Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.7.10.1 (ERK)
95,504 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The betacyte is a genetically engineered insulin-secreting liver cell line that is glucose responsive. Whether this cell is affected by specific beta-cell toxins is unknown. To explore this possibility we exposed these cells and those from the NIT-1 beta-cell line (positive controls) to the toxins streptozotocin (STZ, 2.5-20 mM), alloxan (ALL, 2.5-20 mM), and pentamidine (PENT, 10(-6)-1 mM). STZ and ALL were added for 1 h and pentamidine for 24 h. Insulin secretion from betacytes during a period of 5 h after removal of the toxin was inhibited only by pentamidine; all agents were inhibitory to NIT-1 cells. Glucose metabolism, as determined by a colorimetric MTT reduction assay, was adversely affected in betacytes by ALL (20 mM) and PENT (1 mM), and in NIT-1 cells by STZ (20 mM) as well as by ALL (2.5 mM) and PENT (1 mM). The magnitude of inhibition was less for the betacytes-58 v. 99%. Confluence of cells in culture wells and cell viability as assessed by the fluorochromes propidium iodide and acridine orange was reduced to a lesser extent for the betacytes than for the NIT-1 cells. The metabolic and microscopic effects of the toxins were unchanged in the betacyte from those in the liver cell line, HEP G2, from which the betacyte was engineered. These results of general resistance of the betacyte to beta-cell toxins with differing modes of action offer hope that this cell, or cells created in a similar manner from primary hepatocytes, may be at least partly resistant to the adverse effect of beta-cell toxins involved in autoimmune destruction of the pancreas. This increases the potential of the use of these cells for reversal of diabetes.
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PMID:Effect of beta-cell toxins on genetically engineered insulin-secreting cells. 921 49

We used [18F]-2-fluoro-2-deoxy-D-glucose (FDG) and PET to study regional cerebral glucose utilization in seven patients with fatal familial insomnia (FFI), an inherited prion disease with a mutation at codon 178 of the prion protein gene. Four patients were methionine/methionine homozygotes at codon 129 (symptom duration, 8.5 +/- 1 months) and three were methionine/valine (MET/VAL129) heterozygotes (symptom duration, 35 +/- 11 months). A severely reduced glucose utilization of the thalamus and a mild hypometabolism of the cingulate cortex were found in all FFI patients. In six subjects the brain hypometabolism also affected the basal and lateral frontal cortex, the caudate nucleus, and the middle and inferior temporal cortex. Comparison between homozygous or heterozygous patients at codon 129 showed that the hypometabolism was more widespread in the MET/VAL129 group, which had a significantly longer symptom duration at the time of [18F] FDG PET study. Comparison between neuropathologic and [18F] FDG PET findings in six patients showed that areas with neuronal loss were also hypometabolic. However, cerebral hypometabolism was more widespread than the histopathologic changes and significantly correlated with the presence of protease-resistant prion protein (PrPres). Our findings indicate that hypometabolism of the thalamus and cingulate cortex is the hallmark of FFI, while the involvement of other brain regions depends on the duration of symptoms and some unknown factors specific to each patient. The present data also support the notion that PrPres formation is the cause of neuronal dysfunction in prion diseases.
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PMID:Cerebral metabolism in fatal familial insomnia: relation to duration, neuropathology, and distribution of protease-resistant prion protein. 922 80

To understand the physiological role of low Mr weight phosphotyrosine protein phosphatase (LMW-PTP) in insulin mediated signaling, we established clonal cell lines overexpressing the dominant negative (C12S mutant) LMW-PTP (dnLMW-PTP) from NIH3T3 murine fibroblasts expressing insulin receptor. Upon insulin stimulation we observe an association between the dnLMW-PTP and the beta-subunit of the insulin receptor. This association is dependent on the tyrosine phosphorylation of the insulin receptor since it is not observed in unstimulated cells. Furthermore, in vitro binding experiments between dnLMW-PTP and the insulin receptor reveal that the interaction is mediated by the LMW-PTP catalytic site, as indicated by competition with orthovanadate. DnLMW-PTP overexpression influences both the mitogenic and the metabolic bioeffects of insulin. In particular, in cells overexpressing dnLMW-PTP we observe an increase in the glycogenosynthesis rate and in mitosis as indicated by glucose incorporation into glycogen and thymidine incorporation into DNA, respectively. Moreover, we studied the insulin mediated signal transduction pathways starting from insulin receptor, such as the Src kinase, the p21Ras/ERK, and the PI3K routes. Our findings are consistent with a specific regulation of mitogenesis by LMW-PTP through a pathway involving c-Src kinase but independent by both PI3K and ERK. These data strongly suggest that LMW-PTP acts as a negative regulator of both mitogenetic and metabolic insulin signalling.
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PMID:LMW-PTP is a negative regulator of insulin-mediated mitotic and metabolic signalling. 929 73

