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:2.7.7.49 (reverse transcriptase)
31,746 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Atopic asthma is characterized by chronic inflammation of the bronchial mucosa in which eosinophil- and immunoglobulin E (IgE)-dependent mechanisms are believed to be prominent. Therefore, specific proeosinophilic mediators such as interleukin (IL)-5 and essential cofactors for IgE switching in B-lymphocytes such as IL-4 could play a pivotal role in asthma. However, the exact role that individual inflammatory mediators play in the development of the disease in humans is still unknown. Using semiquantitative reverse transcriptase-polymerase chain reaction amplification in bronchial biopsies from 10 atopic asthmatics, we have tested the hypothesis that IL-4 and IL-5 mRNA expression relative to beta-actin mRNA correlates with validated indicators of disease severity. IL-4 and IL-5 mRNA copies relative to beta-actin mRNA were detected in bronchial biopsies from atopic asthmatics. The numbers of IL-5 mRNA copies relative to beta-actin mRNA correlated with disease severity assessed by the Aas asthma score (r = 0.70, p = 0.01), baseline FEV1 (r = -0.94, p = 0.001), baseline peak expiratory flow rate (r = -0.77, p = 0.01), peak expiratory flow rate variability over 2 wk (r = 0.69, p = 0.028), and the histamine PC20 (r = -0.72, p = 0.018). Conversely, the numbers of IL-4 mRNA copies relative to beta-actin mRNA did not correlate with asthma severity, but they positively correlated with total serum IgE concentrations (r = -0.90, p = 0.001). Our present results support the concept that IL-5 may determine asthma clinical expression and severity, and by inference they support the development of IL-5 targeted therapies.
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PMID:Relationship between IL-4 and IL-5 mRNA expression and disease severity in atopic asthma. 930 82

We have examined the expression of c-met mRNA in tissue from 27 colorectal cancers and ten liver metastases using the reverse transcriptase-polymerase chain reaction method. The expression of c-met mRNA in these tissues was quantified and the copy number of c-met mRNA to 10(8.0) copies of beta-actin mRNA was calculated. Mean copy numbers of c-met mRNA in cancer tissue and normal mucosa were 10(5.5) and 10(4.5) respectively. The c-met expression of cancer was significantly higher than that of normal mucosa (P < 0.0001). In 20 of 22 samples in which c-met expression of both tumor and corresponding normal tissue were examined, c-met was overexpressed in the cancer tissue. No correlation was found between c-met expression and the clinicopathologic background. The mean copy numbers of c-met mRNA in the tissue from the ten liver metastases and normal liver were 10(6.1) and 10(6.2) respectively. Although c-met expression in metastatic tissue was higher than that in the primary cancer tissue, the increase was not statistically significant. In three of four patients with synchronous liver metastases, c-met was overexpressed in the metastatic tissue compared with that in the corresponding primary cancer tissue. These results show that c-met is overexpressed in both primary colorectal cancer and liver metastases and suggest that c-met plays a role in the development of colorectal cancer liver metastases.
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PMID:Expression of c-met proto-oncogene in primary colorectal cancer and liver metastases. 943 98

Synthesis of a number of rat liver proteins, including albumin, fibrinogen, apolipoprotein AI, and transferrin, is elevated in the nephrotic syndrome (NS). Increased synthesis of these proteins is regulated at the transcriptional level and occurs in the context of increased mRNA encoding each protein. Changes in albumin, fibrinogen, apolipoprotein AI, and transferrin mRNA levels in total cellular RNA isolated from the livers of normal rats and rats with passive Heymann nephritis were measured using a kinetically monitored, reverse transcriptase-initiated PCR (kRT-PCR) assay. The kRT-PCR assay rapidly quantitated changes in rat liver mRNA levels with an accuracy comparable to that of more labor-intensive mRNA quantitation methods. The relative levels of beta-actin, apolipoprotein AI, fibrinogen, and albumin mRNAs were very similar in total cellular RNA isolated from rat liver versus H4C3 hepatocytes in culture, suggesting that the H4C3 hepatocyte is an appropriate model for studying expression of genes encoding proteins secreted by the liver. Taken together, the results demonstrate the feasibility of using the kRT-PCR assay for isolation and characterization of a soluble factor responsible for elevated synthesis of hepatocyte mRNAs associated with the nephrotic syndrome.
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PMID:Rat liver transcript profiling in normal and disease states using a kinetic polymerase chain reaction assay. 948 Jul 87

