RESUMO
Transforming growth factor-ß1 (TGF-ß1) is a multifunctional cytokine with anti-inflammatory, immunosuppressive and neuroprotective properties. The hypothalamic-pituitary-adrenal (HPA) axis and immune system exert bidirectional influences on each other, via cortisol and TGF-ß1, but the exact nature of the interaction is not well characterized. The current study examined the effects, in bonnet macaques (Macaca radiata), of two consecutive acute confinement stress periods in an unfamiliar room while mildly restrained, first without and then with dexamethasone pretreatment (0.01 mg/kg IM). Preceding the confinement studies, a non-stress control condition obtained contemporaneous levels of cortisol and TGF-ß1 in both plasma and cerebrospinal fluid (CSF) to match the confinement stress studies. Subjects were reared under either normative or variable foraging demand (VFD) conditions. Since there were no rearing effects at baseline or for any of the conditions tested -- either for cortisol or TGF-ß -- the study analyses were conducted on the combined rearing groups. The stress condition increased both plasma and CSF cortisol levels whereas dexamethasone pretreatment decreased cortisol concentrations to below baseline levels despite stress. The stress condition decreased TGF-ß1 concentrations only in CSF but not in serum. Together the data suggested that stress-induced reductions of a centrally active neuroprotective cytokine occurs in the face of HPA axis activation, potentially facilitating glucocortoid-induced neurotoxicity. Stress-induced reductions of neuroprotective cytokines prompts exploration of protective measures against glucocorticoid-induced neurotoxicity.
RESUMO
BACKGROUND: In multiple myeloma (MM), increased neoangiogenesis contributes to tumor growth and disease progression. Increased levels of endothelial progenitor cells (EPCs) contribute to neoangiogenesis in MM, and, importantly, covary with disease activity and response to treatment. In order to understand the mechanisms responsible for increased EPC levels and neoangiogenic function in MM, we investigated whether these cells were clonal by determining X-chromosome inactivation (XCI) patterns in female patients by a human androgen receptor assay (HUMARA). In addition, EPCs and bone marrow cells were studied for the presence of clonotypic immunoglobulin heavy-chain (IGH) gene rearrangement, which indicates clonality in B cells; thus, its presence in EPCs would indicate a close genetic link between tumor cells in MM and endothelial cells that provide tumor neovascularization. METHODS: A total of twenty-three consecutive patients who had not received chemotherapy were studied. Screening in 18 patients found that 11 displayed allelic AR in peripheral blood mononuclear cells, and these patients were further studied for XCI patterns in EPCs and hair root cells by HUMARA. In 2 patients whose EPCs were clonal by HUMARA, and in an additional 5 new patients, EPCs were studied for IGH gene rearrangement using PCR with family-specific primers for IGH variable genes (VH). RESULTS: In 11 patients, analysis of EPCs by HUMARA revealed significant skewing (> or = 77% expression of a single allele) in 64% (n = 7). In 4 of these patients, XCI skewing was extreme (> or = 90% expression of a single allele). In contrast, XCI in hair root cells was random. Furthermore, PCR amplification with VH primers resulted in amplification of the same product in EPCs and bone marrow cells in 71% (n = 5) of 7 patients, while no IGH rearrangement was found in EPCs from healthy controls. In addition, in patients with XCI skewing in EPCs, advanced age was associated with poorer clinical status, unlike patients whose EPCs had random XCI. CONCLUSION: Our results suggest that EPCs in at least a substantial subpopulation of MM patients are related to the neoplastic clone and that this is an important mechanism for upregulation of tumor neovascularization in MM.
