Thymidine uptake

The most convenient repair assay is thymidine uptake into Go lymphocytes (Evans and Norman, 1968). Since there is little semiconservative DNA synthesis in such cells, and what synthesis there is may be further reduced by the addition of hydroxyurea, background incorporation is essentially zero and all the counts incorporated into an acid-insoluble cell fraction can be assumed to result from repair synthesis. This method is rapid and convenient, but by itself it provides no information as to the number or size of repaired regions obviously, many repaired regions with a few base insertions or a few repaired regions with many base insertions would give similar results in terms of thymidine uptake. Nevertheless, for a quick measure of repair after insult by some suspected DNA-damaging agent, the method is ideal. 

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Production of Mucosal Damage 2.3.1.2.1 Cell culture Stimulated neutrophils are known to be cytotoxic to cells in vitro (Dull et al., 1987 Dallegri et al., 1990 Grisham et al., 1990b). Several in vitro systems have been used to demonstrate oxidative damage to intestinal cells. Xanthine/XO increased Cr release and decreased [ H]thymidine uptake by IEC-18 small intestinal epithelial cell monolayers in a dose-dependent manner (Ma et al., 1991). Rat enterocytes show decreased trypan blue exclusion and increased protein release when incubated with neutrophils stimulated... 

Fig. 12. The relation between hepatocyte shape ( ), 3H-thymidine uptake ( ) expressed as ratio to hepatocytes cultured on polystyrene dishes coated with 1 pg/ml PVLA solution and hepatic function, bile acid secretion (A), expressed as ratio to bile add secretion of hepatocytes cultured on dishes coated with 1 pg/ml of PVLA solution (Reproduced from New Functionality Materials, Vol. B [Ref. 181] through the courtesy of Elsevier Sdence Publisher B.V.)...

Proliferation of CTLL-2 in response to rmlL-2 after exposure to authentic N=0. CTLL 2 cells were exposed to 25% (of saturated solution) authentic N=0, washed, and reexposed to serial dilutions of rmIL 2 for 24 hr pH] Thymidine uptake was determined and is depicted relative to deoxygenated media controls. 

Figure 11. 3H-Thymidine uptake in PBL from S. typhimurium-r/j/ec/ec/ and uninfected calves stimulated with various antigens. Each bar shows the mean of log values of Sist with standard deviations indicated. Statistical analyses were made using Student s t test. (Reproduced with permission from Ref 59. Copyright 1982. Infect. ImmunJ...

Thymidine Uptake Studies. Tritiated thymidine (52 mCi/ anole 0.1 /rCi/ml) was added to cells for 60 min at 37°C. Cells were then washed with PBS, incubated at 4°C for 15 min in the presence of ice cold 5% TCA, rinsed with TCA and scraped from the flasks with a rubber policeman. Cells were again washed with PBS, and solubilized in 0.1N NaOH overnight. Aliquots were assayed for protein (62) and radioactivity (scintillation fluid 100 ml Biosolve (Beckman, Fullerton, CA.), 7g of PPO and 0.6 g of POPOP per liter of toluene). 

A depressed uptake of deoxyuridine brought about by a deficiency of vitamin B12 can be reversed as reflected by a reduction in labeled thymidine uptake by the addition of as little as 1 xg of the vitamin. Folic acid added to a concentration of 50 p.g/ml of the culture will correct abnormal results due to folate deficiency and may partially correct abnormal results due to vitamin B12 deficiency. Abnormal results may occur in bone marrow from patients with iron deficiency and from patients being treated with 5-fluorouracil (B7). 

Insler, G. D. (1981). Population and developmental changes in thymidine uptake kinetics of Hymenolepis diminuta (Cestoda Cyclophyllidea). Comparative Biochemistry and Physiology, 70B 697-702. 

