Loss of the Ca2+/calmodulin-dependent protein kinase type IV in dopaminoceptive neurons enhances behavioral effects of cocaine

The persistent nature of addiction has been associated with activity-induced plasticity of neurons within the striatum and nucleus accumbens (NAc). To identify the molecular processes leading to these adaptations, we performed Cre/loxP-mediated genetic ablations of two key regulators of gene expression in response to activity, the Ca2+/calmodulin-dependent protein kinase IV (CaMKIV) and its postulated main target, the cAMP-responsive element binding protein (CREB). We found that acute cocaine-induced gene expression in the striatum was largely unaffected by the loss of CaMKIV. On the behavioral level, mice lacking CaMKIV in dopaminoceptive neurons displayed increased sensitivity to cocaine as evidenced by augmented expression of locomotor sensitization and enhanced conditioned place preference and reinstatement after extinction. However, the loss of CREB in the forebrain had no effect on either of these behaviors, even though it robustly blunted acute cocaine-induced transcription. To test the relevance of these observations for addiction in humans, we performed an association study of CAMK4 and CREB promoter polymorphisms with cocaine addiction in a large sample of addicts. We found that a single nucleotide polymorphism in the CAMK4 promoter was significantly associated with cocaine addiction, whereas variations in the CREB promoter regions did not correlate with drug abuse. These findings reveal a critical role for CaMKIV in the development and persistence of cocaine-induced behaviors, through mechanisms dissociated from acute effects on gene expression and CREB-dependent transcription.

Hypotension caused by therapeutic doses of venlafaxine: case report and proposed pathophysiological mechanisms

Although venlafaxine is usually associated with modest increases in blood pressure and not so often clinical hypertension, there are a few reported cases of hypotension related to overdoses of this specific antidepressant. The case study of a young female patient with a history of Major Depressive Disorder who initiated treatment with venlafaxine 75 mg/day and developed hypotension when the dosage was titrated up to 225 mg/day is described. The patient did not present comorbid diseases nor use other medication. A temporal association and a dose-dependent relationship between the hypotension and the use of venlafaxine is shown. To the best of the knowledge of the authors, this is the first case report that specifically associates regular doses of venlafaxine with the presence of hypotension. A pathophysiological mechanism is proposed, involving the participation of presynaptic alpha2-adrenergic receptors and the presence of a possible genetic polymorphism of cytochrome P4502D6, which is associated with lower drug metabolization, to explain the relationship between venlafaxine in regular dosage and development of hypotension.

Congenital Hyperinsulinism in Brazilian Neonates: A Study of Histology, KATP Channel Genes, and Proliferation of beta Cells

Congenital hyperinsulinism (CHI) is a rare pancreatic beta-cell disease of neonates, characterized by inappropriate insulin secretion with severe persistent hypoglycemia, with regard to which many questions remain to be answered, despite the important acquisition of its molecular mechanisms in the last decade. The aim of this study was to examine pancreatic histology, beta-cell proliferation (immunohistochemistry with double staining for Ki-67/insulin), and beta-cell adenosine triphosphate-sensitive potassium channels genes from 11 Brazilian patients with severe medically unresponsive CHI who underwent pancreatectomy. Pancreatic histology and beta-cell proliferation in CHI patients were compared to pancreatic samples from 19 age-matched controls. Ten cases were classified as diffuse form (D-CHI) and 1 as focal form (F-CHI). beta-cell nucleomegaly and abundant cytoplasm were absent in controls and were observed only in D-CHI patients. The Ki-67 labeling index (Ki-67-LI) was used to differentiate the adenomatous areas of the F-CHI case (10.15%) from the ""loose cluster of islets`` found in 2 D-CHI samples (2.29% and 2.43%) and 1 control (1.54%) sample. The Ki-67-LI was higher in the F-CHI adenomatous areas, but D-CHI patients also had significantly greater Ki-67-LI (mean value = 2.41%) than age-matched controls (mean value = 1.87%) (P = 0.009). In this 1st genetic study of CHI patients in Brazil, no mutations or new polymorphisms were found in the 33-37 exons of the ABCC8 gene (SUR1) or in the entire exon of the KCNJ11 gene (Kir 6.2) in 4 of 4 patients evaluated. On the other hand, enhanced beta-cell proliferation seems to be a constant feature in CHI patients, both in diffuse and focal forms.