During myocardial ischemia, inhibition of the carnitine-mediated transportation of fatty acid may be beneficial because it facilitates glucose utilization and prevents an accumulation of fatty acid metabolites. We orally administered 3-(2,2,2-trimethyl hydrazinium) propionate (MET), an inhibitor of carnitine synthesis, for 20 days to rats. Then we evaluated left ventricular (LV) function during brief ischemia by using a buffer-perfused isovolumic heart model. After 15 min of reoxygenation after the transient ischemia, LV peak systolic pressure (PSP) almost completely returned to the baseline level in rats given MET (96 +/- 4%), whereas it was only partially (77 +/- 16%) recovered in the placebo-treated rats. We induced myocardial infarction in other rats by ligating the left anterior descending coronary artery. Then the animals were given MET for 20 days, and LV function was compared. In the placebo-treated rats (with myocardial infarction, but without drug treatment), LVPSP was lower than that in the sham group [108 +/- 19 (n = 10) vs. 136 +/- 15 mm Hg (n = 13); p < 0.05], and the time constant (T) of LV pressure decay was elongated (36 +/- 4 vs. 30 +/- 7 ms; p < 0.05). In MET-treated groups, however, neither PSP nor T differed from those in the sham group. In conclusion, inhibition of the carnitine-mediated transportation of fatty acid by MET protected against left ventricular dysfunction in acute and chronic myocardial ischemia.
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PMID:Inhibition of carnitine synthesis protects against left ventricular dysfunction in rats with myocardial ischemia. 933 6

Overexpression of surrogate receptors [epidermal growth factor (EGF) receptor (EGFR) and platelet-derived growth factor receptor] in adipocytes has demonstrated that multiple signaling pathways may lead to GLUT4-mediated glucose uptake. These implicated pathways function independently of IRS-1 phosphorylation and PI3-kinase activation. In addition, we previously demonstrated that EGFR tyrosyl autophosphorylation is required to stimulate GLUT4-mediated glucose transport in 3T3-L1 adipocytes. This observation suggests that signaling molecules that are dependent on EGFR autophosphorylation, such as phospholipase C (PLC), may lie in the signaling pathway to glucose transport. As PLC has been implicated in glucose transport by several clinical and basic mechanistic studies, we investigated whether EGFR signaling may promote glucose transport via modulation of PLC activity. Activation of EGFR overexpressing 3T3-L1 adipocytes leads to a 3.4 +/- 1.2-fold stimulation of PLC activity over basal levels vs. only 1.06 +/- 0.01-fold stimulation by insulin. Pharmacological inhibition of PLC by 50 microM U73122 reduced phosphoinositide accumulation by 79.2 +/- 16.9% and resulted in a concomitant 56.0 +/- 12.7% decrease in EGF-induced glucose transport. This inhibition of glucose transport by U73122 was specific, because the inactive congener, U73343, failed to block EGF-induced glucose transport. Despite the low levels of insulin-induced PLC activity, insulin-stimulated glucose transport activity was similarly inhibited by U73122 (55.9 +/- 13.1% inhibition). Inhibition of PLC activation did not impair either EGF- or insulin-induced activation of glycogen synthase or incorporation of glucose into lipid, supporting the hypothesis that both EGF- and insulin-induced glucose disposal can be independent of GLUT4-mediated glucose transport. The diminution of glucose transport secondary to inhibition of PLC activity was reflected by a decrease in GLUT4 translocation to the plasma membrane upon either EGF or insulin stimulation. These results are consistent with either a permissive or an active role for PLC activity in the translocation of GLUT4 to the plasma membrane.
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PMID:A role for phospholipase C activity in GLUT4-mediated glucose transport. 938 97

