Detection of caprine arthritis-encephalitis- and maedi-visna viruses using the polymerase chain reaction

Summary The polymerase chain reaction (PCR) was used to demonstrate proviral DNA of lentiviruses of small ruminants in cultured cells. Primers for the Taq polymerase were selected in the GAG gene of Icelandic maedi-visna virus and POL gene of caprine arthritis-encephalitis (CAE) virus. Using PCR, proviral DNA of CAE virus was detected at 1 day post infection, 4 days beforeviral protein could be demonstrated using a sensitive immunoblotting protocol and 6 days before the appearance of syncytia. Primers derived from the published sequence of CAE virus successfully primed for the synthesis of homologous virus and Icelandic maedi-visna viruss but not for maedi-visna virus isolated in The Netherlands. In contrast, primers derived from the GAG region of Icelandic maedi-visna virus allowed the amplification of DNA of homologous virus, maedi-visna virus isolated in The Netherlands as well as CAE virus.     

Feulgen-DNA amounts and karyotype lengths of three planarian species of the genusDugesia

Summary Genome sizes of the planariansD. lugubris (2n=8),D. polychroa (2n=8) andD. benazzii (2n=16) were evaluated on metaphase plates by measuring both the Feulgen-DNA contents and the karyotype lengths. In the three species, genome sizes are significantly different; this finding rules out the possibility of a karyotype evolution through simple chromosome rearrangements betweenD. lugubris andD. polychroa. A different Feulgen-DNA content per unit length of karyotype in the three species studied was also found, which suggests that DNA could be differently packed along metaphase chromosomes.     

Epifluorescence microscopy ofAnabaena sp.: Nucleoid configurations and evidence for inclusion-associated DNA

Vital staining of the nucleoid inAnabaena sp. PCC7118 was performed using the double stranded DNA-specific fluorochrome DAPI. In the unicellular mutant, the central epifluorescent zone had a dense skein configuration, while in the filamentous parent strain protrusions, lobes, and distinct isolated elements of the nucleoid were visible. Both variants contrasted with the mainly peripheral, partitioned structure typical of plastids and prochlorophytes. Blue-white emittance from the DNA-DAPI complex was maximum in dividing cells, suggesting that DNA configuration is linked to the cell cycle events. In stationary cultures, epifluorescent cell inclusions were conspicuous: based on this observation, we argue that DNA is associated with carboxysomes in situ.     

杂鳞库蚊复组28S rRNA基因5′端序列研究

对我国三带喙库蚊、环带库蚊和伪杂鳞库蚊共计79只蚊虫的28SrRNA基因5′端序列进行测定,获得的序列长度为301bp,3蚊种的该段序列相同。为设计杂鳞库蚊复组近缘种的PCR鉴别方法打下基础。     

人食管癌癌旁粘膜培养上皮生长特性的研究

人食管癌癌旁组织12例和正常食管粘膜15例组织块在盖玻上用199培养基培养,并掺入放射性氘胸腺嘧啶核苷.用光镜、显微分光光度计,放射自显影,扫描电镜检查移植块生长上皮层的生长速度,细胞核 DNA 含量,DNA 合成和形态学改变。结果是前三周,癌旁粘膜组织块培养上皮的生长率,细胞 DNA 含量和 DNA 合成高于食管正常粘膜.第四周以后两组上皮出现生长率降低,分化增加现象,在此培养中未能见到癌旁粘膜上皮转化为恶性细胞.本实验正常粘膜的培养,其生长的上皮细胞可以作为食管癌致癌因素研究的模型.     

Development of in vitro toxicity tests with cultures of freshly isolated rat hepatocytes

Summary Freshly isolated and cultured hepatocytes were analyzed by two-parameter flow cytometry. The combined analysis of DNA and cellular protein content allowed the contribution of ploidy classes and of subpopulations within a ploidy class to be defined. Analysis of hepatocytes during exposure to dimethylsulfoxide (DMSO), phenobarbital (PB), low oxygen tension (5% O₂) or fetal calf serum (FCS), provided insight into the dynamic response of individual ploidy classes as a function of culture time. By analogy with the age-dependent ploidy shifts in vivo, hepatocyte-cultures shift towards adult animals during exposure to DMSO and towards young animals when cultured at low pO₂ (4% O₂). FCS and phenobarbital disturb this constitutive ploidy balance. FCS increased the 2 N cell population, where stem cells probably respond to the proliferative stimuli provided by growth factors in the serum. Phenobarbital affects the liver-specific 4 N hepatocytes, which agrees with effects seen in liver after exposure in vivo. It is suggested that drug-induced pathological alterations in ploidy in hepatocyte cultures could serve as indicators of compounds, such as liver tumor promoters, which interfere with cell differentiation in liver. The heterotypic cell-cell interaction of freshly isolated hepatocytes with isolated, in vitro cultured, rat liver epithelial cells in co-cultures proved to be a valuable concept in toxicity testing: aldrin epoxidase, an enzyme system involved in xenobiotic metabolism, was stabilized for more than two weeks. After exposure to the three chemicals, 2-acetylaminofluoren, procarbazine and cyproterone-acetate, a preferential toxicity for each compound and cell population was established. Thus heterotypic cell cultures can considerably increase the amount of information available from in vitro studies.The final concept, combining monitoring of cellular DNA (ploidy) and protein content in hepatocyte cultures during and after exposure to a given test compound at tissue oxygen tension with the heterotypic cell-cell interaction, would create a more in vivo-like culture system. This would enhance the predictability of hepatocyte cultures and contribute to a more widespread use of the test system and as a result help to reduce the number of whole-animal tests.