RESUMO
Lowering low-density lipoprotein (LDL) cholesterol levels has been proven to reduce the incidence of cardiovascular and cerebrovascular events and mortality. So far recommendations have not provided information as to a meaningful duration of cholesterol-lowering therapy and were largely guided by economic constraints and limited therapeutic options. In light of the decline in the price of statins, the essential therapeutic agent and the increased efficacy of therapeutic options, treatment can nowadays be geared to target values that can be expected to have an optimal effect even in old age. The most favorable level of LDL-cholesterol for primary prevention is around and below 100â¯mg/dl, provided continuous adherence to these low levels from adolescence onwards. With later onset of cholesterol reduction the existence of initial atheromatous deposits must be expected. Therefore, with age and the manifestation of other risk factors the optimal treatment targets increasingly converge to those for which experience has been gained from secondary prevention. Both measurements of the effect of cholesterol lowering on the volume of atheromatous plaques and of the incidence of vascular events indicate a target for LDL-cholesterol well below 70â¯mg/dl and in the range 50-60â¯mg/dl. At the onset of cholesterol lowering in advanced age, a smaller effect has to be expected but due to the increasing incidence rate of vascular events a higher number of events may be avoided; thus, the efficiency does not necessarily decrease; however, long-term studies indicate that earlier cholesterol lowering provides an advantage for more than a decade, in terms of preventing vascular disease, which tends to increase. Therefore, optimal cardiovascular prevention involves moderate measures to maintain the LDL-cholesterol below 100â¯mg/dl lifelong from childhood on.
Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases , Hiperlipidemias , Idoso , LDL-Colesterol , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Hiperlipidemias/tratamento farmacológico , Prevenção Primária , Fatores de Risco , Prevenção SecundáriaRESUMO
Guidelines for the reduction of cholesterol to prevent atherosclerotic vascular events were recently released by the American Heart Association and the American College of Cardiology. The authors claim to refer entirely to evidence from randomized controlled trials, thereby confining their guidelines to statins as the primary therapeutic option. The guidelines derived from these trials do not specify treatment goals, but refer to the percentage of cholesterol reduction by statin medication with low, moderate, and high intensity. However, these targets are just as little tested in randomized trials as are the cholesterol goals derived from clinical experience. The same applies to the guidelines of the four patient groups which are defined by vascular risk. No major statin trial has included patients on the basis of their global risk; thus the allocation criteria are also arbitrarily chosen. These would actually lead to a significant increase in the number of patients to be treated with high or maximum dosages of statins. Also, adhering to dosage regulations instead of cholesterol goals contradicts the principles of individualized patient care. The option of the new risk score to calculate lifetime risk up to the age of 80 years in addition to the 10-year risk can be appreciated. Unfortunately it is not considered in the therapeutic recommendations provided, despite evidence from population and genetic studies showing that even a moderate lifetime reduction of low-density lipoprotein (LDL) cholesterol or non-HDL cholesterol has a much stronger effect than an aggressive treatment at an advanced age. In respect to secondary prevention, the new American guidelines broadly match the European guidelines. Thus, the involved societies from Germany, Austria and Switzerland recommend continuing according to established standards, such as the EAS/ESC guidelines.
Assuntos
Anticolesterolemiantes/administração & dosagem , Aterosclerose/sangue , Aterosclerose/prevenção & controle , Dietoterapia/normas , Hipercolesterolemia/sangue , Hipercolesterolemia/prevenção & controle , Guias de Prática Clínica como Assunto , Áustria , Cardiologia/normas , Humanos , Fatores de Risco , SuíçaRESUMO
OBJECTIVE: Type III Hyperlipoproteinemia is a rare lipid disorder with a frequency of 1-5 in 5000. It is characterized by the accumulation of triglyceride rich lipoproteins and patients are at increased risk of developping atherosclerosis. Type III HLP is strongly associated with the homozygous presence of the ε2 allele of the APOE gene. However only about 10% of subjects with APOE2/2 genotype develop hyperlipidemia and it is therefore assumed that further genetic and environmental factors are necessary for the expression of disease. It has recently been shown that variation in the APOA5 gene is one of these co-factors. The aim of this study is to investigate the development of cerebrovascular athero?sclerosis in patients with Type III hyperlipopro?teinemia (Type III HLP) and the role of variation in the APOA5 gene as a risk factor. METHODS: 60 patients with type III hyperlipidemia and ApoE2/2 genotype were included in the study after informed consent. The presence of cerebrovascular atherosclerosis was investigated using B-mode ultrasonography of the carotid artery. Serum lipid levels were measured by standard procedures.The APOE genotype and the 1131T>C and S19W SNPs in the APOA5 gene and the APOC3 sstI SNP were determined by restriction isotyping. Allele frequencies were determined by gene counting and compared using Fisher's exact test. Continuous variables were compared using the Mann Whitney test. A p value of 0.05 or below was considered statistically significant. Analysis was performed using Statistica 7 software. RESULTS: The incidence of the APOA5 SNPs, -1131T>C and S19W and the APOC3 sstI SNP were determined as a potential risk modifier. After correction for conventional risk factors, the C allele of the -1131T>C SNP in the APOA5 gene was associated with an increased risk for the development of carotid plaque in patients with Type III HLP with an odds ratio of 3.69. Evaluation of the genotype distribution was compatible with an independent effect of APOA5. CONCLUSIONS: The development of atherosclerosis in patients with Type III HLP is modulated by variation in the APOA5 gene.
