RESUMO
A robust inflammatory response involving tumor necrosis factor-alpha (TNF-alpha) is induced during cisplatin nephrotoxicity. Using chimeric models, Reeves and colleagues now demonstrate that resident kidney cells, rather than infiltrating immune cells, are the major producers of TNF-alpha. Blockade of TNF-alpha attenuates inflammation and associated kidney injury.
Assuntos
Antineoplásicos/efeitos adversos , Cisplatino/efeitos adversos , Insuficiência Renal/induzido quimicamente , Fator de Necrose Tumoral alfa/metabolismo , Animais , Antineoplásicos/farmacologia , Quimera , Cisplatino/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Rim/efeitos dos fármacos , Rim/metabolismo , Rim/patologia , Camundongos , Camundongos Knockout , Modelos Biológicos , Receptores do Fator de Necrose Tumoral/genética , Receptores do Fator de Necrose Tumoral/metabolismo , Insuficiência Renal/metabolismo , Insuficiência Renal/prevenção & controle , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/genéticaRESUMO
PURPOSE: Human cytomegalovirus retinitis, the most common ophthalmic infection of AIDS patients, has been modeled in BALB/c mice infected with murine cytomegalovirus by the supraciliary route. A series of depletion and adoptive transfer studies was performed to determine whether adoptive transfer of T cells protects mice from retinitis caused by murine cytomegalovirus infection after supraciliary inoculation and to determine which subset of T cells is responsible for protection. METHODS: BALB/c mice were thymectomized and T cell-depleted by injection of monoclonal antibodies to CD4, CD8, or both. Murine cytomegalovirus (9 x 10(2) plaque forming units [pfu]) was injected into the supraciliary space. Experimental animals received murine cytomegalovirus-specific T cells or subsets of T cells 2 hours before virus injection, whereas control animals received herpes simplex virus type 1-specific T cells by tail vein injection. Eight days after virus injection, retinal pathology was scored by histopathologic examination of hematoxylin and eosin-stained ocular sections. RESULTS: CD8+ T cell depletion was sufficient for development of retinitis after supraciliary injection of murine cytomegalovirus. Adoptive transfer of murine cytomegalovirus-specific T cells, but not herpes simplex virus type 1-specific T cells, provided protection from retinitis. Additionally, separation of the murine cytomegalovirus-specific T cells into CD8+ and CD4+ subsets before adoptive transfer showed that the CD8+ fraction of the adoptive T cells was responsible for protection. CONCLUSIONS: These results suggest that adoptive transfer of cytomegalovirus-specific T cells or T cell subsets might be used to treat or prevent cytomegalovirus retinitis in immunosuppressed human patients.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Infecções Oculares Virais/prevenção & controle , Infecções por Herpesviridae/prevenção & controle , Imunoterapia Adotiva , Muromegalovirus/fisiologia , Retinite/prevenção & controle , Animais , Infecções Oculares Virais/patologia , Infecções Oculares Virais/virologia , Feminino , Citometria de Fluxo , Infecções por Herpesviridae/patologia , Infecções por Herpesviridae/virologia , Linfonodos/citologia , Depleção Linfocítica , Camundongos , Camundongos Endogâmicos BALB C , Retinite/patologia , Retinite/virologia , TimectomiaRESUMO
PURPOSE: The purpose of this study was to determine whether adoptive transfer of murine cytomegalovirus (MCMV)-immune lymph node cells prevents retinitis in immunosuppressed mice. METHODS: Adult BALB/c mice were thymectomized and T-cell depleted using rat monoclonal antibodies specific for mouse CD4+ and CD8+ T-cells. The level of rat immunoglobulin G in the treated mice was monitored by enzyme-linked immunosorbent assay. Immune cells were labeled with PKH26-GH immediately before adoptive transfer, and flow cytometry was used to determine the percentage of adoptively transferred T-cells (PKH+, fluorescein isothiocyanate [FITC+]) in the spleens of the recipient mice 3 days after transfer. The ability of adoptively transferred cells to protect from retinitis was studied in T-cell-depleted mice injected with MCMV through the supraciliary route. Mice received 4 x 10(7) in vitro-restimulated MCMV-immune cells, 4 x 10(7) freshly isolated MCMV-immune cells, 4 x 10(7) freshly isolated ovalbumin-immune cells, or no cells (control group). RESULTS: The best time to balance depletion of endogenous T-cells with persistence of transferred cells was 3 weeks after T-cell depletion. Both restimulated and freshly isolated MCMV-immune cells conferred protection from retinitis. Freshly isolated ovalbumin-immune lymph node cells did not prevent retinitis, indicating that protection was virus-specific and merely was not because of transfer of antigen-activated lymph node cells. CONCLUSIONS: Adoptive immunotherapy has been used to prevent cytomegalovirus (CMV) infection in patients who have undergone transplantation, and, by extrapolation, the results of these studies suggest that adoptive immunotherapy with human CMV-specific immune cells might be used to prevent or ameliorate CMV retinitis in immunocompromised patients.
