Effects of araC and aphidicolin on DNA chain elongation rate in HeLa S3 cells

Summary The effects of araC and aphidicolin on DNA chain elongation rate were tested. The rate was markedly reduced at low concentrations. Total DNA synthesis was more inhibited, indicating a role of DNA polymerasea in replicon initiation.     

A possibility to achieve genetic transformation in the platyfish-swordtail system (Platypoecilus maculatus — Xiphophorus helleri)

Summary In order to determine how informative homologous donor DNA might be made available to propigment cells of the recipientXiphophorus helleri for transformation, labelled heterologous DNA fromE. coli was injected into the neural crest region or the yolk sac of embryos of the recipient. On the basis of the degradation rate of the donor DNA and the incorporation rate of radioactivity into the recipient DNA, it is concluded that injection into the neural crest region may be a suitable method to make available informative homologous donor DNA for transformation.Supported by DFG through SFB No. 103, and by Stiftung Volkswagenwerk.     

Zur Wirkung von Zn++ auf die Nukleinsäurebiosynthese von Aszites-Tumorzellen

Summary Ascites tumour cells have been employed to study the reactivity of Zn⁺⁺ on nucleic acid biosynthesis. 10^–4 M Zn⁺⁺ caused a selective inhibition of DNA synthesis of intact cells. The rate of RNA- and protein-biosynthesis, however, remained unchanged. The activity of DNA polymerase as well as DNA dependent RNA polymerase was strongly affected by Zn⁺⁺ in vitro.     

Detection of human cytomegalovirus (HCMV) DNA from cell-free plasma of immunosuppressed patients by nested PCR: influence of nucleic acid extraction method

Conclusions Blood cells and plasma preparations from HCMV-seropositive healthy blood donors were all nPCR negative. Detection of HCMV DNA from PBMC and granulocytes (DNAemia) of immunosuppressed patients by nPCR did not correlate with the isolation of infectious virus from these cell populations in cell culture (viremia). However HCMV could be isolated in 60% of cases from other materials of the same patient. HCMV DNA detected in blood cells persisted for up to one year in an asymptomatically infected individual after NTX. The sensitivity of HCMV DNA detection in cell-free plasma (up to 5 fg) depended on the method used for DNA isolation. The rate of HCMV DNA detection in plasma was lower than in leukocytes. In all cases of positive plasma PCR infectious virus could be isolated from any other material of the symptomatically infected patients. Therefore HCMV DNA PCR from plasma of immunosuppressed patients seems to be a suitable and easy alternative to HCMV RT/PCR for routine diagnosis of HCMV disease.     

重庆地区HPV亚型分布和基因芯片技术对宫颈病变筛查意义的研究

目的 了解重庆地区人乳头瘤病毒（human papillomavirus,HPV）感染情况及其亚型分布,探讨HPV的多重感染以及HPV病毒负荷量与不同级别宫颈病变的相关性,并比较HPV-DNA检测杂交捕获法和基因芯片法对宫颈HPV感染的检出效率,评价基因芯片法作为筛查方法的优缺点,为建立最佳筛查方案预防宫颈癌提供科学依据.方法 对2009年在重庆市肿瘤医院肿瘤研究所妇科肿瘤科就诊的360例自愿接受宫颈癌机会性筛查的患者进行液基细胞学和HPV-DNA的检测,其中HPV-DNA检测采用杂交捕获法和基因芯片法,并以基因测序为金标准评价两种检测方法的检测效率.结果 本次共检出27种HPV亚型,检出率前三位依次为：HPV16、HPV58和HPV52.HPV总阳性率为42.50%,其中高危型检出率为34.17%;低危型检出率为15.56%;多重感染率为15.00%.以液基细胞学检查为标准,随宫颈病变程度的增加,HPV感染率、高危型HPV感染率、多重感染率及病毒负荷量逐渐增加,组问差异有显著性,低危型HPV感染率无规律性改变.杂交捕获法和基因芯片法的灵敏度、特异度和约登指数分剐为前者97.56%、98.17%和0.957,后者98.33%、93.40%和0.917.结论 高危亚型HPV感染是宫颈上皮内病变的主要因素,HPV16是重庆地区感染率最高的HPV亚型,其次为HPV58和HPV52.HPV的多重感染和病毒负荷量都与宫颈病变程度有一定的相关性.两种HPV检测方法相比,杂交捕获法特异度较高,而基因芯片法灵敏度较高,基因芯片法可作为新的宫颈病变筛查方法.     

