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Biomedical subjects

M Lorenzo

Publications and source records attributed to M Lorenzo.

At least 73 records · Page 4Linked to original sources
PubMed1996

Transforming growth factor beta 1 induces mitogenesis in fetal rat brown adipocytes.

The presence of transforming growth factor beta 1 (TGF-beta 1) for 24 or 48 h stimulated DNA synthesis, the percentage of cells in the S + G2/M phases of the cell cycle, and cell number, as compared to quiescent cells. The mitogenic capacity of TGF-beta 1 (1 pM) was similar to that shown by 10% fetal calf serum (FCS). TGF-beta 1 for 48 h increased by 5-fold the percentage of cells containing (3H)thymidine-labeled nuclei as compared to quiescent cels. In addition, single fetal brown adipocytes, showing their typical multilocular fat droplets phenotype, become positive for (3H)thymidine-labeled nuclei in response to TGF-beta 1. Moreover, TGF-beta 1 induced the mRNA expression of a complete set of proliferation-related genes, such as c-fos (30 min), c-myc and beta-actin (2 h), and H-ras, cdc2 kinase, and glucose 6-phosphate dehydrogenase (G6PD) at 4 and 8 h, as compared to quiescent cells. Concurrently, TGF-beta 1 for 12 h increased the protein content of proliferating cellular nuclear antigen (PCNA) by 6-fold and p21-ras by 2-fold. Although our results demonstrate that TGF-beta 1 induces the expression of very early genes related to cell proliferation, TGF-beta 1 could be acting either as a mitogen or as a survival factor in induce proliferation to fetal brown adipocytes.

Adipocytes↗
PubMed1996

Laparoscopic cholecystectomy (LC), intraoperative endoscopic sphincterotomy (ES), and common bile duct stones (CBDS) extraction for management of patients with cholecystocholedocholithiasis.

BACKGROUND: A combined method of endoscopic sphincterotomy (ES) with common bile duct stone (CBDS) extraction and laparoscopic cholecystectomy (LC) under general anesthesia for a single-session treatment of patients with colecysto-choledocholithiasis is described. METHODS: From June 1994 to January 1995, 15 consecutive cases considered for elective LC with preoperative diagnosis of CBDS underwent this procedure. Following orotracheal intubation, the patient is turned on the left lateral decubitus for ES and CBDS extraction. Nasobiliary drainage is positioned for per-laparoscopic cholangiogram. Routine LC is finally performed. RESULTS: These two interventions were successfully accomplished in all patients. Mean duration of the operative time for the combined procedure was 97.7 +/- 30.4 min, range 60-140 min. In four (26.6%) cases an accessory trocar with retracting instrument was used to obviate the bowel distension. CONCLUSIONS: No complications of ES or LC were observed. Mean hospital stay was 3 days (range 2-5 days). Routine follow-up (mean 3 +/- 2 months, range 1-12 months) did not reveal biliary-related problems in any of the observed patients.

Adolescent↗
PubMed1996

IGF-I: a mitogen also involved in differentiation processes in mammalian cells.

