Gametogony of Sarcocystis sp. in cell culture

Sexual stages and cystlike bodies of Sarcocystis sp., a protozoan parasite found in muscles of reptiles, birds, and mammals, including man, developed in cell culture. Motile organisms, obtained from leg muscles of wild grackles, were inoculated into cell line cultures of embryonic bovine kidney. Mature micro-and macrogametes and the cystlike forms were found 30 and 42 hours after inoculation respectively. These observations indicate that the parasite is probably a coccidium.     

Gaucher's disease: a genetic disease detected in skin fibroblast cultures

Skin fibroblasts from three adult patients with chronic noncerebral Gaucher's disease, three children of one of the patients, three parents, and six normal individuals were grown in cell culture. Giant fibroblasts containing metachromatic material were seen in all cultures derived from affected individuals and heterozygous carriers but not in those derived from normal individuals.     

Trypanosoma cruzi infection inhibited by peptides modeled from a fibronectin cell attachment domain

The mechanism by which Trypanosoma cruzi, the protozoan parasite that causes Chagas' disease, becomes attached to mammalian cells is not well understood. Fibronectin is thought to participate in the attachment, and in this study the region of fibronectin that interacts with the surface receptors of T. cruzi trypomastigotes was investigated by testing the binding of the amino acid sequence Arg-Gly-Asp-Ser, corresponding to the cell attachment site of fibronectin to T. cruzi trypomastigotes. Peptides with the sequence Arg-Gly-Asp-Ser, but not Arg-Phe-Asp-Ser, Arg-Phe-Asp-Ser-Ala-Ala-Arg-Phe-Asp, Ser-Lys-Pro, Glu-Ser-Gly, or Ala-Lys-Thr-Lys-Pro, bound to the parasite surface and inhibited cell invasion by the pathogen. Monoclonal antibodies to the cell attachment domain of fibronectin also inhibited cell infection by the parasite. The immunization of BALB/c mice with tetanus toxoid-conjugated peptide induced a significant protection against T. cruzi. The data support the notion that the sequence Arg-Gly-Asp-Ser of cell surface fibronectin acts as a recognition site for attachment of the parasites.     

Mammalian cells in culture frequently release type C viruses

Cell cultures commonly used in animal cell research, both cell strains and continuous cell lines from various mammalian species, spontaneously produce type C RNA viruses.     

Two mechanisms for the extinction of gene expression in hybrid cells

When two different mammalian cell types are fused to generate a stable hybrid cell line, genes that are active in only one of the parents are frequently shut off, a phenomenon called extinction. In this study two distinct, complementary mechanisms for such extinction of growth hormone gene expression were identified. In hybrids formed by fusing fibroblasts to pituitary cells, pituitary-specific proteins that bind to the growth hormone promoter were absent. In addition, a negative regulatory element located near the rat growth hormone promoter was specifically activated.     

Migration of Plasmodium sporozoites through cells before infection

Intracellular bacteria and parasites typically invade host cells through the formation of an internalization vacuole around the invading pathogen. Plasmodium sporozoites, the infective stage of the malaria parasite transmitted by mosquitoes, have an alternative mechanism to enter cells. We observed breaching of the plasma membrane of the host cell followed by rapid repair. This mode of entry did not result in the formation of a vacuole around the sporozoite, and was followed by exit of the parasite from the host cell. Sporozoites traversed the cytosol of several cells before invading a hepatocyte by formation of a parasitophorous vacuole, in which they developed into the next infective stage. Sporozoite migration through several cells in the mammalian host appears to be essential for the completion of the life cycle.     

Cell aggregation: role of acid mucopolysaccharides

Factors that induce cell aggregation are released by several types of chick embryo and mammalian cell cultures. These aggregation factors are also present in some serums. The factors in each of the preparations tested were inactivated by treatment with bovine testicular hyaluronidase. Conversely, hyaluronic acid promoted aggregation of only those cells that were aggregated by media containing the factors. These factors appear to be acid mucopolysaccharides, with hyaluronic acid being a major component.     