Assuntos
Infecções Oculares Virais/prevenção & controle , Infecções por Herpesviridae/prevenção & controle , Imunoterapia Adotiva , Linfonodos/imunologia , Muromegalovirus/fisiologia , Retinite/prevenção & controle , Linfócitos T/imunologia , Animais , Anticorpos Monoclonais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Ensaio de Imunoadsorção Enzimática , Infecções Oculares Virais/imunologia , Infecções Oculares Virais/patologia , Feminino , Citometria de Fluxo , Infecções por Herpesviridae/imunologia , Infecções por Herpesviridae/patologia , Tolerância Imunológica , Linfonodos/citologia , Linfonodos/virologia , Depleção Linfocítica , Camundongos , Camundongos Endogâmicos BALB C , Ovalbumina/imunologia , Retinite/imunologia , Retinite/patologia , Retinite/virologia , Fatores de TempoRESUMO
In these studies, DNA PCR was used to identify sites of murine cytomegalovirus (MCMV) latency after inoculation of virus into the supraciliary space of the eye. Reverse transcription (RT) PCR for an immediate early gene and a late gene was used to identify putative sites of virus reactivation after methylprednisolone (steroid)-induced immunosuppression. Ten weeks after inoculation of 5 x 10(2) PFU of MCMV, BALB/c mice were immunosuppressed by intramuscular injection of steroid. Control mice were infected but not immunosuppressed. Two weeks after initiation of immunosuppression, mice were sacrificed. DNA and RNA extracted from homogenized tissues were amplified for immediate early gene 1 (IE1) and late gene, glycoprotein H (gH), DNA and mRNA by PCR and RT-PCR, respectively. Replicating virus was detected in homogenized ocular and non-ocular tissues by plaque assay. In the latently infected PBS-treated control group, viral DNA was detected in the inoculated eye and in several non-ocular tissues; IE1 mRNA was found in most of the DNA-positive tissues, while gH mRNA was amplified only in a few of the MCMV DNA-positive tissues from a single mouse. After immunosuppression, viral DNA and IE1 mRNA were detected at a higher frequency in various tissues of steroid-treated mice. gH mRNA was detected in a significantly higher number of the inoculated eyes, salivary glands and other non-ocular tissues of steroid-treated mice. After immunosuppression, low titers of infectious virus were recovered from the salivary glands of steroid-treated mice, but infectious virus was not recovered from the inoculated eye of either steroid-treated of non-immunosuppressed mice. The DNA PCR results suggest that after inoculation of 5 x 10(2) PFU of MCMV into the supraciliary space of euthymic BALB/c mice, virus becomes latent in the inoculated eye, salivary gland and other extraocular tissues. The RT-PCR results suggest that latent MCMV can be reactivated in multiple tissues by immunosuppression.
