4,5',8-Trimethylpsoralen (psoralen) plus near UV light produces interstrand crosslinks and monoadducts in DNA, both of which are mutagenic. In Escherichia coli, crosslinks are incised by UvrABC excinuclease, an event that can lead to homologous recombination and repair. To determine whether UvrABC incision of crosslinks is a step in the path to mutagenesis as well as repair, the effect of DNA homologous to a target gene on a plasmid was determined. pSV2-gpt DNA was treated with psoralen and transformed into a pair of hosts: one was gpt+, the other was delta (gpt-lac)5. The DNA was extracted and transformed into a tester strain [delta (gpt-lac)5] in which Gpt- mutations in the plasmid were scored. The results show that psoralen-induced mutations were reduced to background levels by the presence of the gpt+ homolog in the host chromosome. delta gpt hosts that were constitutively induced for the SOS response yielded point mutations, whereas noninduced hosts yielded almost exclusively large deletions. Since crosslinks were estimated to be responsible for most of the mutations observed, we conclude that the premutagenic lesion of psoralen crosslinks is recombinagenic and therefore very likely to be the product of UvrABC incision.

ML-236B, a competitive inhibitor of 3-hydroxy-3-methylglutaryl-CoA reductase, significantly reduced both serum cholesterol and phospholipid levels in dogs, when used at a dosage higher than 10 mg/kg per day. Triglyceride levels were not consistently changed, but beta- and pre-beta-lipoproteins were preferentially reduced. Serum cholesterol levels were reduced by 44--45% at the higher dosage of 100--400 mg/kg per day (for 5 weeks) but ML-236B caused no significant changes in the cholesterol content of the liver and aorta and in the activities of serum GOT, GPT, CPK and lecithin : cholesterol acyltransferase. Fecal excretion of neutral sterols was unaffected but that of bile acids was markedly elevated by the drug. Under these conditions, hepatic cholesterol 7alpha-hydroxylase, the rate-limiting enzyme in bile acid biosynthesis, showed no detectable changes.

BACKGROUND

HIV-1 genotypic and phenotypic susceptibility testing (GPT) optimizes antiretroviral selection, but its effect on survival is unknown.

OBJECTIVE

To evaluate the association between GPT and survival.

METHODS

Cohort study.

METHODS

10 U.S. HIV clinics.

METHODS

2699 HIV-infected patients eligible for GPT (plasma HIV RNA level >1000 copies/mL) seen from 1999 through 2005.

METHODS

Demographic characteristics, clinical factors, GPT use, all-cause mortality, and crude and adjusted hazard ratios (HRs) for the association of GPT with survival.

RESULTS

Patients were followed for a median of 3.3 years; 915 (34%) had GPT. Patients who had GPT had lower mortality rates than those who did not (2.0 vs. 2.7 deaths per 100 person-years). In standard Cox models, GPT was associated with improved survival (adjusted HR, 0.69 [95% CI, 0.51 to 0.94]; P = 0.017) after controlling for demographic characteristics, CD4+ cell count, HIV RNA level, and intensity of clinical follow-up. In subgroup analyses, GPT was associated with improved survival for the 2107 highly active antiretroviral therapy (HAART)-experienced patients (2.2 vs. 3.2 deaths per 100 person-years for patients who had GPT vs. those who did not have GPT; adjusted HR, 0.60 [CI, 0.43 to 0.82]; P = 0.002) and for the 921 triple antiretroviral class-experienced patients (2.1 vs. 3.1 deaths per 100 person-years; adjusted HR, 0.61 [CI 0.40 to 0.93]; P = 0.022). Marginal structural models supported associations between GPT and improved survival in the overall cohort (adjusted HR, 0.54; P = 0.001) and in the HAART-experienced group (adjusted HR, 0.56; P = 0.003).

CONCLUSIONS

Use of GPT was not randomized. Residual confounding may exist.

CONCLUSIONS

Use of GPT was independently associated with improved survival among HAART-experienced patients.

BACKGROUND

Centers for Disease Control and Prevention.