Reticular angiomatoid ""malignant"" fibrous histiocytoma-a case report with cytogenetics and molecular genetic analyses

Angiomatoid ""malignant"" fibrous histiocytoma is a rare sarcoma of low malignant potential that occurs most commonly in the extremities of children and young adults. Herein, we present a case of angiomatoid malignant fibrous histiocytoma with unusual histologic features arising in the mediastinum of an 80-year-old man. The tumor exhibited a reticular growth pattern and myxoid stroma. The tumor cells expressed epithelial membrane antigen and desmin. Cytogenetic analysis revealed the translocation t(2;22)(q33;q12). Molecular genetic analysis confirmed the rearrangement of the EWSR1 locus and the presence of the EWSR1/CREB1 fusion. This report expands the clinicopathologic spectrum of angiomatoid malignant fibrous histiocytoma and underscores the value of integrating morphologic, immunophenotypic, and molecular findings in the identification of its unusual morphologic variants. (C) 2011 Elsevier Inc. All rights reserved.

Polymorphisms in innate immunity genes and patients response to dendritic cell-based HIV immuno-treatment

Dendritic cells (DCs)-based vaccine was demonstrated to increase HIV specific cellular immune response; however, in some HIV-infected patients, the response to the vaccine resulted to be not effective. In order to understand if the outcome of the vaccination may be influenced by the host`s genome and natural immunity, we studied the innate immune genome of HIV-infected patients previously vaccinated with DCs. We identified 15 SNPs potentially associated with the response to the immuno-treatment and two SNPs significantly associated with the modulation of the response to the DC vaccine: MBL2 rs10824792 and NOS1 rs693534. These two SNPs were also studied in different ethnic groups (Brazilians, African and Caucasian) of HIV-infected, exposed uninfected and unexposed uninfected subjects. The HIV positive Caucasian patients were also characterized by different disease progressions. Our findings suggest that, independently and/or in addition to other variables. the host`s genome could significantly contribute to the modulation of the response to the DC vaccine. (C) 2009 Elsevier Ltd. All rights reserved.

Vitamin D Receptor and Calcium-Sensing Receptor Gene Polymorphisms in Hypercalciuric Stone-Forming Patients

Background/Aim: Some studies have identified an association of kidney stone formation with vitamin D receptor (VDR) or calcium-sensing receptor (CaSR) polymorphisms. We aimed to evaluate the association between these polymorphisms with urinary calcium excretion (uCa) in calcium-stone-forming patients. Methods: VDR polymorphism, detected by BsmI digestion, and 3 CaSR polymorphisms (G/T at codon 986, G/A at codon 990 and C/G at codon 1011), detected by direct sequencing, were evaluated in 100 hypercalciuric (HCa) and 101 normocalciuric (NCa) calcium-stone-forming patients. Results: The total allelic frequency of VDR polymorphism was: 16% BB, 49% Bb and 35% bb. The prevalence of bb genotype was significantly higher in the HCa when compared to the NCa group (43 vs. 27%). With respect to CaSR polymorphisms, 986S, 990G and 1011E variant alleles were detected, respectively, in 5, 4 and 3% of the whole sample and 5 CaSR haplotypes were identified: 94% ARQ (wildtype), 3% SRQ, 1.5% AGQ, 1.0% ARE and 0.5% AGE. No statistical differences have been observed between NCa and HCa with respect to these CaSR haplotypes. Conclusions: The present study suggested that bb homozygous for VDR polymorphism was overrepresented in hypercalciuric stone formers. Urinary calcium excretion was not associated with CaSR polymorphism in the present sample. Copyright (C) 2009 S. Karger AG, Basel

Genetic association and cellular function of MC1R variant alleles in human pigmentation