The main source of insulin-like growth factor I (IGF-I) postnatally is the liver, under growth hormone stimulation, although IGF-I is already present in embryonic tissues and in fetal serum, when its expression is independent of growth hormone. The extracellular alpha-subunit of the IGF-I receptor (IGF-IR) contains an IGF-I binding domain, and the beta-subunit possesses tyrosine kinase activity, which is greatly enhanced when IGF-I binds to the alpha-subunit and leads to its autophosphorylation. Insulin receptor substrate 1 (IRS-1) is the most well characterized cellular substrate for IGF-I, containing at least 20 potential tyrosine phosphorylation sites. The tyrosine phosphorylated form of IRS-1 acts as a docking protein by associating SH2-containing proteins including the p85 regulatory subunit of phosphatidylinositol-3-kinase (P13-kinase), the protein tyrosine phosphatase SH-PTP2, the SH2- and SH3-containing adaptor protein Nck and the growth factor receptor-bound protein-2 (Grb2/Sem5) protein. Grb2 is found associated with mSOS, a GTP/GDP exchange factor involved in converting the inactive Ras-GDP to the active Ras-GTP. The p85 regulatory subunit of PI3-kinase can be also a direct in vitro substrate of the IGF-IR. Although IRS-1 is the major substrate of the IGF-IR, there is another early phosphotyrosine substrate termed SHC, which also activates Ras via Grb2-mSos complex. Activation of p21-Ras induces a serine/threonine kinase cascade leading to the activation of MAP-kinases. The importance of IGF-I as a mitogen throughout development has been clearly demonstrated in IGF-I and IGF-IR knockout mouse studies and also in transgenic mice over-expressing IGF-I. IGF-I is a mitogen in many cell types in culture such as T lymphocytes, chondrocytes or osteoblasts and it is considered to be a progression factor in mouse fibroblasts. IGF-I is also involved in muscle, neurons and adipogenic differentiation of mesenchymal cells. However, IGF-I induces proliferation and differentiation in fetal brown adipocytes, suggesting that both cellular processes are not necessarily mutually exclusive in fetal cells.

Animals↗
PubMed1996

Initial expression of phosphoenolpyruvate carboxykinase gene in fetal hepatocytes: role of transcription factors.

BACKGROUND/AIMS: Hepatic phosphoenolpyruvate carboxykinase (PEPCK; EC 4.1.1.32) gene is absent in fetal liver. However, this gene can be initially expressed in 20-day-old fetal hepatocyte primary cultures under specific hormonal stimulation. The role of transcriptional factors involved is also studied. METHODS: Primary 20-day-old fetal hepatocytes have been cultured and Northern-blot and nuclear run-on transcription assays have been performed. RESULTS: Fetal hepatocytes in culture initially expressed PEPCK gene by dibutyryl cAMP, in the presence of dexamethasone. Dibutyryl cAMP increased by 8-fold the rate of transcription of PEPCK gene at 30 min, and produced a 50-fold increase in its mRNA content at 3 h. This induction of PEPCK expression by cAMP occurred in the presence of sustained levels of CCAAT/enhancer binding protein (C/EBP) alpha-delta mRNAs, and was accompanied by an increase in the rate of transcription and mRNA content of C/EBP beta gene, and a decrease in the expression of c-myc, in the absence of c-fos expression. In addition, insulin or phorbol esters decreased by 50% the PEPCK rate of transcription and its mRNA accumulation induced by dibutyryl cAMP. This inhibitory effect of insulin or phorbol esters on PEPCK gene expression was accompanied by an increase in the rate of transcription and mRNA content of nuclear factors such as c-fos and c-myc, the expression of C/EBPs remaining essentially unmodified.

Animals↗
PubMed1996

Involvement of Raf-1 kinase and protein kinase C zeta in insulin-like growth factor I-induced brown adipocyte mitogenic signaling cascades: inhibition by cyclic adenosine 3',5'-monophosphate.

In the present study we have examined the signaling cascades involved in insulin-like growth factor I (IGF-I)-induced mitogenesis in fetal rat brown adipocyte primary cultures, a model that constitutively expresses a high number of IGF-I receptors, where IGF-I is a complete mitogen at physiological concentrations. IGF-I rapidly stimulated beta-chain IGF-I receptor autophosphorylation, which peaked at a physiological/mitogenic concentration (1.4 nM) and also stimulated tyrosine phosphorylation of insulin receptor substrate-1 (IRS-1). Tyrosine-phosphorylated IRS-1 bound and subsequently activated phosphatidylinositol 3-kinase by 3.5-fold, whereas the tyrosine-phosphorylated IGF-I receptor was not directly associated with the p85 subunit of the phosphatidylinositol 3-kinase. Moreover, mitogenic concentrations of IGF-I enhanced glucose transport by 2.5-fold. In addition, tyrosine phosphorylation of the 46- and 52-kDa SHC proteins was high in the basal state and doubled after IGF-I treatment, whereas IGF-I enhanced by 4-fold tyrosine phosphorylation of the 66-kDa SHC band. Furthermore, a 2-fold increase in the Ras. GTP active form was induced upon IGF-I stimulation. Downstream from Ras, IGF-I increased both Raf kinase and protein kinase C (PKC) zeta activities by 3.5-fold. (Bu)2cAMP, an inhibitor of IGF-I-induced mitogenesis in fetal brown adipocyte primary cultures, did not block the very early steps of the IGF-I-induced mitogenic cascade, such as IGF-I receptor autophosphorylation, IRS-1 or SHC tyrosine phosphorylation, and Ras activation to its GTP active form. However, (Bu)2cAMP disrupted IGF-I-Raf and IGF-I-PKC zeta signaling pathways by preventing IGF-I-induced Raf-1 kinase and PKC zeta enzymatic activities, respectively. Our results show the first characterization in situ of an IGF-I mitogenic signaling cascade that downstream Ras diverges to the nucleus through two different serine/threonine kinases (Raf-1 kinase and PKC zeta) in mammalian fetal primary cells under physiological conditions. Both kinases represent a point of regulation primarily described for IGF-I-induced, cAMP-inhibited mitogenic pathways.