The inhibitor of cyclin-dependent kinases WAF1/p21 has been shown to mediate cell cycle arrest by p53 and other factors. We have studied its expression in urothelial carcinoma. Immunohistochemistry of paraffin-embedded tissues revealed no detectable p21 protein in normal mucosa, whereas 8 of 17 (47%) carcinomata in situ, 41 of 62 (66%) pTa, 14 of 30 (47%) pT1 and 5 of 15 (33%) muscle-invasive tumours stained positive, usually with a heterogeneous pattern. Expression of p21 was associated with low grade tumours. In contrast, the frequency of p53 accumulation increased with grade and stage as did the frequency of staining for the proliferation marker Ki67. The level of WAF1 mRNA was determined relative to beta-actin mRNA by quantitative reverse transcriptase polymerase chain reaction (RT-PCR) in 15 freshly frozen invasive tumours. In eight samples obtained from normal bladder mucosa, the values ranged from 0.93 to 2.19 arbitrary units (AU) (mean 1.54+/-0.37 AU), but varied widely from non-detectable to 16.21 AU (mean 3.02+/-4.44 AU) in the tumour specimens. In accord with the immunohistochemical findings, WAF1 mRNA expression was elevated over the range found in normal mucosa in 5 of 15 advanced tumours. In addition, RNA analysis revealed a decrease in expression in six tumours. No mutations were observed in the WAF1/p21 gene in these tumours, but two were heterozygous for the codon 31 polymorphism. These data indicate that p21 is frequently expressed in superficial, well differentiated urothelial carcinomas, but less often in muscle-invasive urothelial carcinomas, irrespective of their p53 status. The expression of p21 and its prevalence in low-stage tumours may reflect residual growth-regulatory influences potentially impeding but not necessarily inhibiting tumour development.
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PMID:Frequent and heterogeneous expression of cyclin-dependent kinase inhibitor WAF1/p21 protein and mRNA in urothelial carcinoma. 948 5

The gases nitric oxide (NO) and carbon monoxide (CO) may be involved in hypothalamo-pituitary-adrenal axis (HPA) modulation. In the brain, NO is synthesized by two forms of NO synthase (NOS), a constitutive neuronal form (nNOS) and an inducible form (iNOS). There are also a constitutive heme oxygenase (HO2) and an inducible form (HO1) which generate CO. We have therefore investigated the effect of peripheral lipopolysaccharide (LPS) administration on the gene expression of these enzymes along with interleukin-1beta (IL-1beta) gene expression in the hypothalamus, pituitary and liver. Male Wistar rats (200-250 g body weight) were injected intraperitoneally with endotoxin (Escherichia coli, 055 B5) dissolved in sterile normal saline [250 microg/kg first group, 2.5 mg/kg (second group) and 6.25 mg/kg (third group)] in a final volume of 0.5 ml, or saline alone in the control group. The first and the second groups were studied 1, 3, 8 and 24 h after LPS (n = 4 per group); the third group was studied at 3 h. Total RNA was extracted from the hypothalamus, pituitary and liver, and cDNA was made using standard reverse transcriptase methods. Duplex polymerase chain reaction (PCR) was standardised in order to quantify the expression of a specific gene in relation to the 'house-keeping' gene beta-actin. The specific genes studied were iNOS, nNOS, HO1, HO2 and IL-1beta. The PCR products were separated on agarose gel and densitometric analysis of the bands allowed semi-quantification. In the second group, iNOS and IL-1beta were induced in hypothalamus, pituitary and liver, showing a peak at 3 h (p < 0.001), returning to baseline levels at 24 h. Neuronal NOS was not expressed in the liver under basal conditions or after LPS; in the hypothalamus and pituitary, nNOS was expressed basally but there was no change after LPS. In the first group, iNOS and IL-1beta were again induced in all three tissues studied, but with a delayed time course compared to the second and third groups; the peak change for IL-1beta occurred at 8 h (p < 0.05), again returning to baseline levels at 24 h. The peak for iNOS occurred at 24 h. HO1 and HO2 were expressed in all three tissues under basal conditions; HO1 was increased at 1 h in the liver in the second group, and at 3 h in the pituitary in the third group. There was no change in either HO1 or HO2 in the hypothalamus at any dose at any time point. We conclude that IL-1beta and iNOS are induced in rat hypothalamus and pituitary following various doses of endotoxin. We speculate that while IL-1beta may mediate stimulation of the HPA by endotoxin, NO generation may be involved in the counter-regulation of this response.
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PMID:Endotoxin induces interleukin-1beta and nitric oxide synthase mRNA in rat hypothalamus and pituitary. 950 41