Assuntos
Terapia de Imunossupressão , Muromegalovirus/fisiologia , Proteínas Virais , Ativação Viral , Animais , Sequência de Bases , Primers do DNA , DNA Viral/análise , Olho/virologia , Feminino , Humanos , Proteínas Imediatamente Precoces/biossíntese , Proteínas Imediatamente Precoces/genética , Metilprednisolona/farmacologia , Camundongos , Camundongos Endogâmicos BALB C , Dados de Sequência Molecular , Muromegalovirus/genética , Muromegalovirus/imunologia , RNA Mensageiro/análise , RNA Viral/análise , Ratos , Transcrição Gênica , Proteínas do Envelope Viral/biossíntese , Proteínas do Envelope Viral/genética , Ativação Viral/efeitos dos fármacos , Ativação Viral/genética , Ativação Viral/imunologia , Latência ViralRESUMO
PURPOSE: In primary Sjögren's syndrome (SS), ocular surface changes within the conjunctival epithelium include lymphocytic infiltration, squamous cell metaplasia, and a reduction in goblet cell number. These changes may be the simple result of increased mechanical abrasion secondary to dryness. Alternatively, they may represent a local response to ocular and/or systemic inflammatory processes, perhaps in response to Epstein-Barr viral (EBV) infection, an agent recently implicated in the etiology of SS. To determine whether inflammatory processes or local infection by EBV contribute to the ocular surface pathology of SS, we examined the expression of inflammatory cell surface markers, cytokines, and EBV gene products within the ocular conjunctiva of patients with SS. METHODS: Ocular conjunctival tissue was isolated from patients with primary SS and nondry eye control patients by impression cytology or direct biopsy. These specimens were examined by immunofluorescence microscopy and reverse-transcriptase polymerase chain reaction (RT-PCR) for the expression of various markers. RESULTS: The authors found the frequency of expression of HLA-DR (P < 0.0001), ICAM-1 (P < 0.035), and IL-6 (P < 0.0001) to be significantly elevated in patients with primary SS versus nondry eye control patients. The IL-2 receptor and cytokines IL-1 beta and IL-8 were each found to be expressed with relatively high frequency in both patient populations, whereas mRNAs encoding cytokines IL-2, IFN-gamma, GM-CSF, and TGF-beta were not reproducibly detectable in either population. Messenger RNA encoding a marker for passive-latent EBV infection (EBNA-1) was detected with high frequency in both SS and normal populations. The EBV IL-10 analog BCRF-1 was expressed with low frequency in the SS population; however, these levels were not significantly different from the control population. The expression of two other markers of EBV infection, latent membrane protein (LMP, a lytic and latent marker), and BZLF-1 (putative latent-lytic switch gene) was undetectable in either study population. CONCLUSION: Based on the increased expression of the cell surface molecules HLA-DR and ICAM-1, and the inflammatory cytokine IL-6, the authors propose that local inflammatory processes contribute to the ocular surface changes and ocular surface dryness associated with primary SS.
Assuntos
Túnica Conjuntiva/microbiologia , Citocinas/análise , Herpesvirus Humano 4/isolamento & purificação , Síndrome de Sjogren/microbiologia , Sequência de Bases , Túnica Conjuntiva/imunologia , Citocinas/genética , Primers do DNA , Epitélio/imunologia , Epitélio/microbiologia , Infecções Oculares Virais/imunologia , Infecções Oculares Virais/microbiologia , Infecções por Herpesviridae/imunologia , Infecções por Herpesviridae/microbiologia , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/imunologia , Humanos , Dados de Sequência Molecular , Sondas de Oligonucleotídeos , RNA Mensageiro/análise , Síndrome de Sjogren/imunologiaRESUMO
The lacrimal gland (LG) immunopathology of Sjögren's syndrome (SS) consists of a proliferation of B and CD4 lymphocytes surrounding epithelial structures (Pepose JS, et al: Ophthalmology 1990, 97:1599-1605). Based on the detection of EBV genomes in a greater percentage of SS than normal LG biopsies, we previously postulated that Epstein-Barr virus (EBV) is a risk factor for LG lymphoproliferation in SS (Pflugfelder SC, et al: Ophthalmology 1990, 97:976-984). The purpose of this study was to determine the cellular site(s) of infection, virus type, and antigen expression of EBV infecting normal and SS LGs. EBV DNA was detected by in situ hybridization in intraductal epithelia in 13-33% of lobules in 21% of normal LGs and in cells in areas of B lymphoproliferation as well as the majority of epithelia in 86% of SS LGs. EBV genomic sequences were amplified from 36% of normal and 88% of SS LG biopsies by polymerase chain reaction. Only type 1 EBV sequences were amplified in SS LGs; in contrast EBV nuclear antigen 2-deleted but not type 1 sequences were amplified in normal LGs. Immunohistochemistry with EBV-specific monoclonal antibodies was performed on normal and SS LGs. No EBV antigens were detected in normal LGs. In contrast, latent antigens (latent membrane protein, EBV nuclear antigen 2) were detected in lymphocytes in areas of B lymphoproliferation, and early and late lytic cycle antigens were observed in epithelia in SS LGs. These studies suggest that EBV may play a role in the LG B lymphoproliferation and epithelial pathologic changes observed in SS.