Etude de la consommation d'oxygène et de la teneur en acide désoxyribonucléique du foie à divers âges chez le rat

Summary The quantitative study of whole liver DNA and liver nuclei DNA at various ages, and the simultaneous study of oxygen uptake shows the following facts:(1) The rate of oxygen consumption to the fresh weight or to the DNA or to diploid chromosomes group is the same for the adult or the young aged 8 days. The values of oxygen consumption calculated for one cell or for one nucleus (taking notice of the fact that the mean level of liver nuclei DNA varies with the age) at 8 days is only the half of the adult value. Even at two months the value is 20% lower than in the adult.(2) In older animals there is a decrease of oxygen uptake which is more noticeable on a cellular scale than with the classical rate to fresh or dry weight. We affirm the fact that the diminution of the oxygen uptake of each cell is compensated by the increase of the number of cells.     

Kinetic analysis,size profiling,and bioenergetic association of DNA released by selected cell lines in vitro

Although circulating DNA (cirDNA) analysis shows great promise as a screening tool for a wide range of pathologies, numerous stumbling blocks hinder the rapid translation of research to clinical practice. This is related directly to the inherent complexity of the in vivo setting, wherein the influence of complex systems of interconnected cellular responses and putative DNA sources creates a seemingly arbitrary representation of the quantitative and qualitative properties of the cirDNA in the blood of any individual. Therefore, to evaluate the potential of in vitro cell cultures to circumvent the difficulties encountered in in vivo investigations, the purpose of this work was to elucidate the characteristics of the DNA released [cell-free DNA (cfDNA)] by eight different cell lines. This revealed three different forms of cfDNA release patterns and the presence of nucleosomal fragments as well as actively released forms of DNA, which are not only consistently observed in every tested cell line, but also in plasma samples. Correlations between cfDNA release and cellular origin, growth rate, and cancer status were also investigated by screening and comparing bioenergetics flux parameters. These results show statistically significant correlations between cfDNA levels and glycolysis, while no correlations between cfDNA levels and oxidative phosphorylation were observed. Furthermore, several correlations between growth rate, cancer status, and dependency on aerobic glycolysis were observed. Cell cultures can, therefore, successfully serve as closed-circuit models to either replace or be used in conjunction with biofluid samples, which will enable sharper focus on specific cell types or DNA origins.     

Changes in Plasmodium berghei berghei in mice maintained at high temperatures]

The study of the evolution of Plasmodium berghei berghei is made in mice kept in a high temperature (35 degrees C) throughout the experiment. Some of these mouse parasites (less than 30%) show a gigantic atypical morphology. In the parasite growing in animals kept at 35 degrees C, the amount of DNA is higher than DNA rate of the parasites growing in control mice (20-22 degrees C). There is no evidence of any relation between the increase of DNA amount and the morphological modification of these parasites.     

Effect of epidermal growth factor on growth and maturation of fetal and neonatal rat small intestine in organ culture

Small intestinal explants from pre- and post-natal rats were incubated in an organ culture system in the absence and presence of epidermal growth factor (EGF). The rate of synthesis of small intestinal DNA and protein as well as the activity of lactase and alkaline phosphatase increased rapidly between 17 and 20-day gestational age, whereafter they declined. The maximal incorporation of 3H-thymidine and 14C-alanine into DNA and protein, respectively, was significantly stimulated by EGF (100 ng/ml). EGF had no effect on the activity of either lactase or alkaline phosphatase in the small intestinal explants.     