Assuntos
Apolipoproteínas A/genética , Predisposição Genética para Doença , Hiperlipoproteinemia Tipo III/genética , Arteriosclerose Intracraniana/genética , Polimorfismo de Nucleotídeo Único , Adulto , Idoso , Apolipoproteína A-V , Artérias Carótidas/diagnóstico por imagem , Artérias Carótidas/patologia , Doenças das Artérias Carótidas/sangue , Doenças das Artérias Carótidas/diagnóstico por imagem , Doenças das Artérias Carótidas/genética , Feminino , Genótipo , Humanos , Hiperlipoproteinemia Tipo III/sangue , Arteriosclerose Intracraniana/sangue , Arteriosclerose Intracraniana/patologia , Masculino , Pessoa de Meia-Idade , UltrassonografiaRESUMO
The -1131T>C polymorphism in the newly identified apolipoprotein A5 (APOA5) gene has been associated with elevated plasma triglycerides. We determined its incidence in 915 patients attending a lipid outpatient clinic. The frequency of the C allele was significantly higher in patients with triglycerides above the 90th percentile and patients with type III hyperlipidemia compared to those with hypercholesterolemia. The C allele was associated with increased plasma triglycerides and decreased plasma HDL cholesterol, conditions associated with an increased risk of coronary heart disease. The effects on plasma lipids were only observed in overweight (BMI>25) patients and were greater in patients who were also carriers of a least one epsilon4 allele in the APOE gene. Thus additional genetic and/or metabolic factors are required in order for the triglyceride raising and HDL lowering effect of the -1131T>C polymorphism in APOA5 to be expressed.
Assuntos
Apolipoproteínas/genética , Hiperlipidemias/genética , Polimorfismo de Nucleotídeo Único/genética , Triglicerídeos/sangue , Alelos , Apolipoproteína A-V , Apolipoproteínas A , Índice de Massa Corporal , Colesterol/sangue , Doença das Coronárias/sangue , Doença das Coronárias/genética , Feminino , Frequência do Gene , Genótipo , Humanos , Hiperlipidemias/sangue , Incidência , Masculino , Pessoa de Meia-IdadeRESUMO
Synthetic GH secretagogues stimulate GH release through binding to a recently cloned specific GH secretagogue receptor (GHS-R). The endogenous ligand of this receptor may be part of a new endocrine pathway controlling GH secretion. Two different receptor variants, type 1a and 1b, have been described that differ in their 3'-terminal amino acids. We investigated the genomic structure and transcriptional regulation of the human GHS-R. An 18-kb genomic clone including sequences encoding for the two GHS-R variants was isolated. Sequencing revealed that the two variants originate from specific RNA processing of a single gene that spans approximately 4.1 kb. The transcription start site was defined by 5'-inverse PCR analysis at position -227. RT-PCR analysis points to differential transcriptional initiation and processing. Type 1a is encoded by two exons; 2152 bp of intronic sequence are removed by splicing at position 796/797 relative to the translation start site. Type 1b is encoded by a single exon. A putative polyadenylation signal consensus motif was identified at position +4118; 2.7 kb of the 5'-flanking region were sequenced, and putative transcription factor binding sites were identified. Transcriptional regulation was investigated by transient transfections using promoter fragments ranging in size from 168-1745 bp; 1745 bp of the GHS-R promoter directed significant levels of luciferase expression in GH(4) rat pituitary cells, whereas no activity was detected in monkey kidney COS-7 cells, human endometrium Skut-1B cells, mouse hypothalamic LHRH neuronal GT1-7 cells, or mouse corticotroph pituitary AtT20 cells. A minimal 309-bp promoter allowed pituitary-specific expression. Its activity in COS-7 cells was enhanced by cotransfection of the pituitary-specific transcription factor Pit-1. We did not find any regulation of the GHS-R promoter by forskolin, somatostatin, insulin-like growth factor I, or 12-O-tetraphorbol 12-myristate 13-acetate. Thyroid hormone and estrogen lead to a significant stimulation; glucocorticoids lead to a significant inhibition. Further mapping suggests a thyroid hormone-responsive element, an estrogen-responsive element, and a glucocorticoid-responsive element located between -309 and the translation start codon. These studies demonstrate the nature of the human GHS-R gene and identify its 5'-flanking region. Furthermore, pituitary-specific activity of the promoter and regulation by various hormones are demonstrated.
