Specific cellular microenvironments for spatiotemporal regulation of StAR and steroid synthesis.

For many years, research in the field of steroid synthesis has aimed to understand the regulation of the rate-limiting step of steroid synthesis, i.e. the transport of cholesterol from the outer to the inner mitochondrial membrane, and identify the protein involved in the conversion of cholesterol into pregnenolone. The extraordinary work by B Clark, J Wells, S R King, and D M Stocco eventually identified this protein and named it steroidogenic acute regulatory protein (StAR). The group's finding was also one of the milestones in understanding the mechanism of nonvesicular lipid transport between organelles. A notable feature of StAR is its high degree of phosphorylation. In fact, StAR phosphorylation in the acute phase is required for full steroid biosynthesis. As a contribution to this subject, our work has led to the characterization of StAR as a substrate of kinases and phosphatases and as an integral part of a mitochondrion-associated multiprotein complex, essential for StAR function and cholesterol binding and mitochondrial transport to yield maximum steroid production. Results allow us to postulate the existence of a specific cellular microenvironment where StAR protein synthesis and activation, along with steroid synthesis and secretion, are performed in a compartmentalized manner, at the site of hormone receptor stimulation, and involving the compartmentalized formation of the steroid molecule-synthesizing complex.

Study of the Genetic Variants in BRCA1/2 and Non-BRCA Genes in a Population-Based Cohort of 2155 Breast/Ovary Cancer Patients, Including 443 Triple-Negative Breast Cancer Patients, in Argentina.

Gene/s sequencing in hereditary breast/ovary cancer (HBOC) in routine diagnosis is challenged by the analysis of panels. We aim to report a retrospective analysis of BRCA1/2 and non-BRCA gene sequencing in patients with breast/ovary cancer (BOC), including triple-negative breast cancer (TNBC), in our population. In total 2155 BOC patients (1900 analyzed in BRCA1/2 and 255 by multigenic panels) gave 372 (17.2.6%) and 107 (24.1%) likely pathogenic/pathogenic variants (LPVs/PVs), including BRCA and non-BRCA genes, for the total and TNBC patients, respectively. When BOC was present in the same proband, a 51.3% rate was found for LPVs/PVs in BRCA1/2. Most of the LPVs/PVs in the panels were in BRCA1/2; non-BRCA gene LPVs/PVs were in CDH1, CHEK2, CDKN2A, MUTYH, NBN, RAD51D, and TP53. TNBC is associated with BRCA1/2 at a higher rate than the rest of the breast cancer types. The more prevalent PVs in BRCA1/2 genes (mostly in BRCA1) do not rule out the importance to panels of genes, although they are certainly far from shedding light on the gap of the 85% predicted linkage association of BOC with BRCA1/2 and are still not elucidated.

Angiotensin II-upregulated MAP kinase phosphatase-3 modulates FOXO1 and p21 in adrenocortical H295R cells.

MAPK phosphatases (MKP) downregulate the activity of mitogen-activated protein kinases (MAPK), such as ERK1/2, and modulate the processes regulated by these kinases. ERK1/2 participate in a wide range of processes including tissue-specific hormone-stimulated steroidogenesis. H295R cells are a suitable model for the study of human adrenal cortex functions, particularly steroid synthesis, and respond to angiotensin II (Ang II) triggering ERK1/2 phosphorylation in a transient fashion. MKP-3 dephosphorylates ERK1/2 and, as recently reported, forkhead box protein 1 (FOXO1). Here, we analyzed MKP-3 expression in H295R cells and its putative regulation by Ang II. Results showed the expression of MKP-3 full length (L) and a short splice variant (S), and the upregulation of both isoforms by Ang II. L and S messenger and protein levels increased 30 min after Ang II stimulation and declined over the next 3 h, a temporal frame compatible with ERK1/2 dephosphorylation. In addition, FOXO1 activation is known to include its dephosphorylation and nuclear translocation. Therefore, we analyzed the effect of Ang II on FOXO1 modulation. Ang II induced FOXO1 transient phosphorylation and translocation and also the induction of p21, a FOXO1-dependent gene, whereas MKP-3 knock-down reduced both FOXO1 translocation and p21 induction. These data suggest that, through MKP-3, Ang II counteracts its own effects on ERK1/2 activity and also triggers the activation of FOXO-1 and the induction of cell cycle inhibitor p21. Taken together, the current findings reveal the participation of MKP-3 not only in turn-off but also in turn-on signals which control important cellular processes.

Angiotensin II stimulation promotes mitochondrial fusion as a novel mechanism involved in protein kinase compartmentalization and cholesterol transport in human adrenocortical cells.