Assuntos
Células Endoteliais/fisiologia , Mieloma Múltiplo/genética , Mieloma Múltiplo/fisiopatologia , Células-Tronco Multipotentes/fisiologia , Neovascularização Patológica/fisiopatologia , Fatores Etários , Células Clonais , Feminino , Rearranjo Gênico , Humanos , Cadeias Pesadas de Imunoglobulinas/genética , Prognóstico , Receptores Androgênicos/análise , Inativação do Cromossomo XRESUMO
OBJECTIVES: To investigate the long-term prognostic significance of baseline plasma MMP-1 levels in a group of well-characterized male patients with known or suspected coronary artery disease, including those presenting with acute coronary syndrome. BACKGROUND: MMP-1 is an interstitial collagenase that is considered the primary enzyme responsible for collagen degradation. In addition, MMP-1 can lead to platelet activation through the PAR1 pathway that is independent of thrombin. METHODS: Baseline plasma MMP-1 levels were measured in 364 male patients who were referred for coronary angiography and followed prospectively for five years for the development of all-cause mortality. RESULTS: After adjustment for a variety of baseline clinical, angiographic and laboratory parameters, baseline plasma MMP-1 levels (analyzed as a continuous variable) were an independent predictor of all-cause mortality at 5 years (HR, 1.49; 95% CI, 1.23-1.80; P < 0.0001). Furthermore, in 3 additional multivariate models that included a wide variety of contemporary biomarkers with established prognostic efficacy (i.e., ST2, GDF-15, Cystatin C, hs-CRP, Myeloperoxidase, NT-proBNP, TIMP-1, Adiponectin, RDW, hemoglobin, and Erythropoietin), MMP-1 remained an independent predictor of all-cause mortality at 5 years. Similar results were obtained when the analyses were restricted to the subpopulation of patients presenting with acute coronary syndrome. CONCLUSIONS: Elevated levels of MMP-1 are associated with an increased risk of long-term all-cause mortality in patients with known or suspected coronary disease that is independent of a variety of clinical, angiographic, and laboratory variables, including a whole host of contemporary biomarkers with established prognostic efficacy representing multiple different pathophysiologic processes.
Assuntos
Angiografia Coronária , Doença da Artéria Coronariana/sangue , Metaloproteinase 1 da Matriz/sangue , Síndrome Coronariana Aguda/sangue , Síndrome Coronariana Aguda/mortalidade , Idoso , Biomarcadores/sangue , Colágeno/metabolismo , Doença da Artéria Coronariana/mortalidade , Humanos , Estimativa de Kaplan-Meier , Masculino , Pessoa de Meia-Idade , Ativação Plaquetária , Prognóstico , Trombina/metabolismoRESUMO
The pharmacokinetics of digoxin and metildigoxin were investigated in geriatric patients on maintenance treatment. Minimum serum glycoside concentrations were determined on 3 consecutive days, and the elimination rate over a withdrawal period of 4 to 6 days was studied. In patients with serum creatinine levels of less than or equal to 1.3 mg/dl, the oral standard dose D1) of digoxin necessary for a minimum serum concentration of 1.0ng/ml was 1.4 times higher than that of metildigoxin. There was no significant difference in the elimination rate of both glycosides. The pharmacokinetics of metildigoxin were further investigated in patients with elevated serum creatinine levels. The standard dose was best correlated with the creatinine clearance, calculated from the serum creatinine, age, weight and sex of the patients. The apparent volume of distribution of metildigoxin decreased with the drug's total body clearance.
Assuntos
Creatinina/sangue , Digoxina/análogos & derivados , Digoxina/metabolismo , Medigoxina/metabolismo , Idoso , Digoxina/sangue , Feminino , Humanos , Cinética , MasculinoRESUMO
BACKGROUND/AIMS: The endoscopic and histologic findings of gastroesophageal reflux disease are usually indistinct. The current study was designed to define accurately the histology in gastroesophageal reflux disease and to develop a hypothesis that reflux produces immunohistochemical changes. METHODS: The study was based on the examination of endoscopic esophageal biopsy specimens obtained from 20 patients with evidence of reflux with 24-hour pH-meter monitoring and from 20 control subjects without clinical or endoscopic reflux. The pathogenesis of reflux esophagitis was discussed by comparing the histopathologic changes with determined Ki-67, p53 and Bcl-2 immunoreactivity. RESULTS: In this study, the presence of esophagitis was determined endoscopically in only 55% of the patients with gastroesophageal reflux disease, while microscopic esophagitis was detected in 60% of them. No correlation was found between presence of endoscopic esophagitis and microscopic esophagitis in the patients with gastroesophageal reflux disease. There was a significant difference between control cases and the patients according to histological parameters, which included basal activity (p=0.006), height of papillae (p=0.006), intraepithelial neutrophils (p=0.000), intraepithelial eosinophils (p=0.006), congestion (p=0.001), and dilated intercellular spaces (p=0.006). Immunohistochemically, there was a significant difference in the expression of p53 and Ki-67 between the three study groups (patients with/without microscopic esophagitis, controls) (p<0.05). However, there was no difference in Bcl-2 between the patients with reflux and control cases. CONCLUSIONS: In this study, we considered that microscopic esophagitis does not always accompany reflux, and the lack of reliable diagnostic histologic criteria is still a serious problem for pathologists. Immunohistochemically, an increase in cell proliferative activity and p53 protein accumulation to repair oxidative DNA damage related to reflux were observed. However, the close Bcl-2 immunoreactivity in all groups that was indicated by a weak positivity suggests that the inhibition of apoptosis may not be involved in reflux esophagitis.