Sepsis precipitates a systemic inflammatory stimulus that causes systemic release of cytokines and sequestration of polymorphonuclear neutrophils, resulting in degranulation of matrix metalloproteinases (MMPs), which causes extracellular matrix basement membrane degradation. One of the important anti-inflammatory properties of tetracyclines is their ability to inhibit MMPs. In this study, we focused on the regulation of MMPs in sepsis and their reduction by treatment with nonantimicrobial chemically modified tetracyclines (CMTs), which retain their anti-inflammatory activity. Sepsis was induced by cecal ligation and puncture (CLP) method. At 24 h and 1 h before CLP, some rats received CMT-3 (25 mg/kg), another group of rats received hydroxamate (H; an inhibitor of MMP; 25 mg/kg), and untreated rats received saline by gavage. At 0 h, 0.5 h, 1.5 h, and 24 h after CLP, blood and liver samples were collected. Plasma and liver MMP-9 by zymography and Western immunoblotting, plasma nitric oxide by measuring nitrate level, plasma glutamic oxaloacetic transaminase (GOT) and glutamic pyruvic transaminase (GPT) by enzymatic method, and liver gelatinase by radiolabeled gelatin lysis assay and 24 h mortality were determined. Plasma MMP-9 (92 kDa), nitrate, and GOT and GPT levels were elevated compared with the time 0 level and reached peak at 1.5 h CLP and remained high for 24 h. Both CMT-3 and H treatment reduced GOT,GPT, 92-kDa gelatinase, and nitrate levels throughout the 24 h. CMT-3 and H are equally effective in sepsis treatment. The 24-h mortality for CLP rats was 30%, whereas pretreatment with CMT-3 and H resulted in 0% mortality. Hepatic MMP-9 and gelatinase activity increased significantly after CLP, and pretreatment with CMT-3 and H inhibited these expressions. These results indicate the beneficial effect of CMT-3 in preventing the increase in GOT, GPT, NO, MMP-9, gelatinase activity, and the ensuing septic shock.

Several enzymes with the capacity to degrade glutamate have been suggested as possible neuroprotectants. We initially evaluated the kinetic properties of glutamate pyruvate transaminase (GPT; also known as alanine aminotransferase), glutamine synthetase, and glutamate dehydrogenase under physiologic conditions to degrade neurotoxic concentrations of glutamate. Although all three enzymes initially degraded glutamate rapidly, only GPT was able to reduce toxic (500 microM) levels of glutamate into the physiologic (<20 microM) range. Primary cultures of fetal murine cortical neurons were subjected to paradigms of either exogenous or endogenous glutamate toxicity to evaluate the neuroprotective value of GPT. Neuronal survival after exposure to added glutamate ranging from 100 to 500 microM was improved significantly in the presence of GPT >> or =1 U/ml). Cultures were also exposed to the glutamate transporter inhibitor L-trans-pyrrolidine-2,4-dicarboxylate (PDC), which produces neuronal injury by elevating extracellular glutamate. GPT significantly reduced the toxicity of PDC. This reduction was associated with a reduction in the PDC-dependent rise in the medium concentration of glutamate. These results suggest that enzymatic degradation of glutamate by GPT can be an alternative to glutamate receptor blockade as a strategy to protect neurons from excitotoxic injury.

Activated inflammatory leukocytes generate a variety of reactive oxygen and nitrogen species (RONS) that may have roles in mutagenesis and carcinogenesis. The purpose of the present study was to explore the relationship between inflammatory leukocyte activation and mutagenesis using co-culture systems. We investigated the mutagenic potentials of 12-O-tetradecanoylphorbol-13-acetate (TPA)-stimulated differentiated HL-60 (human promyelocytic leukemia cells), and RAW 264.7 cells (murine macrophages) stimulated with lipopolysaccharide (LPS) and interferon (IFN)-gamma by co-culturing each cell line with AS52 cells, a transgenic Chinese hamster ovary cell line. HL-60 cells rapidly generated superoxide (O(2)(-)) 15 min to 1 h (peak at 30 min) following TPA stimulation. RAW 264.7 cells stimulated with LPS and IFN-gamma produced O(2)(-), nitric oxide (NO) and peroxynitrite (ONOO(-)) continuously for 5-25 h. There was a 2.0-fold increase in the mutation frequency of the gpt gene in AS52 cells co-cultured with TPA stimulated HL-60 cells, when compared with non-treated cells. Importantly, this increase in mutation frequency was significantly suppressed by antioxidants, such as superoxide dismutase (SOD) and diphenylene iodonium (DPI), an NADPH oxidase inhibitor (inhibition rates: IRs = 18.2 and 35.1%, respectively). Similarly, co-culture of AS52 cells with LPS/IFN-gamma-stimulated RAW 264.7 cells also increased the mutation frequency of the gpt gene by 2.6-fold, and this increase in mutation frequency was suppressed by SOD, DPI and N(5)-(1-iminoethyl)-L-ornithine dihydrochloride (L-NIO), an specific iNOS inhibitor (IRs = 58.3, 70.8 and 70.8%, respectively). In co-culture experiments, activated HL-60 and RAW 264.7 cells increased 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels in AS52 cells when compared with non-treated controls (1.7- and 1.6-fold, respectively). Treatment of AS52 cells with hydrogen peroxide (H(2)O(2), 100 micro M), ONOO(-) (100 micro M) and SIN-1 (100 micro M), a ONOO(-) generator, also increased the mutation frequency of the gpt gene (4.6-, 5.4- and 2.8-fold, respectively). Taken together, these results support the hypothesis that RONS, derived from activated inflammatory leukocytes, are mutagenic in the biological systems, and that RONS generation inhibitors are potentially anti-mutagenic, and thus may be useful in cancer preventive strategies.