Adipose Tissue, Brown↗
PubMed1996

p21ras induced differentiation-related gene expression in fetal brown adipocyte primary cells and cell lines.

Transfection of primary rat fetal brown adipocytes with constructs of SV40 large T antigen, alone and together with lys12-mutated H-ras gene, gave permanent cell lines showing an immortalized or transformed phenotype, respectively, all of them selected by the expression of the uncoupling protein (UCP), a tissue-specific marker. Primary brown adipocytes and immortalized cell lines respond to insulin-like growth factor I (IGF-I) by increasing their lipid content and the mRNA expression of both the adipogenic marker fatty acid synthase (FAS) and the thermogenic marker UCP. IGF-I-induced differentiation-related gene expression at 24 h in both primary and immortalized brown adipocytes was mediated by an increase in p21ras.GTP active protein content. Transformed cell lines overexpressing exogenous p21ras (mainly in its ras.GTP active form) constitutively showed a higher lipid content and a higher FAS and UCP mRNA expression compared to primary and immortalized cells. These transformed cells were IGF-I independent with respect to their studied differentiation-related parameters. Additionally, transient transfection of primary brown adipocytes with the transforming ras gene induced UCP and FAS mRNA expression as well as cotransactivated UCP-chloramphenicol acetyltransferase fusion gene. Moreover, IGF-I transactivation of UCP promoter was partially precluded by cotransfection with the dominant-negative ras gene. Our results strongly suggest that IGF-I/p21ras induces adipogenic- and thermogenic-related gene expression in brown adipocytes.

Adipocytes↗
PubMed1995

Differentiation of rat brown adipocytes during late foetal development: role of insulin-like growth factor I.

Rat brown adipocytes at day 22 of foetal development showed greater size, higher mitochondria content and larger amounts of lipids, as determined by flow cytometry, than 20-day foetal cells. Simultaneously, an inhibition on the percentage of brown adipocytes into S+G2/M phases of the cell cycle was observed between days 20 and 22 of foetal development. The expression of several adipogenesis-related genes, such as fatty acid synthase, malic enzyme, glucose-6-phosphate dehydrogenase and insulin-regulated glucose transporter, increased at the end of foetal life in brown adipose tissue. In addition, the lipogenic enzyme activities and the lipogenic flux increased during late foetal development, resulting in mature brown adipocytes showing a multilocular fat droplet phenotype. Concurrently, brown adipocytes induced the expression of the uncoupling protein (UP) mRNA and UP protein, as visualized by immunofluorescence. The three isoforms of CCAAT enhancer-binding proteins (C/EBPs) were expressed at the mRNA level in brown adipose tissue at day 20. C/EBP alpha decreased and C/EBP beta and delta increased their expression between days 20 and 22 of foetal development, respectively. Brown adipose tissue constitutively expressed insulin-like growth factor I (IGF-I) and IGF-I receptor (IGF-IR) mRNAs. Moreover, IGF-IR mRNA content increased between days 20 and 22 in parallel with the occurrence of tissue differentiation.

