OBJECTIVE: This study aimed to examine the effect of vitrification on apoptosis and survival in human preantral follicles after thawing.METHODS: This experimental study was conducted at an acute tertiary care hospital from March 2012 to April 2013. Ovaries were sliced into 5×5×1-mm pieces and divided into the following three groups: preantral follicle isolation, ovarian tissue vitrification-warming followed by follicle isolation, and immunohistochemistry of fresh ovarian tissue. For statistical analyses, the Student t-test, chi-square test, Kruskal-Wallis test, and Kaplan-Meier survival analysis were used.RESULTS: A total of 161 preantral follicles (70% secondary) were collected from ovarian cortex tissue of six women between 30 and 37 years of age who underwent oophorectomy due to cervical cancer or breast cancer. There were no significant differences in the follicular morphology of fresh preantral follicles and vitrified follicles after thawing. The mean Fas ligand (FasL) mRNA expression level was 0.43±0.20 (relative to β-actin) in fresh preantral follicles versus 0.51±0.20 in vitrified follicles (p=0.22). The mean caspase-3 mRNA expression level in fresh preantral follicles was 0.56±0.49 vs. 0.27±0.21 in vitrified follicles (p=0.233). One vitrified-thawed secondary follicle grew and developed to an antral follicle within 6 days of culture.CONCLUSION: Vitrification did not affect preantral follicle morphology or mRNA expression of the apoptosis markers FasL and caspase-3. Further studies are required to establish whether vitrification affects the outcomes of in vitro culture and the maturation of preantral follicles.
Apoptosis ; Breast Neoplasms ; Caspase 3 ; Fas Ligand Protein ; Female ; Humans ; Immunohistochemistry ; In Vitro Techniques ; Ovariectomy ; Ovary ; RNA, Messenger ; Tertiary Healthcare ; Uterine Cervical Neoplasms ; Vitrification

Objectives: One of the most widely used pesticides today is chlorpyrifos (CPF). Cytochrome P450 (CYP)2B6, the most prominent catalyst in CPF bioactivation, is highly polymorphic. The objective of our study was to evaluate the role of CYP2B6*6, which contains both 516G>T and 785A>G polymorphisms, in CPF toxicity, as represented by the concentration of 3,5,6-trichloro-2-pyridinol (TCPy), among vegetable farmers in Central Java, Indonesia, where CPF has been commonly used. Methods: A cross-sectional study was conducted among 132 vegetable farmers. Individual socio-demographic and occupational characteristics, as determinants of TCPy levels, were obtained using a structured interviewer-administered questionnaire and subsequently used to estimate the cumulative exposure level (CEL). TCPy levels were detected with liquid chromatography-mass spectrometry. CYP2B6*6 gene polymorphisms were analyzed using a TaqMan® SNP Genotyping Assay and Sanger sequencing. Linear regression analysis was performed to analyze the association between TCPy, as a biomarker of CPF exposure, and its determinants. Results: The prevalence of CYP2B6*6 polymorphisms was 31% for *1/*1, 51% for *1/*6, and 18% for *6/*6. TCPy concentrations were higher among participants with CYP2B6*1/*1 than among those with *1/*6 or *6/*6 genotypes. CYP2B6*6 gene polymorphisms, smoking, CEL, body mass index, and spraying time were retained in the final linear regression model as determinants of TCPy. Conclusions The results suggest that CYP2B6*6 gene polymorphisms may play an important role in influencing susceptibility to CPF exposure. CYP2B6*6 gene polymorphisms together with CEL, smoking habits, body mass index, and spraying time were the determinants of urinary TCPy concentrations, as a biomarker of CPF toxicity.

Objective: The objective of our study was to determine the genotype frequencies of CYP2C19*2 and *3 gene polymorphisms among Javanese farmers exposed to chlorpyrifos (CPF) in Central Java, Indonesia. Methods: This cross-sectional study was conducted from July to October 2020 in Central Java, Indonesia, involving 151 vegetable farmers aged 18-65 years who used CPF for at least one year. CYP2C19*2 and *3 gene polymorphisms were analyzed using PCR-RFLP. Direct calculations were applied to calculate allele and genotype frequencies. The difference in genotype frequencies among the sex and cumulative exposure level (CEL) group was performed using the Chi-square test. In contrast, the proportion difference of allele frequencies was analyzed using Z-test. Results: The frequency of CYP2C19 genotypes *1/*1, *1/*3, *1/*2, and *2/*2 were 64 %, 7 %, 23 % and 6 %. We observed no significant difference in the genotype distribution according to sex group and CEL group. Conclusion In summary, the prevalence of toxicologically relevant CYP2C19 polymorphisms was determined in the Javanese agricultural population. The CYP2C19 genotype may be helpful as an essential biomarker of genetic susceptibility towards CPF exposure. Nevertheless, further studies to confirm the role of CYP2C19 in this context are still needed.
Cytochrome P-450 CYP2C19 ; Pesticides