We have examined MC1R variant allele frequencies in the general population of South East Queensland and in a collection of adolescent dizygotic and monozygotic twins and family members to define statistical associations with hair and skin color, freckling, and mole count. Results of these studies are consistent with a linear recessive allelic model with multiplicative penetrance in the inheritance of red hair. Four alleles, D84E, R151C, R160W, and D294H, are strongly associated with red hair and fair skin with multinomial regression analysis showing odds ratios of 63, 118, 50, and 94, respectively. An additional three low-penetrance alleles V60L, V92M, and R163Q have odds ratios 6, 5, and 2 relative to the wild-type allele. To address the cellular effects of MC1R variant alleles in signal transduction, we expressed these receptors in permanently transfected HEK293 cells. Measurement of receptor activity via induction of a cAMP-responsive luciferase reporter gene found that the R151C and R160W receptors were active in the presence of NDP-MSH ligand, but at much reduced levels compared with that seen with the wild-type receptor. The ability to stimulate phosphorylation of the cAMP response element binding protein (CREB) transcription factor was also apparent in all stimulated MC1R variant allele-expressing HEK293 cell extracts as assessed by immunoblotting. In contrast, human melanoma cell lines showed wide variation in the their ability to undergo cAMP-mediated CREB phosphorylation. Culture of human melanocytes of known MC1R genotype may provide the best experimental approach to examine the functional consequences for each MC1R variant allele. With this objective, we have established more than 300 melanocyte cell strains of defined MC1R genotype.

TGFB1 and IL8 gene polymorphisms and susceptibility to visceral leishmaniasis

Visceral leishmaniasis (VL) or Kala-azar is a serious protozoan infectious disease caused by an obligate intracellular parasite. Cytokines have a major role in determining progression and severity of clinical manifestations in VL. We investigated polymorphisms in the TGFB1 and IL8 genes, which are cytokines known to have a role in onset and severity of the disease. Polymorphisms at TGFB1 -509 C/T and +869 T/C, and IL8 -251 A/T were analyzed by a PCR-RFLP technique, in 198 patients with VL, 98 individuals with asymptomatic infection positive for a delayed-type hypersensitivity test (DTH+) and in 101 individuals with no evidence of infection (DTH-). The presence of the T allele in position -509 of the TGFB1 gene conferred a two-fold risk to develop infection both when including those with clinical symptoms (DTH+ and VL, grouped) or when considering DTH+ only, respectively p = 0.007, OR = 1.9 [1.19-3.02] and p = 0.012, OR = 2.01 [1.17-3.79], when compared with DTH- individuals. In addition, occurrence of hemorrhage was associated with TGFB1 -509 T allele. We suggest that the -509 T allele of the TGFB1 gene, a cytokine with a biologically relevant role in the natural history of the disease, may contribute to overall susceptibility to infection by Leishmania and to severity of the clinical disease. (C) 2011 Elsevier B.V. All rights reserved.

Genetic Analysis of Age-at-Onset for Cardiovascular Risk Factors in a Brazilian Family Study

Background/Aims: Statistical analysis of age-at-onset involving family data is particularly complicated because there is a correlation pattern that needs to be modeled and also because there are measurements that are censored. In this paper, our main purpose was to evaluate the effect of genetic and shared family environmental factors on age-at-onset of three cardiovascular risk factors: hypertension, diabetes and high cholesterol. Methods: The mixed-effects Cox model proposed by Pankratz et al. [2005] was used to analyze the data from 81 families, involving 1,675 individuals from the village of Baependi, in the state of Minas Gerais, Brazil. Results: The analyses performed showed that the polygenic effect plays a greater role than the shared family environmental effect in explaining the variability of the age-at-onset of hypertension, diabetes and high cholesterol. The model which simultaneously evaluated both effects indicated that there are individuals which may have risk of hypertension due to polygenic effects 130% higher than the overall average risk for the entire sample. For diabetes and high cholesterol the risks of some individuals were 115 and 45%, respectively, higher than the overall average risk for the entire population. Conclusions: Results showed evidence of significant polygenic effects indicating that age-at-onset is a useful trait for gene mapping of the common complex diseases analyzed. In addition, we found that the polygenic random component might absorb the effects of some covariates usually considered in the risk evaluation, such as gender, age and BMI. Copyright (C) 2008 S. Karger AG, Basel

Influence of Single-Nucleotide Polymorphisms on C-Reactive Protein Levels in Chronic Kidney Disease Before and After Kidney Transplantation