Assuntos
Infecções por Herpesviridae/patologia , Herpesvirus Humano 4/isolamento & purificação , Aparelho Lacrimal/patologia , Síndrome de Sjogren/microbiologia , Síndrome de Sjogren/patologia , Adulto , Idoso , Antígenos Virais/genética , Sequência de Bases , Cadáver , Proteínas de Ligação a DNA/genética , Antígenos Nucleares do Vírus Epstein-Barr , Herpesvirus Humano 4/imunologia , Humanos , Imuno-Histoquímica , Hibridização In Situ , Aparelho Lacrimal/microbiologia , Pessoa de Meia-Idade , Dados de Sequência Molecular , Reação em Cadeia da PolimeraseRESUMO
Passive administration of antibody against herpes simplex virus type 1 (HSV-1) has been shown to protect against stromal keratitis and death from encephalitis. Although the exact mechanism by which passively-transferred antibody protects is not known, one of the features of protection by passively-transferred antibody is interference with the ability of the virus to spread within the nervous system. In the experiments reported herein, studies were performed to determine if 8D2, a monoclonal antibody against a type-common epitope of glycoprotein D, could protect mice from retinal necrosis following uniocular anterior chamber inoculation of HSV-1. Mice were protected from retinal necrosis when the antibody was administered 2 hours before virus inoculation or 24 hours after virus inoculation. When antibody was injected 2 hours before virus inoculation, the titer of virus at day 1 p.i. in the injected eyes of antibody-treated and control mice was the same, but by 3 days p.i., the titer of virus in the antibody-treated mice was significantly lower than that recovered from control mice. The titers of virus in the brains and in the uninoculated eyes of antibody-treated mice were also significantly lower than in control mice. The results of these studies suggest that passively-transferred antibody protects against retinal necrosis by limiting spread of virus to the CNS or replication of virus within the CNS.
Assuntos
Anticorpos Monoclonais/administração & dosagem , Infecções Oculares Virais/prevenção & controle , Imunização Passiva , Retinite/prevenção & controle , Proteínas do Envelope Viral/administração & dosagem , Animais , Câmara Anterior/microbiologia , Especificidade de Anticorpos/imunologia , Encéfalo/microbiologia , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Retinite/patologia , Simplexvirus/isolamento & purificaçãoRESUMO
Members of the herpesvirus family, cytomegalovirus (CMV), Epstein-Barr virus (EBV), and herpes simplex virus (HSV), have been recognized as causal agents of chorioretinal inflammatory diseases. We investigated the use of the polymerase chain reaction for the detection of CMV, HSV, and EBV genomes in aqueous, subretinal fluid, and vitreous specimens in patients with clinically diagnosed CMV retinitis. Cytomegalovirus but not HSV or EBV genomic sequences were detected in all of these clinical specimens. We also investigated 18 normal aqueous and eight normal vitreous specimens obtained from patients undergoing cataract or vitrectomy surgery. Cytomegalovirus, HSV, and EBV DNA were not detected in any of the normal aqueous specimens. There was one weakly positive CMV normal vitreous, but none was HSV or EBV positive by the polymerase chain reaction. These results indicate that the polymerase chain reaction may be useful as a rapid and sensitive diagnostic technique to aid in the confirmation of clinical observations.
