Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Previous work has demonstrated heterogeneous effects of methylating agents on induction of DNA damage inducible genes in Escherichia coli. These studies employed E. coli mutants that have fusions of the lac operon to genes induced by treatment with sublethal levels of alkylating agents. These mutants were selected from random insertions of the Mu-dl (Apr lac) phage by screening for induction of beta-galactosidase activity in the presence of methylmethanesulfonate or N-methyl-N'-nitro-N-nitrosoguanidine. The current report extends these findings by analyzing gene expression caused by mechlorethamine, chloroethylnitrosoureas and cis-diamminedichloroplatinum(II) (cis-DDP). The results demonstrate heterogeneous effects by these agents on gene expression. While 1-(2-chloroethyl)-3-cyclohexyl-1-nitrosourea induces alkA, other nitrosoureas, mechlorethamine, and cis-DDP do not cause expression of this gene. Further, while all nitrosoureas caused expression of aidC, mechlorethamine and cis-DDP did not. Lastly, cis-DDP caused marked expression of a sulA fusion mutant while not inducing any of the other E. coli fusion mutants.
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PMID:Gene expression caused by alkylating agents and cis-diamminedichloroplatinum(II) in Escherichia coli. 304 78

The effects of cis-diamminedichloroplatinum(II) (cis-DDP) and trans-DDP adducts on mammalian transcription in vivo have been investigated. A plasmid containing the beta-galactosidase (beta-gal) reporter gene was modified with either of the two platinum compounds and transfected into human or hamster cell lines. A 2-3 fold higher level of transcription was observed in both cell lines from plasmids containing trans-DDP adducts as compared to plasmids modified by cis-DDP. This difference in transcriptional activity was not decreased in human and rodent nucleotide excision repair deficient cell lines, indicating that more efficient excision repair of the trans-DDP adducts was not the cause of its lower ability to block transcription in this assay. For this conclusion to be valid, it is assumed that trans-DDP adducts are repaired primarily by the nucleotide excision repair pathway, as is the case with the adducts of cis-DDP. The possibility that trans-DDP adducts are preferentially bypassed by RNA polymerase was examined by monitoring the elongation of beta-gal mRNA on damaged templates in vivo. Nascent beta-gal mRNA transcripts were recovered from excision repair deficient xeroderma pigmentosum A cells transfected with platinated plasmids, and the extent of RNA synthesis was measured by using ribonuclease protection. Fourfold more trans-DDP than cis-DDP adducts were required to inhibit transcription elongation by 63%. RNA polymerase II bypassed cis- and trans-DDP DNA adducts with efficiencies of 0-16% and 60-70%, respectively. These data provide insight into the differential toxicity of the two platinum isomers.
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PMID:DNA adducts of cis-diamminedichloroplatinum(II) and its trans isomer inhibit RNA polymerase II differentially in vivo. 757 87