Assuntos
Regulação da Expressão Gênica , Receptores de Superfície Celular/genética , Receptores Acoplados a Proteínas G , Sequência de Aminoácidos , Animais , Sequência de Bases , Linhagem Celular , Códon , Proteínas de Ligação a DNA/farmacologia , Estradiol/farmacologia , Expressão Gênica , Regulação da Expressão Gênica/efeitos dos fármacos , Glucocorticoides/farmacologia , Haplorrinos , Humanos , Luciferases/genética , Camundongos , Dados de Sequência Molecular , Regiões Promotoras Genéticas , Biossíntese de Proteínas , Ratos , Receptores de Superfície Celular/química , Receptores de Superfície Celular/fisiologia , Receptores de Grelina , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Análise de Sequência de DNA , Fator de Transcrição Pit-1 , Fatores de Transcrição/farmacologia , Transcrição Gênica , Transfecção , Tri-Iodotironina/farmacologiaRESUMO
Hypothalamic corticotropin-releasing hormone (CRH) is the principal regulator of the hypothalamus-pituitary-adrenal axis in mammals. In addition, immunoreactive CRH is also present at peripheral sites, where it is thought to act as a proinflammatory peptide. However, the source of peripheral CRH has remained obscure. Human lymphocytes were shown to produce immunoreactive CRH, yet the data on CRH mRNA expression in these cells are equivocal. More recently, Vaughan et al. discovered a new member of the CRH family, termed urocortin. Urocortin was shown to act through the same receptors as CRH. The current study was designed to investigate both mRNA and protein expression of CRH and urocortin in human lymphocytes. Using a commercial CRH(1-41) radioimmunoassay, we demonstrate that normal human lymphocytes and Jurkat T lymphoma cells produce significant amounts of immunoreactive peptide. However, no CRH mRNA was detectable by RT-PCR in these cells. In contrast, a band of the correct size and sequence was amplified with urocortin-specific primers. Immunocytochemical analysis of human lymphocytes using antibodies that could distinguish between CRH and urocortin revealed significant expression of urocortin but not of CRH, consistent with our RT-PCR data. We conclude that human lymphocytes produce urocortin, but not CRH.
Assuntos
Hormônio Liberador da Corticotropina/biossíntese , Linfócitos/metabolismo , Células Cultivadas , Hormônio Liberador da Corticotropina/genética , Expressão Gênica , Humanos , Células Jurkat/metabolismo , Reação em Cadeia da Polimerase , RNA Mensageiro/análise , UrocortinasRESUMO
The immunosuppressive effects of glucocorticoids (GC) have led to their wide application in the treatment of inflammatory and autoimmune states. However, long term GC treatment is associated with severe side-effects. The development of agents displaying a more favorable ratio of wanted and unwanted GC effects, is, therefore, a major goal of pharmacological and clinical research. In this study, the progesterone receptor agonist medroxyprogesterone acetate (MPA), which also binds to the glucocorticoid receptor (GR), was tested with regard to its immunosuppressive properties. Using a recently established electroporation protocol, we show that MPA (but not progesterone) can suppress a human interleukin-2 (IL-2) promoter-luciferase construct to the same extent as the synthetic GC dexamethasone in normal human lymphocytes. MPA also markedly suppressed IL-2 (as well as IL-1 and IL-6) release, as assessed by specific enzyme-linked immunosorbent assays. In contrast, a highly dexamethasone-inducible glucocorticoid response element-driven promoter construct was only marginally stimulated by MPA in both normal human lymphocytes and HeLa cells. RT-PCR and Western blot analysis of normal human lymphocytes revealed that they do not express progesterone receptor messenger ribonucleic acid and protein, respectively. In contrast, the GR protein was clearly detectable in all samples and was shown to mediate the effects of MPA in transfected Jurkat T lymphoma cells. Our data indicate that 1) MPA can transrepress the human IL-2 gene in normal human lymphocytes in the absence of significant trans-activation; and 2) this effect is mediated by GR. Because of its dissociative GC activity, MPA is a highly promising substance for the treatment of inflammatory/autoimmune states.
