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Biomedical subjects

I Huvar

Publications and source records attributed to I Huvar.

13 recordsLinked to original sources
PubMed2000

Ras activation in normal white blood cells and childhood acute lymphoblastic leukemia.

Ras is an important cellular switch, relaying growth-promoting signals from the plasma membrane to the nucleus. In cultured cells, Ras is activated by various hematopoietic cytokines and growth factors, but the activation state of Ras in peripheral WBCs and bone marrow cells has not been studied nor has Ras activation been assessed in cells from patients with acute lymphoblastic leukemia (ALL). Using an enzyme-based method, we assessed Ras activation in peripheral WBCs, lymphocytes, and bone marrow cells from normal subjects and from children with T-cell ALL (T-ALL) and B-lineage ALL (B-ALL). In normal subjects, we found mean Ras activations of 14.3, 12.5, and 17.2% for peripheral blood WBCs, lymphocytes, and bone marrow cells, respectively. All three of these values are higher than we have found in other normal human cells, compatible with constitutive activation of Ras by cytokines and growth factors present in serum and bone marrow. In 9 of 18 children with T-ALL, Ras activation exceeded two SDs above the mean of the corresponding cells from normal subjects, whereas in none of 11 patients with B-ALL did Ras show increased activation; activating genetic mutations in ras occur in less than 10% of ALL patients. Thus, Ras is relatively activated in peripheral blood WBCs, lymphocytes, and bone marrow cells compared with other normal human cells, and Ras is activated frequently in T-ALL but not in B-ALL. Increased Ras activation in T-ALL compared with B-ALL may contribute to the more aggressive nature of the former disease.

Adolescent↗
PubMed1998

Inhibition of phosphatidylinositol 3-kinase activity by adenovirus-mediated gene transfer and its effect on insulin action.

Phosphatidylinositol 3-kinase (PI 3-K) is implicated in cellular events including glucose transport, glycogen synthesis, and protein synthesis. It is activated in insulin-stimulated cells by binding of the Src homology 2 (SH2) domains in its 85-kDa regulatory subunit to insulin receptor substrate-1 (IRS-1), and, others. We have previously shown that IRS-1-associated PI 3-kinase activity is not essential for insulin-stimulated glucose transport in 3T3-L1 adipocytes, and that alternate pathways exist in these cells. We now show that adenovirus-mediated overexpression of the p85N-SH2 domain in these cells behaves in a dominant-negative manner, interfering with complex formation between endogenous PI 3-K and its SH2 binding targets. This not only inhibited insulin-stimulated IRS-1-associated PI 3-kinase activity, but also completely blocked anti-phosphotyrosine-associated PI 3-kinase activity, which would include the non-IRS-1-associated activity. This resulted in inhibition of insulin-stimulated glucose transport, glycogen synthase activity and DNA synthesis. Further, Ser/Thr phosphorylation of downstream molecules Akt and p70 S6 kinase was inhibited. However, co-expression of a membrane-targeted p110(C) with the p85N-SH2 protein rescued glucose transport, supporting our argument that the p85N-SH2 protein specifically blocks insulin-mediated PI 3-kinase activity, and, that the signaling pathways downstream of PI 3-kinase are intact. Unexpectedly, GTP-bound Ras was elevated in the basal state. Since p85 is known to interact with GTPase-activating protein in 3T3-L1 adipocytes, the overexpressed p85N-SH2 peptide could titrate out cellular GTPase-activating protein by direct association, such that it is unavailable to hydrolyze GTP-bound Ras. However, insulin-induced mitogen-activated protein kinase phosphorylation was inhibited. Thus, PI 3-kinase may be required for this action at a step independent of and downstream of Ras. We conclude that, in 3T3-L1 adipocytes, non-IRS-1-associated PI 3-kinase activity is crucial for insulin's metabolic signaling, and that overexpressed p85N-SH2 protein inhibits a variety of insulin's ultimate biological effects.

3T3 Cells↗
PubMed1997

A decrease in the intracellular guanosine 5'-triphosphate concentration is necessary for granulocytic differentiation of HL-60 cells, but growth cessation and differentiation are not associated with a change in the activation state of Ras, the transforming principle of HL-60 cells.