In addition to its acute effects on contractile function of vascular and cardiac muscle, ET-1 influences cell growth, proliferation and migration. ET-1 stimulates DNA synthesis, as indicated by increased H-thymidine uptake [31-33], stimulates cellular Na /H exchange [33] and increases the expression of the proto-oncogenes, c-fos and c-myc [31] in vascular smooth muscle cells. ET-1 also induces cardiac gene expression [34]. 

Fig. 16 Comparison of cell viability of H9C2 cardiocytes by H-Thymidine uptake criteria. Viability of hypoxic cells (24 h) treated with IL was compared to Normoxic cells p — NS), PL treated hypoxic cells p < 0.01) and untreated hypoxic cells p < 0.01). (From Ref. " l)...

Organic and aqueous feverfew powdered leaf extracts were found to inhibit IL-1-induced prostaglandin E2 release from synovial cells, IL-2-induced thymidine uptake by lymphoblasts, and mitogen-induced uptake of thymidine by peripheral blood mononuclear cells (PBMCs) (23). Parthenolide also inhibited thymidine uptake by PBMCs. Both parthenolide and the extracts were cytotoxic to the PBMCs and synovial cells thus, the anti-inflammatory effects of feverfew may be secondary to cytotoxicity. These results reflect those of previous researchers who found parthenolide and other sesquiterpene lactones to be cytotoxic to cultures of human fibroblasts, human laryngeal carcinoma cells, and human cells transformed with simian virus 40 (24). 

Determine thymidine uptake by liquid scintillation counting as a measure of drug-induced cytostatic or toxicity. 

In an attempt to define whether chalone acts at the phase (Bullough s antephase) or earlier in the cell cycle, Baden and Sviokla (B3) did experiments using both the mitotic arrest technique with Colcemid and scintillation counting of tritiated thymidine uptake into DNA isolated from epidermis exposed to chalone. The number of experiments was small (3) but they found no difference in thymidine incorporation while confirming the decreases in mitotic figures arrested by colchicine. This supported the concept of a G2 effect. Epinephrine in their experiments had no effect. 

In regard to anoxia or hypoxia as a major mechanism, most recent evidence has been unfavorable, particularly for the action of amino thiols. For instance, a correlation was observed between radioprotective activity and depressive effect of thymidine uptake in DNA synthesis, whereas anoxia did not Inhibit the uptake No relation was shown between radioprotective activity and ability to Increase oxygen uptake in spleen, kidney, muscle, or red blood cells and the intracellular ox-red state in rats was unaltered even after massive doses of radiation Amino thiols, nitrite, and PAPP did not affect... 

Chakradeo PP, Kayal J J, Bhide SV. 1993. Effect of benzo(a)pyrene and methyl(acetoxymethyl) nitrosamine on thymidine uptake and induction of aryl hydrocarbon hydroxylase activity in human fetal oesophageal cells in culture. Cell Biol Int 17(7) 671-676. 

Tsutsumi S, Amagai T, Kawaguchi M, et al. 1980. Effects of strontium chloride (SrCl2) on 3H-thymidine uptake into rat lymphocytes. Bull Tokyo Dent Coll 21(4) 253-259. 

No. Inhibition of pH] Thymidine Uptake (% Intrinsic Activity, EC50 or IC5Q in nMP % of Control in Limbic DOPA Levels 10 mg/kg ifP % of Control in Limbic 5-HTP Levels 10 mg/kg ifP... 

Kelley, M., Cooper, C., Matticoli, A., and Greway, A. (2005) The detection of anti erythropoietin antibodies in human serum and plasma. Part II. Validation of a semi quantitative 3H thymidine uptake assay for neutralizing antibodies. Journal oflmmuno logical Methods, 300, 179 191. 

Diamino-l,2,4-thiadiazole possesses radioprotective activity3, a correlation has been established466 between this property and a depressive effect on thymidine uptake on myelocytes, and on the rate of DNA synthesis in bone marrow and intestinal epithilium. The radioprotective activity of 5-amino-3-methylthio-l,2,4-thiadiazole has also been reported.467... 

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