Assuntos
Refluxo Gastroesofágico/metabolismo , Refluxo Gastroesofágico/patologia , Antígeno Ki-67/biossíntese , Proteínas Proto-Oncogênicas c-bcl-2/biossíntese , Proteína Supressora de Tumor p53/biossíntese , Adulto , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-IdadeRESUMO
Angiogenesis governs the progression of multiple myeloma (MM). Circulating endothelial cells (CECs) contribute to angiogenesis and comprise mature ECs and endothelial progenitor cells (EPCs). The present study sought to characterize CECs and their relation to disease activity and therapeutic response in 31 consecutive patients with MM. CECs, identified as CD34(+)/CD146(+)/CD105(+)/CD11b(-) cells, were 6-fold higher in patients compared to controls and correlated positively with serum M protein and beta(2)-microglobulin. Circulating EPCs displayed late colony formation/outgrowth and capillary-like network formation on matrigel; these processes were inhibited after effective thalidomide treatment. Co-expression of vascular endothelial growth factor receptor-2 (KDR) and CD133 characterized EPCs in MM, and KDR mRNA elevations correlated with M protein levels. In vitro exposure of ECs to thalidomide or its derivative CC-5013 inhibited gene expression of the receptors for transforming growth factor-beta and thrombin. Thus, elevated levels of CECs and EPCs covary with disease activity and response to thalidomide, underscoring the angiogenic aspect of MM and suggesting that angioblastlike EPCs are a pathogenic biomarker and a rational treatment target in MM. The results also highlight the anti-angiogenic properties of thalidomide and CC-5013 and further elucidate possible mechanisms of their effectiveness against MM. (Blood. 2005;105:3286-3294).
Assuntos
Biomarcadores Tumorais , Endotélio Vascular/patologia , Mieloma Múltiplo/patologia , Células Neoplásicas Circulantes/patologia , Células-Tronco/patologia , Talidomida/análogos & derivados , Adulto , Idoso , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/uso terapêutico , Células Cultivadas , Endotélio Vascular/efeitos dos fármacos , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Lenalidomida , Masculino , Pessoa de Meia-Idade , Mieloma Múltiplo/tratamento farmacológico , Neovascularização Patológica/patologia , Talidomida/farmacologia , Talidomida/uso terapêutico , Veias Umbilicais/citologiaRESUMO
The biological availability of digitoxin from Lanicor was compared with that from two different galenical preparations of Card-Lamuran (Card-Lamuran and MF708d both containing equal amounts of active ingredients: 0,125 mg digitoxin and 10 mg raubasine). The patients who were kept on their individually adjusted oral digitoxin maintenance dosage received the three preparations in a randomised order. Additionally, the equivalent dosages of Lanicor and Lanitop were determined from the data on their biological availability. In 24 patients with heart failure (mean age 70.5 years), radioimmunoassay of the glycoside concentration in the serum was performed. The patients were cardially well compensated with Lanicor and it could be assumed that there would be no change in the daily maintenance dosage for the entire period of the study (42 days). Our results show that digitoxin had the same bio-availability from Lanicor and the two different galenical preparations of Card-Lamuran and MF708d. Patients can therefore safely be switched from one of these preparations to the other. On average, doses of Lanicor 1.55 times higher than those of Lanitop must be given to obtain the same serum glycoside concentrations. The variation of this factor was no greater than the variation in serum concentrations of digitoxin during continued maintenance therapy with Lanicor. The mean serum concentrations of digitoxin under maintenance therapy in our geriatric patients (mean value 2.1 mg/ml) were higher than the digitoxin concentrations published in the literature for younger patients (average 1.4 ng/ml). The calculated daily maintenance doses providing a digitoxin concentration of 1.4 mg/ml were ca. 0.3 mg Lanicor and ca. 0.2 mg Lanitop. This is somewhat less than generally assumed. This agrees with the clinical experience that the glycoside maintenance dosage in elderly patients is generally less than in middle-aged patients.