Docetaxel has been reported to show promising anti-tumour activity in pancreatic ductal cancer (PC). This study was conducted to evaluate the activity and toxicity of moderate-dose (60 mg m(-2)) docetaxel in Japanese chemo-naive patients with measurable metastatic PC. The patients had a performance status of 0-2. They received docetaxel intravenously over a 1- to 2-h period without any premedication for hypersensitivity reactions. This treatment was repeated every 3-4 weeks with dose adjustments based on the toxic effects observed. Twenty-one patients were eligible and treated with docetaxel. The median number of courses was 2 (range, 1-4). None of the patients achieved an objective response; seven showed no change and 13 showed progressive disease. In one patient, the response was not assessable because of early death. The median survival time for all patients was 118 days. The main grade 3-4 toxicities by patient were leucocytopenia (67%) and neutropenia (86%). Other grade 3-4 toxicities included anaemia (10%), thrombocytopenia (5%), nausea/vomiting (29%), anorexia (29%), GOT/GPT increase (10%), alkaline phosphatase increase (14%), malaise/fatigue (33%) and alopecia (24%). In conclusion, docetaxel, administered on this schedule, did not show significant anti-tumour activity in patients with metastatic PC.

OBJECTIVE

Kawasaki disease (KD) is an acute systematic vasculitis in children which causes coronary arterial lesions and hydrops of gallbladder. Our objective is to correlate the clinical significance and influence on disease outcome of patients with gallbladder abnormalities in Kawasaki dissease.

METHODS

Children who met KD diagnosis criteria and were admitted for IVIG treatment were retrospectively enrolled for analysis. Patients with abdominal sonography were divided into 2 groups based on the absence (Group A, N = 61) or presence (Group B, N = 16) of gallbladder abnormalities (GBA), defined as hydrops or acalculous cholecystitis. Between the two groups, clinical features, demographic data (including admission days, coronary artery lesions, IVIG resistance), and laboratory data before/after IVIG treatment were collected for analysis.

RESULTS

The presence of sonographic gallbladder abnormalities is correlated with higher levels of serum CRP, GPT, and neutrophils. It also points to an increased number of IVIG resistance rates in group B. There was no significant statistical difference among clinical features, age, gender, admission days, or coronary artery lesions between the two groups.

CONCLUSIONS

Sonographic gallbladder abnormalities are associated with higher CRP, GPT, neutrophil and IVIG resistance in KD. It can be used as a predictor of IVIG resistance in patients with KD.