1. We examined the possibility of functional and molecular expression of volume-regulated Cl- channels in vascular smooth muscle using the whole-cell patch-clamp technique and quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) on cells from canine pulmonary and renal arteries. 2. Decreasing external osmolarity induced cell swelling, which was accompanied by activation of Cl--dependent outward-rectifying membrane currents with an anion permeability sequence of SCN- > I- > Br- > Cl- > aspartate-. These currents were sensitive to block by DIDS, extracellular ATP and the antioestrogen compound tamoxifen. 3. Experiments were performed to determine whether the molecular form of the volume-regulated chloride channel (ClC-3) is expressed in pulmonary and renal arteries. Quantitative RT-PCR confirmed expression of ClC-3 in both types of smooth muscle. ClC-3 expression was 76.4 % of beta-actin in renal artery and 48.0 % of beta-actin in pulmonary artery. 4. We conclude that volume-regulated Cl- channels are expressed in vascular smooth muscle cells and exhibit functional properties similar to those found in other types of cells, presumably contributing to the regulation of cell volume, electrical activity and, possibly, myogenic tone.
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PMID:Functional and molecular expression of volume-regulated chloride channels in canine vascular smooth muscle cells. 959 97

A reverse transcriptase polymerase chain reaction (rt-PCR) for quantification of gene expression has been optimized for analysis of folylpolyglutamate synthase (FPGS) and thymidylate synthase (TS), using beta-actin as an internal standard (house-keeping gene). Total RNA was isolated from tumor tissue, reversely transcribed to cDNA and PCR amplified with primers specific for TS, FPGS and beta-actin in separate vials. PCR products were separated and quantified by high-pressure liquid chromatography (HPLC) without addition of radioactive or fluorescent markers, which minimizes labor and occupational hazards. The day-to-day variation in the HPLC analysis was 2.7% and the within sample variations for rt-PCR/HPLC analysis of TS and FPGS were 18.5% for both assays. This method provides a tool for convenient gene expression analysis in clinical biopsies.
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PMID:Rapid quantitative PCR determination of relative gene expression in tumor specimens using high-pressure liquid chromatography. 959 Oct 43

The production of immunoglobulin E (IgE) antibody is largely dependent on the ratio between interleukin-4 (IL-4) (a T helper 2 (Th2)-type cytokine) and interferon-gamma (IFN-gamma) (a T helper 1 (Th1)-type cytokine). Interleukin-5 (IL-5) (also a Th2-type cytokine) is an important eosinophil differentiation factor and also co-stimulates B-cell growth and differentiation. The present study was designed to evaluate and compare the expression of IFN-gamma, IL-4 and IL-5 mRNA in the nasal mucosal membrane of sensitized Brown-Norway (BN) rats. Fourteen BN rats were divided into two groups: non-sensitized (control) and sensitized. The sensitized group was injected with ovalbumin (OA) intraperitoneally on three consecutive days. Twenty-one days later, rats were exposed to 1% OA aerosol. Twenty-four hours after exposure to aerosol, nasal mucosa was extracted from both groups and reverse transcriptase-polymerase chain reaction (RT-PCR) was performed. The densities of the bands of IL-4, IL-5 and IFN-gamma mRNA were expressed as percentages against beta-actin mRNA. Our results showed that the mean values for IL-4 and IL-5 mRNA were increased significantly in sensitized rats compared with control rats. In contrast, the mean value for IFN-gamma mRNA was significantly lower in sensitized rats compared with those of the control group. Our data therefore suggest that sensitization of rat nasal mucous membranes results in the predominant expression of Th2-type cytokines.
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PMID:Expression of interferon-gamma, interleukin-4 and interleukin-5 mRNA in the nasal mucosal membrane of rats with allergic rhinitis. 965 23