Assuntos
Disponibilidade Biológica , Biofarmácia , Digitoxina/sangue , Fatores Etários , Idoso , Creatinina/sangue , Digitoxina/administração & dosagem , Digitoxina/análogos & derivados , Digitoxina/uso terapêutico , Formas de Dosagem , Combinação de Medicamentos , Feminino , Insuficiência Cardíaca/tratamento farmacológico , Humanos , Assistência de Longa Duração , Masculino , Metilação , Pessoa de Meia-Idade , Radioimunoensaio , Ioimbina/administração & dosagemRESUMO
Single doses of beta-methyl-digoxin 0.4 mg were given to groups of 17-18 healthy volunteers as an intravenous infusion lasting 2 hours, or orally as Lanitop Liquidum or Lanitop tablets. The serum glycoside concentration and urinary glycoside excretion were measured over 8 and 32 h. The absolute bioavailability from the oral preparations in comparison with the infusion was lower for the first 8 h than for the entire 32 h of the investigation; the relative bioavailability from tablets was the same as from the solution for both periods. For both periods the area under the serum concentration/time curve and the urinary glycoside excretion were significantly lower after administration of the tablets than after intravenous infusion. Taking the average of both parameters, the absolute bioavailability of beta-methyl-digoxin was about 80% from the solution and about 70% from the tablets. In 18 patients undergoing intravenous or oral therapy with beta-methyl-digoxin steady state glycoside concentration were compared in a cross-over study of intravenous maintenance therapy with Lanitop ampoules or oral treatment with Lanitop tablets. For a standard daily dose of 0.2 mg beta-methyl-digoxin the serum concentrations were 1.35 +/- 0.10 ng/ml during both intravenous and oral administration. The intra-individual variation in glycoside concentration after changing from intravenous to oral maintenance therapy, or vice versa, was about the same as during continued intravenous or oral administration. It is concluded that the rate of rise of serum concentration after a single dose may be a useful indicator of the rate of absorption, but that the area under the serum concentration/time curve and the urinary glycoside excretion up to 32 h are unsuitable for determining equivalent doses of different formulations or routes of administration of digitalis glycosides.
Assuntos
Digoxina/análogos & derivados , Administração Oral , Adulto , Disponibilidade Biológica , Digoxina/administração & dosagem , Digoxina/metabolismo , Esquema de Medicação , Feminino , Humanos , Infusões Parenterais , Absorção Intestinal , Cinética , Masculino , Pessoa de Meia-IdadeRESUMO
The aim of the present investigation was to estimate the ratio of the intravenous doses of beta-methyl-digoxin and digoxin required to produce identical serum glycoside concentrations in man. 20 patients on intravenous maintenance therapy were changed from beta-methyl-digoxin to the identical dose of digoxin or vice versa. Each drug was given for 7 days. Serum concentrations 13% higher were found during administraton of beta-methyl-digoxin. Assuming a half life of 60 h after withdrawal, the dose of digoxin producing the same minimum serum concentration was estimated to be 1.16 times higher than that of beta-methyl-digoxin. 18 healthy volunteers received 0.4 mg beta-methyl- digoxin, and 23 the same dose of digoxin, as an intravenous infusion over 2 h. The serum concentrations and urinary glycoside excretion were measured over a period of 32 hrs. During the first hour after the infusion the serum concentration of digoxin declined more rapidly than that of beeta-methyl-digoxin. Thereafter, the ratio of the serum concentrtions did not change appreciably up to the end of the investigation. The area under the serum concentration/time curve was about 13% greater for beta-methyl-digoxin than for digoxin; this difference was not significant. The average renal clearance was 96 +- 9 ml for beta-methyl-digoxin, 151 +- 13 ml for digoxin. Since the total body clearance of digoxin is only about 1.16 times higher than that of beta-methyl-digoxin, the lower renal clearance of beta-methyl-digoxin must partly be compensated by higher extrarenal clearance. From the ratios of the areas under the serum concentration/time curves after single doses of beta-methyl-digoxin and digoxin, and the minimum serum concentrations during maintenance therapy, it was concluded that the dose of digoxin to produce the same average serum concentrations would be about 1.15 times higher than that of beta-methyl-dogoxin. In comparison wtih the large variations in individual dosage of digoxin and beta-methyl-digoxin, this difference is too small to be of practical importance.