Propagation of poliovirus,measles, and vaccinia in guinea pig spleen cell strains

A new mammalian cell strain was established from guinea pig spleen tissues and has shown interesting virus sensitivity. Several viruses have been successfully propagated in the cells. For one of these, poliovirus, it represents one of the two successful attempts at propagating the virus in nonprimate or nonhuman cultures.     

Expression of the familial hypercholesterolemia gene in heterozygotes: mechanism for a dominant disorder in man

Studies in ctltured fibroblasts indicate that the primary genetic abnormality in familial hypercholesterolemia involves a deficiency in a cell surface receptor for low density lipoproteins (LDL). In normal cells, binding of LDL to this receptor regulates cholesterol metabolism by suppressing cholesterol synthesis and increasing LDL degradation. In cells from heterozygotes, a 60 percent reduction in LDL receptors leads to a concentration-dependent defect in regulation, so that attainment of equal rates of cholesterol synthesis and LDL degradation in normal and heterozygous cells requires a two- to threefold higher concentration of LDL in the heterozygote. The identification of this genetic regulatory defect in fibroblasts of heterozygotes makes available an in vitro system for studying the effects of a dominant mutation on gene expression in mammalian cells.     

Heterogeneity of normal human diploid fibroblasts: isolation and characterization of one phenotype

Cultures of human diploid fibroblasts contain cells that respond to exposure to the first component of complement (C1) by initiating DNA synthesis and growth. The plasma membranes of these cells have specific binding sites for the C1q subcomponent of C1. A fluorescence-activated cell sorter was used to isolate a subset of cells with a high affinity for C1q, and the growth and synthesis activities of these high-affinity cells were studied after numerous replications in vitro. These cells synthesize DNA and grow faster than the parent cultures and low-affinity cells, and they produce two to three times as much protein. About 40 percent of their total protein synthesis activity is directed to collagen production, unusually high proportions of collagen types III and V being produced. These properties and the high affinity of the cells for C1q are retained for at least six cell transfers. This phenotype has the properties expected of fibroblasts in healing wounds and inflamed tissues.     

Hurler's syndrome: demonstration of an inherited disorder of connective tissue in cell culture

Skin fibroblasts from three patients with Hurler's syndrome were grown in tissue culture and shown to contain metachromatic granules when stained for mucopolysaccharides with toluidine blue O. Similar inclusions were observed in cultures of fibroblasts from other members of the families, who appeared to be clinically normal but who were, judged from studies of pedigree, heterozygous or hemizygous for the abnormal gene.     

Persistence without pathology in phosphoglycan-deficient Leishmania major

Leishmania infections involve an acute phase of replication within macrophages, typically associated with pathology. After recovery parasites persist for long periods, which can lead to severe disease upon reactivation. Unlike the role of host factors, parasite factors affecting persistence are poorly understood. Leishmania major lacking phosphoglycans (lpg2-) were unable to survive in sand flies and macrophages, but retained the ability to persist indefinitely in the mammalian host without inducing disease. The L. major lpg2- thus provides a platform for probing parasite factors implicated in persistence and its role in disease and immunity.     

New method for manipulation,maintenance, and cloning of single mammalian cells in vitro

Individual mammalian cells can be isolated with the aid of easily handled glass beads to which the cells have become attached. Procedures for the preparation of cell cultures on beads, and for the recognition and manipulation of beads carrying single monocytes, HeLa cells, and Detroit-98 cells are described. Data obtained with Brucella infected monocytes, illustrating the efficiency of the method, are presented.     

慢病毒载体介导成纤维细胞高效稳定表达绿色荧光蛋白

 试验尝试建立稳定表达外源基因的人成纤维细胞系。取成年男性包皮皮肤的皮下组织,分离培养人皮肤成纤维细胞,分别采用脂质体和慢病毒载体介导转染人皮肤成纤维细胞表达绿色荧光蛋白。结果显示,来自成年人包皮皮下组织的成纤维细胞呈梭形,具有快速的增殖和稳定的生长性能。脂质体介导转染的人皮肤成纤维细胞绿色荧光蛋白表达不稳定,阳性细胞表达率低,转染后的人成纤维细胞变得更为细长,细胞生长速度降低。慢病毒载体介导人皮肤成纤维细胞高效稳定表达绿色荧光蛋白,转染后细胞生长性能和形态没有变化,经过多次传代和冻存复苏对人皮肤成纤维细胞绿色荧光蛋白的表达没有影响,通过流式细胞仪对慢病毒介导表达绿色荧光蛋白的人皮肤成纤维细胞检测显示,绿色荧光蛋白表达阳性率为99.85%,并且表达绿色荧光蛋白的人皮肤成纤维细胞细胞均一程度为76.05%。试验证实了慢病毒载体能够高效稳定介导人皮肤成纤维细胞表达绿色荧光蛋白。     

