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PubMed1982

Persistent Varicella-Zoster virus infection in a human rhabdomyosarcoma cell line and recovery of a plaque variant.

Varicella-zoster virus (VZV) has been found to persistently infect the human rhabdomyosarcoma cell line A204. Infectious center assays and fluorescent antibody staining demonstrated continuous production of infectious VZV and viral antigen. The level of infection determined by fluorescent antibody staining was variable, and usually only a small percentage of the cells were capable of producing plaques in permissive fibroblasts. The extent of infection was similar in cell cultures passaged at split ratios of 1:2 or 1:10 and grown at 33 or 37 degrees C. VZV recovered from A204 cells several months after establishment of the persistent infection had markedly increased syncytia-forming activity as compared with the parental VZV grown in human diploid fibroblast cells and the three monkey kidney-derived cell lines Vero, CV-1, and MA104. The expression of this altered phenotype continued after serial passage of the cell-associated virus in human diploid fibroblast and Vero cells. Consequently, we designated the reisolated VZV as plaque variant A. The buoyant densities of VZV plaque variant A and VZV DNAs in CsCl gradients were indistinguishable.

Antigens, Viral↗

PubMed1967

Properties of Venezuelan equine encephalomyelitis virus grown in vivo.

One intracerebral passage of either the parent egg seed (PES) or an attenuated variant (10t) of the Trinidad strain of Venezuelan equine encephalomyelitis (VEE) virus in young adult mice produced progeny that were no longer differentiated unequivocally on the basis of plaque size. Plaques averaging about 2 mm in diameter, which was somewhat smaller than those formed by the PES virus and larger than those of the 10t strain, were formed by both strains. Seven serial passages of the PES virus in mouse brain failed to alter its virulence appreciably. In contrast, passage in mouse brain progressively changed the properties of the attenuated 10t strain. A substrain was isolated that possessed virulence similar to that of the PES virus and formed small plaques similar to those of the 10t strain. These findings showed a unique dissociation between the plaque size and virulence of the 10t strain. The new substrain differed from the PES virus and the 10t strain in its capacity for growth in mouse tissues after intraperitoneal inoculation. The substrain multiplied poorly in splenic tissue, which supports growth of the PES and 10t strains, but grew to high titers in the brain, which does not support appreciable growth of the 10t strain.

Animals↗

PubMed1991

Growth characteristics of human laryngeal carcinoma in athymic mice.

We report the growth characteristics of seven human laryngeal carcinomas transplanted into athymic mice. Six of the tumors were harvested from surgical pathology specimens and one was from a laryngeal carcinoma cell line. Tumor tissue measuring 2-3 cubic millimeters was implanted subcutaneously into the flank of the mice. Time intervals between implantation and active growth were recorded and tumor volume was calculated weekly. Mice were sacrificed at various time intervals. For three tumors, second and third passages of the tumor were performed. Successful tumor growth was achieved in 6/7 (85.7%) tumors. Primary implantation of tumor was performed in a total of 26 mice, 17 of which grew successfully. The average time interval between tumor implantation and measurable tumor growth for the five zenografts derived directly from patients was 36 days (range 25-40 days). The mean doubling time was 16 days (range 7-30 days). The overall take rate per animal for serial passage of tumor was 36 out of 43 (83.7%). The lag phase and the mean tumor doubling time decreased with each successive passage. At autopsy most tumors were confined to the subcutaneous structures by a well-formed capsule. Lymph node metastases and disseminated metastases were not seen. We conclude that the athymic mouse model is suitable to establish primary tumor growth of human laryngeal cancer.

Animals↗

PubMedDate not supplied

Murine virus contaminant of Trypanosoma cruzi experimental infection.

The possibility that some virus contaminants could be altering host response to Trypanosoma cruzi experimental infection was investigated. Data obtained showed that CBA/J mice infected with stocks of parasite maintained in mice (YIUEC) presented higher level of parasitemia and shorter survival times than those infected with a stock (YITC) which was also maintained in mice but had been previously passaged in cell culture. Mouse antibody production tests, performed with the filtered plasma of mice infected with YIUEC, indicated the presence of mouse hepatitis virus (MHV) while no virus was detected when testing the plasma of YITC infected mice. Filtered plasma of YIEUC infected mice was shown to contain a factor able to enhance the level of parasitemia and to reduce the mean survival time of mice challenged with 10(5) YITC. This factor, that could be serially passaged to naïve mice was shown to be a coronavirus by neutralization tests.

