LUCIANI RENATA SILVEIRA DE CARVALHO
Projetos de Pesquisa
Unidades Organizacionais
Instituto Central, Hospital das Clínicas, Faculdade de Medicina - Médico
LIM/42 - Laboratório de Hormônios e Genética Molecular, Hospital das Clínicas, Faculdade de Medicina
LIM/42 - Laboratório de Hormônios e Genética Molecular, Hospital das Clínicas, Faculdade de Medicina
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conferenceObject Copy Number Variants in Patients with Congenital Hypopituitarism Associated with Complex Phenotypes(2015) CORREA, F.; FRANCA, M.; CANTON, A.; OTTO, A.; COSTALONGA, E.; BRITO, V; CARVALHO, L.; COSTA, S.; ARNHOLD, I; JORGE, A.; ROSENBERG, C.; MENDONCA, B.- Ketoconazole Treatment Decreases the Viability of Immortalized Pituitary Cell Lines Associated with an Increased Expression of Apoptosis-Related Genes and Cell Cycle Inhibitors(2015) GUZZO, M. F.; CARVALHO, L. R.; BRONSTEIN, M. D.Ketoconazole, which was initially developed as an antifungal agent, is a potent inhibitor of adrenal steroidogenesis and has therefore been used in the management of Cushing's disease. Surprisingly, the reduction of cortisol levels during ketoconazole treatment is not accompanied by the expected elevation in plasma adrenocorticotrophic hormone (ACTH) at the loss of negative cortisol feedback from corticotrophic cells, suggesting a direct effect of ketoconazole on these cells. To characterize the direct effects of ketoconazole, we evaluated its invitro effect on cell viability using the pituitary tumoural cell lines AtT-20 (which secretes ACTH), GH3 (which secretes growth hormone and prolactin) and T3.1 (which secretes -subunit) and we also determined the expression levels of genes involved in apoptosis and DNA replication by the quantitative reverse transcription polymerase chain reaction (qRT-PCR). We also evaluated ACTH levels in AtT-20 cells during ketoconazole treatment. We observed a ketoconazole concentration-dependent decrease in pituitary cell viability and reduced ACTH levels in AtT-20 cells after removal of the drug. We also observed increased expression of cell death receptors (e.g. Fas, tumour necrosis factor receptor) and caspases (e.g., caspase-6, caspase-7, caspase-9), suggesting activation of the apoptosis pathway. In addition, we observed increased gene expression of the cell cycle inhibitors p21 and p27 in GH3 cells and increased expression of p21 in T3.1 cells. In conclusion, our findings suggest that ketoconazole significantly reduces cell viability in a concentration-dependent manner in pituitary tumour cell lines and is associated with an increase in apoptosis- and cell cycle regulation-related gene expression.
- FGFR1 and PROKR2 rare variants found in patients with combined pituitary hormone deficiencies(2015) CORREA, Fernanda A.; TRARBACH, Ericka B.; TUSSET, Cintia; LATRONICO, Ana Claudia; MONTENEGRO, Luciana R.; CARVALHO, Luciani R.; FRANCA, Marcela M.; OTTO, Aline P.; COSTALONGA, Everlayny F.; BRITO, Vinicius N.; ABREU, Ana Paula; NISHI, Mirian Y.; JORGE, Alexander A. L.; ARNHOLD, Ivo J. P.; SIDIS, Yisrael; PITTELOUD, Nelly; MENDONCA, Berenice B.The genetic aetiology of congenital hypopituitarism (CH) is not entirely elucidated. FGFR1 and PROKR2 loss-of-function mutations are classically involved in hypogonadotrophic hypogonadism (HH), however, due to the clinical and genetic overlap of HH and CH; these genes may also be involved in the pathogenesis of CH. Using a candidate gene approach, we screened 156 Brazilian patients with combined pituitary hormone deficiencies (CPHD) for loss-of-function mutations in FGFR1 and PROKR2. We identified three FGFR1 variants (p.Arg448Trp, p.Ser107Leu and p.Pro772Ser) in four unrelated patients (two males) and two PROKR2 variants (p. Arg85Cys and p. Arg248Glu) in two unrelated female patients. Five of the six patients harbouring the variants had a first-degree relative that was an unaffected carrier of it. Results of functional studies indicated that the new FGFR1 variant p.Arg448Trp is a loss-of-function variant, while p.Ser107Leu and p.Pro772Ser present signalling activity similar to the wild-type form. Regarding PROKR2 variants, results from previous functional studies indicated that p.Arg85Cys moderately compromises receptor signalling through both MAPK and Ca2+ pathways while p.Arg248Glu decreases calcium mobilization but has normal MAPK activity. The presence of loss-of-function variants of FGFR1 and PROKR2 in our patients with CPHD is indicative of an adjuvant and/or modifier effect of these rare variants on the phenotype. The presence of the same variants in unaffected relatives implies that they cannot solely cause the phenotype. Other associated genetic and/or environmental modifiers may play a role in the aetiology of this condition.