To find out the active principles against ethanol-induced toxicity in mice, Andrographis paniculata Nees. (Ap) was chosen and isolated andrographolide (ANDRO) and arabinogalactan proteins (AGPs). ANDRO was detected by HPTLC, FTIR and quantified by HPLC (10mg/g of Ap powder). AGPs was detected by beta-glucosyl Yariv staining of SDS-PAGE gel, FTIR and quantified by single radial gel diffusion assay with beta-glucosyl Yariv reagent (0.5mg/g Ap powder). The mice are pretreated intra-peritoneally (i.p.) with different doses (62.5, 125, 250, and 500mg/kg) of body weight of mice] of ANDRO and AGPs for 7 days and then ethanol (7.5g/kg of body weight) was injected, i.p. Besides, silymarin was used as standard hepatoprotective agent for comparative study with ANDRO and AGPs. The ameliorative activity of ANDRO and AGP against hepatic renal alcohol toxicity was measured by assessing GOT, GPT, ACP, ALP and LP levels in liver and kidney. It has been observed that pretreatment of mice with ANDRO and AGPs at 500mg/kg of body weight and 125mg/kg of body weight respectively could able to minimize the toxicity in compare to ethanol treated group as revealed by the different enzymatic assay in liver and kidney tissues and the results were comparable with silymarin. Hence, out of several ill-defined compounds present in Ap, ANDRO and AGPs are the potential bioactive compounds responsible for protection against ethanol-induced toxicity.

Piperine, an active alkaloidal constituent of the extract obtained from Piper longum and Piper nigrum, was evaluated for its antihepatotoxic potential in order to validate its use in traditional therapeutic formulations. This plant principle exerted a significant protection against tert-butyl hydroperoxide and carbon tetrachloride hepatotoxicity by reducing both in vitro and in vivo lipid peroxidation, enzymatic leakage of GPT and AP, and by preventing the depletion of GSH and total thiols in the intoxicated mice. Silymarin, a known hepatoprotective drug was tested simultaneously for comparison. Piperine showed a lower hepatoprotective potency than silymarin.

To further analyze the VDJ recombination defect in lymphoid pre-B cells from mice with severe combined immune deficiency (scid mice), we have assayed the ability of Abelson murine leukemia virus (A-MuLV) transformed pre-B cells from scid mice to rearrange a recombination substrate in which inverted VH to DJH joins activate a selectable (gpt) gene. In unselected populations, substrate rearrangements occurred frequently, but were aberrant and probably analogous to the aberrant rearrangements observed at endogenous scid Ig gene loci. In contrast, populations of scid pre-B lines selected for gpt activity within the substrate contained mostly "normal" VH to DJH joins within the introduced substrate. These findings demonstrate that scid pre-B cells can make normal joins at low efficiency and are discussed with respect to the potential mechanism of the scid defect and the occurrence of Igs in leaky scid mice.

OBJECTIVE

To investigate the effect of partial splenic embolization (PSE) on platelet values in liver cirrhosis patients with thrombocytopenia and to determine the effective embolization area for platelet values improvement.

METHODS

Blood parameters and liver function indicators were measured on 10 liver cirrhosis patients (6 in Child-Pugh grade A and 4 in grade B) with thrombocytopenia (platelet values < 80 x 10(3)/microL) before embolization. Computed tomography scan was also needed in advance to acquire the splenic baseline. After 2 to 3 d, angiography and splenic embolization were performed. A second computed tomography scan was made to confirm the embolization area after 2 to 3 wk of embolization. The blood parameters of patients were also examined biweekly during the 1 year follow-up period.

RESULTS

According to the computed tomography images after partial splenic embolization, we divided all patients into two groups: low (< 30%), and high >> or = 30%) embolization area groups. The platelet values were increased by 3 times compared to baseline levels after 2 wk of embolization in high embolization area group. In addition, there were significant differences in platelet values between low and high embolization area groups. GPT values decreased significantly in all patients after 2 wk of embolization. The improvement in platelet and GPT values still persisted until 1 year after PSE. In addition, 3 of 4 (75%) Child-Pugh grade B patients progressed to grade A after 2 mo of PSE. The complication rate in < 30% and>> or = 30% embolization area groups was 50% and 100%, respectively.

CONCLUSIONS

Partial splenic embolization is an effective method to improve platelet values and GPT values in liver cirrhosis patients with thrombocytopenia and the>> or = 30% embolization area is meaningful for platelet values improvement. The relationship between the complication rate and embolization area needs further studies.