Animals↗

PubMed1975

Ibaraki disease and its relationship to bluetongue.

Ibaraki disease, an epizootic disease of cattle in Japan resembling bluetongue, is characterized by fever and lesions affecting the mucous membranes, the skin, the musculature and vascular system. Degeneration of striated muscular tissue is observed in the oesophagus, larynx, pharynx, tongue and the skeletal muscles. Oedema and haemorrhage are marked in the mouth, lips, abomasum, around the coronets, etc., and are occasionally followed by degeneration of the epithelium leading to erosions or ulcerations. Severe lesions affecting the oesophageal and laryngopharyngeal musculature cause difficulty in swallowing which in turn produces dehydration and emaciation, and occasionally the aspiration pneumonia, which constitute the major causes of death of affected animals. These clinical and pathological findings indicate the similarity of the disease to bluetongue in sheep and cattle. Ibaraki disease was first recognised in Japan in 1959 and 1960. Seasonally its occurrence is limited to late summer and autumn, and geographically to the central and western parts of Japan, roughly south of 37 degrees north latitude. It is absent from the higher altitudes. The seasonal and geographical incidence suggests the possibility of an arthropod vector; but direct evidence for such a vector is still lacking. Serological data suggest the presence of Ibaraki virus on Bali Island in Indonesia and in Taiwan. The disease can be transmitted serially in calves by the intravenous inoculation of blood obtained at the height of a febrile reaction. Ibaraki virus can be isolated in bovine cell cultures from both natural and experimentally produced cases of the disease. The virus multiplies and induces cytopathic effects in primary cultures of bovine, sheep and hamster lung origin, and L cells; but it does not grow in primary cultures of horse and swine kidney nor in HeLa cell cultures. The virus is readily passaged serially in 4 to 5-day-old eggs by yolk-sac inoculation and incubation at 33.5 degrees C. It multiplies in the brains of mice of any age after incracerebral inoculation but younger mice give a better viral growth and develop encephalitis. No evidence has been obtained that rabbits and guinea pigs are susceptible to Ibaraki virus...

Animals↗

PubMed1996

Isolation of the equine granulocytic ehrlichiosis agent, Ehrlichia equi, in tick cell culture.

The equine granulocytic ehrlichiosis agent, Ehrlichia equi, is closely related or identical to the human granulocytic ehrlichiosis (HGE) agent. Both are suspected of being transmitted by ticks. We have successfully isolated E. equi in a cell line, IDE8, derived from a putative vector, the tick Ixodes scapularis. Peripheral blood leukocytes from an experimentally infected horse were inoculated onto IDE8 monolayers. Cultures were incubated in a candle jar at 34 degrees C in tick cell culture medium with NaHCO3 and an organic buffer [3-(N-morpholino)-propanesulfonic acid] (MOPS). Within 2 weeks, infected cells were detected in Giemsa-stained culture samples, and the organisms subsequently spread to uninfected cells in the cultures. E. equi was passaged serially by transferring a portion of an infected culture to new cell layers every 2 to 3 weeks. The identity of the organisms was confirmed by PCR using oligonucleotide primers specific for E. equi and the HGE agent and by immunocytology. Homologous equine antibodies and human anti-HGE convalescent serum recognized E. equi grown in tick cell culture. Electron microscopy revealed electron-lucent and -dense ehrlichia-like forms developing within host cell endosomes. E. equi passaged twice in tick cell culture retained infectivity and pathogenicity for the equine host, as demonstrated by intravenous inoculation of a suspension of infected tick cells and subsequent reisolation from peripheral blood, in fulfillment of Koch's postulates. The horse developed severe clinical signs, i.e., fever, inappetence, thrombocytopenia, icterus, and limb edema, typical of granulocytic equine ehrlichiosis, within 1 week.

Animals↗

PubMed1981

Generation of transforming viruses in cultures of chicken fibroblasts infected with an avian leukosis virus.