We found that when human promyelocytic leukemic cells (HL-60 cells) were induced to differentiate along the granulocytic lineage by two diverse mechanisms (starvation for an essential amino acid or treatment with DMSO), there was a marked decrease in the intracellular guanosine 5'-triphosphate (GTP) concentration with no change in the guanosine 5'-diphosphate (GDP) concentration. Differentiation was prevented by guanine or guanosine in a dose-dependent manner. We showed that: (a) guanine had to be converted to a nucleotide because it did not prevent differentiation of hypoxanthine-guanine phosphoribosyltransferase-deficient HL-60 cells; (b) the effect of guanine correlated with a return of the cytosolic GTP:GDP ratio to normal; and (c) other purine bases were not effective. We hypothesized that the decreased GTP:GDP ratio in differentiating HL-60 cells might decrease the relative amount of GTP bound to Ras, a key regulatory GTP-binding protein important to cell growth and differentiation. Consistent with data showing that HL-60 cells harbor an activating N-Ras mutation, we found that the percentage of Ras molecules in the GTP-bound state was high in proliferating HL-60 cells (27 +/- 3%) compared with other cultured mammalian cells (< 1%); however, we found no change in the activation state of Ras when cells ceased to proliferate and differentiated in response to DMSO, amino acid deprivation, or inhibitors of guanylate synthesis. We conclude that: (a) a decrease in the intracellular GTP concentration is necessary for HL-60 cells to undergo granulocytic differentiation; and (b) although a high degree of Ras activation contributes to the malignant phenotype of the cell, there is no change in the activation state of Ras during granulocytic differentiation.

Amino Acids↗
PubMed1997

[Hemostasis using vasopressin analogs during conization of the uterine cervix and minor vaginal operations].

Peroperative haemorrhage is a frequent complication of conization of the portio uteri by surgery-by a "cold knife" scalpel or another method (LEEP, laser, etc.). In a clinical study comprising 41 patients with the diagnosis CIN I-III or laceration of the portio uteri classical conization by means of a scalpel was made with concurrent haemostasis with terlipressin (TLP) by paracervical administration (400 micrograms-REMESTYP 2 amp.). Concurrently the blood loss in ml was followed, the Hb level and other effects of the preparation. From the results ensues that side-effects after local administration of TLP were observed only in 12.2%. Peroperative blood losses were on average under 15 ml. Local paracervical TLP administration appears effective from the clinical aspect with regard to the low frequency of side-effects and the favourable haemostatic effect.

Blood Loss, Surgical↗
PubMed1996

Enhanced tumorigenic behavior of glioblastoma cells expressing a truncated epidermal growth factor receptor is mediated through the Ras-Shc-Grb2 pathway.

A mutant epidermal growth factor receptor (DeltaEGFR) containing a deletion of 267 amino acids from the extracellular domain is common in human glioblastomas. We have previously shown that the mutant receptor fails to bind EGF, is constitutively phosphorylated, and confers upon U87MG glioblastoma cells expressing it (U87MG. DeltaEGFR), an increased ability to form tumors in mice. Here we demonstrate that the constitutively phosphorylated DeltaEGFR enhances growth of glioblastoma cells through increased activity of Ras: 1) there was an increase in the proportion of Ras present in the GTP-bound form, and 2) introduction of neutralizing anti-Ras 259 antibodies into U87MG and U87MG.DeltaEGFR cells by microinjection inhibited DNA synthesis to the same low level in both cell populations. We also show that the truncated EGF receptor constitutively associates with the adapter proteins Shc and Grb2 which are involved in the recruitment of Ras to activated receptors. Several derivatives of DeltaEGFR containing single, or multiple mutations at critical autophosphorylation sites were constructed and used to demonstrate that the major Shc binding site is Tyr-1148, and that Grb2 association occurs primarily through Tyr-1068. We conclude that the increased tumorigenic potential of glioblastoma cells expressing the truncated EGF receptor is due at least in part to Ras activation presumably involving the Shc and Grb2 adapter proteins.

Adaptor Proteins, Signal Transducing↗
PubMed1996

Regulation of gene expression by cGMP-dependent protein kinase. Transactivation of the c-fos promoter.