Liver is the main detoxifying organ and therefore the target of high concentrations of genotoxic compounds, such as environmental carcinogens and anticancer drugs. Here, we investigated the usefulness of lovastatin, which is nowadays widely used for lipid lowering purpose, as a hepatoprotective drug following the administration of the anthracycline derivative doxorubicin in vivo. To this end, BALB/c mice were exposed to either a single high dose or three consecutive low doses of doxorubicin. Acute and subacute hepatotoxicities were analyzed with or without lovastatin co-treatment. Lovastatin protected the liver against doxorubicin-induced acute pro-inflammatory and pro-fibrotic stress responses as indicated by an attenuated mRNA expression of tumor necrosis factor alpha (TNFα) and connective tissue growth factor (CTGF), respectively. Hepatoprotection by lovastatin was due to a reduced induction of DNA damage following doxorubicin treatment. The statin also mitigated subacute anthracycline-provoked hepatotoxicity as shown on the level of doxorubicin- and epirubicin-stimulated CTGF mRNA expression as well as histopathologically detectable fibrosis and serum concentration of marker enzymes of hepatotoxicity (GPT/GLDH). Kidney damage following doxorubicin exposure was not detectable under our experimental conditions.Moreover, lovastatin showed multiple inhibitory effects on doxorubicin-triggered hepatic expression of genes involved in oxidative stress response, drug transport, DNA repair, cell cycle progression and cell death. Doxorubicin also stimulated the formation of ceramides. Ceramide production, however, was not blocked by lovastatin, indicating that hepatoprotection by lovastatin is independent of the sphingolipid metabolism. Overall, the data show that lovastatin is hepatoprotective following genotoxic stress induced by anthracyclines. Based on the data, we hypothesize that statins might be suitable to lower hepatic injury following anthracycline-based anticancer therapy.

We have investigated the effects of the calcium antagonist nifedipine on antigen-induced bronchoconstriction in vivo and in vitro. Eight grass-pollen-sensitive asthmatics were given either nifedipine (20 mg sublingually) or placebo 30 min before antigen challenge. The fall in forced expiratory volume in one second after pretreatment with placebo was 42.8 +/- 10.1%. After nifedipine this fall was significantly reduced to 26.5 +/- 11.7% (p less than 0.005). Two in vitro models of allergic asthma have been studied: actively sensitized guinea pig tracheal strips (GPT) and passively sensitized human bronchial muscle (HBM). Contraction of GPT by acetylcholine, histamine, and antigen challenge was unaffected by nifedipine 10(-4)M. Contraction of HBM by acetylcholine, histamine and grass pollen antigen challenge was significantly reduced by nifedipine 10(-4)M and 10(-6)M. The magnitude of the reduction in contraction to antigen challenge was comparable to the inhibition of acetylcholine and histamine responses. It would appear most likely that nifedipine exerts its effect mainly on bronchial muscle contractility rather than by stabilizing mast cells.

The enzyme which initiates the dolichol pathway of protein N-glycosylation, dolichol-P-dependent N-acetylglucosamine-1-P transferase (GPT), is encoded by the ALG7 gene. Essential for viability, ALG7 has been evolutionarily conserved and shown to be involved in a variety of functions. ALG7 is an early growth-response gene in yeast, and downregulation of ALG7 expression results in diminished N-glycosylation and secretion of Xenopus oocyte proteins. We have now investigated the consequences of diminished GPT activity in yeast using mutant ALG7 genes with deletions in the 3' untranslated region (3' UTR). We show that a 2.5- to 4-fold reduction in GPT activity gave rise to distinct phenotypes, whose severity was inversely related to the level of GPT activity. These phenotypes included hypersensitivity to tunicamycin, enlarged cell size, extensive aggregation, lack of a typical stationary (G0) arrest, and defective spore germination. We conclude that yeast cells are sensitive to GPT dosage, and that attenuation of GPT activity interferes with various functions in the yeast life cycle.