Various "housekeeping" genes are often used as endogenous controls in gene expression experiments. We have cloned from swine, three genes commonly used as endogenous controls in other species and have characterized their relative levels of expression in various porcine tissues and their response to various cell activators. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and beta-actin were readily detected by northern hybridization in various tissues and in alveolar macrophages. The expression of hypoxanthine phosphoribosyltransferase (HPRT) was detected only by northern hybridization of poly-A+ enriched RNA and by reverse transcriptase-polymerase chain reaction (RT-PCR), making it more suitable for highly sensitive detection methods. Expression of GAPDH varied less among tissues than did beta-actin, making it more useful control for comparisons of gene expression between tissues with northern hybridizations. Various treatments of cultured alveolar macrophages differentially affected levels of beta-actin and GAPDH, while HPRT expression was unchanged in alveolar macrophages or spleen cells similarly treated. Therefore, while HPRT can be used as the endogenous control with sensitive detection methods such as RT-PCR, less sensitive detection methods require a more abundant gene such as GAPDH.
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PMID:Regulation of hypoxanthine phosphoribosyltransferase, glyceraldehyde-3-phosphate dehydrogenase and beta-actin mRNA expression in porcine immune cells and tissues. 967 36

The inositol 1,4,5-trisphosphate receptor (InsP3R) is an intracellular Ca2+ channel that plays a role in the regulation of insulin secretion. In rat isolated pancreatic islets the expression of types I, II and III InsP3R mRNA was identified by reverse transcriptase-polymerase chain reaction and confirmed by cDNA cloning and sequencing. The islet ratios of types I, II and III InsP3R mRNA to beta-actin mRNA were 0.08 +/- 0.02, 0.08 +/- 0.03 and 0.25 +/- 0.04 respectively. Types I, II and III InsP3R mRNA were also expressed in rat (RINm5F) and mouse (betaHC9) pancreatic beta-cell lines, and rat cerebellum. Type III InsP3R mRNA was quantitatively the most abundant form in rat islets and RINm5F cells. In betaHC9 cells, types II and III InsP3R mRNA were expressed at similar levels, and in much greater abundance than type I mRNA. Type III was the least abundant InsP3R mRNA in cerebellum. Culture of betaHC9 cells for 5 days at 2.8 and 25 mM glucose, or RINm5F cells for 7 days at 5.5 and 20 mM glucose, resulted in significantly enhanced expression of type III, but not types I and II, InsP3R mRNA in the cells at the higher glucose concentrations. During short-term (0.5-2 h) incubations, betaHC9 cell type III InsP3R mRNA levels increased in response to glucose in a time- and concentration-dependent manner. Actinomycin D inhibited the glucose response. Alpha-ketoisocaproic acid also stimulated betaHC9 cell type III InsP3R mRNA expression in a concentration-dependent manner, whereas 2-deoxyglucose and 3-O-methylglucose were without effect. The different levels of expression of mRNA for three InsP3R isoforms in islets and insulinoma cells, and the influence of glucose and alpha-ketoisocaproic acid on the expression of type III mRNA, suggests that nutrient metabolism plays a role in the regulation of this gene and that the function of InsP3R subtypes may be unique with each playing a distinct role in beta-cell signal transduction and insulin secretion.
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PMID:Characterization of inositol 1,4,5-trisphosphate receptor isoform mRNA expression and regulation in rat pancreatic islets, RINm5F cells and betaHC9 cells. 972 61


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