Assuntos
Glucocorticoides/farmacologia , Linfócitos/efeitos dos fármacos , Acetato de Medroxiprogesterona/farmacologia , Western Blotting , Células Cultivadas , Expressão Gênica/efeitos dos fármacos , Glucocorticoides/efeitos adversos , Células HeLa , Humanos , Interleucina-2/biossíntese , Interleucina-2/genética , Interleucina-6/biossíntese , Células Jurkat , Linfócitos/imunologia , Acetato de Medroxiprogesterona/metabolismo , Regiões Promotoras Genéticas , RNA Mensageiro/análise , Receptores Androgênicos/fisiologia , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Receptores de Progesterona/genética , Receptores de Progesterona/fisiologia , Proteínas Recombinantes de Fusão , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Ativação Transcricional/efeitos dos fármacos , TransfecçãoRESUMO
The effects of fluvastatin and bezafibrate on lipids, lipoproteins, and apoproteins (apo) were investigated in a multicenter randomized, double-blind, parallel-group study. After 8 weeks of strictly controlled (computer-based assessment) dietary stabilization, patients with primary hypercholesterolemia (low-density lipoprotein cholesterol [LDL-C] > or = 160 mg/dL; triglycerides < or = 300 mg/dL) were enrolled into a 6-week placebo phase. Altogether, 131 patients were randomized to receive either fluvastatin at 40 mg once daily (n = 64; mean age 53 years) or bezafibrate at 400 mg once daily (n = 67; mean age 52 years) for 12 weeks. Compliance with the diet was monitored (3-day food records) after 6 and 12 weeks. Fluvastatin led to significant reductions in LDL-C (-23%), total cholesterol (-17%), LDL-C/high-density lipoprotein cholesterol (HDL-C) (-24%) and apo B (-19%). Fluvastatin significantly increased LpA-I (+8%) and apo E (+20%). Bezafibrate produced significant reductions in LDL-C (-17%), total cholesterol (-13%), LDL-C/HDL-C (-24%), triglycerides (-28%), apo B (-15%), and LpA-I (-10%) and significantly increased HDL-C (+12%), apo A-I (+9%), apo A-II (+30%), apo E (+14%), and Lp(a) (+3%). No clinically notable increases in levels of liver enzymes or creatine phosphokinase were observed with either treatment. Both treatments were well tolerated. There was a low incidence of adverse events that tended to be mild and included headache, muscular pain, angina, and dyspepsia. The frequency of adverse events was similar in both treatment groups, and no significant differences in dietary behavior were observed. In conclusion, fluvastatin is a well tolerated 3-hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitor for the treatment of primary hypercholesterolemia. Effects of fluvastatin on LpA-I occur irrespective of changes in HDL-C.
Assuntos
Anticolesterolemiantes/uso terapêutico , Bezafibrato/uso terapêutico , Ácidos Graxos Monoinsaturados/uso terapêutico , Hipercolesterolemia/tratamento farmacológico , Indóis/uso terapêutico , Lipídeos/sangue , Adulto , Idoso , Análise de Variância , Anticolesterolemiantes/efeitos adversos , Bezafibrato/efeitos adversos , Método Duplo-Cego , Ácidos Graxos Monoinsaturados/efeitos adversos , Feminino , Fluvastatina , Humanos , Inibidores de Hidroximetilglutaril-CoA Redutases , Hipercolesterolemia/sangue , Indóis/efeitos adversos , Masculino , Pessoa de Meia-Idade , Resultado do TratamentoRESUMO
Somatostatin exerts inhibitory effects on virtually all endocrine and exocrine secretions. The somatostatin receptor subtype 2 (sst2) acts as a critical molecule for growth hormone regulation and cell proliferation. We investigated the structure and regulation of the human sst2 gene. A genomic clone including the sst2 gene was isolated, 1.5 kb of the promoter was sequenced and putative transcription factor binding sites were identified. The transcription start site was located 93 nucleotides upstream of the translation start site. The nucleotide sequences of the complete gene and 0.5 kb of 3' region were determined. A possible polyadenylation signal was identified. Transcriptional regulation was investigated by transient transfections using various promoter fragments. A -1100 sst2 promoter directed significant levels of luciferase expression in GH4 rat pituitary cells and Skut1-B endometrium cells whereas only low activity was detected in JEG3 chorion carcinoma cells or COS-7 monkey kidney cells. A minimal -252 promoter allowed cell specific expression. We did not find any regulation of the sst2 promoter by somatostatin, forskolin, TRH, TPA, T3, and 17beta-estradiol. Glucocorticoids lead to a significant inhibition of sst2 promoter activity. Further mapping suggest a glucocorticoid-responsive element between -905 and -707 and between -252 and -163. These studies demonstrate the nature of the human sst2 gene and identify its 5' and 3' flanking regions. Furthermore, specific activity of the promoter and regulation by various hormones is demonstrated.