The likely future increase in atmospheric CO(2) and associated changes in climate will affect global patterns of plant production. Models integrate understanding of the influence of the environment on plant physiological processes and so enable estimates of future changes to be made. Moreover, they allow us to assess the consequences of different assumptions for predictions and so stimulate further research. This paper is a review of the sensitivities of one such model, Hybrid6.5, a detailed mechanistic model of terrestrial primary production. This model is typical of its type, and the sensitivities of the global distribution of predicted production to model assumptions and possible future CO(2) levels and climate are assessed. Sensitivity tests show that leaf phenology has large effects on mean C(3) crop and needleleaved cold deciduous tree production, reducing potential net primary production (NPP) from that obtained using constant maximum annual leaf area index by 32.9% and 41.6%, respectively. Generalized Plant Type (GPT) specific parameterizations, particularly photosynthetic capacity per unit leaf N, affect mean predicted NPP of higher C(3) plants by -22.3% to 27.9%, depending on the GPT, compared to NPP predictions obtained using mean parameter values. An increase in atmospheric CO(2) concentrations from current values to 720 ppm by the end of this century, with associated effects on climate from a typical climate model, is predicted to increase global NPP by 37.3%. Mean increases range from 43.9-52.9% across different C(3) GPTs, whereas the mean NPP of C(4) grass and crop increases by 5.9%. Significant uncertainties concern the extent to which acclimative processes may reduce any potential future increase in primary production and the degree to which any gains are transferred to durable, and especially edible, biomass. Experimentalists and modellers need to work closely together to reduce these uncertainties. A number of research priorities are suggested. 'The green leaf or, to be more precise, the microscopic green grain of chlorophyll, is the focus, the point in the world to which solar energy flows on one side while all the manifestations of life on earth take their source on the other side.' Kliment Arkadievich Timiryazev The conclusions of a century of plant physiology, speech at Moscow University, 12 January 1901.

Teratocarcinoma (TCC) stem cells can function as vehicles for the introduction of specific recombinant genes into mice. Because most genes do not code for a selectable marker, we investigated the transformation efficiency of vectors with a linked selectable gene. In one series, TCC cells first selected for thymidine kinase deficiency were treated with DNA from the plasmid vector PtkH beta 1 containing the human genomic beta-globin gene and the thymidine kinase gene of herpes simplex virus. A high transformation frequency was obtained after selection in hypoxanthine-aminopterin-thymidine medium. Hybridization tests revealed that the majority of transformants had intact copies of the human gene among three to six total copies per cell. These were associated with cellular DNA sequences as judged from the presence of additional new restriction fragments and from stability of the sequences in tumors produced by injecting the cells subcutaneously. Total polyadenylate-containing RNA from cell cultures of two out of four transformants examined showed hybridization to the human gene probe: one RNA species resembled mature human beta-globin mRNA transcripts; the others were of larger size. In differentiating tumors, various tissues, including hematopoietic cells of TCC provenance could be found. In a second model set of experiments, wild-type TCC cells were used to test a dominant-selection scheme with pSV-gpt vectors. Numerous transformants were isolated, and their transfected DNA was apparently stably integrated. Thus, any gene of choice can be transferred into TCC stem cells even without mutagenesis of the cells, and selected cell clones can be characterized. Cells of interest may then be introduced into early embryos to produce new mouse strains with predetermined genetic changes.

Mice were administered a single dose of carbon tetrachloride (CCl(4)) to induce acute liver injury. We found that lactate dehydrogenase (LDH) and glutamic pyruvic transaminase (GPT) levels in serum, as well as the level of thiobarbituric acid reaction substances (TBARS) in liver homogenate increased significantly in a manner both dose dependent and time dependent after CCl(4) administration. Such results suggest that the liver is susceptible to CCl(4) treatment and that lipid peroxidation is associated with CCl(4)-induced liver injury. The spin-trapping electron paramagnetic resonance (EPR) method was used to detect nitric oxide (NO) level in liver. The chemiluminescence method was also employed to measure the NO(2)(-)/NO(3)(-) concentration in serum. The NO levels in liver tissues and NO(2)(-)/NO(3)(-) concentration in serum were found to decrease significantly both in a dose-dependent manner and in time course after CCl(4) treatment. The nitric oxide synthase (NOS) II activity in the liver, in contrast, was found to increase significantly. Our study suggests that not only should the expression of NOS be analyzed but NO organ and blood concentration must be measured in the study of diseases involving nitric oxide. L-arginine treatment had no significant effect on the liver function of CCl(4)-treated mice. It was found that NO donor sodium nitroprusside (SNP; 50 or 100 microg/kg) treatment resulted in decreases of LDH, GPT, and TBARS levels, leading to a protective effect on CCl(4)-treated mice. On the other hand, N(G)-nitro-L-arginine methyl ester (L-NAME, 100 or 300 mg/kg) treatment caused more severe liver damage. Moreover, we have found in an in vitro EPR study that SNP could scavenge lipid peroxyl radical LOO&z.rad;. The above results together suggest that NO may protect CCl(4)-induced liver injury through scavenging lipid radical, inhibiting the lipid peroxidation chain reaction. On the basis of our analysis, we put forth two explanations for the stated discrepancy between NOS II and NO production: (i) NO was used up gradually in terminating lipid peroxidation and (ii) NADPH was depleted (on the basis of correlation evidence only).