During serial passages of an avian leukosis virus (the transformation-defective, src deletion mutant of Bratislava 77 avian sarcoma virus, designated tdB77) in chicken embryo fibroblasts, viruses which transformed chicken embryo fibroblasts in vitro emerged. Chicken embryo fibroblasts infected with these viruses (SK770 and Sk780) had a distinctive morphology, formed foci in monolayer cultures, and grew independent of anchorage in semisolid agar. Bone marrow cells were not transformed by these viruses. Another virus (SK790) with similar properties emerged during serial subcultures of chicken embryo fibroblasts after a single infection with tdB77. The 50S to RNAs isolated from these viruses contained a tdB77-sized genome (7.6 kilobases), 8.7- and 5.7-kilobase RNAs, and either a 4.1-kilobase RNA or a 4.6-kilobase RNA. These RNAs did not hybridize with cDNA's representing the src, erb, mac, and myb genes of avian acute transforming viruses. Cells transformed by any one of the Sk viruses (SK770, SK780, or SK790) synthesized two novel gag-related polyproteins having molecular weights of 110,000 (p110) and 125,000 (p125). We investigated the compositions of these proteins with monospecific antiviral protein sera. We found that p110 was a gag-pol fusion protein which contained antigenic determinants, leaving 49,000 daltons which was antigenically unrelated to the structural and replicative proteins of avian leukosis viruses. An analysis of the SK viral RNAs with specific DNA probes indicated that the 5.7-kilobase RNA contained gag sequences but lacked pol sequences and, therefore, probably encoded p125. The transforming ability, the deleted genome, and the induced polyproteins of the SK viruses were reminiscent of the properties of several replication-defective acute transforming viruses.

Animals↗

PubMed2001

Smooth muscle actin expression by human articular chondrocytes and their contraction of a collagen-glycosaminoglycan matrix in vitro.

Recent studies have demonstrated that human articular chondrocytes can express the gene for a contractile muscle actin, alpha-smooth muscle actin (SMA), in situ. One objective of this work was to evaluate the SMA-content of isolated human articular chondrocytes using Western blot analysis and to correlate the amount of SMA in the cells with passage number and the number of days in culture. A second objective was to determine if articular cartilage-derived cells expressing the gene for SMA in vitro also continue to express type II collagen. A final aim of the current study was to determine if SMA-containing cartilage-derived cells were capable of contracting a collagen glycosaminoglycan analog of extracellular matrix in vitro. Articular chondrocytes were isolated from 13 patients undergoing total joint arthroplasty. Cells were serially passaged through passage 7. Samples were allocated for Western blot analysis of SMA. Cells in monolayer culture were also stained immunohistochemically for SMA and type II collagen. Cells from passage 3 and 7 were seeded into a porous type I collagen-glycosaminoglycan matrix and the diameter of the scaffolds measured every other day for 21 days. Immunohistochemistry of the articular cartilage samples revealed SMA in the articular chondrocytes in situ with a greater percentage of cells staining positive in the superficial half (60 +/- 1.2%; mean +/- SEM) of the cartilage than in the basal half (28 +/- 1.3%). There was an increasing amount of SMA in the cells in monolayer culture with passage number and a meaningful correlation of the SMA content with the days in culture (linear regression analysis; R2 = 0.72). Double staining for SMA and type II collagen showed that type II collagen-expressing cells in monolayer could also express SMA. SMA-containing cells were found to contract the collagen glycosaminoglycan matrix, with the cells containing more SMA (passage 7 cells) displaying more matrix contraction than those with a lesser amount of SMA (passage 3 cells). The results indicate that control of the expression of SMA may be important when employing articular chondrocytes, expanded in monolayer culture, for implantation alone or in a cell-seeded matrix for cartilage repair procedures.

Actins↗

PubMed1999

Markedly different rates and resistance profiles exhibited by seven commonly used and newer beta-lactams on the selection of resistant variants of Enterobacter cloacae.

Seven beta-lactam antibiotics (cefepime, cefoperazone, ceftazidime, ceftriaxone, cefamandole, imipenem and meropenem) were tested for their potential to select resistance in standard and clinical strains of Enterobacter cloacae (n = 9). The strains were subcultured daily with the test antibiotics at doubling concentrations starting at 0.125 x MIC. Development of resistance throughout the passages was detected by a disc diffusion test. Ceftazidime, ceftriaxone and cefamandole selected resistance at a faster rate than cefoperazone, cefepime and meropenem. Imipenem did not select resistance in the nine strains tested and was the only antibiotic that eradicated all the strains during selection. The resistance patterns of strains selected by meropenem, cefepime and the other cephalosporins were markedly different, although cross-resistance to the early generation cephalosporins was common. The resistance phenotypes of most strains remained stable upon serial passages in antibiotic-free medium. The findings of this study highlight the importance of the choice of antibiotic for therapy not only on the basis of its antibacterial activity, but also on its potential to select resistance to itself and other antibiotics.