In order to design a feasible somatic cell gene delivery system for the treatment of type I diabetes, a suitable cell type needs to be determined. We have previously shown that the stable transfection of the full-length insulin cDNA into the human liver cell line, (HEP G2ins) resulted in synthesis, storage and acute regulated release of insulin to analogues of cAMP, but not to the physiological stimulus glucose. In attempting to explain the lack of glucose responsiveness of the HEP G2ins cells we have stably transfected these cells with the human islet glucose transporter GLUT 2 (HEP, G2ins/g cells). The HEP G2ins/g cell clones exhibit glucose-stimulated insulin secretion and glucose potentiation of the secretory response to nonglucose secretagogues. While glucose responsiveness commenced at a lower concentration than normal islets, a secretion curve approaching normal physiological conditions was generated. Immunoelectron microscopy revealed the presence of insulin-containing granules, similar in size and appearance to those of the normal beta cell. These results demonstrate that while it is most likely that the HEP G2ins/g cell line predominantly secretes insulin via the constitutive pathway, significant acute regulated release was seen in response to glucose, and thus represents significant progress in the creation of a genetically engineered 'artificial beta cell' from a human hepatocyte cell line.
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PMID:Gene therapy of diabetes: glucose-stimulated insulin secretion in a human hepatoma cell line (HEP G2ins/g). 942 44

The intracellular mechanisms used by insulin and insulin-like growth factors to block programmed cell death are unknown. To identify receptor structures and signaling pathways essential for anti-apoptotic effects on cells, we have created a chimeric receptor (colony-stimulating factor-1 receptor/insulin receptor chimera (CSF1R/IR)) connecting the extracellular, ligand-binding domain of the colony-stimulating factor-1 (CSF-1) receptor to the transmembrane and cytoplasmic domains of the insulin receptor. Upon activation with CSF-1, the CSF1R/IR phosphorylates itself and intracellular substrates in a manner characteristic of normal insulin receptors. CSF-1 treatment protected cells expressing the CSF1R/IR from staurosporine-induced apoptosis. A chimeric receptor (CSF1R/IRDelta960) with a deletion of 12 amino acids from its juxtamembrane domain was constructed and expressed. CSF-1-treated cells expressing the CSF1R/IRDelta960 are unable to phosphorylate IRS-1 and Shc (Chaika, O. V., Chaika, N., Volle, D. J., Wilden, P. A. , Pirrucello, S. J., and Lewis, R. E. (1997) J. Biol. Chem. 272, 11968-11974). CSF-1 stimulated glucose uptake, mitogen-activated protein kinases, and IRS-1-associated phosphatidylinositol 3' kinase in cells expressing the CSF1R/IR but not in cells expressing the CSF1R/IRDelta960. Surprisingly, the CSF1R/IRDelta960 was as effective as the CSF1R/IR in mediating CSF-1 protection of cells from staurosporine-induced apoptosis. These observations indicate that the anti-apoptotic effects of the insulin receptor cytoplasmic domain can be mediated by signaling pathways distinct from those requiring IRS-1 and Shc.
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PMID:Anti-apoptotic signaling by a colony-stimulating factor-1 receptor/insulin receptor chimera with a juxtamembrane deletion. 950 32