Purin- und Pyrimidinbasen der Desoxyribonukleinsäure des Ehrlichschen Mäuse-Ascites-Karzinomas

Summary The proportion of purines and pyrimidines in the DNA of the Ehrlich mouse ascites carcinoma is examined and compared with the base proportion of the DNA from liver and kidney of the normal mouse. The following findings were obtained: The DNA of the ascites tumor contains more guanine and less cytosine and thymine than the DNA of the normal mouse. For guanine this difference is statistically significant. The changed base compound is discussed with regard to the changed growing rate in tumors and the altered chromosome structure in the Ehrlich ascites carcinoma.

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Durchgeführt mit Unterstützung der Deutschen Forschungsgemeinschaft.     

Gene knockout and knockin by zinc-finger nucleases: current status and perspectives

Zinc-finger nucleases (ZFNs) are engineered site-specific DNA cleavage enzymes that may be designed to recognize long target sites and thus cut DNA with high specificity. ZFNs mediate permanent and targeted genetic alteration via induction of a double-strand break at a specific genomic site. Compared to conventional homology-based gene targeting, ZFNs can increase the targeting rate by up to 100,000-fold; gene disruption via mutagenic DNA repair is similarly efficient. The utility of ZFNs has been shown in many organisms, including insects, amphibians, plants, nematodes, and several mammals, including humans. This broad range of tractable species renders ZFNs a useful tool for improving the understanding of complex physiological systems, to produce transgenic animals, cell lines, and plants, and to treat human disease.     

Acylphosphatase overexpression triggers SH-SY5Y differentiation towards neuronal phenotype

An acylphosphatase (AcPase) overexpression study was carried out on SH-SY5Y neuroblastoma cells, using a green fluorescent fusion protein (AcP-GFP), with GFP acting as a reporter protein. The cellular proliferation rate was significantly reduced by overexpression of AcPase by a factor of ten. In contrast, clones transfected with two inactive AcPase mutants showed a growth rate comparable to control cells. This suggests that AcPase catalyzes the proliferative down-regulation. AcPase-overexpressing clones showed a physiological mortality rate as assessed by an MTT reduction test and by evaluation of necrotic markers. DNA fragmentation analysis and assays of caspase-3 and poly (ADP-ribose) polymerase (PARP)-active fragments showed no evidence of any apoptotic pattern. AcPase overexpression led to a marked increase in PARP activity as well as Bcl-2 content; these are commonly up-regulated during differentiative processes in neuronal cells. In fact, the typical differentiation marker, growth-associated-protein 43, was significantly up-regulated. Microscopic observations also showed a clear increase in the differentiative phenotype in AcPase-overexpressing cells. Our results clearly show that AcPase plays a primary causative role in neuronal differentiation.Received 3 May 2004; accepted 25 May 2004     

Metabolic activity of adipose tissue in the goat during gestation and at the beginning of lactation]

In 5 French Alpine goats, omental adipose tissue acetyl-CoA carboxylase, glucose-6-phosphate deshydrogenase, malic enzyme and lipoprotein lipase activities significantly decreased during the third month of gestation, whereas plasma non-esterified fatty acid and triacyglycerol contents increased. This probably reflects an early decreasing rate of adipose tissue anabolism during gestation in the Goat. At the third week of lactation, anabolic activities relative to DNA content of adipose tissue were extremely low, and the tissue weight relative to DNA was lower than during gestation. Metabolic alterations of omental adipose tissue in early lactation do not seem to be related to milk production level. These results could contribute to a better control of the kinetic of body lipid stores during the reproductive cycle in high milk yielding ruminants.     

Effect of epidermal growth factor on growth and maturation of fetal and neonatal rat small intestine in organ culture

Summary Small intestinal explants from pre- and post-natal rats were incubated in an organ culture system in the absence and presence of epidermal growth factor (EGF). The rate of synthesis of small intestinal DNA and protein as well as the activity of lactase and alkaline phosphatase increased rapidly between 17 and 20-day gestational age, whereafter they declined. The maximal incorporation of³H-thymidine and¹⁴C-alanine into DNA and protein, respectively, was significantly stimulated by EGF (100 ng/ml). EGF had no effect on the activity of either lactase or alkaline phosphatase in the small intestinal explants.Acknowledgment. The project was supported by grants from the Veterans Administration Research Service. The authors wish to thank Dr. M. C. Geokas, Chief, Department of Medicine, Veterans Administration Medical Center, Martinez, CA, for providing us with excellent laboratory facilities and for his encouragement in this study.     