In steroid-producing cells, cholesterol transport from the outer to the inner mitochondrial membrane is the first and rate-limiting step for the synthesis of all steroid hormones. Cholesterol can be transported into mitochondria by specific mitochondrial protein carriers like the steroidogenic acute regulatory protein (StAR). StAR is phosphorylated by mitochondrial ERK in a cAMP-dependent transduction pathway to achieve maximal steroid production. Mitochondria are highly dynamic organelles that undergo replication, mitophagy and morphology changes, all processes allowed by mitochondrial fusion and fission, known as mitochondrial dynamics. Mitofusin (Mfn) 1 and 2 are GTPases involved in the regulation of fusion, while dynamin-related protein 1 (Drp1) is the major regulator of mitochondrial fission. Despite the role of mitochondrial dynamics in neurological and endocrine disorders, little is known about fusion/fission in steroidogenic tissues. In this context, the present work aimed to study the role of angiotensin II (Ang II) in protein subcellular compartmentalization, mitochondrial dynamics and the involvement of this process in the regulation of aldosterone synthesis. We demonstrate here that Ang II stimulation promoted the recruitment and activation of PKCε, ERK and its upstream kinase MEK to the mitochondria, all of them essential for steroid synthesis. Moreover, Ang II prompted a shift from punctate to tubular/elongated (fusion) mitochondrial shape, in line with the observation of hormone-dependent upregulation of Mfn2 levels. Concomitantly, mitochondrial Drp1 was diminished, driving mitochondria toward fusion. Moreover, Mfn2 expression is required for StAR, ERK and MEK mitochondrial localization and ultimately for aldosterone synthesis. Collectively, this study provides fresh insights into the importance of hormonal regulation in mitochondrial dynamics as a novel mechanism involved in aldosterone production.

BRCA1 and BRCA2 Mutations Other Than the Founder Alleles Among Ashkenazi Jewish in the Population of Argentina.

In Ashkenazi Jewish (AJ) high risk families 3 mutations [2 in BRCA1 (c. 68_69del and c.5266dup) and 1 in BRCA2 (c.5946del)] account for the majority of high risk breast and ovarian cancer cases in that ethnic group. Few studies with limited number of genotyped individuals have expanded the spectrum of mutations in both BRCA genes beyond the 3 mutation panel. In this study, 279 high risk individual AJ were counseled at CEMIC (Centro de Educación Médica e Investigaciones Clínicas), and were genotyped first for the 3 recurrent mutation panel followed by Next Generation Sequencing (NGS) of BRCA1 BRCA2 in 76 individuals who tested negative for the first genotyping step. Of 279 probands (259 women), 55 (50 women) harbored one of the 3 mutations (19.7%); Of 76 fully sequenced cases (73 women), 6 (5 women) (7.9%) carried a pathogenic mutation: in BRCA1, c.2728C>T - p.(Gln910*); c.5407-?_(*1_?)del and c.5445G>A - p.(Trp1815*); in BRCA2, c.5351dup - p.(Asn1784Lysfs*3); c.7308del - p.(Asn2436Lysfs*33) and c.9026_9030del - p.(Tyr3009Serfs*7). Of 61 mutation carriers the distribution was as follows: 11 cancer free at the time of genotyping, 34 female breast cancer cases with age range 28-72 years (41.6 ± 9.3), 3 male breast cancer cases with age range 59-75 years (65 ± 7.3), 6 breast and ovarian cancer cases with age range 35-60 years (breast 40.4 ± 5.2; ovary 47.8 ± 7.2) and 7 ovarian cancer cases with age range 41-77 years (60.6 ± 13.3). This information proved highly useful for counseling, treatment, and prevention for the patient and the family. In conclusion comprehensive BRCA1/2 testing in AJ high risk breast ovarian cancer cases adds valuable clinically relevant information in a subset of cases estimated up to 7% and is therefore recommended.

BRCA1 and BRCA2 mutations and clinical interpretation in 398 ovarian cancer patients: comparison with breast cancer variants in a similar population.

