Induction of complete and mosaic sex-linked recessive lethal mutations by cigarette smoke filtrate in Drosophila melanogaster.

The genotoxic effects of cigarette smoke filtrate (SF) on the germ-line stages were examined in Drosophila melanogaster using the sex-linked recessive lethal test, which detects a broad spectrum of genetic alterations and proved to show correlations between mutagenicity and carcinogenicity of the tested chemicals. SF was extracted from fiberglass filter cartridges; each used in smoking 15 cigarettes. The proper SF concentrations (0.2 µL) in 0.45% NaCl saline were injected intraperitoneally in 2- to 3-day-old wild-type males, alongside with controls injected with 0.2 µL of saline. The genotoxicity effects of SF were examined in all spermatogenesis stages of treated males. Results showed that SF was toxic with an median lethal dose value of approximately 0.2% and induced significant sterility effects. The mutagenicity of SF (0.2%) was significantly stage specific and induced complete sex-linked recessive lethal mutations in the broods representing the spermatocytes and late and early spermatogonia, and induced mosaic mutations in the untreated progeny in the brood representing late spermatogonia. These results indicated, for the first time, that SF induces mosaic mutations, which could result from DNA instabilities and labile permutations that can be replicated and passed to future generations before being fixed into mutations in the untreated progeny of treated males, or originating from mutations that result in increasing hyperplasia of the gonad that subsequently produce the actual mutations in later cell cycles. Such delayed mutagenic effects of SF indicated that SF and, consequently, cigarette smoking have much greater genotoxicity than what was previously predicted.

Effect of methotrexate on the survival of human lymphocyte cultures carrying MTHFR 677 (C>T) and MTHFR 1298 (A>C) mutations.

The correlations between the presence of MTHFR 677 (C>T) and MTHFR 1298 (A>C) mutations in human lymphocytes and the sensitivity of lymphocytes to methotrexate (MTX) were examined in cultures derived from 82 unrelated women, genotyped for these mutations by polymerase chain reaction-restriction fragment length polymorphism. Lymphocytes heterozygous for the mutant allele, MTHFR 677T, were significantly more sensitive to methotrexate than those carrying the homozygous wild-type allele, MTHFR677C, and those carrying either the mutant or the wild-type alleles in the polymorphic MTHFR 1298 site. In addition, the lymphocyte cultures carrying the mutant MTHFR 1298C allele were not different in their sensitivity to MTX from those cultures carrying the wild-type allele, MTHFR 1298A. This demonstrated that the polymorphic site MTHFR C677, but not MTHFR1298, could be considered as a useful pharmacogenetic determinant in planning and designing the effective personal MTX chemotherapeutic doses and regimes.

Associations of variants in MTHFR and MTRR genes with male infertility in the Jordanian population.

Folate pathway is expected to play an important role in spermatogenesis since it is involved in DNA synthesis, repair and methylation. The purpose of this study was to examine the association between male infertility and the MTHFR (C677T and A1298C) and MTRR (A66G) polymorphisms. A group of 300 males was recruited in this study from different Jordanian infertility clinics. Of these, 150 cases of infertile men that included oligozoospermia cases (n=45), severe oligozoospermia (n=71) and azoospermia (n=34) were studied. The other 150 males were age matched fertile controls. Genotyping of MTHFR and MTRR polymorphisms was performed using PCR-RFLP technique. The results showed an association between MTHFR 677TT genotype and male infertility (P<0.05). However, the distribution of MTHFR A1298C and MTRR A66G genotypes were not different between the fertile and infertile groups (P>0.05). In addition, none of the examined polymorphisms was related to any of the semen parameters in the infertile group. In conclusion, this study showed that MTHFR C677T polymorphism is associated with male infertility in Jordanians.

Assessment of DNA damage using chromosomal aberrations assay in lymphocytes of waterpipe smokers.

OBJECTIVES: The aim of this study was to investigate the genotoxicity of waterpipe smoking in the lymphocytes of waterpipe smokers using chromosomal aberrations (CAs) assay. MATERIALS AND METHODS: Fifty waterpipe smokers and 18 healthy non-smokers volunteered to participate in the study. Additionally, 18 heavy cigarette smokers were recruited for comparison. Chromosomal aberrations (CAs) assay was used to evaluate DNA damage in the lymphocytes. RESULTS: The results showed that similarly to cigarette smoking, waterpipe smoking significantly increased the frequencies of CAs (p < 0.01). In addition, the frequencies of CAs increased with more waterpipe use. CONCLUSIONS: Waterpipe smoking causes DNA damage to lymphocytes and the damage increases with more waterpipe use.

Assessment of genotoxicity of waterpipe and cigarette smoking in lymphocytes using the sister-chromatid exchange assay: a comparative study.