Introduction. We sought to evaluate 2 sing] e-nucleotide polymorphisms (SNPs) in the C-reactive protein (CRP) gene promoter region for their effects on CRP levels in chronic kidney disease (CKD) patients before and after a successful kidney transplantation. Methods. Fifty CKD patients were evaluated before and at the first and second years after the graft. Two SNPs were studied, a bi-allelic (G -> A) at the -409 and a tri-allelic (C -> T -> A) variation at the -390 position in the CRP gene. Results. All patients presented the -409GG genotype. At the -390 position, the ""A"" allele was not found; there were 15 ""CC"" patients, 11 ""TT"" patients, and 24 ""CT"" patients. CRP levels were different among patients with various genotypes (P < .019). Also the presence of the allele ""T"" was sufficient to determine differences in CRP levels both in pretransplantation (P = .045) and at 1 year posttransplantation (P = .011), but not at the second year (P = .448). Conclusion. SNPs at the -390 position of the CRP gene promoter region influence CRP basal levels in such a way that the ""C"" allele correlated with the lowest and the ""T"" with the highest. We did not observe this influence in our patients at the second year posttransplantation.

Primary retroperitoneal lipoma: A soft tissue pathology heresy? Report of a case with classic histologic, cytogenetics, and molecular genetic features

Adipose tissue tumors of the retroperitoneum showing no identifiable cytologic atypia are usually classified as lipoma-like well-differentiated liposarcoma. Whether a subset of these tumors represents true examples of retroperitoneal lipoma remains a controversial subject, because the diagnostic liposarcoma cells may be of difficult identification, even after extensive sampling. Herein, we describe a large retroperitoneal lipoma with classic histopathologic, cytogenetic, molecular cytogenetic, and molecular genetic features. Extensive morphologic inspection showed no evidence of cytologic atypia. Cytogenetic analysis performed on fresh tissue material revealed the classic lipoma chromosome t(3;12)(q27;q14-15). Fluorescence in situ hybridization on multiple sections excluded the presence of MDM2 and CDK4 amplification, but showed HMGA2 balanced rearrangement in most cells. Reverse-transcriptase polymerase chain reaction followed by sequencing analysis confirmed the presence of the HMGA2-LPP fusion gene, a characteristic and the most common fusion product found in lipoma. The patient has been followed for 2.5 years without evidence of recurrence or metastasis. These results indicate that retroperitoneal lipomata do exist, but their diagnosis must rely on stringent histologic, cytogenetic, and molecular genetic analysis.

Understanding the genetic diversity, spatial genetic structure and mating system at the hierarchical levels of fruits and individuals of a continuous Theobroma cacao population from the Brazilian Amazon

Understanding the mating patterns of populations of tree species is a key component of ex situ genetic conservation. In this study, we analysed the genetic diversity, spatial genetic structure (SGS) and mating system at the hierarchical levels of fruits and individuals as well as pollen dispersal patterns in a continuous population of Theobroma cacao in Para State, Brazil. A total of 156 individuals in a 0.56 ha plot were mapped and genotyped for nine microsatellite loci. For the mating system analyses, 50 seeds were collected from nine seed trees by sampling five fruits per tree (10 seeds per fruit). Among the 156 individuals, 127 had unique multilocus genotypes, and the remaining were clones. The population was spatially aggregated; it demonstrated a significant SGS up to 15m that could be attributed primarily to the presence of clones. However, the short seed dispersal distance also contributed to this pattern. Population matings occurred mainly via outcrossing, but selfing was observed in some seed trees, which indicated the presence of individual variation for self-incompatibility. The matings were also correlated, especially within ((r) over cap (p(m)) = 0.607) rather than among the fruits ((r) over cap (p(m)) = 0.099), which suggested that a small number of pollen donors fertilised each fruit. The paternity analysis suggested a high proportion of pollen migration (61.3%), although within the plot, most of the pollen dispersal encompassed short distances (28m). The determination of these novel parameters provides the fundamental information required to establish long-term ex situ conservation strategies for this important tropical species. Heredity (2011) 106, 973-985; doi:10.1038/hdy.2010.145; published online 8 December 2010