Assuntos
Digoxina/análogos & derivados , Digoxina/sangue , Idoso , Digoxina/administração & dosagem , Digoxina/urina , Feminino , Meia-Vida , Humanos , Injeções Intravenosas , Rim/metabolismo , Masculino , Pessoa de Meia-Idade , Fatores de TempoRESUMO
OBJECTIVE: To investigate the relation between the degree of ST segment resolution (STR) after thrombolysis and the pressure derived collateral flow index (CFIp), determined using an intracoronary pressure measurement technique in patients with recent myocardial infarction. METHODS: 33 patients were studied. TIMI grade III flow was achieved in the infarct related artery by thrombolysis. A surface ECG was obtained on admission and 90 minutes later. The sum of ST segment elevations was measured by summing all leads with ST elevation on the baseline ECG and on the 90 minute ECG (after thrombolysis) and calculating the percentage recovery. The study population was divided into two groups, with good STR (> or = 50%; group 1) or poor STR (< 50%; group 2). After angiography, a fibreoptic pressure monitoring guidewire was advanced to the stenosis to be dilated. The CFIp was determined as the ratio [coronary wedge pressure - central venous pressure]/[mean aortic pressure - central venous pressure]. RESULTS: The mean STR on the surface ECG was 54.6% and mean (SD) CFIp was 0.25 (0.12) (range 0.10-0.41). There was an inverse correlation between the individually calculated percentage of STR and CFIp (r = -0.64, p < 0.01). The mean CFIp was lower in patients with a good STR than in those with a poor STR (0.18 (0.07) v 0.27 (0.10), p < 0.02). CONCLUSIONS: Although TIMI grade III flow was achieved after thrombolysis, a worse STR on the surface ECG was associated with higher CFIp measured in the infarct related artery. CFIp appears to reflect the degree of microvascular obstruction by quantifying impedance of the microvasculature.
Assuntos
Circulação Coronária/fisiologia , Infarto do Miocárdio/fisiopatologia , Terapia Trombolítica , Pressão Sanguínea/fisiologia , Circulação Colateral/fisiologia , Eletrocardiografia , Feminino , Humanos , Masculino , Microcirculação/fisiologia , Pessoa de Meia-Idade , Infarto do Miocárdio/terapiaRESUMO
BACKGROUND/AIMS: To evaluate the involvement of ET-1 in ischemia-reperfusion (I/R) injury. METHODS: Superior artery occlusion was performed in Wistar albino rats for 30 min followed by 2-hour (early reperfusion; ER) or 24-hour (late reperfusion; LR) reperfusion periods. RESULTS: Intestinal transit was found to be reduced in the ER and LR groups (19.0 +/- 2.5%; p < 0.001 and 72.7 +/- 6.0%; p < 0.05, respectively) compared to the control group (85. 8 +/- 2.5%), while treatment with the ET receptor antagonist bosentan (BOS; 10 mg/kg i.v.) abolished this delay in LR. Myeloperoxidase activity showed a significant increase in ER (7.07 +/- 85.70 U/g; p < 0.001) compared to control (281.16 +/- 43.23 U/g), but BOS had no effect on this increase. The protein oxidation level was found to be higher in LR (5.92 +/- 0.77 nmol/mg protein; p < 0. 05) compared to the control (3.77 +/- 0.45 nmol/mg protein), and was reversed by BOS treatment. CONCLUSION: The results of the present study imply that I/R delays intestinal transit involving an endothelin-dependent mechanism.
Assuntos
Endotelinas/metabolismo , Motilidade Gastrointestinal , Intestinos/fisiopatologia , Animais , Anti-Hipertensivos/uso terapêutico , Bosentana , Modelos Animais de Doenças , Antagonistas dos Receptores de Endotelina , Endotelinas/efeitos dos fármacos , Feminino , Motilidade Gastrointestinal/efeitos dos fármacos , Glutationa/metabolismo , Mucosa Intestinal/metabolismo , Intestinos/irrigação sanguínea , Peroxidação de Lipídeos , Masculino , Peroxidase/metabolismo , Ratos , Ratos Wistar , Traumatismo por Reperfusão/tratamento farmacológico , Traumatismo por Reperfusão/metabolismo , Traumatismo por Reperfusão/fisiopatologia , Sulfonamidas/uso terapêuticoRESUMO
Exposure of primate infants to adverse rearing conditions during the first half year of life can result in enduring behavioral, neuroendocrine, and immunologic abnormalities. However, the effects of differential rearing on cytokines, some of which can regulate immune and inflammatory responses and modulate activity of the central nervous system and the hypothalamic-pituitary-adrenal (HPA) axis, are largely unexamined. The present study explored the relationship between circulating levels of transforming growth factor-beta 1 (TGF-beta 1) and cortisol in macaques reared either normally or under conditions of variable foraging demand (VFD). Under VFD rearing, for a period of 4 months, the infants' mothers intermittently had to expend more time and effort to obtain food than did the mothers of normally reared control subjects. Two years after cessation of the rearing experience, exposure to a moderate stressor (confinement in an unfamiliar room for 90 min) induced elevated levels of serum TGF-beta 1 and plasma cortisol in VFD-reared monkeys compared to normally reared controls. The correlation between TGF-beta 1 and cortisol levels was substantially higher in the normally reared subjects. Examination of the relationship between HPA axis and immune function will improve our understanding of the pathophysiological consequences of adverse rearing.