Assuntos
Receptores de Somatostatina/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Células COS , Clonagem Molecular , Códon de Iniciação , Regulação da Expressão Gênica , Biblioteca Genômica , Glucocorticoides/farmacologia , Haplorrinos , Humanos , Dados de Sequência Molecular , Neoplasias Hipofisárias/genética , Placenta , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Ratos , Análise de Sequência de DNA , Transcrição Gênica/genética , Células Tumorais CultivadasRESUMO
The immunosuppressive effects of glucocorticoids are largely due to transcriptional inhibition of immunologically relevant genes, such as the interleukin-2 (IL-2) gene. These effects are mediated by the intracellular glucocorticoid receptor (GR). In humans, alternative splicing of the GR precursor mRNA gives rise to two receptor isoforms, termed GRalpha and GRbeta. We previously demonstrated that GRbeta could antagonize GRalpha-mediated transactivation of a glucocorticoid-responsive element (GRE)-driven reporter gene in COS-7 cells. The present study was designed to analyze the roles of the two GR isoforms on glucocorticoid-mediated transrepression of the IL-2 gene. Using a recently developed transfection technique, we demonstrate that in primary human lymphocytes, stimulation of a 548 bp IL-2 promoter-luciferase reporter construct by phorbol ester and calcium ionophore is reversed by dexamethasone to a similar extent as in Jurkat T lymphoma cells transfected with a GRalpha expression vector. Transfection of a GRbeta expression vector alone did not result in IL-2 promoter repression in response to glucocorticoids. Furthermore, GRbeta did not antagonize the repressive effects of GRalpha on IL-2 promoter activity. Surprisingly, overexpression of GRbeta in Jurkat cells did not cause significant inhibition of GRalpha-induced transactivation of a GRE-dependent luciferase reporter gene either. We conclude that the transrepressive effect of glucocorticoids on IL-2 gene transcription is exclusively mediated by GRalpha. GRbeta can neither antagonize GRalpha-mediated transactivation nor transrepression in Jurkat cells, indicating a cell type-specific pattern of GRbeta-mediated antiglucocorticoid activity.
Assuntos
Dexametasona/farmacologia , Regulação da Expressão Gênica/fisiologia , Glucocorticoides/farmacologia , Interleucina-2/genética , Receptores de Glucocorticoides/fisiologia , Cálcio/metabolismo , Células Cultivadas , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Ionóforos/farmacologia , Células Jurkat , Linfócitos , Proteínas Recombinantes de Fusão , Acetato de Tetradecanoilforbol/farmacologia , Ativação Transcricional/efeitos dos fármacos , Ativação Transcricional/fisiologia , TransfecçãoRESUMO
It is well established that steroidogenesis in the adrenal cortex is regulated by extraadrenal factors, such as ACTH and angiotensin II. However, over the last years, it has become increasingly clear that paracrine and autocrine mechanisms are also important for steroid synthesis in the adrenal gland. The current study was designed to analyze whether the pleiotropic cytokine leukemia inhibitory factor (LIF) and/or its receptor (LIF-R) are expressed in the normal human adrenal cortex, and whether they may play a role in regulating steroidogenesis. Using LIF- and LIF-R-specific primers, we show by RT-PCR that both mRNAs are expressed in this tissue, as well as in the NCI-H295 adrenal carcinoma cell line. The correct sequences of the PCR products were verified by restriction enzyme analysis and DNA sequencing. Immunohistochemistry, employing specific antibodies against LIF and LIF-R, reveals expression of both proteins in the normal human adrenal cortex. Finally, we show that LIF can significantly enhance basal and ACTH-induced production of cortisol and aldosterone in NCI-H295 cells. In summary, we show for the first time that LIF and its receptor are expressed in the normal human adrenal cortex. Our stimulation experiments indicate that the intraadrenal LIF/LIF-R system may participate in regulating adrenal steroidogenesis.
Assuntos
Córtex Suprarrenal/metabolismo , Inibidores do Crescimento/metabolismo , Interleucina-6 , Linfocinas/metabolismo , Receptores de Citocinas/metabolismo , Esteroides/biossíntese , Neoplasias das Glândulas Suprarrenais/genética , Neoplasias das Glândulas Suprarrenais/metabolismo , Aldosterona/biossíntese , Sequência de Bases , Primers do DNA/genética , Expressão Gênica , Inibidores do Crescimento/genética , Humanos , Hidrocortisona/biossíntese , Fator Inibidor de Leucemia , Subunidade alfa de Receptor de Fator Inibidor de Leucemia , Linfocinas/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Citocinas/genética , Receptores de OSM-LIF , Células Tumorais CultivadasRESUMO
The molecular mechanisms leading to increased cellular proliferation rates and, thus, tumor formation in the anterior pituitary gland are poorly understood. The cyclin-dependent kinase inhibitor p27Kip1 is a key molecule regulating the G1 phase of the cell cycle in many cell types. Furthermore, it was shown that p27 knock-out mice develop pro-opiomelanocortin-positive pituitary tumors. In an effort to clarify the role of p27 in the normal and tumorous human pituitary, we studied the expression of p27 by immunohistochemistry, using a highly specific mouse monoclonal anti-human p27 antibody. Normal pituitaries and 54 pituitary adenomas (twelve somatotrope adenomas, nine prolactinomas, twelve corticotrope adenomas, three TSH-producing tumors, six gonadotrope adenomas, six null cell adenomas, and six oncocytomas) were analyzed. p27 expression was determined semiquantitatively with regard to both the percentage of positive cells and the intensity of the staining. Normal human pituitaries showed strong expression of p27 in most nuclei. In contrast, the levels of p27 were reduced in the majority of the tumors analyzed. Twenty-two tumors (six somatotrope adenomas, five prolactinomas, four corticotrope adenomas, two TSH-producing tumors, two gonadotrope adenomas, and three null cell adenomas) were completely p27-negative. In 18 tumors, p27 expression was found in < or = 10% of the cells. In the other ten tumors, 11-80% of the cells were p27-positive. In summary, we were able to demonstrate reduced expression levels of the cell-cycle inhibitor p27 in tumors derived from all pituitary cell types. Our data indicate that p27 may be an important regulator of cellular proliferation in the anterior pituitary, the underexpression of which could play a role in pituitary tumorigenesis.