Anti-Bacterial Agents↗

PubMed1999

Evidence for clonal outgrowth of androgen-independent prostate cancer cells from androgen-dependent tumors through a two-step process.

Prostate cancers require androgen for growth but progress to an androgen-independent stage under the selective pressure of androgen ablation therapy. Here we describe a novel human prostate cancer xenograft (LAPC-9) propagated by serial passage in male severe combined immunodeficient mice that expresses prostate-specific antigen and wild-type androgen receptor. In response to castration, LAPC-9 cells undergo growth arrest and persist in a dormant, androgen-responsive state for at least 6 months. After prolonged periods of androgen deprivation, spontaneous androgen-independent outgrowths develop. Thus, prostate cancers progress to androgen independence through two distinct stages, initially escaping dependence on androgen for survival and, subsequently, for growth. Through the use of serial dilution and fluctuation analysis, we provide evidence that the latter stage of androgen independence results from clonal expansion of androgen-independent cells that are present at a frequency of about 1 per 10(5)-10(6) androgen-dependent cells. We conclude that prostate cancers contain heterogeneous mixtures of cells that vary in their dependence on androgen for growth and survival and that treatment with antiandrogen therapy provides selective pressure and alters the relative frequency of these cells, thereby leading to outgrowths of androgen-independent cancers.

Androgens↗

PubMed1987

Growth and continuous passage of COMMA-D mouse mammary epithelial cells in hormonally defined serum-free medium.

Growth of the mouse mammary epithelial cell line designated COMMA-D has been studied in serum-free medium (SFM) formulated with Ham's F12 and Dulbecco's modified Eagle's medium (1/1) containing 15 mM 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid, 2 mM glutamine, gentamicin (50 micrograms/ml; basal medium) and supplemented with insulin (10 micrograms/ml), transferrin (10 micrograms/ml), selenous acid (10 ng/ml), epidermal growth factor (20 ng/ml; EGF), 10 nM 3,5,3'-triiodothyronine, 50 microM ethanolamine, 1.0 nM 17 beta-estradiol, 65 microM glutathione, and ovalbumin (100 micrograms/ml). COMMA-D cells were able to undergo serial passage and continued to exhibit dome formation after 20 passages in SFM. Cells seeded at low density in SFM underwent four population doublings at low passage number in 1 week compared to six doublings for cells grown in medium containing insulin, transferrin, selenium, EGF, and 1% fetal bovine serum. After many passages in SFM, the growth rates of cells were similar to those in serum-supplemented medium used for stock culture. Deletion of insulin or EGF from SFM resulted in cell growth similar to that of cells seeded in basal medium alone. When cells were seeded in basal medium without added supplements, addition of insulin or EGF resulted in 29 and 22%, respectively, of the number of cells grown in SFM for 5 days. However, when insulin and EGF were combined in basal medium, the cell number at 5 days was 83% of that in SFM. When insulin was deleted from SFM, COMMA-D cells became responsive to insulin-like growth factors I and II. The growth-promoting characteristics of EGF and transforming growth factor alpha were compared in SFM and were not distinguishable, showing identical dose-response curves. When incorporation of [3H]thymidine was used as an assay of cell growth, saturating levels of basic fibroblast growth factor (20 ng/ml) showed a stimulation 1.35 times greater than EGF (20 ng/ml). When EGF and fibroblast growth factor were combined, the stimulation was 1.75 times greater than EGF alone suggesting that COMMA-D cells are responsive to multiple classes of growth factors. COMMA-D cells seeded in basal medium supplemented with insulin, transferrin, and selenous acid have been used to detect mitogenic activity present in extracts of hypothalamus, uterus, and pituitary. The results show that COMMA-D cells can be grown long term in a hormonally defined serum-free medium and that maximal mitogenic effects were seen only with the addition of two or more growth factors.

Animals↗

PubMed1975

Marek's disease virus (Kekava strain) replication in chickens, chick embryos and cell cultures.

In the course of 12 passages of Marek's disease virus (MDV) strain Kekava (MDV-Kekava) in chickens, the morbidity varied greatly (from 23 to 50 percent). MDV-Kekava produced plaques in cultures of chick embryo kidney and adult chicken kidney cells and chick embryo fibroblasts (CEF). The virus adaptation to the cultures was very slow. MDV-Kekava induced the formation of pocks on the chorioallantoic membranes (CAM) of chick embryos but the proportion of embryos with CAM lesions did not exceed 24 percent. Serial passaging of the virus in chick embryos beyond the 5th passage was unsuccessful. The results of virus isolation in chickens, cell cultures and chick embryos indicate the possibility of a long-term latent virus carrier state in chickens without development of tumours.