To identify mechanisms by which GH receptors (GHR) mediate downstream events representative of growth and metabolic responses to GH, stimulation by GH of c-fos and egr-1 expression and glucose transport activity were examined in Chinese hamster ovary (CHO) cells expressing mutated GHR. In CHO cells expressing wild-type GHR(GHR(1-638)), GH stimulated the expression of c-fos and egr-1, and stimulated 2-deoxyglucose uptake, responses also mediated by endogenous GHR in 3T3-F442A cells. Deletion of the proline-rich box 1 of GHR (GHR(deltaP)) abrogated all of these responses to GH, indicating that box 1, a site of association of GHR with the tyrosine kinase JAK2, is crucial for these GH-stimulated responses. As the C-terminal half of the cytoplasmic domain of GHR is required for GH-stimulated calcium flux and for stimulation of spi-2.1 transcription, GHR lacking this sequence (GHR(1-454)) were examined. Not only did GHR(1-454) mediate stimulation of c-fos and egr-1 expression and 2-deoxyglucose uptake, but they also mediated GH-stimulated transcriptional activation via Elk-1, a transcription factor associated with the c-fos Serum Response Element. Thus, the C-terminal half of the cytoplasmic domain of GHR is not required for GH-stimulated c-fos transcription, suggesting that increased calcium is not required for GH-stimulated c-fos expression. In CHO cells lacking all but five N-terminal residues of the cytoplasmic domain (GHR(1-294)), GH did not induce c-fos or egr-1 expression or stimulate 2-deoxyglucose uptake. Further, in 3T3-F442A fibroblasts with endogenous GHR, GH-stimulated c-fos expression and 2-deoxyglucose uptake were reduced by the tyrosine kinase inhibitors herbimycin A, staurosporine, and P11. Herbimycin A and staurosporine inhibit JAK2 and tyrosyl phosphorylation of all proteins stimulated by GH, whereas P11 inhibits the GH-dependent tyrosyl phosphorylation of only some proteins, including extracellular signal regulated kinases ERK1 and -2, but not JAK2. Taken together, these results implicate association of GHR with JAK2 and GH-stimulated tyrosyl phosphorylation of an additional cellular protein in GH-stimulated glucose transport and c-fos and egr-1 expression. These studies also indicate that, in contrast to spi-2.1, the N-terminal half of the cytoplasmic domain of GHR is sufficient to mediate stimulation of c-fos and egr-1 expression and Elk-1 activation, supporting multiple mechanisms for GH signaling to the nucleus.
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PMID:Regulation of glucose transport and c-fos and egr-1 expression in cells with mutated or endogenous growth hormone receptors. 952 72

Vascular endothelial growth factor (VEGF) stimulates nitric oxide (NO) production by endothelial cells in vitro and in vivo. However, the impact of VEGF on inducible nitric oxide synthase (iNOS) activity and NO synthesis in cultured mesangial cells is not known. Therefore, we measured nitrite accumulation in cytokine-stimulated, rat mesangial cells (RMC) in response to graded concentrations of VEGF. Addition of VEGF (10-50 ng/ml) did not alter RMC viability or NO production in either normal (5.6 mM) or high (33.3 mM) glucose conditions. Exposure of RMC to VEGF did not modify the effects of L-arginine (20 mM) or L-NAME (1 mM) on nitrite accumulation in normal or high glucose media. The steady state abundance of iNOS mRNA and the cytosolic content of iNOS protein were unaffected by addition of VEGF. Cultured RMC expressed the high-affinity tyrosine kinase VEGF receptors, flt and flk/KDR, and the levels were not modulated by incubation in normal or high glucose media. We conclude that VEGF does not regulate proliferation or NO production in cultured RMC. These findings suggest that disturbances in the normal interaction between VEGF and NO are not involved in the pathogenesis of abnormal mesangial cell structure or function in diabetic nephropathy.
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PMID:Effect of vascular endothelial growth factor on nitric oxide production by cultured rat mesangial cells. 957 Nov 72

To prove that perfused liver can be preserved at room temperature (22 degrees C), we made the experiment, in which HTK was basic solution, perfluorocarbon acted as oxygen carrier and lipid acted as energy substrate in the homoiothermy condition, pig liver organs were perfused extracorporeally through the V. portal with perfusion solution. In fixed period of time the perfusion solution from the liver was taken and analysed to determine for liver biochemical function and observe the concentration and size of hepatocellular mitochondria. In oxygen carrier perfusion solution the ammonia concentration was low, urea concentration was high, damage to mitochondria was minimum and by addition of lipid emulsion the concentration of glucose can be maintained. The experimental and control data were obviously significant. The result demonstrated that oxygen carrier (perfluorocarbon) as oxygen supplier can provides enough oxygen to liver cell, at the sametime, lipid emulsion intralipid) provides energy so as the perfused liver can be preserved at high temperature, i.e. 22 degrees C at relative long time (40 hours) and still has the function of removing toxic substance such as ammonia and converting it to urea, meanwhile maintain the normal structure of mitochondria. The preservation of perfused liver at homoiothermy is possible.
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PMID:[Experimental study on homoiothermic extracorporeal liver preservation]. 959 56


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