Assuntos
Adenoma/genética , Proteínas de Ciclo Celular , Regulação da Expressão Gênica no Desenvolvimento , Proteínas Associadas aos Microtúbulos/genética , Neoplasias Hipofisárias/genética , Proteínas Supressoras de Tumor , Adenoma/metabolismo , Anticorpos Monoclonais , Inibidor de Quinase Dependente de Ciclina p27 , Quinases Ciclina-Dependentes/antagonistas & inibidores , Inibidores Enzimáticos/química , Humanos , Imuno-Histoquímica , Proteínas Associadas aos Microtúbulos/biossíntese , Hipófise/citologia , Hipófise/patologia , Neoplasias Hipofisárias/metabolismoRESUMO
Due to their immunosuppressive effects, glucocorticoids (GC) are widely used in the treatment of inflammatory and autoimmune states. However, long-term GC treatment is associated with severe side effects. To increase the ratio of wanted and unwanted GC effects, is, therefore, a desirable goal, which could be achieved by either developing new "dissociating" GC or by combining conventional GC therapy with substances that selectively interfere with glucocorticoid receptor (GR) function. Vitamin B6 was previously shown to inhibit GR transactivation in non-immune cells. In the present study, we tested whether vitamin B6 would also interfere with GR function in immune cells and/or with transrepression in non-immune cells. Normal human lymphocytes and Jurkat T lymphoma cells were transfected with luciferase reporter constructs under the control of the interleukin-2 (IL-2) and the leukemia inhibitory factor (LIF) promoter, respectively. Cells were stimulated with phorbol ester, ionomycin, and different concentrations of dexamethasone, either in the absence (a vitamin B6-free medium was especially prepared for this study) or presence of vitamin B6. Both promoters were strongly induced in response to phorbol ester and ionomycin. Dexamethasone inhibited this effect in a dose-dependent manner both in the presence and absence of vitamin B6. Similar results were obtained at the protein level (IL-2- and LIF-specific ELISAs). Induction of a glucocorticoid response element (GRE)-driven promoter construct by dexamethasone in lymphoid cells was only marginally reduced by vitamin B6. In contrast, GR-mediated transactivation was strongly inhibited by vitamin B6 in HeLa cells, while GR-mediated transrepression of a matrix metalloproteinase 9 (MMP9) promoter construct was not affected. Our data indicate that vitamin B6 does not interfere with GC action in immune cells (wanted GC effects) while selectively inhibiting GR-dependent transactivation in non-immune cells (unwanted GC effects). Combination of GC treatment with supraphysiological doses of vitamin B6 may, thus, reduce the side effects of this type of immunosuppressive therapy, provided that the observed effects can be reproduced at subtoxic vitamin B6 concentrations in vivo.
Assuntos
Dexametasona/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Glucocorticoides/farmacologia , Regiões Promotoras Genéticas/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Vitamina B 6/farmacologia , Citocinas/sangue , Humanos , Células Jurkat , Linfócitos/efeitos dos fármacos , Linfócitos/imunologia , Plasmídeos , Acetato de Tetradecanoilforbol/farmacologia , Ativação Transcricional/efeitos dos fármacosRESUMO
Histological analysis of surgically removed adrenal masses often fails to differentiate between benign and malignant tumors. In normal cells, the telomeric ends of the chromosomes are shortened with each cell division, leading to chromosome destabilization and cellular senescence after a critical number of cell cycles. In tumor cells, telomere shortening is prevented by a specific DNA polymerase, called telomerase. In an effort to clarify the role of telomerase in the pathogenesis of adrenal tumors, and to test whether its activity could serve as marker of malignancy, we measured telomerase activity in 41 human adrenal tissue samples that were classified both by the clinical course and by histological examination. Telomerase activity was determined by TRAP ELISA and expressed as high (>50% of positive control telomerase activity), medium (31-50%), low (11-30%), very low (< or = 10%), or absent (0%). The 8 normal adrenal tissue samples showed very low levels of telomerase activity. Mean telomerase activity also very low in 3/3 incidentalomas, 6/6 Cushing adenomas, 6/6 Conn adenomas, 7/7 adrenocortical carcinomas, 8/8 benign pheochromocytomas, and 2/3 malignant pheochromocytomas. In contrast, one malignant pheochromocytoma showed high telomerase activity. These data indicate that telomerase activity may not be a suitable marker for malignancy in the adrenal gland. Our results also challenge the current dogma of close correlation between cell dedifferentiation and telomerase activity.