The mechanism of adenine toxicity in an hpt gpt strain of Escherichia coli that is extremely sensitive to adenine inhibition was investigated. Adenine-resistant derivatives had secondary mutations in adeninephosphoribosyltransferase or the purR repressor. Growth studies with various purine salvage pathway mutants and the ability of guanosine to prevent adenine toxicity indicated that adenine exerts its toxic effects by depleting guanine nucleotide pools. In the presence of adenine, ATP pools increased twofold in wild-type cells and stabilized after 5 min. In contrast, ATP pools continued to rise in hpt gpt cells up to 25 min and increased sevenfold after adenine addition. hpt gpt cells were shown to have higher levels of adeninephosphoribosyltransferase than did the wild-type cells. In response to adenine addition, GTP pools dropped three- to fourfold in all strains tested. Although GTP levels returned to near normal values in wild-type cells after 35 min, no restoration of GTP pools was observed in the hpt gpt strain during this period. Measurements of guanine pools before and after the addition of adenine indicated that guaninephosphoribosyltransferase plays an important role in maintaining GTP pools by converting the free guanine to GMP during guanine nucleotide depletion.

Copper in drinking water has been associated with Non-Indian Childhood Cirrhosis (NICC), a form of early childhood liver cirrhosis. This epidemiological study examines the exposition of infants to increased copper concentrations through drinking water from public water supplies in Berlin, Germany, and if this dietary copper intake can cause liver damage in early childhood. In total, water samples from 2944 households with infants were tested for copper. Mean copper concentrations in the two different types of collected composite samples were 0.44 and 0.56 mg/l, respectively. Families having a copper concentration at or above 0.8 mg/l in one or both of the composite samples (29.9% of all sampled households) and a defined minimum ingestion of tap water of their infant were recommended to undergo a paediatric examination. Nearly every of the 541 recommended infants were examined by a local paediatrician and of these 183 received a blood serum analysis, too. None of the infants had clear signs of a liver disease although a few serum parameters lay outside the accompanying reference range and abdominal ultrasound imaging gave slightly unusual results in five cases. Additionally, no signs of a negative health effect could be found in the statistical analysis of the serum parameters GOT, GPT, GGT, total bilirubin, serum copper, or ceruloplasmin in relation to estimated daily and total copper intakes of the infants from tap water. No dose relation of serum parameters and estimated copper intakes could be established. From the results of the study, no confirmed indication of a liver malfunction in infants whose food had been prepared using tap water with an elevated copper concentration could be found and, therefore, no indication of a hazard due to copper pipes connected to public water supplies could be detected.

BACKGROUND

Gexia-Zhuyu Tang (GZT), also called Gexiazhuyu decoction (GXZYD), is a traditional Chinese herbal medicine for chronic liver diseases such as cirrhosis and liver fibrosis.

OBJECTIVE

In this study, we have investigated the affects of GZT on a rat model of dimethylnitrosamine (DMN)-induced liver fibrosis.

METHODS

In this study, the protective effects of GZT on DMN-induced liver fibrosis were measured using a rat model. Following 5 weeks of DMN-treatment (8 mg/kg, i.p., given 3 consecutive days each week), oral administration of GZT at 1.8 g/kg daily via oral gavage for 2weeks beginning at week 13.

RESULTS

Both body and liver weights were significantly decreased. The reductions in body and liver weights corresponded with increasing liver damage severity. Furthermore, GZT-treatment remarkably decreased the levels of serum GOT (glutamate oxaloacetate transaminase) and GPT (glutamic pyruvic transaminase), and the mRNA expression levels of collagen alpha-1(I) and alpha-smooth muscle actin (alpha-SMA) in DMN-induced hepatic fibrosis. In addition, hepatic stellate cells (HSCs) play a major role in various types of liver fibrosis through initial myofibroblast transformation. The proliferation of HSCs was inhibited by GZT. Treatment with GZT also induced HSC apoptosis in a dose- and time-dependent manner. GZT treatment induced HSC apoptosis by facilitating Ca(2+) release from the mitochondria within 6h. Subsequently, caspases 3 and 12 were elevated by 72 h after treatment.