Adipocytes↗
PubMed1995

Transforming growth factor beta 1 induces differentiation-specific gene expression in fetal rat brown adipocytes.

Fetal rat brown adipocytes show a low number of transforming growth factor beta 1 (TGF-beta 1) binding sites of high affinity, revealing the presence of type I, II and III TGF-beta 1 receptors and a minor-labeled species of approximately 140 kDa. The culture of cells in the presence of TGF-beta 1 induced the expression of the tissue-specific gene uncoupling protein in a dose- and time-dependent manner. In addition, TGF-beta 1 up-regulates the expression of genes involved in adipogenesis such as fatty acid synthase, glycerol 3-phosphate dehydrogenase, malic enzyme and glucose 6-phosphate dehydrogenase, as well as induces the expression of fibronectin (specific target gene for TGF-beta 1). Our results suggest that TGF-beta 1 is a major signal involved in initiating and/or maintaining the thermogenic and adipogenic differentiation of rat fetal brown adipocytes.

Adipose Tissue, Brown↗
PubMed1995

Technique of total hepatectomy in the rat.

We report a novel one-stage technique of total hepatectomy in the rat, in which the liver is replaced by an autologous prosthesis. This prosthesis is obtained from a donor rat, and consists of the subhepatic vena cava and the left renal vein harvested en block. This graft is sutured with the subdiaphragmatic cava and linked with cuffs to the subhepatic vena cava and the portal vein in the donor rat. This procedure is associated with a very low operative mortality. Rats with glucose and plasma-expander infusions survived for a mean time of 23 +/- 9 hr. This technique of total hepatectomy can be considered a valid and reproducible model of true anhepaty for metabolic and survival studies with liver support systems.

Animals↗
PubMed1995

cAMP inhibits IGF-I-induced mitogenesis in fetal rat brown adipocytes: role of p21 ras.

Dibutyryl cAMP (DBcAMP) inhibited insulin-like growth factor I (IGF-I)-induced DNA synthesis of rat fetal brown adipocyte primary cultures. Its mitogenic inhibitory capacity, measured as [3H]thymidine incorporation into acid-precipitable material, was dose-dependent, being maximal at 0.5 mM. The entry of cells into S + G2/M phases of the cell cycle and the increase in cell number induced by IGF-I were partially inhibited by DBcAMP at 24 h and totally prevented at 48 h. DBcAMP inhibited the mRNA expression of c-myc induced by IGF-I at 2 h. Moreover, DBcAMP inhibited in parallel H-ras mRNA expression, p21 ras, and proliferating cellular nuclear antigen protein content induced by IGF-I at 24 and 48 h of culture, respectively, suggesting the involvement of p21 ras in the progression of fetal brown adipocytes through the S phase of the cell cycle and DNA synthesis.

1-Methyl-3-isobutylxanthine↗
PubMed1995

Laparoscopic cholecystectomy in obese patients compared with nonobese patients.

Obese patients treated by laparoscopic cholecystectomy currently appear to be the largest risk subgroup amenable to consistent scientific evaluation. Here we report our experience and compare the results in obese patients with those obtained in nonobese patients undergoing the laparoscopic procedure. Laparoscopic cholecystectomy in obese patients was technically more difficult with significantly longer operating time (p < 0.01), but intraoperative and postoperative technical complications were not significant in the groups analyzed. Obese patients present significant anesthesiological complications (p < or = 0.001). The results of this experience and the literature review indicate that the therapeutic advantages proved in nonobese patients can be extended to the obese population.

Adult↗
PubMed1994

Glucose-6-phosphate dehydrogenase gene expression in fetal hepatocyte primary cultures under nonproliferative and proliferative conditions.