Tobacco smoking is a major world health problem. Recently, waterpipe smoking has become more popular in many countries. Although the genotoxicity associated with cigarette smoking has been extensively investigated, studies evaluating such toxicity in waterpipe users are still lacking. In this study, we examined the genotoxicity of waterpipe smoking in lymphocytes compared with the genotoxicity of cigarette smoking. Genotoxicity was evaluated using the sister chromatid exchanges (SCEs) assay. Fifty waterpipe smokers and 18 healthy nonsmokers participated in this study. Additionally, 18 heavy cigarette smokers (CS) were recruited for comparison. The results show that waterpipe smoking and cigarette smoking significantly increase the frequencies of SCEs (P < 0.01) compared with those of nonsmokers, indicating the genotoxic effect of tobacco smoking. In addition, frequencies of SCEs were significantly higher among waterpipe smokers compared with CS (P < 0.01), indicating that waterpipe smoking is more genotoxic than cigarette smoking. Moreover, the frequency of SCEs increased with the extent of waterpipe use. In conclusion, waterpipe smoking is genotoxic to lymphocytes and the magnitude of its genotoxicity is higher than that induced by regular cigarette smoking.

Ancient founder mutation is responsible for Imerslund-Gräsbeck Syndrome among diverse ethnicities.

BACKGROUND: Imerslund-Gräsbeck syndrome (IGS) was described just over 50 years ago by Olga Imerslund and Ralph Gräsbeck and colleagues. IGS is caused by specific malabsorption of cobalamin (Cbl) due to bi-allelic mutations in either the cubilin gene (CUBN) or the human amnionless homolog (AMN). Mutations in the two genes are commonly seen in founder populations or in societies with a high degree of consanguineous marriages. One particular mutation in AMN, c.208-2A>G, causing an out-of-frame loss of exon 4 in the mRNA, is responsible for some 15% of IGS cases globally. We present evidence that this founder mutation causes a substantial percentage of cases among diverse ethnicities and that the mutation is as old as human civilization. METHODS: Partial genotyping indicated a founder event but its presence in diverse peoples of Arabic, Turkish, Jewish, and Hispanic ancestry suggested that the mutation might be recurrent. We therefore studied the flanking sequence spanning 3.5 Mb to elucidate the origin of the haplotype and estimate the age of the mutation using a Bayesian inference method based on observed linkage disequilibrium. RESULTS: The mutation's distribution, the size of the shared haplotype, and estimates of growth rate and carrier frequency indicated that the mutation was a single prehistoric event. Dating back to the ancient Middle East around 11,600 BC, the mutation predates the advent of writing, farming, and the monotheistic religions of the region. CONCLUSIONS: This mutation causes over 50% of the IGS cases among Arabic, Turkish, and Sephardic Jewish families, making it a primary target for genetic screening among diverse IGS cases originating from the Middle East. Thus, rare founder mutations may cause a substantial number of cases, even among diverse ethnicities not usually thought to be related.

Methylenetetrahydrofolate reductase polymorphisms C677T and A1298C as maternal risk factors for Down syndrome in Jordan.

BACKGROUND: The activity of the methylenetetrahydrofolate reductase (MTHFR) enzyme is regulated by the two polymorphisms C677T and A1298C, which reduce enzyme activity and result in hypomethylation of chromosomes that increase the risk of nondisjunction. These polymorphisms are suggested to be risk factors for Down syndrome (DS) in some populations. AIM: The aim of this study was to test if C677T and A1298C polymorphisms are correlated to maternal risk of DS in Jordan. METHODS: The proportions of C677T and A1298C polymorphisms were examined in 53 case mothers who delivered DS children and 29 controls. The median age of case mothers was 35 years when delivering their affected children. Polymerase chain reaction-restriction fragment length polymorphism was used for genotyping. RESULTS: The frequency of MTHFR C677T allele in all DS mothers was 3.2-fold higher than in the controls (odds ratio [OR] = 3.12, 95% confidence interval [CI]: 1.303-7.677). Also, the proportion of 677T in the older case mothers was different from the controls, but was significantly higher in younger case mothers than in the controls (OR = 4.2, 95% CI: 1.61-10.97, p = 0.003). The proportions of 677CT and 677TT genotypes in younger cases are, respectively, 10- and 9-fold higher than in the controls. The proportions of MTHFR A1298C are significantly different among all case groups and the controls (χ(2) = 4.27, p = 0.127), but there was a significant difference between young case mothers and both older case mothers group and the controls (OR = 1.25, 95% CI: 0.405-3.85, p = 0.698). CONCLUSIONS: There is strong association between MTHFRC677T and maternal risk of DS in Jordanian mothers younger than 35 years old and the MTHFR1298C allele has a lesser but additive risk effect in MTHFR677T/1298C compound heterozygotes.