Animals↗

PubMed1976

The serial cultivation of suspended BHK-21/13 cells in serum-free Waymouth medium.

A simple medium system was developed to obtain growth of BHK-21 cells in shaker cultures in the absence of serum. These cells have now undergone over 80 serial passages in serum-free Waymouth medium and have been recovered from the frozen state after storage for over 1 month in medium containing 10% dimethyl sulfoxide (DMSO) and 1% bovine serum albumin (BSA). Various amounts of exogenous lipid in the form of sodium oleate were added to cultures of cells growing in serum-free Waymouth medium. Concentrations of 10-50 mug of sodium oleate/ml had no detrimental effects on the cells as measured by trypan blue uptake. Furthermore, the cells were serially passed ten times in the presence of 10 mug sodium oleate/ml. Depletion of calf serum from the growth medium and addition of known quantities of lipids to the system provides a means of revealing subtle changes in lipid synthesis and lipid turnover during cellular growth.

Cell Line↗

PubMed1998

Experimental induction of alopecia areata-like hair loss in C3H/HeJ mice using full-thickness skin grafts.

Alopecia areata (AA)-like hair loss in C3H/HeJ mice provides an excellent model for human AA disease research. The potential to induce mouse AA in normal haired C3H/HeJ mice at an early age or serially passage the AA phenotype was investigated by exchange of full-thickness skin grafts. Skin grafts from normal male and female C3H/HeJ, or severe combined immunodeficient C3H/SmnC Prkdc(scid)/J, mice onto AA-affected C3H/HeJ mice became inflamed and lost hair (28 of 28). Successful grafts from AA-affected C3H/HeJ mice induced hair loss in histocompatible C3H/OuJ mice (four of 13) and normal C3H/HeJ mice dependent on age (four of 17 at <31 d and 15 of 15 at >70 d). The AA phenotype was serially transmitted from induced AA mice to normal C3H/HeJ mice (nine of nine). Grafts from AA-affected C3H/HeJ mice onto C3H/SmnC Prkd(scid)/J mice resulted in depigmented hair fiber regrowth and perifollicular neutrophil and eosinophil infiltrates but no hair loss (15 of 15). Sham grafting did not induce AA (none of 10). The finding that AA can be serially transferred from AA-affected C3H/HeJ mice to normal littermates and C3H/ OuJ mice, indicates that an immune response against hair follicles can be induced with suitable stimuli. Conversely, skin grafts from normal C3H/HeJ, or C3H/SmnC Prkd(scid)/J, mice rapidly lose hair due to lymphocyte, but not neutrophil and eosinophil, mediated inflammation. This AA induction method reproducibly provides large numbers of AA-affected mice to study the pathogenesis and treatment of human AA.

Alopecia Areata↗

PubMed2004

Expression of psoriasis-associated fatty acid-binding protein in senescent human dermal microvascular endothelial cells.

Aging is associated with the progressive pathophysiologic modification of endothelial cells. In vitro endothelial cell senescence is accompanied by proliferative activity failure and by perturbations in gene and protein expressions. Moreover, this cellular senescence in culture has been proposed to reflect processes that occur in aging organisms. In order to observe the changing patterns of protein expression in senescent human dermal microvascular endothelial cells (HDMECs), proteins obtained from both early- and late-passaged HDMECs were separated by two-dimensional electrophoresis, visualized by silver staining, and quantified by image processing. Proteins of interest were extracted by in-gel digestion with trypsin and quantified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS), by searching the National Center for Biotechnology Information protein-sequence database. More than 2000 spots were detected by 2D electrophoresis within a linear pH range of 3-10. Twenty-two major differentially expressed spots were observed in serially passaged HDMECs and identified with high confidence by MALDI-TOF-MS. One of these spots was found to be a 14-15 kDa psoriasis-associated fatty acid-binding protein (PA-FABP) with high affinity for long-chain fatty acids. The expression of PA-FABP was confirmed to be elevated in senescent HDMECs (passage 20) by fluorescence-activated cell sorting (FACS), confocal laser microscopy, and by immunohistochemistry in aged human skin tissue. Our results suggest that the overexpression of FABP in cultured senescent HDMECs is closely related to skin aging.

Antibody Specificity↗

PubMed1975

In vivo behavior of revertants from Stahylococcus aureus L-phase variants compared with the parent strain.