Assuntos
Hidrocortisona/sangue , Estresse Psicológico/sangue , Estresse Psicológico/imunologia , Fator de Crescimento Transformador beta/sangue , Animais , Feminino , Macaca radiata , Masculino , Fator de Crescimento Transformador beta1RESUMO
Apolipoprotein B (apoB) is required for the assembly and secretion of triglyceride-rich lipoproteins. ApoB synthesis is constitutive, and post-translational mechanisms modulate its secretion. Transforming growth factor beta (TGF-beta) increased apoB secretion in both differentiated and nondifferentiated Caco-2 cells and decreased secretion in HepG2 cells without affecting apolipoprotein A-I secretion. TGF-beta altered apoB secretion by changing steady-state mRNA levels and protein synthesis. Expression of SMAD3 and SMAD4 differentially regulated apoB secretion in these cells. Thus, SMADs mediate dissimilar secretion of apoB in both the cell lines by affecting gene transcription. We identified a 485-bp element, 55 kb upstream of the apob gene that contains a SMAD binding motif. This motif increased the expression of chloramphenicol acetyltransferase in Caco-2 cells treated with TGF-beta or transfected with SMADs. Hence, TGF-beta activates SMADs that bind to the 485-bp intestinal enhancer element in the apob gene and increase its transcription and secretion in Caco-2 cells. This is the first example showing differential transcriptional regulation of the apob gene by cytokines and dissimilar regulation of one gene in two different cell lines by TGF-beta. In this regulation, the presence of cytokine-responsive motif in the tissue-specific enhancer element confers cell-specific response.
Assuntos
Apolipoproteínas B/genética , Apolipoproteínas B/metabolismo , Regulação da Expressão Gênica , Fator de Crescimento Transformador beta/metabolismo , Motivos de Aminoácidos , Northern Blotting , Células CACO-2 , Cloranfenicol O-Acetiltransferase/metabolismo , Proteínas de Ligação a DNA/biossíntese , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/farmacologia , Flavonoides/farmacologia , Humanos , Plasmídeos/metabolismo , RNA/metabolismo , RNA Mensageiro/metabolismo , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Proteína Smad3 , Proteína Smad4 , Fatores de Tempo , Distribuição Tecidual , Transativadores/biossíntese , Transcrição Gênica , Transfecção , Transgenes , Células Tumorais CultivadasRESUMO
The transforming growth factor-beta (TGF-beta) family of cytokines regulates vascular development and inflammatory responses. We have recently shown that exposure of human umbilical vein endothelial cells (HUVECs) to hypoxia (1% O(2)) increases gene expression and bioactivation of TGF-beta2 and induces its downstream effectors, Smad proteins (Smads), to associate with DNA. In the present study, we show that hypoxia-induced TGF-beta2 gene expression is dependent on thrombospondin-1-mediated bioactivation of latent TGF-beta. Blocking TGF-beta2 but not TGF-beta1 in hypoxic endothelial cell cultures inhibited induction of the TGF-beta2 gene, indicating that an autocrine mechanism driven by bioactivation of TGF-beta2 leads to its gene expression in hypoxic HUVECs. Exposure of HUVECs to hypoxia resulted in phosphorylation and nuclear transportation of Smad2 and Smad3 proteins as well as stimulation of transcriptional activities of Smad3 and the transcription factor hypoxia-inducible factor-1alpha and culminated in up-regulation of TGF-beta2 gene expression. Autocrine regulation of TGF-beta2 production in hypoxia may involve cross-talk between Smad3 and HIF-1alpha signaling pathways, and could be an important mechanism by which endothelial cells respond to hypoxic stress.