Metabolic status of temperature induced swollen conidia ofAspergillus nidulans

Summary There is an active synthesis of DNA, RNA, protein, and carbohydrate in conidia incubated at 48°C. DNA increases more or less 3 times; while RNA, protein and carbohydrate are synthesized at a much faster rate. Conidia do not germinate at this temperature.Acknowledgments. Grateful thanks are due to Professor B. M. Johri for valuable suggestions. R. K. S. acknowledges with thanks a Senior Research Fellowship of the Council of Scientific and Industrial Research, New Delhi.     

Die DNS-Synthese in Leber und Niere junger Ratten. Autoradiographische Untersuchungen mit3H-Thymidin

Summary Autoradiographic studies following a single i.p. injection of³H-thymidine were performed in liver and kidney parenchyma of new-born and young adult rats at different ages (1, 2, 4, 7, 12, 18, 24, 30, 60 and 120 days). In 1-day-old animals the tritium index (i.e. percentage of DNA synthesizing nuclei) of both organs is lower than in the rat embryo. From 1–4 days fluctuations occur, then the tritium index rises at 7 and 12 days. But now an exponential decrease is observed up to day 120. This depression and fluctuation of the tritium index probably depends on a postpartal functional transposition and especially in the liver on a structural transformation. The mean grain density in the labelled nuclei increases from 1 up to 24 days, which is likely connected with a very high amount of endogenous thymidine in new-born rats. From day 24 the mean grain density can be taken as a relative measure for the rate of DNA synthesis. The reduction in the rate of DNA synthesis from 24–120 days is explained as a consequence of the commutation from the rapid to the slow mode in cellular proliferation.

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Mit Unterstützung der Deutschen Forschungsgemeinschaft.     

Alternariol, a dibenzopyrone mycotoxin of Alternaria spp., is a new photosensitizing and DNA cross-linking agent

The mycotoxin alternariol (3,4',5-trihydroxy-6'-methyldibenzo [a] pyrone) but not alternariol monomethyl ether (3,4'-dihydroxy-5-methoxy-6'-methyldibenzo [a] pyrone) is phototoxic to Escherichia coli in the presence of near UV light (320-400 nm). The phototoxicity bioassays with a DNA repair-deficient mutant of E. coli suggested that DNA may be the molecular target for photo-induced toxicity of alternariol. Interactions between alternariol and double-stranded, supercoiled DNA suggest that alternariol interacts with DNA by intercalation. No DNA breakage was detected in this system; however, alternariol forms a complex and cross-links double-stranded DNA in near UV light. These results suggest that alternariol is a new phototoxic, DNA-intercalating agent and is a DNA cross-linking mycotoxin in near UV light.     

Electron microscopy of mitochondrial DNA in Podospora anserina and the presence of a multimeric range of circular DNA molecules from senescent cultures]

Mitochondrial DNA from young cultures of race s of Podospora anserina was isolated. Its density in Cesium chloride density equilibrium gradients was 1.694 g/cc. Examination by the electron microscope revealed that ca 1% of this DNA consisted of circles, 31 micrometer in contour length; the remaining DNA was composed of linear molecules ranging in length from 2 to 33 micrometer. In DNA of similar density obtained from senescent cultures of the same race s, about 11% of the molecules consisted of a multimeric set of circles ranging in size from 0.9 to 15 micrometer, with most being in the 1.8 and 2.7 micrometer classes. The similarity of these DNA molecules with the mitochondrial DNA from rho(-) yeast mutants is discussed.     

The nature of DNA synthesis by isolated nuclei from cells of a rat tumour

Summary The DNA synthetic activity of nuclei isolated from a solid rat tumour was determined. The nuclei had DNA synthetic properties similar to nuclei from other sources but the time course of the reactions was different.