Propagation of pure fetal and maternal mesenchymal stromal cells from terminal chorionic villi of human term placenta.

Long term propagation of human fetal mesenchymal stromal cells (MSC) in vitro has proven elusive due to limited availability of fetal tissue sources and lack of appropriate methodologies. Here, we have demonstrated the presence of fetal and maternal cells within the tips of terminal chorionic villi (TCV) of normal human term placenta, and we have exploited inherent differences in the adhesive and migratory properties of maternal vs. fetal cells, to establish pure MSC cultures of both cell types. The origin and purity of each culture was confirmed by X-Y chromosome-specific fluorescence in situ hybridization (FISH) and short tandem repeat (STR) genotyping. This is the first demonstration of fetal and maternal cells in the TCV of human term placenta and also of deriving pure fetal MSC cultures from them. The concomitant availability of pure cultures of adult and fetal MSC from one tissue provides a good system to compare genetic and epigenetic differences between adult and fetal MSCs; and also to generate new models of cell based therapies in regenerative medicine.

De novo 7p partial trisomy characterized by subtelomeric FISH and whole-genome array in a girl with mental retardation.

Chromosome rearrangements involving telomeres have been established as one of the major causes of idiopathic mental retardation/developmental delay. This case of 7p partial trisomy syndrome in a 3-year-old female child presenting with developmental delay emphasizes the clinical relevance of cytogenetic diagnosis in the better management of genetic disorders. Application of subtelomeric FISH technique revealed the presence of interstitial telomeres and led to the ascertainment of partial trisomy for the distal 7p segment localized on the telomeric end of the short arm of chromosome 19. Whole-genome cytogenetic microarray-based analysis showed a mosaic 3.5 Mb gain at Xq21.1 besides the approximately 24.5 Mb gain corresponding to 7p15.3- > pter. The possible mechanisms of origin of the chromosomal rearrangement and the clinical relevance of trisomy for the genes lying in the critical regions are discussed.

Mutational analysis of the betaglycan gene-coding region in susceptibility for ovarian failure.

BACKGROUND: Elevation of FSH is frequently a consequence of impaired ovarian follicle growth. Down-regulation of the FSH levels by inhibins is mediated through its receptor betaglycan in the gonadotrophs. Understanding of germline status of the betaglycan gene (TGFBR3) is essential for ovarian failure pathophysiology. METHODS: Sequence analysis was performed for the coding region of TGFBR3 gene in a cohort of 196 ovarian failure cases that include 133 premature ovarian failure (POF) cases, 63 primary amenorrhoea (PA) cases compared with 200 controls. RESULTS: Forty-six variants including six novel exonic variants and 16 novel intronic variants were revealed. Two variants were missense: (i) p.Iso184Val in a control and (ii) p.Pro775Ser in a POF case. Genotypic distribution of three variants (c.382-81C>T, c.382-77T>C and c.1200G>A) was significantly different in the patients as compared with the controls. Five variants c.382-81C>T, c.382-77T>C, c.566-216G>A, c.1200G>A and c.2022T>C were chosen for haplotyping. The CCAAT haplotype was significantly higher in the patient population as compared with the controls (P = 0.00007). CONCLUSION: This study establishes the first mutational report of the TGFBR3 gene in correlation with ovarian failure. Significant diversity of genotype distribution and haplotype analysis suggested susceptibility of the TGFBR3 gene for ovarian failure aetiology.

Screening of the galactose-1-phosphate uridyltransferase gene in Indian women with ovarian failure.

The present study was aimed at mutational screening of the gene coding for galactose-1-phosphate uridyltransferase in females with premature ovarian failure within an Indian population. A case-control-based study approach was used. It included females with premature ovarian failure (n = 108), primary amenorrhoea (n = 37) and secondary amenorrhoea (n = 9), and a control group of 136 women with a normal ovarian pattern. Gene sequencing analysis for the presence of mutations in the promoter and the coding regions of GALT has shown the absence of any mutation. A hexanucleotide deletion was found in the third intronic region of GALT in both cases and controls. These data support the hypothesis that there is no significant association between GALT mutations and ovarian failure, and hence the present authors conclude that there is no relationship between ovarian failure and GALT polymorphisms in Indian women.

Prevalence of chromosome defects in azoospermic and oligoastheno-teratozoospermic South Indian infertile men attending an infertility clinic.

