ABSTRACT
Background: An abnormality in pulse amplitude and frequency of gonadotropin releasing hormone [GnRH] secretion is the most characteristics of polycystic ovarian syndrome [PCOS]. On the other hand, arginine-phenylalanine-amide [RFamide]-related peptide-3 [RFRP3] inhibits the secretion of GnRH in mammalian hypothalamus. The current study performed in order to investigate the expression of RFRP3 mRNA in the dorsomedial hypothalamic nucleus [DMH] after the induction of PCOS in a rat model of constant light exposure, and the possible role of parity on occurrence of PCOS
Materials and Methods: In the experimental study, female nulliparous [n=12] and primiparous [n=12] rats were randomly subdivided into control and PCOS subgroups [n=6]. PCOS were induced by 90 days exposure to constant light. After 90 days, blood, brain, and ovaries were sampled. Serum levels of follicle stimulating hormone [FSH], luteinizing hormone [LH], and testosterone were evaluated. In addition, six adult female ovariectomized rats as a control of real-time polymerase chain reaction [PCR] tests were prepared and in the DMH of all rats, the relative mRNA expression of RFRP3 was assessed
Results: Histological evaluation of ovaries represented the polycystic features. In addition, serum concentrations of testosterone in the PCOS subgroups were more than the controls [P<0.05]. Furthermore, the relative expression of RFRP3 mRNA in PCOS subgroups was lower than the controls [P<0.05]
Conclusion: Constant light model of the PCOS-induced rats decreased the gene expression of RFRP3 in the DMH that suggests the decrease of RFRP3 may reduce its inhibitory effect on GnRH during the PCOS pathogenesis. This effect was stronger in the nulliparous rats than the primiparous
ABSTRACT
Objective: The phenylalanine hydroxylase [PAH] locus has high linkage disequilibrium. Haplotypes related to this locus may thus be considered sufficiently informative for genetic diagnosis and carrier screening using multi-allelic markers. In this study, we present an efficient method for haplotype analysis of PAH locus using multiplexing dyes. In addition, we explain how to resolve the dye shift challenge in multiplex short tandem repeat [STR] genotyping
Materials and Methods: One hundred family trios were included in this descriptive study. The forward primer of a tetra-nucleotide STR and the reverse primer of a variable number tandem repeat [VNTR] were labeled with three different non-overlapping dyes 5-carboxyfluorescein [FAM], 6-carboxy-N,N,N',N'-tetramethylrhodamine [HEX] and 6-carboxy-N,N,N',N'-tetramethylrhodamine [TAMRA]. The polymerase chain reaction [PCR] products from each family trio were multiplexed for capillary electrophoresis and results were analyzed using Peak Scanner software
Results: Multiplexing trio products decreased the cost significantly. The TAMRA labeled products had a significant predictable shift [migrated at a slower electrophoretic rate] relative to the HEX and FAM labeled products. Through our methodology we achieve, the less inter-dye shift than intra-dye shift variance. Correcting the dye shift in the labeled products, according to the reference allele size, significantly decreased the inter-dye variability [P<0.001]
Conclusion: Multiplexing trio products helps to detect and resolve the dye shift accurately in each family, which otherwise would result in diagnostic error. The dye system of FAM, HEX and TAMRA is more feasible and cheaper than other dye systems
ABSTRACT
Objective: methylmalonic acidura [MMA] is a rare autosomal recessive inborn error of metabolism. In this study we present a novel nucleotide change in the mutase [MUT] gene of two unrelated Iranian pedigrees and introduce the methods used for its functional analysis
Materials and Methods: two probands with definite diagnosis of MMA and a common novel variant in the MUT were included in a descriptive study. Bioinformatic prediction of the splicing variant was done with different prediction servers. Reverse transcription- polymerase chain reaction [RT-PCR] was done for splicing analysis and the products were analyzed by sequencing
Results: the included index patients showed elevated levels of propionylcarnitine [C3]. Urine organic acid analysis confirmed the diagnosis of MMA, and screening for mutations in the MUT revealed a novel C to G variation at the 3' splice acceptor site in intron 12. In silico analysis suggested the change as a mutation in a conserved sequence. The splicing analysis showed that the C to G nucleotide change at position -3 in the acceptor splice site can lead to retention of the intron 12 sequence
Conclusion: this is the first report of a mutation at the position -3 in the MUT intron 12 [c.2125-3C>G]. The results suggest that the identified variation can be associated with the typical clinical manifestations of MMA