- Role of GLI2 in hypopituitarism phenotype(2015) ARNHOLD, Ivo J. P.; FRANCA, Marcela M.; CARVALHO, Luciani R.; MENDONCA, Berenice B.; JORGE, Alexander A. L.GLI2 is a zinc-finger transcription factor involved in the Sonic Hedgehog pathway. Gli2 mutant mice have hypoplastic anterior and absent posterior pituitary glands. We reviewed the literature for patients with hypopituitarism and alterations in GLI2. Twenty-five patients (16 families) had heterozygous truncating mutations, and the phenotype frequently included GH deficiency, a small anterior pituitary lobe and an ectopic/undescended posterior pituitary lobe on magnetic resonance imaging and postaxial polydactyly. The inheritance pattern was autosomal dominant with incomplete penetrance and variable expressivity. The mutation was frequently inherited from an asymptomatic parent. Eleven patients had heterozygous non-synonymous GLI2 variants that were classified as variants of unknown significance, because they were either absent from or had a frequency lower than 0.001 in the databases. In these patients, the posterior pituitary was also ectopic, but none had polydactyly. A third group of variants found in patients with hypopituitarism were considered benign because their frequency was >= 0.001 in the databases. GLI2 is a large and polymorphic gene, and sequencing may identify variants whose interpretation may be difficult. Incomplete penetrance implies in the participation of other genetic and/or environmental factors. An interaction between Gli2 mutations and prenatal ethanol exposure has been demonstrated in mice dysmorphology. In conclusion, a relatively high frequency of GLI2 mutations and variants were identified in patients with congenital GH deficiency without other brain defects, and most of these patients presented with combined pituitary hormone deficiency and an ectopic posterior pituitary lobe. Future studies may clarify the relative role and frequency of GLI2 alterations in the aetiology of hypopituitarism.
conferenceObject A Novel OTX2 Mutation, P.H230L, Causes Hypopituitarism with Incomplete Penetrance: Exome Sequencing to Identify Modifier Genes(2015) MADEIRA, J.; MOREIRA, M.; FRANCA, M.; OTTO, A.; CORREA, F.; ARNHOLD, I.; MENDONCA, B.; FANG, Q.; MA, Q.; LI, J.; GERGICS, P.; CAMPER, S.; CARVALHO, L.conferenceObject HESX1 Mutations Cause Hypopituitarism with Different Clinical Features(2015) BENEDETTI, A. Figueredo; FANG, Q.; GREGORY, L.; MENDONCA, B. Bilharinho de; ARNHOLD, I; DATTANI, M.; CAMPER, S.; LI, J.; MA, Q.; SADEGHI-NEJAD, A.; CARVALHO, L.conferenceObject A Homozygous Point Mutation in the GH1 Promoter (-161T > C) Leads to Reduced GH Expression in Siblings with Isolated GH Deficiency (IGHD)(2015) CARVALHO, L.; MADEIRA, J.; MARTIN, R.; MONTENEGRO, L.; FRANCA, M.; COSTALONGA, E.; CORREA, F.; OTTO, A.; ARNHOLD, I; FREITAS, H.; MACHADO, U.; MENDONCA, B.; JORGE, A.- Frequent development of combined pituitary hormone deficiency in patients initially diagnosed as isolated growth hormone deficiency: a long term follow-up of patients from a single center(2015) OTTO, Aline P.; FRANCA, Marcela M.; CORREA, Fernanda A.; COSTALONGA, Everlayny F.; LEITE, Claudia C.; MENDONCA, Berenice B.; ARNHOLD, Ivo J. P.; CARVALHO, Luciani R. S.; JORGE, Alexander A. L.Children initially diagnosed with isolated GH deficiency (IGHD) have a variable rate to progress to combined pituitary hormone deficiency (CPHD) during follow-up. To evaluate the development of CPHD in a group of childhood-onset IGHD followed at a single tertiary center over a long period of time. We retrospectively analyzed data from 83 patients initially diagnosed as IGHD with a mean follow-up of 15.2 years. The Kaplan-Meier method and Cox regression analysis was used to estimate the temporal progression and to identify risk factors to development of CPHD over time. From 83 patients initially with IGHD, 37 (45 %) developed CPHD after a median time of follow up of 5.4 years (range from 1.2 to 21 years). LH and FSH deficiencies were the most common pituitary hormone (38 %) deficiencies developed followed by TSH (31 %), ACTH (12 %) and ADH deficiency (5 %). ADH deficiency (3.1 +/- A 1 years from GHD diagnosis) presented earlier and ACTH deficiency (9.3 +/- A 3.5 years) presented later during follow up compared to LH/FSH (8.3 +/- A 4 years) and TSH (7.5 +/- A 5.6 years) deficiencies. In a Cox regression model, pituitary stalk abnormalities was the strongest risk factor for the development of CPHD (hazard ratio of 3.28; p = 0.002). Our study indicated a high frequency of development of CPHD in patients initially diagnosed as IGHD at childhood. Half of our patients with IGHD developed the second hormone deficiency after 5 years of diagnosis, reinforcing the need for lifelong monitoring of pituitary function in these patients.