The prevalence of chromosomal abnormalities in azoospermic and oligoastheno-teratozoospermic infertile men of South Indian origin undergoing assisted reproductive technologies was evaluated. In addition, the study aimed to investigate new abnormal karyotypes involving autosomes in azoospermia and sex chromosomes in oligoastheno-teratozoospermic individuals that are supposed to be rare. Metaphase chromosomes of 744 infertile men, including 272 men with azoospermia and 472 men with oligoastheno-teratozoospermia (OAT), were analysed using Giemsa-trypsin-Giemsa banding and fluorescence in-situ hybridization (FISH) wherever necessary. Chromosomal abnormalities were observed in 59 (7.9%) individuals of the total studied population. Among these, 30 out of 272 (11.0%) azoospermic men and 29 out of 472 (6.1%) infertile men with OAT showed chromosomal abnormalities. A strong and statistically significant association (OR = 1.89; P = 0.0235) of chromosomal abnormalities and sex chromosome abnormalities (OR = 4.29; P = 0.001) with azoospermia when compared with OAT was observed. In addition, six autosomal abnormalities associated with azoospermia and two abnormalities involving Y chromosome, which include a novel karyotype (mos 46,XY/51,XYYYYYY) in OAT individuals, were detected.

N-acetyl transferase 2 polymorphism and advanced stages of endometriosis in South Indian women.

Aylamine-N-acetyl transferase is a phase II detoxification enzyme encoded by the gene NAT2. Single nucleotide polymorphism (SNP) changes from the wild type NAT2 *4 allele result in allelic variants *5, *6 and *7. Homozygotes for the NAT2 *4 wild type are fast acetylators; heterozygotes with one wild-type allele and a variant NAT2 *5, *6 or *7 allele have reduced enzyme activity and individuals with two variant alleles are slow acetylators. Previous studies have implicated NAT2 as a susceptibility factor in endometriosis. This study investigated the NAT2 allele frequencies and genotype distributions in 252 unrelated women with endometriosis and 264 controls of South Indian origin. No differences were found between the frequencies of fast and slow acetylators in cases (34.9% and 65.1%) and controls (33.3% and 66.7%). Two NAT2 genotypes *7/*7 (1.2%) and *5/*6/*7 (1.6%) were detected in endometriosis cases only. Four new combinations, 6D (481 + 590 mutation), 7C (590 + 857), 7D (590 + 803 + 857) and 7E (481 + 590 + 803 + 857) were detected, which have not been reported earlier. Similar genotype and phenotype results were obtained in 33 affected sister-pairs. The case-control data from this study suggest there is no association between endometriosis and NAT2 in South Indian women; however, two new variant genotypes and seven SNP combinations were also identified in cases only, which suggests that the gene may still have some as yet undetermined role in the disease.

CYP1A1, GSTM1 and GSTT1 genetic polymorphism is associated with susceptibility to polycystic ovaries in South Indian women.

Polymorphic variants in the phase I enzyme, cytochrome P450 gene, may lead to increased toxification, whereas polymorphisms in the phase II enzyme, glutathione S-transferase genes, may result in impaired detoxification. Alterations in the activities of phase I drug metabolizing enzymes and phase II detoxification enzymes may cause abnormal functioning and formation of follicular cysts in the ovaries and thus causing an imbalance in the hormone profiles. This study aimed to investigate the relationship between genetic polymorphisms of CYP1A1 (T6235C), GSTM1 and GSTT1 in South Indian women with polycystic ovaries (PCO) using polymerase chain reaction-restriction fragment length polymorphism. The frequencies of variants of these genes were studied in 180 women with confirmed PCO and in 72 healthy fertile women with successful pregnancy record. No significant difference was found between the frequencies of GSTM1 and GSTT1 null genotypes in PCO cases and healthy controls. However, CYP1A1 Msp I homozygous mutants were strongly associated (P = 0.0139) with increased susceptibility to PCO. Three genotype combinations, CYP1A1 (mt/mt) with GSTM1 [-] and GSTT1 [-], CYP1A1 (wt/mt) with GSTM1 [-] and GSTT1 [-] and CYP1A1 (mt/mt) with GSTM1 [-], GSTT1 [+], were also observed in women with PCO. In conclusion, the presence of hyperinducible CYP1A1 (T6235C) mutant genotype and its mutants in combination with GSTM1 and GSTT1 null genotypes might cause an imbalance between phase I and phase II enzymes, and therefore may represent a risk factor for PCO.