-1), 13 produced 15-acetyldeoxinivalenol (0.1- 1.9 mg kg-1), 10 produced Fusarenone X (0.1- 2.4 mg kg-1) and 7 produced zearalenone (0.1- 0.6 mg kg-1). These results suggest that F. graminearum strains isolated from the wheat growing regions in Argentina belong to DON chemotype. Although some strains produced both deoxinivalenol and nivalenol, nivalenol was produced in lower levels. The natural occurrence of nivalenol in wheat affected by head-blight collected in the main production area during two years (2001-2002) was also determined. From 19 samples 13 were contaminated with deoxinivalenol in a range of 0.3 to 70 mg kg-1and 2 samples with both deoxinivalenol (7.5 and 6.7 mg kg-1) and nivalenol (0.05 and 0.1 mg kg-1), respectively. This is the first report of natural occurrence of nivalenol in wheat cultivate in Argentina.

O principal causador de giberela no trigo na Argentina e sua capacidade de produzir tricotecenos foram estudados por GC e HPLC em amostras altamente infectadas. A espécie predominante foi Fusarium graminearum, sendo que de um total de 33 isolados, 10 produziram deoxinivalenol (0,1-29 mg kg -1), 13 produziram deoxinivalenol (1,0-708 mg kg-1) e nivalenol (0,1-6,2 mg kg-1), 12 produziram 3-acetildeoxinivalenol (0,1-14 mg kg-1), 13 produziram 15-acetildeoxinivalenol (0,1-1,9 mg kg-1), 10 produziram fusarenona X (0,1- 2,4 mg kg-1) e 7 produziram zearalenona (0,1- 0,6 mg kg-1). Esses resultados sugerem que as cepas de F. graminearum isoladas de trigo cultivado na Argentina pertencem ao quimiotipo DON. Embora algumas cepas tenham produzido tanto DON quanto NIV, NIV foi produzido em quantidade inferior ao DON. A ocorrência natural de nivalenol em trigo afetado pela giberela coletado na principal área de produção durante dois anos (2001-2002) foi também determinada. De 19 amostras, 13 estavam contaminadas com deoxinivalenol na faixa de 0,3 a 70 mg kg-1 e 2amostras continham tanto deoxinivalenol (7,5 e 6,7 mg kg-1) quanto nivalenol (0,05 e 0,1 mg kg-1), respectivamente. Esse é o primeiro relato da ocorrência de nivalenol em trigo cultivado na Argentina.

Molecular markers of heavy metal toxicity--a new paradigm for health risk assessment.

Last decade has witnessed increased interest in studies dealing with molecular markers of health and disease expression of genes. Specific toxicant "signatures" have been detected using genome base technologies such as microarrays. Further toxins have been classified on the basis of these signatures. Knowledge on these signatures has helped in the identification of novel drug candidates. This review discusses the gene expression studies recently made on arsenic, cadmium, mercury, chromium, lead, copper, nickel, manganese, and other essential elements. Toxicogenomics standards and their organizations have also been briefly described. Although this information can not be considered as complete, recent reports from different laboratories on bacteria, fish, laboratory animals and humans have been summarized. It is expected that toxicogenomics data presented in this review will be helpful in planning and excretion of human health risk assessment programs.

The prospect of application of toxicogenetics/pharmcogenetics theory and methods in forensic practice / 法医学杂志

Individual response to drugs, toxicants, environmental chemicals and allergens varies with genotype. Some respond well to these substances without significant consequences, while others may respond strongly with severe consequences and even death. Toxicogenetics and toxicogenomics as well as pharmacogenetics explain the genetic basis for the variations of individual response to toxicants by sequencing the human genome and large-scale identification of genome polymorphism. The new disciplines will provide a new route for forensic specialists to determine the cause of death.

Gene Expression Profiling of 6-MP (6-mercaptopurine) in Liver

The KFDA (Korea Food & Drug Administration) has performed a collaborative toxicogenomics project since 2003. Its aim is to construct a toxicology database of 12 compounds administered to mice at initial phase. We chose 6-MP (6-mercaptopurine) which has been used in the treatment of childhood leukemia. It was administered at low (0.224 mg/kg) and at high (2.24 mg/kg) dose (5 mice per group) intraperitonealy to the postnatal 6 weeks mice, then the serum and liver were collected at the indicated time (6, 24 and 72 h) after scarification. Serum biochemical markers for liver toxicity were measured and histopathologic studies also were carried out. The gene expression profiling was carried out by using Applied Biosystems 1700 Full Genome Expression Mouse. By self-organization maps (SOM), we identified groups with unique gene expression patterns, some of them are supposed to be related to 6-MP induced toxicity, including lipid metabolism abnormality, inflammatory response, oxidative stress, ATP depletion and cell death. The potential toxic effects appearing as gene expression changes are dependent of the time of 6-MP but independent of the dosage of it. This study would contribute to establishment of international database as well as national one about hepatotoxicity.

XPERNATO-TOX: an Integrated Toxicogenomics Knowledgebase

Toxicogenomics combines transcriptome, proteome and metabolome profiling with conventional toxicology to investigate the interaction between biological molecules and toxicant or environmental stress in disease caution. Toxicogenomics faces the problems of comparison and integration across different sources of data. Cause of unusual characteristics of toxicogenomic data, researcher should be assisted by data analysis and annotation for getting meaningful information. There are already existing repositories which claim to stand for toxicogenomics database. However, those just contain limited abilities for toxicogenomic research. For supporting toxicologist who comes up against toxicogenomic data flood, now we propose novel toxicogenomics knowledgebase system, XPERANTO-TOX. XPERANTO-TOX is an integrated system for toxicogenomic data management and analysis. It is composed of three distinct but closely connected parts. Firstly, Data Storage System is for reposit many kinds of '-omics' data and conventional toxicology data. Secondly, Data Analysis System consists of analytical modules for integrated toxicogenomics data. At last, Data Annotation System is for giving extensive insight of data to researcher.

GraPT: Genomic InteRpreter about Predictive Toxicology

Toxicogenomics has recently emerged in the field of toxicology and the DNA microarray technique has become common strategy for predictive toxicology which studies molecular mechanism caused by exposure of chemical or environmental stress. Although microarray experiment offers extensive genomic information to the researchers, yet high dimensional characteristic of the data often makes it hard to extract meaningful result. Therefore we developed toxicant enrichment analysis similar to the common enrichment approach. We also developed web-based system graPT to enable considerable prediction of toxic endpoints of experimental chemical.

El salto de la doble hélice. Consecuencias del Proyecto Genoma Humano en la medicina del siglo XXI (Segunda parte) / The leap of the double helix. Consequences of the Human Genome Project in medicine of the 21st century (Part II

El Proyecto Genoma Humano, iniciado en octubre de 1990, ha permitido desentrañar, 12 años después, la secuencia nucleotídica del ADN humano. Este hecho ha producido un avance singular en la medicina moderna, posibilitando a través de la detección de las variaciones nucleotídicas en la secuencia del ADN, el desarrollo de estudios genéticos, la determinación de pronósticos y guías terapéuticas con fármacos, y el desarrollo de nuevas drogas gracias al avance de la farmacogenómica. Todo esto permitiría, en un futuro cercano, la predicción de respuestas a maniobras terapéuticas en los procedimientos de anestesia, cuidados críticos y tratamiento del dolor. Este desarrollo introduce también problemas éticos, específicamente en los campos de la terapia génica y la clonación.

Current Progress of Next Generation Battery of Toxicology-Cellular and Molecular Toxicology, and Toxicogenomics

No abstract available.