Genetic polymorphims of estrogen receptor alpha −397 PvuII (T>C) and −351 XbaI (A>G) in a portuguese population: prevalence and relation with breast cancer susceptibility

Estrogen receptor alpha (ERα), that mediates the biologic effects of estrogen in estrogen-sensitive tissues like breast, is genetically polymorphic. To evaluate the association between ?397 PvuII (T>C) and ?351 XbaI (A>G) restriction fragment length polymorphisms (RFLPs) in intron 1 of ERα gene and susceptibility of breast cancer, we undertook a case–control study in BRCA1 185delAG and 5382insC/BRCA2 6174delT negative Portuguese women. The study population consisted of 107 patients with histological diagnosis of breast cancer and 121 women with no history of breast cancer. Genomic DNA was extracted from blood samples and genotyping analyses were performed by PCR–RFLP. XbaI polymorphism was associated with a significant reduced risk of breast cancer for carriers of the x allele in homozygozity (OR 0.178; 95 % CI 0.070–0.456; P < 0.001) or heterozigozity (OR 0.223; 95 % CI 0.089–0.561; P = 0.001). The PvuII polymorphism was associated with a non-significantly reduced risk. The combined analysis of PvuII and XbaI polymorphisms revealed none synergistic effect of the two genotypes, except for simultaneous carriers of pp and xx genotypes, that have a reduced risk of breast cancer (OR 0.226; 95 % CI 0.049–1.035; P = 0.044). The combination of PvuII and XbaI genotypes into haplotypes showed that carriers of two copies of the px (ppxx) haplotype had a reduced risk of breast cancer (OR 0.405; 95 % CI 0.194–0.843; P = 0.014), compared with PX (PPXX + PPXx + PpXX + PpXx) haplotypes. PvuII and XbaI polymorphisms were in linkage disequilibrium both in cases (D = 0.044, r² = 0.049, X² = 5.216, P = 0.022) and controls (D = 0.090, r² = 0.139, X² = 16.819, P < 0.001), but not in the entire sample population analyzed as a whole (D = 0.087, r² = 0.0076, X² = 1.733, P = 0.188). In conclusion, in this case–control study we found that ERα gene XbaI polymorphism may modify individual susceptibility for breast cancer in this population.     

Models for antigen receptor gene rearrangement. I. Biased receptor editing in B cells: implications for allelic exclusion.

Recent evidence suggests that lymphocyte Ag receptor gene rearrangement does not always stop after the expression of the first productively rearranged receptor. Light chain gene rearrangement in B cells, and alpha-chain rearrangement in T cells can continue, which raises the question: how is allelic exclusion maintained, if at all, in the face of continued rearrangement? In this and the accompanying paper, we present comprehensive models of Ag receptor gene rearrangement and the interaction of this process with clonal selection. Our B cell model enables us to reconcile observations on the kappa:lambda ratio and on kappa allele usage, showing that B cell receptor gene rearrangement must be a highly ordered, rather than a random, process. We show that order is exhibited on three levels: a preference for rearranging kappa rather than lambda light chain genes; a preference to make secondary rearrangements on the allele that has already been rearranged, rather than choosing the location of the next rearrangement at random; and a sequentiality of J segment choice within each kappa allele. This order, combined with the stringency of negative selection, is shown to lead to effective allelic exclusion.     

Insulin-like growth factor I receptor signaling is required for exercise-induced cardiac hypertrophy