The culture of fetal hepatocytes at high cell density for 64 h in medium supplemented with 5 mM glucose produced an induction of glucose-6-phosphate dehydrogenase (G6PD) mRNA in a time-dependent manner. Insulin and triiodothyronine (T3), separately, increase G6PD mRNA expression, producing an additive effect at 64 h when combined. Glucagon and, to a greater extent, dibutyryl-cAMP decreased the G6PD mRNA expression observed in the presence of 5 mM glucose and T3. Dexamethasone repressed the G6PD mRNA expression induced by glucose and insulin and decreased this expression when induced by T3, regardless of the presence of insulin. At low cell density, EGF in the presence of dexamethasone induced in parallel DNA synthesis, G6PD mRNA content, and specific activity, while EGF failed to increase these parameters at high cell density. In addition, G6PD expression in proliferative fetal hepatocytes was unresponsive to lipogenic hormones.

Animals↗
PubMed1994

[Does Bolande's malignant tumor exist?].

Congenital mesoblastic nephroma (CMN) is a rare renal neoplasm which Bolande differentiated from Wilms tumour given its benign nature. We describe 7 cases of CMN which have been treated over the last ten years, in order to highlight to what extent a tumour which is generally considered to be benign can have an aggressive behaviour. 5 patients were diagnosed during their neonatal period (3 of them prenatally) and 2 after the third month of life. Clinical presentation, simple abdominal radiology, echography and abdominal CT scan, which showed characteristic images, led us to a final diagnosis. Tumoral resection was carried out in the 6 typical CMN. The atypical or aggressive CMN was urgently operated under critical circumstances due to spontaneous tumoral rupture. The pathological study showed a CMN of cellular variety with a sarcomatous component. Despite adjuvant chemotherapy, there was a tumoral recurrence which required a surgical second-look and subsequent chemotherapeutical treatment. Bolande's tumour is generally benign, but there is an aggressive cellular variant that fits into the intermediate zone of a pathological spectrum. This aggressive variant have different clinical, radiological and anatomo-pathological features and therefore it involves radical surgical treatment and additional chemotherapy with a rigorous follow-up in the short and the long term.

Abdomen↗
PubMed1993

Regulation of malic enzyme gene expression by nutrients, hormones, and growth factors in fetal hepatocyte primary cultures.

The culture of fetal hepatocytes for 64 h in medium supplemented with 5 mM glucose, T3, insulin, and dexamethasone resulted in the coordinate precocious expression of malic enzyme mRNA, protein, and specific activity. T3 was the main inducer; meanwhile, insulin exerted a small synergistic effect when added with T3. Dexamethasone had a potentiation effect on the T3 response of malic enzyme mRNA expression regardless of the presence of insulin. This effect of dexamethasone on T3 response of malic enzyme mRNA expression was time (64 h) and glucose dependent. Glucagon, and to a greater degree dibutyryl-cAMP, repressed malic enzyme mRNA as well as protein expression by T3 and dexamethasone, in the absence of insulin. Glucose and other carbon sources such as lactate-pyruvate or dihydroxyacetone induced the abundance of malic enzyme mRNA in the absence of hormones. Insulin and T3 produced a high accumulation of malic enzyme mRNA in lactate-pyruvate medium, this effect being decreased by dexamethasone. EGF suppressed the induction produced by T3 and dexamethasone on malic enzyme mRNA, while the expression of beta-actin mRNA remained essentially unmodified.

Animal Nutritional Physiological Phenomena↗
PubMed1993

IGF-I is a mitogen involved in differentiation-related gene expression in fetal rat brown adipocytes.