BACKGROUND: Ovarian cancer is the leading cause of death worldwide among gynecologic malignancies. The recent approval of inhibitors of poly (ADP-ribose) polymerase (iPARP) in the treatment of ovarian cancer in the presence of a BRCA1/2 mutation has sparked the analysis of women with such diagnosis, which can further benefit from the detection of carriers in the family. Germline sequence and large rearrangements for BRCA1/2 were tested in 398 consecutive epithelial ovarian cancer (EOC) patients. The aim of this study was to identify the frequency and spectrum of germline BRCA1/2 pathogenic alterations in a cohort of patients with ovarian serous carcinoma, with a view to adequately selecting patients for prevention through family counseling and correlating this frequency with platinum sensitivity as a guidance to identify patients eligible for iPARP in our population. RESULTS: A total of 96 patients carried a pathogenic germline mutation, accounting for an overall 24.1% mutation incidence. Among mutation carriers, BRCA1 showed 62.5% incidence, BRCA2 rendered 36.5%, and one patient exhibited a mutation in both genes. Three pathogenic mutations were recurrent mutations detected five, three, and four times and represented 12.5% of the mutated samples. Worth highlighting, a 50% mutation incidence was detected when breast and ovarian cancer coexisted in the same patient. Novel mutations amounted to 9.4% of the total mutations, as compared to 4.7% in breast cancer. Forty out of 60 BRCA1 mutations were beyond the ovarian cancer cluster region (OCCR), in stark contrast with 22 out of 36 BRCA2 mutations being inside the OCCR. Taken together, germline BRCA1/2 mutations in EOC patients showed a distinct mutational spectrum compared to our previously published data on breast cancer patients. CONCLUSIONS: In sum, our study provides novel data on ovarian BRCA1/2 mutation prevalence worldwide, enhances adequate patient selection for family counseling and prevention, and sheds light on the benefits of iPARP treatment.

Role of Protein Phosphorylation and Tyrosine Phosphatases in the Adrenal Regulation of Steroid Synthesis and Mitochondrial Function.

In adrenocortical cells, adrenocorticotropin (ACTH) promotes the activation of several protein kinases. The action of these kinases is linked to steroid production, mainly through steroidogenic acute regulatory protein (StAR), whose expression and activity are dependent on protein phosphorylation events at genomic and non-genomic levels. Hormone-dependent mitochondrial dynamics and cell proliferation are functions also associated with protein kinases. On the other hand, protein tyrosine dephosphorylation is an additional component of the ACTH signaling pathway, which involves the "classical" protein tyrosine phosphatases (PTPs), such as Src homology domain (SH) 2-containing PTP (SHP2c), and members of the MAP kinase phosphatase (MKP) family, such as MKP-1. PTPs are rapidly activated by posttranslational mechanisms and participate in hormone-stimulated steroid production. In this process, the SHP2 tyrosine phosphatase plays a crucial role in a mechanism that includes an acyl-CoA synthetase-4 (Acsl4), arachidonic acid (AA) release and StAR induction. In contrast, MKPs in steroidogenic cells have a role in the turn-off of the hormonal signal in ERK-dependent processes such as steroid synthesis and, perhaps, cell proliferation. This review analyzes the participation of these tyrosine phosphates in the ACTH signaling pathway and the action of kinases and phosphatases in the regulation of mitochondrial dynamics and steroid production. In addition, the participation of kinases and phosphatases in the signal cascade triggered by different stimuli in other steroidogenic tissues is also compared to adrenocortical cell/ACTH and discussed.

Rotura renal no traumática. Posoperatorio de transplante renal. A propósito de un caso. / Non-traumatic kidney rupture. Kidney transplant postoperative period. A case report

Se presenta el caso de un paciente de 40 años de edad, que desarrolla una rotura del injerto renal a los 5 días del postoperatorio de trasplante renal con donante cadavérico en nuestra Institución. Se discuten los factores de riesgo para esta complicación, las causas más frecuentes en el trasplante renal, el diagnóstico y el manejo actual de esta grave complicación.

The case of a 40 year old male patient with a rupture of the kidney graft after 5 days of a kidney transplant from a cadaveric donor is presented. Common risk factors for this complication, frequent causes in kidney transplant, diagnosis, and current management of this serious complication are discussed

Role of intramitochondrial arachidonic acid and acyl-CoA synthetase 4 in angiotensin II-regulated aldosterone synthesis in NCI-H295R adrenocortical cell line.

Although the role of arachidonic acid (AA) in angiotensin II (ANG II)- and potassium-stimulated steroid production in zona glomerulosa cells is well documented, the mechanism responsible for AA release is not fully described. In this study we evaluated the mechanism involved in the release of intramitochondrial AA and its role in the regulation of aldosterone synthesis by ANG II in glomerulosa cells. We show that ANG II and potassium induce the expression of acyl-coenzyme A (CoA) thioesterase 2 and acyl-CoA synthetase 4, two enzymes involved in intramitochondrial AA generation/export system well characterized in other steroidogenic systems. We demonstrate that mitochondrial ATP is required for AA generation/export system, steroid production, and steroidogenic acute regulatory protein induction. We also demonstrate the role of protein tyrosine phosphatases regulating acyl-CoA synthetase 4 and steroidogenic acute regulatory protein induction, and hence ANG II-stimulated aldosterone synthesis.

