RESUMO
Mutations in the retina-specific ABC transporter (ABCR) gene are responsible for autosomal recessive Stargardt disease (arSTGD). Mutation detection efficiency in ABCR in arSTGD patients ranges between 30% and 66% in previously published studies, because of high allelic heterogeneity and technical limitations of the employed methods. Conditions were developed to screen the ABCR gene by double-gradient denaturing-gradient gel electrophoresis. The efficacy of this method was evaluated by analysis of DNA samples with previously characterized ABCR mutations. This approach was applied to mutation detection in 44 Italian arSTGD patients corresponding to 36 independent genomes, in order to assess the nature and frequency of the ABCR mutations in this ethnic group. In 34 of 36 (94.4%) STGD patients, 37 sequence changes were identified, including 26 missense, six frameshift, three splicing, and two nonsense variations. Among these, 20 had not been previously described. Several polymorphisms were detected in affected individuals and in matched controls. Our findings extend the spectrum of mutations identified in STGD patients and suggest the existence of a subset of molecular defects specific to the Italian population. The identification of at least two disease-associated mutations in four healthy control individuals indicates a higher than expected carrier frequency of variant ABCR alleles in the general population. Genotype-phenotype analysis in our series showed a possible correlation between the nature and location of some mutations and specific ophthalmoscopic features of STGD disease.
Assuntos
Transportadores de Cassetes de Ligação de ATP/genética , Degeneração Macular/genética , Mutação , Transportadores de Cassetes de Ligação de ATP/química , Adolescente , Adulto , Idoso , Alelos , Sequência de Bases , Estudos de Casos e Controles , Criança , Análise Mutacional de DNA , Primers do DNA/genética , Eletroforese em Gel de Poliacrilamida , Feminino , Genótipo , Humanos , Itália , Degeneração Macular/patologia , Masculino , Pessoa de Meia-Idade , Fenótipo , Polimorfismo GenéticoRESUMO
In a preview study we found that luteinizing granulosa cells from follicles of patients with polycystic ovary syndrome (PCOS) have a reduced capacity to synthesize progesterone in vitro. Because the 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4-isomerase (3 beta-HSD) is an important enzyme for the biosynthesis of progesterone, the reduced capacity of PCO luteinizing granulosa cells to synthesize progesterone in vitro may be due to reduced 3 beta-HSD gene expression. A reverse transcriptase polymerase chain reaction for 3 beta-HSD was performed and the relative intensity of signals for 3 beta-HSD was evaluated using computer-assisted densitometry. Cells from polycystic ovaries expressed less 3 beta-HSD in follicles < or 10 mm (p < 0.05) and in follicles > or 16 mm (p < 0.05) than cells from normal ovaries. Furthermore, after human chorionic gonadotropin stimulus (50 ng/ml), cells from polycystic ovaries expressed less 3 beta-HSD in follicles > or = 16 mm (p < 0.01) than cells from normal ovaries. The data show that there is a specific change in the gene expression of 3 beta-HSD in PCO granulosa cells resulting in a suppressed capacity to secrete progesterone.
Assuntos
3-Hidroxiesteroide Desidrogenases/genética , Expressão Gênica , Células da Granulosa/enzimologia , Síndrome do Ovário Policístico/enzimologia , Células Cultivadas , Gonadotropina Coriônica/farmacologia , Feminino , Células da Granulosa/efeitos dos fármacos , Humanos , Progesterona/metabolismo , RNA/análise , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
In a clinically hyperthyroid patient a TSH-value of 0.75 mU/L was measured. Upon retesting TSH was <0.1 mU/L. The patient's sample initially had been preceded, in the pipetting device, by a grossly hypothyroid patient's sample. Analogous problems were subsequently documented in 10 cases when samples of other hyperthyroid patients were preceded by samples with modestly to markedly elevated TSH. The spurious elevation of TSH (0.12 to 1.18 vs. <0.1 mU/L) appears to be due to a carryover effect from the respective previous sample within the used automatic pipetting system. This carryover was much larger than previously suggested, and occurred unpredictably.
Assuntos
Artefatos , Automação/métodos , Doença de Graves/sangue , Tireotropina/sangue , Feminino , Bócio/sangue , Bócio/diagnóstico , Doença de Graves/diagnóstico , Humanos , Hipotireoidismo/sangue , Hipotireoidismo/diagnóstico , Radioimunoensaio , Tiroxina/sangue , Tri-Iodotironina/sangueRESUMO
Ovarian hyperstimulation syndrome (OHSS) is a severe complication of ovarian stimulation for assisted reproductive techniques. Clinical manifestations are massive extravascular fluid accumulation and haemoconcentration. Vascular endothelial growth factor (VEGF) has been demonstrated to mediate the development of OHSS. Intravenous albumin at the time of oocyte aspiration has been suggested as an effective prophylactic treatment against the occurrence of severe OHSS. Here it is reported that in cultured human luteinizing granulosa cells, VEGF mRNA expression was enhanced by human albumin and maximum expression was observed in cultured granulosa cells obtained from patients with serum oestradiol concentrations >2000 pg/ml on the day of human chorionic gonadotrophin injection (P < 0. 05).