CONCLUSIONS

Our studies indicate that GZT exhibited both hepatoprotective and antifibrogenic effects in DMN-induced hepatic injury. These findings suggest that GZT may be useful in preventing the development of hepatic fibrosis.

AS52 cells are Chinese hamster ovary (CHO) cells that carry a single functional copy of the bacterial gpt gene and allow the isolation of 6-thioguanine-resistant (6TGr)mutants arising from mutation at the chromosally integrated gpt locus. The gpt locus in AS52 cells is extremely stable, giving rise to 6TGr mutants at frequencies comparable to the endogenous CHO hprt locus. In this study, we describe the spectrum of spontaneous mutations observed in AS52 cells by Southern blot and DNA sequence analyses. Using the polymerase chain reaction (PCR) and the Thermus aquaticus (Taq) polymerase, we have enzymatically amplified 6TGr mutant gpt sequences in vitro. The PCR product was then sequenced without further cloning manipulations to directly identify gpt structural gene mutations. Deletions predominant among the 62 spontaneous 6TGr-AS52 mutant clones analyzed in this study. Of these, 79% (49/62) of the mutations were identified as deletions either by Southern blotting, PCR amplification or DNA sequence analysis. Among these deletions is a predominant 3-base deletion that was observed in 31% (19/62) of the mutants. These data provide a basis for future comparisons of induced point mutational spectra derived in the AS52 cell line, and demonstrate the utility of PCR in the generation of DNA sequence spectra derived from chromosomally integrated mammalian loci.

Recombination was measured in Chinese hamster ovary (CHO-K1) cells and in the X-ray-sensitive mutants xrs1 and xrs7, which show a defect in DNA double-strand break repair. To assay recombination, pairs of derivatives of the plasmid pSV2gpt were constructed with nonoverlapping deletions in the gpt gene region and cotransferred into the different cell types. Recombination efficiencies, measured as the transformation frequency with a pair of deletion plasmids relative to that with the complete pSV2gpt plasmid, were about 6% in both CHO-K1 and the xrs mutants for plasmids linearized at a site outside the gpt gene. However, these efficiencies were substantially enhanced by the introduction of a double-strand break into the homologous region of the gpt gene in one of a pair of deletion plasmids before cotransfer. This enhancement was apparently only about half as great for the xrs cells as for CHO-K1, but variation in the data was considerable. A much larger difference between CHO-K1 and the xrs mutants was found when the DNA concentration dependence of transformation was explored. While the transformation frequency of CHO-K1 increased linearly with DNA concentration, no such increase occurred with the xrs mutants irrespective of whether complete plasmids or pairs of deletion plasmids were transferred. The fraction of cells taking up DNA, assayed autoradiographically, was similar in all cell types. Therefore we suggest that while homologous recombination of plasmid molecules may not be substantially reduced in the xrs mutants,processes involved in the stable integration of plasmid DNA into genomic DNA are significantly impaired.

Liver, being primary site for biotransformation of foreign compounds is vulnerable to various chemical assaults. Ginseng has a wide range of pharmacological and therapeutical action. In the present study an attempt has been made to study the cadmium chloride (CdCl(2)) induced toxicity in liver and its possible protection by Panax ginseng. Swiss albino mice were divided into four groups: (i) Control group--only vehicle (double distilled water) (ii) Ginseng treated group - 10mg/kg b.wt. orally (iii) CdCl(2) treated group - 1.0mg/kg b.wt. CdCl(2) i.p. (iv) Combination group--Ginseng root extract (10mg/kg b.wt.) and CdCl(2) (1.0mg/kg b.wt.). Activities of alkaline phosphatase, GOT, GPT were measured in serum and lipid peroxidation (LPO) and GSH content were measured in liver. The results indicated a significant increase in LPO, GOT, GPT activities and decrease in GSH and serum alkaline phosphatase activities after CdCl(2) treatment. Ginseng alone did not show any significant alterations except a significant decrease in LPO level. Combined treatment of Ginseng and CdCl(2) showed significant decrease in LPO, GOT, GPT and elevation in GSH and serum alkaline phosphatase as compared to CdCl(2) treated group. Thus, Ginseng is found to be protective against cadmium-induced hepatic injuries.