The in vivo behavior of revertants of Staphylococcus aureus L-phase variants were studied in mice. The Giorgio strain of S. aureus was induced into L-phase variants by methicillin. After 13 serial passages in methicillin agar and six passages in antibiotic-free agar, reversion occurred. Six revertants, all phage untypable, coagulase negative, and mannitol negative, were inoculated intravenously into Swiss albino mice in comparison with the parent strain. Inoculation of 10(8) colony-forming units resulted in 38% mortality at 28 days for the Giorgio group and 0 to 2% for the revertants. The geometric mean titer of staphylococci in the kidneys was 3.5 x 10(5) for the Giorgio group and 0.5 x 10(0) to 1.0 x 10(2) for the revertants. Serial colony counts of blood, lungs, spleen, liver, and kidneys in mice inoculated with 10(7) colony-forming units showed faster clearance and failure of the revertant to colonize the kidneys. It is concluded that the in vivo behavior of the L-phase revertants was vastly different from that of the parent Giorgio strain.

Animals↗

PubMed1996

Construction of a type 1 human immunodeficiency virus that maintains a primer binding site complementary to tRNA(His).

The initiation of human immunodeficiency virus type 1 reverse transcription occurs by extension of a tRNA(Lys3) primer bound near the 5' end of the viral RNA genome which is designated the primer binding site (PBS). Sequences within the viral genome upstream of the PBS which are complementary to the anticodon loop (USUU) and the T psi C loop and arm (AGGGTm psi) of tRNA(Lys3) are postulated to play a role in maintaining the selective use of tRNA(Lys3) in reverse transcription. To investigate this possibility, proviral genomes which contain a PBS complementary to the 3'-terminal 18 nucleotides of tRNA(His) [pHXB2(His)] as well as sequences upstream of this PBS which are complementary to either the anticodon loop [CCACAA; pHXB2(His-AC)] or T psi C loop [GACCGAGG; pHXB2(His-T psi C)] of tRNA(His) were constructed. Infectious virus was recovered upon transfection into COS-1 cells of pHXB2(His), pHXB2(His-AC), or pHXB2(His-T psi C). The appearance of infectious virus after cocultivation with SupT1 cells was delayed for the proviruses containing a PBS complementary to tRNA(His) compared with that obtained by transfection of the wild-type provirus [pHXB2(WT)]. However, by several passages in SupT1 cells, the mutant viruses demonstrated replication kinetics similar to those of the wild-type virus. A DNA sequence analysis of the PBS region from integrated proviruses revealed that by day 15 of culture, the PBS of viruses derived from pHXB2(His) and pHXB2(His-T psi C) reverted back to the wild-type PBS complementary to tRNA(Lys3). In contrast, viruses derived from pHXB2(His-AC) maintained a PBS complementary to tRNA(His) for over 4 months in culture encompassing 12 serial passages. This study, then, is the first report of a stable human immunodeficiency virus type 1 which utilizes an alternative tRNA primer and suggests that interactions between the primer tRNA anticodon loop and viral sequences upstream of the PBS contribute to the specificity of the tRNA primer used in reverse transcription.

Animals↗

PubMedDate not supplied

[Replicative properties of HIV-1 strains, resistant to 3'-azido-2'3'-dideoxythymidine].

The replicative activity of two strains of human immunodeficiency virus type I, 1974 and 1978, resistant to 3'-azido-2',3'-dideoxythymidine (AZT), was investigated. Assessment of AZT sensitivity by the standard method showed the 50% effective dose (ED50) for strain 1974 to be 0.1 mumole and for strain 1978, 1 mumole. Both strains had the slow/low. 3 phenotype. The level of virus-specific antigens in these strains was 3-7 times lower than in AZT-sensitive strain IIIB. In order to study the effects of AZT on virus replication, the strains were passaged in Jurkat-tat cells in the presence of 3.5 mumole of AZT. Virus activity was assessed from the level of intracellular (S-12) and extracellular (P-100) antigens, cell viability, and syncytium formation. Cell viability increased by 10%, syncytium formation dropped 5 times, and antigen accumulation increased 1.5-2 times in the culture with strain 1974. For strain 1978 these parameters were as follows: 17% decrease of cell viability, increase of syncytium formation by 1.7 times, and no changes in the level of viral antigen. After four serial passages in the presence of AZT ED50 for strain 1974 was 0.5 mumole and at least 1 mumole for strain 1978.

Cell Survival↗