Assuntos
Humor Aquoso/microbiologia , DNA Viral/análise , Herpesviridae/isolamento & purificação , Reação em Cadeia da Polimerase , Corpo Vítreo/microbiologia , Síndrome da Imunodeficiência Adquirida/microbiologia , Anticorpos Antivirais/sangue , Autorradiografia , Citomegalovirus/genética , Citomegalovirus/isolamento & purificação , Infecções por Citomegalovirus/microbiologia , Herpesviridae/genética , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/isolamento & purificação , Humanos , Reação em Cadeia da Polimerase/métodos , Retinite/microbiologia , Simplexvirus/genética , Simplexvirus/isolamento & purificaçãoRESUMO
In BALB/c mice, acute retinal necrosis occurs in the uninoculated eye 8 to 10 days following uniocular anterior chamber inoculation of the KOS strain of herpes simplex virus type 1 (HSV-1). Retinitis in the uninjected eye can be prevented if HSV-1-specific immune effector cells that have been restimulated with virus in vitro are administered intravenously within 1 day of anterior chamber inoculation of virus. We explored further the mechanism of protection afforded by these activated immune effector cells. The results of our studies revealed that optimal protection from retinitis required in vitro restimulation, since infusion of 50 x 10(6) HSV-1-primed but nonrestimulated cells could not protect as well as 10 x 10(6) activated cells. Analysis of both restimulated and nonrestimulated cells showed that only in vitro-restimulated cells were cytotoxic to HSV-1-infected syngeneic target cells. From these studies, we concluded that the ability to kill virus-infected target cells contributed to optimal protection achieved by intravenous administration of activated immune effector cells. Furthermore, T-cell subset depletion of activated immune effector cells demonstrated that both L3T4+ and Lyt-2+ T cells in the transfer inoculum contributed to protection. Additional studies revealed that although the transferred immune effector cells reached the injected eye within 24 h, virus replication in the injected eye was not affected. In the uninjected eye, virus titers were low, consistent with protection of this eye from retinitis. Taken together, the virus recovery results suggest that the interaction of virus with intravenously administered HSV-1-specific immune effector cells which limits virus spread and/or replication of virus probably occurred within the central nervous system and prevented the second wave of virus from entering the uninoculated eye.
Assuntos
Imunoterapia Adotiva , Ceratite Dendrítica/imunologia , Retina/patologia , Simplexvirus/imunologia , Linfócitos T/imunologia , Animais , Feminino , Ceratite Dendrítica/patologia , Ceratite Dendrítica/prevenção & controle , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Necrose , Retinite/imunologia , Retinite/prevenção & controle , Subpopulações de Linfócitos T/imunologia , Células VeroRESUMO
Herpetic ocular disease is one of the major causes of corneal blindness. Clinical diagnosis of corneal disease is based principally on corneal appearance. However, abnormal morphology of the corneal epithelium (CE) is not an indicator for the presence of a herpes virus. Further, it has not been established if herpes viruses are present in normal corneal epithelial tissue. In these studies, the polymerase chain reaction was used to evaluate normal and diseased corneal epithelium for the presence of herpes simplex virus type 1 (HSV-1), Epstein-Barr virus (EBV) and cytomegalovirus (CMV) genomic sequences. Thirty-two normal corneal epithelium specimens obtained from cadavers shortly after death were analyzed for HSV-1, EBV and CMV genomic sequences. Three of the 32 normal CE specimens were positive for amplified EBV DNA, 1 was positive for HSV-1 DNA, and none was positive for CMV DNA. We also tested eight herpetic dendritic lesions of which 3 were HSV-1 culture and PCR positive. The remaining five dendritic lesions were HSV-1 culture and PCR negative. Since these lesions were not evaluated for other herpesviruses, the etiology of these dendritic lesions is unknown. Six corneal epithelium samples from HIV-infected donors were negative for EBV, CMV and HSV-1 amplified sequences. Positive EBV, CMV and HSV-1 serology on all normal donors and on donors with clinically apparent disease did not correlate with positive PCR results. The results of these studies suggest that EBV and HSV-1 DNA can be amplified from a small percentage of apparently normal corneal epithelium.