Síndrome de birth-hogg-dube. A propósito de un caso. / Birth-hogg-dube syndrome. A case report

Se describe el caso de una paciente de 48 años de edad, portadora de múltiples lesiones quísticas pulmonares y una tumoración benigna en riñón izquierdo denominada Síndrome de Birt-Hogg-Dube. El objetivo del presente trabajo es reportar esta rara entidad y discutir sobre la necesidad de realizar controles periódicos en los pacientes que la padecen, dada la mayor frecuencia de transformación maligna de esta patología, así como la complicación del cuadro pulmonar con neumotórax espontáneo

We report the case of a 48-year-old female patient who presented multiple lung cystic lesions and a benign tumor in the left kidney called Birt-Hogg-Dube syndrome. The purpose of this paper is to report this rare disease and discuss the need for periodical control once the problem is diagnosed. This condition presents an increased risk of renal neoplasm and spontaneous pneumothorax is the most common complication of the pulmonary pathology.

Tyrosine phosphatases as key regulators of StAR induction and cholesterol transport: SHP2 as a potential tyrosine phosphatase involved in steroid synthesis.

The phospho-dephosphorylation of intermediate proteins is a key event in the regulation of steroid biosynthesis. In this regard, it is well accepted that steroidogenic hormones act through the activation of serine/threonine (Ser/Thr) protein kinases. Although many cellular processes can be regulated by a crosstalk between different kinases and phosphatases, the relationship of Ser/Thr phosphorylation and tyrosine (Tyr)-dephosphorylation is a recently explored field in the regulation of steroid synthesis. Indeed in steroidogenic cells, one of the targets of hormone-induced Ser/Thr phosphorylation is a protein tyrosine phosphatase. Whereas protein tyrosine phosphatases were initially regarded as household enzymes with constitutive activity, dephosphorylating all the substrates they encountered, evidence is now accumulating that protein tyrosine phosphatases are tightly regulated by various mechanisms. Here, we will describe the role of protein tyrosine phosphatases in the regulation of steroid biosynthesis, relating them to steroidogenic acute regulatory protein, arachidonic acid metabolism and mitochondrial rearrangement.

¿Puede el sodio en el baño de diálisis modificar la ganancia de peso interdiálisis? / can sodium in the dialysis bath modify the inter-dialysis weight gain?

El control de la tensión arterial y del volumen del liquido extracelular (VLEC) es un objetivo primordial de la hemodiálisis. La concentración de Na+ en el baño de diálisis es una herramienta que puede utilizarse para conseguir este objetivo. Objetivo: Corroborar si la reducción de Na+ en el baño de diálisis, logra reducir la ganancia de peso inter diálisis. Resultados: Se realizó un estudio prospectivo, longitudinal y experimental, tomando 3 períodos estacionales idénticos, de 3 meses cada uno (diciembre-enero- febrero), durante 3 años consecutivos. Analizado el período 2008/2009 (Na+: 136 mEq/L) respecto del período 2006/2007 (Na+:138 mEq/L), se observó una reducción neta global promedio de 530g (p < 0.003) en la GPPI. Realizando el mismo análisis del período 2007/2008 (Na+ : 37 mEq/L ) respecto del período 2006/2007 (Na+ :138 mEq/L), se observó una reducción neta global promedio de 616g (p < 0.015). Conclusión: Se observó una reducción franca y significativa en la ganancia de peso promedio inter diálisis con la reducción de 2 mEq/L en el baño de diálisis, Con la reducción de 1 mEq/L se observó una reducción promedio similar aunque con una mayor variabilidad entre pacientes.

Introduction: Blood pressure and extracellular fluid (ECF) volume control is a major objective in hemodialysis. Na+ concentration in the dialysis bath is a tool that can be used in order to meet this objective. Objective: To check if Na+ reduction in the dialysis bath can reduce interdialysis weight gain. Results: A prospective, longitudinal, and experimental study considering 3 identical seasonal periods of 3 months each (December-January-February) was carried out during 3 consecutive years. When analyzing the 2008/2009 period (Na+: 136 mEq/L) with regard to the 2006/2007 period (Na+: 138 mEq/L), an average global net reduction of 530g (p < 0.003) was observed in the interdialysis average weight gain. When performing the same analysis for the 2007/2008 period (Na+ : 137 mEq/L) with regard to the 2006/2007 period (Na+: 138 mEq/L), an average global net reduction of 616g (p < 0.015) was observed. Conclusion: A clear and significant reduction was observed in the inter-dialysis average weight gain with the 2 mEq/L reduction in the dialysis bath. With the 1 mEq/ L reduction, a similar average reduction was observed although it showed greater inter-patient variability.