Cholangiocarcinoma is a virtually incurable tumor, resistant to current surgical, chemotherapy, and radiotherapy interventions. We applied the gene therapy strategy of toxin gene conversion of nontoxic prodrug to chemotherapeutic drug in combination with radiation therapy to the treatment of cholangiocarcinoma. In this regard, 5-fluorouracil (5-FU) is an accepted radiosensitizing and chemotherapeutic agent presently used in cancer therapy. The Escherichia coli enzyme cytosine deaminase (CD) converts the prodrug 5-fluorocytosine (5-FC) to 5-FU. Therefore, our goal was to express the CD gene in the human cholangiocarcinoma cell line, SK-ChA-1, assess the cytotoxicity of intracellular production of 5-FU, and determine any enhanced cell killing by the addition of external beam radiation. The susceptibility of SK-ChA-1 cells to recombinant adenoviral infection was determined by fluorescence-activated cell sorting analysis. We used the recombinant adenoviral vector AdCMVLacZ, encoding the E. coli beta-galactosidase reporter gene under control of the human cytomegalovirus (CMV) promoter, to infect SK-ChA-1 and HeLa cells at 10 and 100 plaque forming units (pfu)/cell, followed by FACS analysis. To evaluate CD-mediated conversion of 5-FC to 5-FU and subsequent cytotoxicity, SK-ChA-1 cells were infected with the recombinant adenovirus AdCMVCD, which encodes CD. Cells were then plated in 96-well microtiter plates and exposed to varying concentrations of 5-FC. Cell proliferation assays (tetrazolium salt conversion to formazan colorimetric assay) were performed beginning 2-8 days after plating. We evaluated the effects of external beam radiation using a single 8 Gy 60Co dose to AdCMVCD infected cells, with prior exposure to 5-FC for 2-3 days. MTS assays were performed following radiation treatment. Radiation dose-response analysis, via clonogenic assay, was used as a more sensitive assay to confirm the interaction of the treatment conditions. s.c. SK-ChA-1 tumors in athymic nude mice were established, which then received three intratumoral injections of 1 x 10(9) pfu AdCMVCD. Mice received i.p. injections of 400 mg/kg of 5-FC twice daily for 7 days beginning the day of initial AdCMVCD injection (day -2). The radiation treatment group received 10 Gy of 60Co exposure to their tumor on day 0. SK-ChA-1 cells were efficiently transduced (48.7 and 99.2%) by 10 and 100 pfu/cell of AdCMVLacZ, respectively. From 37.9 to 84.4% of SK-ChA-1 cells were killed following infection with 10 pfu/cell AdCMVCD and 8 days of exposure to various concentrations of 5-FC (5, 10, 30, 50, and 100 microg/ml). Higher 5-FC concentrations and longer duration of exposure resulted in greater cell killing. Radiation treatment (8 Gy) enhanced cell killing by greater than 70% when combined with 10 or 20 microg/ml of 5-FC. Radiation dose-response analysis with clonogenic assay confirmed enhanced SK-ChA-1 cell cytotoxicity as a result of radiation treatment following AdCMVCD infection and 5-FC exposure, with radiobiological parameters alpha = 0.44 and D0 = 0.96. Combined treatment of SK-ChA-1 tumors with AdCMVCD, 5-FC, and radiation in animals resulted in significantly greater survival, time to tumor regrowth, and doubling time compared to the nonradiation treatment group (P = 0.03, 0.015, and 0.002, respectively). Significantly greater change in tumor size, smaller ratio of final tumor size to original tumor size, and smaller final tumor size were observed in the radiation treatment group compared to the no radiation treatment group (P = 0.02, 0.03, and 0.03, respectively). Human cholangiocarcinoma cells were transduced with a recombinant adenovirus in vitro at high efficiency and were susceptible to CD-mediated intracellular 5-FU production. Radiobiological survival curve parameters confirmed an interactive cytotoxic effect when viral infection and prodrug therapy were combined with external beam radiation exposure. (ABSTRACT TRUNCATED)
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PMID:Molecular chemotherapy combined with radiation therapy enhances killing of cholangiocarcinoma cells in vitro and in vivo. 933 Oct 94

1. Recent studies have revealed an association between coronary risk factors and both the number and function of bone marrow-derived endothelial progenitor cells (EPC). We investigated the effect of oxidized low-density lipoprotein (ox-LDL) on the senescence of EPC, leading to cellular dysfunction. 2. Endothelial progenitor cells were isolated from human peripheral blood and characterized. The exposure of cultured EPC to ox-LDL (10 microg/mL) significantly accelerated the rate of senescence compared with control during 20 days in culture as determined by acidic beta-galactosidase staining. Oxidized LDL-induced EPC senescence was significantly inhibited by pretreatment with either lectin-like ox-LDL receptor-1 (LOX-1) antibody (Ab) or atorvastatin (P < 0.01). 3. Because cellular senescence is critically influenced by telomerase, which elongates telomeres, we measured telomerase activity using a polymerase chain reaction-ELISA-based assay. Oxidized LDL significantly diminished telomerase activity to approximately 50%, an effect that was significantly abolished by pretreatment with either LOX-1 Ab or atorvastatin (P < 0.01). 4. We examined whether ox-LDL-induced EPC senescence translates into EPC dysfunction. An MTS assay disclosed an inhibitory effect of ox-LDL on EPC proliferation. In a Matrigel assay, EPC treated with ox-LDL were less likely to participate in network formation compared with controls. 5. In conclusions, ox-LDL accelerates the onset of EPC senescence, which may be related to telomerase inactivation. Oxidized LDL-induced EPC senescence leads to the impairment of proliferative capacity and network formation.
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PMID:Oxidized low-density lipoprotein induces endothelial progenitor cell senescence, leading to cellular dysfunction. 1523 25