Assuntos
Córnea/microbiologia , DNA Viral/análise , Amplificação de Genes , Ceratite Dendrítica/microbiologia , Reação em Cadeia da Polimerase , Simplexvirus/isolamento & purificação , Adulto , Idoso , Idoso de 80 Anos ou mais , Anticorpos Antivirais/análise , Sequência de Bases , Southern Blotting , Citomegalovirus/genética , Citomegalovirus/isolamento & purificação , Epitélio/microbiologia , Feminino , Imunofluorescência , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/isolamento & purificação , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Hibridização de Ácido Nucleico , Sondas de Oligonucleotídeos , Valor Preditivo dos Testes , Simplexvirus/genéticaRESUMO
Epstein-Barr virus (EBV) has been implicated in several ocular diseases; however, detection of the EBV genome in ocular tissues has not been documented. We report the detection of amplified EBV genomic sequences in 11 of 26 normal lacrimal gland DNA samples by using the polymerase chain reaction. Serum was available for 19 of the lacrimal gland donors. All 19 were EBV seropositive, although of the 19 lacrimal gland-seropositive patients, EBV sequences were detected in only 10 of the samples. Further, amplified EBV sequences were not detected in circulating lymphocyte DNA from normal seropositive volunteers, most likely because of the low frequency of circulating EBV-infected B cells. Amplification of EBV from cadaver lacrimal gland DNA was possible with minute quantities of DNA, whereas peripheral blood mononuclear cell DNA from normal volunteers did not amplify EBV sequences. Interestingly, the peripheral blood mononuclear cell polymerase chain reactions contained approximately 100 times more DNA than the lacrimal gland polymerase chain reactions. We conclude that the lacrimal gland may be a site for EBV persistence and that positive EBV serology is not an indicator of which individuals may have EBV harbored within their lacrimal glands.
Assuntos
Herpesvirus Humano 4/isolamento & purificação , Aparelho Lacrimal/microbiologia , Sequência de Bases , Sondas de DNA , DNA Viral/genética , Desoxirribonuclease BamHI , Genes Virais , Herpesvirus Humano 4/genética , Humanos , Linfócitos/microbiologia , Dados de Sequência Molecular , Reação em Cadeia da PolimeraseRESUMO
CD21, the receptor for the C3d complement fragment, has been reported to be the Epstein-Barr virus (EBV) receptor in B lymphocytes and cultured squamous epithelial cells. This receptor has previously been found to be expressed in the epithelia of nonocular mucosal-associated lymphoid tissues (MALT) which are also sites of persistent EBV infection. We recently found evidence of persistent EBV infection in human ocular MALT and hypothesized that the epithelia in these tissues may also express the complement (C3d)/EBV receptor. To test this hypothesis, histologic sections of human lacrimal gland, ocular surface tissue (conjunctiva, limbus, and cornea), and conjunctival impression cytology specimens were stained by immunohistochemical techniques using three different anti-CD21 monoclonal antibodies (HB-5, B2, OKB7). HB-5 stained lacrimal gland ductal and all ocular surface epithelia, except limbus. B2 stained lacrimal gland ductal and limbal epithelia, and OKB7 stained only the limbal and corneal epithelia. The intensity of limbal and corneal epithelial staining with all anti-CD21 antibodies correlated with the level of epithelial differentiation, with the weakest staining noted in the cells which are thought to be the stem and transient amplifying cells of the cornea. These results suggest that external ocular tissues, similar to other MALT, have CD21-positive epithelia which may be potential targets for EBV infection.
Assuntos
Olho/metabolismo , Herpesvirus Humano 4/metabolismo , Aparelho Lacrimal/metabolismo , Receptores de Complemento/análise , Receptores Virais/análise , Anticorpos Monoclonais , Antígenos de Diferenciação de Linfócitos B/análise , Túnica Conjuntiva/metabolismo , Córnea/metabolismo , Epitélio/metabolismo , Imunofluorescência , Humanos , Técnicas Imunoenzimáticas , Aparelho Lacrimal/imunologia , Receptores de Complemento 3d , Esclera/metabolismoRESUMO
Infection of nonpermissive primary hamster embryo cells with equine herpesvirus type 3 (EHV-3; multiplicity of infection = 10 pfu/cell) resulted in an abortive infection and the development of several hundred foci of rapidly growing cells. Five of these foci were chosen at random for the establishment of transformed cell lines, designated EVD-1 (equine venereal disease) through 5. These transformed cell lines exhibited altered biological properties typical of transformed cells, including immortality, growth to high saturation density, colony formation in soft agar, reduced serum requirements, aneuploid karyotype, and oncogenicity in syngeneic animals. Subsequently, five corresponding tumor cell lines (EVD-1T through 5T) with similar biological properties were established. All EHV-3 transformed and tumor cell lines have been shown to express EHV-3-specific proteins by indirect immunofluorescence assays employing rabbit antisera to EHV-3 infected equine cells. None of the transformed cell lines were found to release infectious virus by infectious center or cocultivation assay or to contain viral particles by electron microscopy.