Assuntos
Neoplasias das Glândulas Suprarrenais/enzimologia , Glândulas Suprarrenais/enzimologia , Telomerase/fisiologia , Adenoma/enzimologia , Adenoma/fisiopatologia , Neoplasias das Glândulas Suprarrenais/diagnóstico , Glândulas Suprarrenais/fisiopatologia , Carcinoma Adrenocortical/enzimologia , Carcinoma Adrenocortical/fisiopatologia , Benzidinas/química , Biomarcadores Tumorais , Compostos Cromogênicos/química , Histocitoquímica , Humanos , Células Jurkat , Hibridização de Ácido Nucleico , Feocromocitoma/enzimologia , Feocromocitoma/fisiopatologia , Reação em Cadeia da Polimerase , Telomerase/análiseRESUMO
BACKGROUND: Growth hormone secretagogues (GHS) are highly potent synthetic peptides which release growth hormone (GH) by activation of a growth hormone-releasing hormone-independent signal cascade. A specific growth hormone secretagogue receptor (GHS-R) has been isolated, its endogenous ligand is still unknown. It might represent another major endocrine pathway controlling GH secretion. To gain insight into the specific function of the human GHS-R we studied the gene structure. Two variants, type 1a and 1b, have been described, but their specific functions are unknown. METHODS AND RESULTS: A specific probe for the GHS-R was cloned following reverse transcription and PCR amplification of pituitary mRNA. A genomic human placenta library was screened for the GHS-R gene. Positive clones were identified and further characterized by Southern blotting and sequencing. A genomic clone of 18 kb in size was determined to include the coding sequence of both GHS-R variants. Here we show that GHS-R type 1a and type 1b are encoded by a single gene. Sequencing of the immediate 5'-flanking region suggests a number of transcription factor binding sites, but their functional significance remains to be investigated. CONCLUSION: A genomic clone encoding for the two known variants of the human GHS-R was isolated. Further studies will determine physiological relevance and regulation of GHS-R. This will facilitate studies of GHS as diagnostic and therapeutic agents in GH disorders.
Assuntos
Proteínas de Ligação ao GTP/genética , Receptores de Superfície Celular/genética , Receptores Acoplados a Proteínas G , Humanos , Receptores de Grelina , Mapeamento por RestriçãoRESUMO
Familial hypercholesterolemia is one of the most common hereditary metabolic disorders, untreated with grave cardiovascular consequences. A general practitioner will see at least one affected individual each month, but will rarely be aware of the diagnosis, though it is easily suspected: an LDL-cholesterol ≥â 190â mg/dl, a family history of premature cardiovascular disease, or clinical signs as arcus lipoides, tendinous xanthomata, or a thickened Achilles' tendon must draw the attention to familial hypercholesterolemia. Because of the burden of high cholesterol levels from childhood on therapy should be initiated early enough, which has become greatly ameliorated since the introduction of statins. In conjunction with additional risk factors, notably low HDL-cholesterol or elevated lipoprotein(a) the cardiovascular sequelae can be dramatic and may call for more intense therapies. However, often the routine of successful cholesterol lowering covers the diagnosis nowadays, so that a heritable metabolic disorder is not suspected, which, however, prevents an effective prevention in relatives, particularly the children of the patient.