The functional impairment associated with atherogenic factors, including hypertension, constitutes a limitation to the ability of endothelial progenitor cells (EPCs) to repair. In addition, estrogens have been shown to play a role in reendothelialization after vascular injury. We investigated the effects of estrogens on differentiation and senescence of EPCs derived from bone marrow (BM-EPCs) in spontaneously hypertensive rats (SHR/Izm). Bone marrow (BM) cells were obtained from the tibias and femurs of age-matched, male SHR/Izm and Wistar-Kyoto rats (WKY/Izm). The number of differentiated, adherent BM-EPCs derived from SHR/Izm was significantly smaller than the number derived from WKY/Izm. 17beta-Estradiol (E2) significantly increased the number of adherent BM-EPCs from SHR/Izm, and this effect was significantly attenuated by pharmacological phosphatidylinositol 3-kinase (PI3-K) blockers. Immunoblotting analysis revealed that E2 treatment led to phosphorylation of Akt. Senescence, as assessed by acidic beta-galactosidase staining, occurred at a significantly greater rate in the BM-EPCs from SHR/Izm than in those from WKY/Izm, but E2 treatment dramatically delayed the senescence of BM-EPCs from SHR/Izm. A polymerase chain reaction (PCR)-ELISA based assay revealed that telomerase activity in BM-EPCs from SHR/Izm was significantly lower than in those from WKY/Izm, but that E2 treatment significantly augmented it. Both MTS and colony forming unit assay revealed that E2 treatment significantly augmented the functional activity in BM-endothelial cell (EC)-like cells from SHR/Izm compared to that in control BM-EC-like cells (no treatment). In conclusion, the differentiation of BM-EPCs derived from SHR/Izm was significantly decreased compared with that of BM-EPCs from WKY/Izm. In addition, the rate of senescence was significantly greater in the BM-EPCs from SHR/Izm than in those from WKY/Izm. Estrogen was shown to augment differentiation and delay the onset of senescence in BM-EPCs from SHR/Izm.
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PMID:Effect of estrogen on differentiation and senescence in endothelial progenitor cells derived from bone marrow in spontaneously hypertensive rats. 1641 50

The ability of endothelial progenitor cells (EPCs) to participate in endothelial repair is impaired by angiotensin II (Ang II) and other atherogenic factors. Therefore, we investigated the effects of Ang II on the differentiation and senescence of EPCs derived from bone marrow (BM-EPCs) in an Ang II-infusion rat model. Wistar rats (n=40) were infused with Ang II or vehicle, either alone or in combination with an Ang II type 1 receptor (AT(1)R) blocker (valsartan). Bone marrow cells were obtained from the tibias and femurs. Rats of the Ang II treatment group had a significantly lower number of differentiated, adherent BM-EPCs than those of the non-treated control group. Addition of valsartan restored the level of attached, differentiated BM-EPCs to the level in the non-treated controls. The number of senescent BM-EPCs, as assessed by acidic beta-galactosidase staining, was significantly greater in the Ang II-alone group than the control group, and addition of valsartan dramatically delayed the senescence of BM-EPCs in the Ang II-alone group. A polymerase chain reaction (PCR)-ELISA-based assay revealed that telomerase activity was significantly lower in BM-EPCs from the Ang II-alone group than in those from the control group, and addition of valsartan significantly augmented this activity. An MTS assay revealed that Ang II treatment significantly decreased the functional activity in BM-EPCs, and this effect was significantly reversed by valsartan. In conclusion, Ang II decreased the differentiation and accelerated the senescence of BM-EPCs via AT(1)R.
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PMID:Endothelial progenitor cell differentiation and senescence in an angiotensin II-infusion rat model. 1694 Jul 8

Protein transduction domains (PTDs) are versatile peptide sequences that facilitate cell delivery of several cargo molecules including proteins. PTDs usually consist of short stretches of basic amino acids that can cross the plasma membrane and gain entry into cells. Traditionally, to assess PTD mediated protein delivery, PTD-fusion proteins have been used as purified proteins. To overcome the requirement for a protein purification step, we used a secretory signal peptide to allow PTD-CRE fusion proteins to be exported from transfected mammalian cells. PTD induced protein transduction into cells was assessed by a CRE-mediated recombination event that resulted in beta-galactosidase expression. Several PTDs were tested including the prototypic TAT, different TAT variants, Antp, MTS and polyarginine. A negative correlation was observed between the cationic charge on the PTD and the extent of secretion. Poor secretion was found when the PTD charge was greater than +5. One TAT-CRE protein variant had a 14-fold enhancement above CRE alone when added to cells in the presence of chloroquine. This PTD domain also enhanced gene expression after plasmid delivery. These data illustrate that some secreted PTD proteins may be useful reagents to improve protein delivery in mammalian systems and a novel approach to enhancing the response to DNA transfections.
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PMID:Comparison of protein transduction domains in mediating cell delivery of a secreted CRE protein. 1817 54