Assuntos
Albuminas/uso terapêutico , Fatores de Crescimento Endotelial/biossíntese , Células da Granulosa/efeitos dos fármacos , Linfocinas/biossíntese , Síndrome de Hiperestimulação Ovariana/tratamento farmacológico , Células Cultivadas , Avaliação Pré-Clínica de Medicamentos , Feminino , Células da Granulosa/metabolismo , Humanos , Fase Luteal , Síndrome de Hiperestimulação Ovariana/metabolismo , Síndrome de Hiperestimulação Ovariana/patologia , Fator A de Crescimento do Endotélio Vascular , Fatores de Crescimento do Endotélio VascularRESUMO
Our study compared 84 patients with polycystic ovary syndrome (PCOS) with 84 control patients who had normal ovaries and who were matched for the main determinants of success in in-vitro fertilization (IVF) and embryo transfer. Serum concentrations of oestradiol and progesterone on the day of human chorionic gonadotrophin (HCG) injection were significantly higher in PCOS than in normal patients (oestradiol 2016 +/- 1.8 pg/ml versus 1456 +/- 40.9 pg/ml, P < 0.01; progesterone 1.6 +/- 0.1 ng/ml versus 1.2 +/- 0.1 ng/ml, P = 0.03). Furthermore despite oocytes from PCOS patients having a reduced fertilization rate compared with normal patients (61.8 +/- 4.1% versus 73.5 +/- 4.3%, P = 0.03), the differences in pregnancy rate (22.6 versus 19%) and miscarriage (31.5 versus 18.7%) were not statistically significant. In PCOS patients, a critical breakpoint was identified at serum progesterone concentrations of 1.2 ng/ml on the day of HCG injection. The PCOS patients with progesterone > or = 1.2 ng/ml showed a higher pregnancy and miscarriage rate than PCOS patients with progesterone < 1.2 ng/ml (26.6 versus 17.9%, P < 0.01; and 41.7% versus 14.3%, P < 0.01 respectively). These findings suggest that premature progesterone production does not have an adverse effect on pregnancy rate in PCOS, but on the contrary, may be a predictor for success in IVF/embryo transfer.
Assuntos
Gonadotropina Coriônica/administração & dosagem , Fertilização in vitro , Infertilidade Feminina/terapia , Síndrome do Ovário Policístico/complicações , Progesterona/sangue , Adulto , Androgênios/sangue , Biomarcadores/sangue , Índice de Massa Corporal , Estradiol/sangue , Feminino , Hormônio Foliculoestimulante/sangue , Humanos , Infertilidade Feminina/etiologia , Hormônio Luteinizante/sangue , Gravidez , Valores de ReferênciaRESUMO
The underlying cause of anovulation and miscarriage in polycystic ovary syndrome (PCOS) is unknown. Progesterone may play an important role in oocyte fertilization and embryo implantation. Therefore, in this study we analyse the endocrine function of luteinizing granulosa cells to synthesize progesterone in vivo and in vitro in PCOS and normal patients participating in an in-vitro fertilization programme. Human luteinizing granulosa cells were obtained from 10 patients with normal ovaries (controls) and 10 patients with PCOS by follicular aspiration of individual follicles of each patient and pooled in an attempt to obtain three groups: cells from follicle sizes < or =10,>10< or =15 and > or =16. Serum concentrations of oestradiol and progesterone on the day of human chorionic gonadotrophin (HCG) injection were significantly higher (P < 0.01 and P < 0.05) in PCOS patients than in controls. After HCG stimulation, in-vitro progesterone production was enhanced in granulosa cells of the control group and concentrations increased with follicular size as expected. However, the concentration of progesterone of PCOS patients did not increase with follicular size and there was a significant difference between normal and PCOS groups in follicles >10< or =15 mm (P < 0.05) and > or =16 mm (P < 0.01). Oestradiol production was increased in follicles > or =16 mm in both groups, although this did not reach significance. In summary, it seems that PCOS granulosa cells demonstrate an abnormal capacity to synthesize progesterone in vivo and in vitro. The understanding of granulosa cell function in PCOS may explain the anovulation and miscarriage that occurs in these patients.