Assuntos
Doenças Cardiovasculares/diagnóstico , Doenças Cardiovasculares/terapia , Predisposição Genética para Doença/prevenção & controle , Hiperlipoproteinemia Tipo II/diagnóstico , Hiperlipoproteinemia Tipo II/terapia , Doenças Cardiovasculares/etiologia , Humanos , Hiperlipoproteinemia Tipo II/complicaçõesRESUMO
OBJECTIVE: Genomewide association studies (GWAS), conventional association studies and the characterization of families with ApoA5 deficiency have shown that variation in the apolipoprotein A5 (APOA5) gene is associated with plasma triglyceride levels. The aim of this study was to determine the frequency of rare variants in the APOA5 gene in patients with various forms of hypertriglyceridemia. METHODS: The DNA sequence of the exons plus exon/intron boundaries of the APOA5 gene of 291 patients with triglycerides above the 95th percentile for age and sex (98 of whom had triglycerides above 875 mg/dl), 111 patients with APOE2/2 genotype of whom 100 had Type III Hyperlipidemia and 108 probands with triglycerides below the 25th percentile for age and sex was determined. RESULTS: Twenty four variants were detected of which eight have been previously reported. There were nine patients with triglycerides above 875 mg/dl and nine patients with moderately elevated triglycerides who were carriers of at least one deleterious mutation in the APOA5 gene. Of the patients with Type III HLP, three (3%) were carriers of rare variants and there was a single rare variant detected in the group of probands with triglycerides below the 25th percentile for age and sex. CONCLUSION: Rare mutations in the APOA5 gene are more frequent in patients with elevated triglycerides than in those with Type III HLP.
Assuntos
Apolipoproteínas A/genética , Análise Mutacional de DNA , Hipertrigliceridemia/genética , Mutação , Adulto , Idoso , Apolipoproteína A-V , Biomarcadores/sangue , Estudos de Casos e Controles , Éxons , Feminino , Frequência do Gene , Predisposição Genética para Doença , Alemanha , Humanos , Hipertrigliceridemia/sangue , Íntrons , Masculino , Pessoa de Meia-Idade , Fenótipo , Triglicerídeos/sangue , Regulação para CimaRESUMO
OBJECTIVE: Genomewide association studies (GWAS) have shown that variation in the lipoprotein lipase gene (LPL) is associated with plasma triglyceride levels but that common variants account for only 1.25% of the variance. The aim of this study was to determine the frequency of rare variants in the LPL gene in patients with various forms of hypertriglyceridemia. METHODS: The DNA sequence of the exons plus exon/intron boundaries of the LPL gene of 313 patients with triglycerides above the 95th percentile for age and sex (107 of whom had triglycerides above 875 mg/dl) and 121 patients with Type III hyperlipidemia was determined. RESULTS: Twenty rare variants were detected of which seven have been previously reported. All of the rare variants were present as heterozygotes. Sixteen were missense mutations, two were short deletion mutants and there were single nonsense and insertion mutations. Fifteen of the missense mutations resulted in an amino acid change. There were 13 patients (12.1%) with triglycerides above 875 mg/dl and 10 patients (4.9%) with moderately elevated triglycerides, who were carriers of at least one rare, non-synonymous mutation in the LPL gene. Of the patients with Type III HLP, two were carriers of rare variants. CONCLUSION: Rare mutations in the LPL gene are frequent in patients with elevated triglycerides.
Assuntos
Hiperlipoproteinemia Tipo III/genética , Hipertrigliceridemia/genética , Lipase Lipoproteica/genética , Mutação , Polimorfismo de Nucleotídeo Único , Triglicerídeos/sangue , Adulto , Idoso , Biomarcadores/sangue , Análise Mutacional de DNA , Feminino , Frequência do Gene , Predisposição Genética para Doença , Alemanha , Heterozigoto , Humanos , Hiperlipoproteinemia Tipo III/sangue , Hiperlipoproteinemia Tipo III/enzimologia , Hipertrigliceridemia/sangue , Hipertrigliceridemia/enzimologia , Masculino , Pessoa de Meia-Idade , Fenótipo , Regulação para CimaRESUMO
OBJECTIVE: This prospective study compares MRI of atherosclerotic plaque in the abdominal aorta at 3 T with that at 1.5 T in patients suffering from hereditary hyperlipidaemia, a major risk factor for atherosclerosis. METHODS: MRI of the abdominal aorta at 1.5 and 3 T was performed in 21 patients (mean age 58 years). The study protocol consisted of proton density (PD), T(1), T(2) and fat-saturated T(2) weighted black blood images of the abdominal aorta in corresponding orientation. Two independent radiologists performed image rating. First, image quality was rated on a five-point scale. Second, atherosclerotic plaques were scored according to the modified American Heart Association (AHA) classification and analysed for field strength-related differences. Weighted κ statistics were calculated to assess interobserver agreement. RESULTS: Interobserver agreement was substantial for nearly all categories. MRI at 3 T offered superior image quality in all contrast weightings, most significantly in T(1) and T(2) weighted techniques. Plaque burden in the study collective was unexpectedly moderate. The majority of plaques were classified as AHA III lesions; no lesions were classified above AHA V. There was no significant influence of the field strength regarding the AHA classification. CONCLUSION: Abdominal aortal plaque screening is basically feasible at both field strengths, whereas the image quality is rated superior at 3 T. However, the role of the method in clinical practice remains uncertain, since substantial findings in the high-risk collective were scarce.