Nefropatía por virus BK en paciente transplantado renal. Primer caso diagnóstico en Bahía Blanca / BK virus nephropathy in a kidney transplant patient. First case diagnosed in Bahía Blanca

Se presenta el caso de un paciente de 65 años de edad, que desarrolla una nefropatía por virus BK, 10 meses posteriores al trasplante renal con donante vivo no relacionado. Se discuten los factores de riesgo para adquirir esta entidad, el diagnóstico, así como las escasas herramientas terapéuticas con las cuales contamos en la actualidad para el tratamiento de esta enfermedad intercurrente relativamente nueva.

We report the case of a 65 year-old male patient, who received a renal transplant from a non-related live donor and, after 10 months, developed a BK virus nephropathy. In this work we discuss risk factors and diagnosis, as well as the few therapeutics tools available to treat this relatively new intercurrent disease.

Glomerulopatia a cambios mínimos en un paciente con esclerosis múltiple tratado con interferón / Glomerulopathy at minimal changes in a patient with multiple sclerosis treated with interferon

La glomerulopatía a cambios mínimos (MCGN) es una enfermedad glomerular primaria que, en pequeño porcentaje, se asocia a otras patologías y al uso de drogas. Si bien, es una causa rara de nefropatía, el interferón (IFN) es una de las medicaciones que pueden afectar al riñón de manera idiosincrática. Las formas más frecuentes son la glomerulopatía membranosa y la glomerulosclerosis focal y segmentaria asociadas a insuficiencia renal aguda. Se presenta el caso de un paciente que siendo portador de una esclerosis múltiple, recibió tratamiento con IFN durante 2 años. Nueve meses posteriores a la suspensión de la droga, desarrolla síndrome nefrótico secundario a MCGN asociado a función renal normal, y actualmente, está en tratamiento. Hasta el presente no se han descripto antecedentes de complicaciones renales causadas por IFN, que se desarrollaron 10 meses después de la suspensión de la medicación.

Minimal-change glomerulopathy (MCGN) is a primary glomerular disease associated, in a few cases, with other pathologies and with drug use. Although it is not often the cause of nephropathy, interferon (IFN) is one of the drugs that can induce kidney disease. Membranous glomerulopathy and focal segmental glomerulosclerosis are the most commonly described forms among IFN-induced nephropathy that are associated to acute renal failure. In this paper, we present a patient with multiple sclerosis who was given a 2 year course of IFN. Nine months after interrupting the administration of the drug, the patient develops a nephrotic syndrome underlying a MCGM associated to normal renal failure. At present, the patient is under treatment. So far, there is no history of renal complications caused by IFN 10 months after treatment interruption.

Concerted regulation of free arachidonic acid and hormone-induced steroid synthesis by acyl-CoA thioesterases and acyl-CoA synthetases in adrenal cells.

Although the role of arachidonic acid (AA) in the regulation of steroidogenesis is well documented, the mechanism for AA release is not clear. Therefore, the aim of this study was to characterize the role of an acyl-CoA thioesterase (ARTISt) and an acyl-CoA synthetase as members of an alternative pathway in the regulation of the intracellular levels of AA in steroidogenesis. Purified recombinant ARTISt releases AA from arachidonoyl-CoA (AA-CoA) with a Km of 2 micro m. Antibodies raised against recombinant acyl-CoA thioesterase recognize the endogenous protein in both adrenal tissue and Y1 adrenal tumor cells by immunohistochemistry and immunocytochemistry and Western blot. Stimulation of Y1 cells with ACTH significantly stimulated endogenous mitochondrial thioesterases activity (1.8-fold). Nordihydroguaiaretic acid (NDGA), an inhibitor of AA release known to affect steroidogenesis, affects the in vitro activity of recombinant ARTISt and also the endogenous mitochondrial acyl-CoA thioesterases. ACTH-stimulated steroid synthesis in Y1 cells was significantly inhibited by a synergistic effect of NDGA and triacsin C an inhibitor of the AA-CoA synthetase. The apparent IC50 for NDGA was reduced from 50 micro m to 25, 7.5 and 4.5 micro m in the presence of 0.1, 0.5 and 2 micro m triacsin C, respectively. Our results strongly support the existence of a new pathway of AA release that operates in the regulation of steroid synthesis in adrenal cells.