The cAMP/cAMP-dependent protein kinase (A-kinase) and Ca2+/calmodulin-dependent protein kinase (Cam-kinase) signal transduction pathways are well known to regulate gene transcription, but this has not been demonstrated directly for the cGMP/cGMP-dependent protein kinase (G-kinase) signal transduction pathway. Here we report that transfection of G-kinase into G-kinase-deficient cells causes activation of the human c-fos promoter in a strictly cGMP-dependent manner. The effect of G-kinase appeared to be mediated by several sequence elements, most notably the serum response element (SRE), the AP-1 binding site (FAP), and the cAMP response element (CRE). The magnitude of G-kinase transactivation of the fos promoter was similar to that of A-kinase, but there were significant differences between G-kinase and A-kinase activation of single enhancer elements and of a chimeric Gal4-CREB transcription factor. Our results indicate that G-kinase transduces signals to the nucleus independently of A-kinase or Ca2+, although it may target some of the same transcription factors as A-kinase and Cam-kinase.

Animals↗
PubMed1996

Ras-GTP levels are elevated in human NF1 peripheral nerve tumors.

Neurofibromin, the gene product of NF1, is a Ras GTPase Activating Protein. The absence of neurofibromin leads to increased levels of Ras-GTP, which contributes to the proliferation of NF1 neurogenic sarcoma cell lines. Whether this pathogenic mechanism is applicable to benign and malignant peripheral nerve tumours from NF1 and non NF1 patients is not known, due to lack of a tissue based assay. We have adapted a colorimetric enzymatic assay for determining levels of Ras bound guanine nucleotides in tissues. Ras-GTP levels were increased in NF1 neurogenic sarcomas (15 times) and benign NF1 neurofibromas (four times), compared to non NF1 schwannomas. Neurofibromin was not expressed in NF1 sarcomas, in support of its important negative Ras regulatory role in the pathogenesis of NF1 peripheral nerve tumors.

Base Sequence↗
PubMed1994

[Hysteroscopic sterilization using Ovabloc].

The authors give an account on 29 sterilizations made by the hysteroscopic route, using the preparation OVABLOC. It is a reversible block of the oviducts by occlusive material. Under hysteroscopic control into the inner orifice of the oviduct by means of a special injecting device under pressure a two-component mixture of silicone and a catalyst is instilled which causes within 5 min. hardening of the mass and the formation of an elastic relief filling of the entire oviduct. In 25 of the women (86.2%) the procedure was accomplished without complication, in the 3 (10.3%) the procedure failed on account of technical difficulties and in one case (3.5%) during hysteroscopy the uterine wall was perforated. On X-ray check-up after three months of 25 successfully operated women in 24 the position of the tubal occlusion was unaltered (the material is due to dispersed silver particles contrasting on X-ray); in one case it was loosened. The advantages of the method are that it is sparing, well tolerated and easily reversible, the disadvantage is that it is relatively laborious and expensive.

Female↗
PubMed1994

[Hysteroscopic resection of myomas].

The authors give an account of their initial experience with hysteroscopic resection of myomas. During the investigation period (January 1991-August 1992) 34 of these operations were performed. In all instances at the same time laparoscopy was performed. The operation was free from complications in 26 patients (76.5%), in three instances (8.8%) peroperative haemorrhage developed, in 4 (11.8%) postoperative haemorrhage and in one instance (2.9%) perforation of the uterine wall. In 29 (85.3%) thus operated patients the menstrual cycle became normal within three months, after this period of time amenorrhoea persisted in three patients (8.8%). Two women (5.9%) reported irregular bleeding after the operation. To improve the success of the operation accurate indication is necessary, preferably based on previous diagnostic hysteroscopy, and improvement of the surgical technique.

Female↗
PubMed1993

[Control of labor with prostaglandins after previous cesarean section].

The authors studied the possibilities of vaginal labour management by induction after previous caesarean section. They compared the effectiveness of oxytocin and prostaglandins in uterus action. A total of 48 patients after previous caesarean section were induced, 26 by oxytocin and 22 by prostaglandins. The success of the induction was 30.7% with oxytocin and 95.4% with prostaglandins. Compared were also both the incidence of complications during labour and the effect of the uterotonic used on the fetus. It follows from the results that local application of prostaglandins is more advantageous than overall application and carries practically no risk for the patient in labour.

Administration, Intravaginal↗