Fetal rat brown adipocytes at time zero of culture constitute a population of cells of broad spectrum, as estimated by cell size, endogenous fluorescence and lipid content, and show an intrinsic mitogenic competence. They express constitutively early growth-related genes such as c-myc, c-fos, and beta-actin, tissue specific-genes such as the uncoupling protein (UCP) and the lipogenic marker malic enzyme (ME). Fetal brown adipocytes bear a high expression of insulin-like growth factor receptor (IGF-IR), and show a high affinity IGF-I specific-binding to its receptor, and a high number of binding sites per cell. After cell quiescence, insulin-like growth factor I (IGF-I) was as potent as 10% FCS in inducing DNA synthesis, cell number increase, and the entry of cells into the cell-cycle. In addition, IGF-I or 10% FCS for 48 h increased the percentage of [3H]thymidine-labeled nuclei as compared to quiescent cells. Single cell autoradiographic microphotographs show typical multilocular fat droplets brown adipocytes, resulting positive to [3H]thymidine-labeled nuclei in response to IGF-I. IGF-I increased mRNA expression of the early-response genes c-fos (30 min), c-myc (2 and 24 h), and H-ras (4 and 24 h). 10% FCS also increased c-fos and c-myc, but failed to increase H-ras as an early event. IGF-I or 10% FCS, however, similarly increased the mRNA late expression of c-myc, H-ras, c-raf, beta-actin, and glucose 6-phosphate dehydrogenase (G6PD) at 72 h, as compared to quiescent cells. IGF-I or FCS maintained at 24 h or increased at 48 and 72 h UCP mRNA expression. The results demonstrate that IGF-I is a mitogen for fetal rat brown adipocytes, capable of inducing the expression of early and late growth-regulated genes, and of increasing the lipogenic marker ME and the tissue-specific gene UCP, suggesting the involvement of IGF-I in the differentiation as well as in the proliferation processes.

Adipose Tissue, Brown↗
PubMed1993

Platelet derived growth factor/tyrosine kinase receptor mediated proliferation.

PDGF heterodimer of A and B chains, a complete mitogen for 3T3 mouse fibroblasts, exemplifies those growth factors interacting with membrane associated tyrosine kinase receptors. Its binding to the PDGF-receptors results in receptor dimerization and subsequent activation of tyrosine kinase activity in the cytoplasmic protein domain, autophosphorylation of the receptor being the first event in the transduction cascade. Before the ligand-receptor complex is internalized and degraded, receptor stimulation is transmitted to the general transduction network, in which several tyrosine kinase substrates are activated by phosphorylation and changes the cytoplasmic biochemistry. These changes include cytoplasmic alkalinization, increases in the intracellular concentration of cyclic-AMP and Ca2+ and activation of protein kinase C through the degradation of phosphoinositides. The known substrates recruited by the PDGF-receptor association are phosphatidylinositol-3'-kinase, ras-GTPase-activating protein, phospholipase C-gamma, serine-threonine kinase Raf-1 and src and src-related tyrosine kinases. Upon binding of PDGF to its receptor, transactivation of transcriptional and nuclear factors such as c-fos and c-myc genes and dephosphorylation of c-jun occurs, V-sis, the oncogen of the simian sarcoma virus (SSV), is highly homologous to the c-sis/PDGF-B gene that encodes the homodimer of the B-chain of the PDGF receptor. Cells transformed by SSV have been studied as a model system for the autocrine stimulation of the PDGF receptor.

Animals↗
PubMed1992

Hormonal regulation of malic enzyme and glucose-6-phosphate-dehydrogenase expression in fetal brown-adipocyte primary cultures under non-proliferative conditions.

The expression of malic enzyme and glucose-6-phosphate (Glc6P) dehydrogenase was investigated in primary cultures of fetal brown adipocytes after the prolonged presence (6 d or 10 d) of various hormones under non-proliferative conditions. The presence of triiodothyronine for 6 d and 10 d resulted in maturation of the triiodothyronine regulatory mechanism of malic-enzyme expression at the mRNA level. However, triiodothyronine had no effect on Glc6P dehydrogenase expression. Insulin increased malic-enzyme and Glc6P dehydrogenase expression at the mRNA and protein level after 6 d and 10 d of culture. The joint presence of triiodothyronine and insulin produced an additive effect on malic-enzyme expression at the mRNA and protein level after 6 d and 10 d of culture, by two independent mechanisms. Noradrenaline prevented the effect at the protein level after 6 d, but not after 10 d, probably due to loss of the beta-adrenergic response of brown adipocytes after prolonged culture. Triiodothyronine overexpressed the Glc6P dehydrogenase mRNA induced by the presence of insulin at 6 d and 10 d of culture. There was no adrenergic regulation of Glc6P dehydrogenase expression in cultured fetal brown adipocytes, regardless of the time of culture.

Adipose Tissue, Brown↗