Histidase activity in cultivated human amniotic fluid cells

Epithelial and fibroblast cells were obtained from cultures of amniotic fluid cells. Epithelial cells demonstrated high activities of histidase. In contrast, histidase activity was not detected in fibroblasts derived from the same original culture. This observation indicates that cultures of amniotic fluid cells consist of cells with different biochemical properties as well as morphological characteristics.     

Phenotypic expression of transformation: induction in cell culture by a phorbol ester

A biologically potent, tumor-promoting phorbol ester from Croton tiglium L. induces the appearance of transformed clones in a population of contact-inhibited fibroblasts of the mouse cell line 3T3. Mixed populations of cells with 10,000 3T3 cells and 100 virus-transformed cells were exposed to the phorbol ester and, in comparison with untreated cells, showed a marked increase in the numbers of transformed clones that grew. Exposure of 3T3 cultures to the carcinogen 7,12-dimethylbenz(a)anthracene alone or prior to exposure to the phorbol ester did not cause any increase in the number of transformed clones.     

拟胚体对成体细胞参与胚胎嵌合的影响

试验尝试构建成体细胞同小鼠早期胚胎的嵌合共生胚胎.取成年人皮肤组织的成纤维细胞,慢病毒转染皮肤成纤维细胞标记EGFP荧光蛋白,同小鼠早期8-细胞胚胎进行嵌合.结果显示慢病毒高效标记人皮肤成纤维细胞表达EGFP荧光蛋白,通过皮肤成纤维细胞与小鼠胚胎干细胞形成的拟胚体环境作用后,皮肤成纤维细胞成功与小鼠胚胎形成嵌合胚,嵌合囊胚形成率为38.08%,表达EGFP荧光蛋白的皮肤成纤维细胞能够嵌合到小鼠胚胎的不同部位.嵌合胚在胚胎干细胞分离培养环境下进行培养,嵌合到小鼠内细胞团的皮肤成纤维细胞参与小鼠内细胞团的组成,参与小鼠内细胞团组成的胚胎占嵌合胚比率为1.74%.小鼠胚胎干细胞拟胚体环境作用可以成功介导人皮肤成纤维细胞同小鼠早期胚胎形成嵌合共生体系.     

An analog of myristic acid with selective toxicity for African trypanosomes.

Trypanosoma brucei, the protozoan parasite responsible for African sleeping sickness, evades the host immune response through the process of antigenic variation. The variant antigen, known as the variant surface glycoprotein (VSG), is anchored to the cell surface by a glycosyl phosphatidylinositol (GPI) structure that contains myristate (n-tetradecanoate) as its only fatty acid component. The utilization of heteroatom-containing analogs of myristate was studied both in a cell-free system and in vivo. Results indicated that the specificity of fatty acid incorporation depends on chain length rather than on hydrophobicity. One analog, 10-(propoxy)decanoic acid, was highly toxic to trypanosomes in culture although it is nontoxic to mammalian cells.     

Interferon inducers in vitro: difference in sensitivity to inhbitiros of RNA and protein synthesis

Interferon can be induced by diverse agents in a variety of mammalian cell cultures through apparently two mechanisms. One results in an early (2 to 10 hours) appearance of interferon and is relatively resistant to inhibition by actinomycin, puromycin, or fluorophenylalanine. A second mechanism results in a late (18 to 24 hours) appearance of interferon and is more sensitive to inhibition by these inhibitors. The molecular basis for each mechanism is unclear. Since each interferon inducer may have multiple effects on the cell, the differences observed may not necessarily reflect a fundamental difference in the mechanism of interferon stimulation.