The receptors for IGF-I (IGF-IR) and insulin (IR) have been implicated in physiological cardiac growth, but it is unknown whether IGF-IR or IR signaling are critically required. We generated mice with cardiomyocyte-specific knockout of IGF-IR (CIGF1RKO) and compared them with cardiomyocyte-specific insulin receptor knockout (CIRKO) mice in response to 5 wk exercise swim training. Cardiac development was normal in CIGF1RKO mice, but the hypertrophic response to exercise was prevented. In contrast, despite reduced baseline heart size, the hypertrophic response of CIRKO hearts to exercise was preserved. Exercise increased IGF-IR content in control and CIRKO hearts. Akt phosphorylation increased in exercise-trained control and CIRKO hearts and, surprisingly, in CIGF1RKO hearts as well. In exercise-trained control and CIRKO mice, expression of peroxisome proliferator-activated receptor-gamma coactivator-1alpha (PGC-1alpha) and glycogen content were both increased but were unchanged in trained CIGF1RKO mice. Activation of AMP-activated protein kinase (AMPK) and its downstream target eukaryotic elongation factor-2 was increased in exercise-trained CIGF1RKO but not in CIRKO or control hearts. In cultured neonatal rat cardiomyocytes, activation of AMPK with 5-aminoimidazole-4-carboxamide-1-beta-D-ribofuranoside (AICAR) prevented IGF-I/insulin-induced cardiomyocyte hypertrophy. These studies identify an essential role for IGF-IR in mediating physiological cardiomyocyte hypertrophy. IGF-IR deficiency promotes energetic stress in response to exercise, thereby activating AMPK, which leads to phosphorylation of eukaryotic elongation factor-2. These signaling events antagonize Akt signaling, which although necessary for mediating physiological cardiac hypertrophy, is insufficient to promote cardiac hypertrophy in the absence of myocardial IGF-I signaling.     

Detection of peroxisomes in human liver and kidney fixed with formalin and embedded in paraffin: the use of catalase and lipid beta-oxidation enzymes as immunocytochemical markers

Summary We describe the immunocytochemical localization of four peroxisomal enzymes by light microscopy in human liver and kidney processed routinely by formalin fixation and paraffin embedding. Monospecific antisera against catalase and three enzymes of peroxisomal lipid -oxidation (acyl-CoA oxidase, bifunctional protein (enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase) and 3-ketoacyl-CoA thiolase) were used in conjunction with either the indirect immunoperoxidase method or the protein A—gold technique followed by silver intensification. The sections of formalin-fixed paraffin-embedded tissue had to be deparaffinized and subjected to controlled proteolysis in order to obtain satisfactory immunostaining. Under the conditions employed, peroxisomes were distinctly visualized in liver parenchymal cells with no reaction in bile duct epithelial or sinusoidal lining cells. In the kidney, peroxisomes were confined to the proximal tubular epithelial cells with negative staining of glomeruli, distal tubules and collecting ducts. A positive immunocytochemical reaction was obtained even in paraffin blocks stored for several years. The method offers a simple approach for detection of peroxisomes and evaluation of their various enzyme proteins in material processed routinely in histopathology laboratories and should prove useful in the investigation of the role of peroxisomes in human pathology for both prospective and retrospective studies.     

Models for antigen receptor gene rearrangement. II. Multiple rearrangement in the TCR: allelic exclusion or inclusion?

This series of papers addresses the effects of continuous Ag receptor gene rearrangement in lymphocytes on allelic exclusion. The previous paper discussed light chain gene rearrangement and receptor editing in B cells, and showed that these processes are ordered on three different levels. This order, combined with the constraints imposed by a strong negative selection, was shown to lead to effective allelic exclusion. In the present paper, we discuss rearrangement of TCR genes. In the TCR alpha-chain, allelic inclusion may be the rule rather than the exception. Several previous models, which attempted to explain experimental observations, such as the fractions of cells containing two productive TCRalpha rearrangements, did not sufficiently account for TCR gene organization, which limits secondary rearrangement, and for the effects of subsequent thymic selection. We present here a detailed, comprehensive computer simulation of TCR gene rearrangement, incorporating the interaction of this process with other aspects of lymphocyte development, including cell division, selection, cell death, and maturation. Our model shows how the observed fraction of T cells containing productive TCRalpha rearrangements on both alleles can be explained by the parameters of thymic selection imposed over a random rearrangement process.