EBV-associated Nuclear Antigen Research Articles

Forensic application of Epstein-Barr virus genotype: Correlation between viral genotype and geographical area

Using 50 forensic blood samples, the latent membrane protein 2A (LMP-2A) gene of Epstein-Barr virus (EBV) DNA was amplified to find a geographic correlation among the EBV genotypes. EBV DNA was detected in nine samples. From a phylogenetic analysis using 18 reported sequences as a reference, six EBV subtypes (Ia, Ib, Ic IIa, IIb, and IIc) were found. Japanese isolates were included in subtypes Ia or IIa. All the Asian reference isolates, except isolate D6, were included in subtype Ia or IIa. Mediterranean, an Alaskan and other African isolates were included in types Ib, Ic, IIb and IIc. The EBV genotype in the LMP-2A gene was thus demonstrated as being correlated with the host's geographical location. Typing in the EBV-associated nuclear antigen 2 gene was not related to that in the LMP-2A gene. Detection of the EBV genotype in the LMP-2A gene may be useful for determining the geographical origins of unidentified cadavers.

Regulation of tumor proliferation and tumor differentiation in gastrointestinal stroma tumors

One patient with infectious mononucleosis (IM) was studied from the probable time of Epstein-Barr virus (EBV) infection (38 days before the onset of clinically overt disease), during the incubation and acute phases, until 6 months after clinical remission. Analysis of spontaneous outgrowth of EBV-carrying lymphoblastoid cells, by limiting dilution on feeder layer cultures, showed that virus containing B lymphocytes are already present early during the incubation period. Also low interferon serum levels were detected early after infection, and only before the onset of clinical disease. All other studied clinical laboratory and virus-associated variables were within normal range during the incubation phase, but changed to a pattern characteristic of IM in parallel to the clinical symptoms. During the acute disease EBV-associated nuclear antigen (EBNA)-positive cells could be directly detected among the lymphocytes, and antibodies to EBV antigens appeared. Lymphocytes stained by monoclonal antibodies, detecting Ia-like determinants (activated cells) and supressor cells, increased dramatically, in parallel to a strong increase of functional suppressor cell activity, measured by inhibition of blastogenesis and PWM-induced immunoglobulin production. During the acute phase there was also a decrease of spontaneous cytotoxicity against the NK-sensitive cell line K562, while cytotoxicity (spontaneous) against an autologous EBV-positive lymphoblastoid cell line (LCL) was detected only during this phase. These reactions correlated to the presence of blasts, and the autologous reaction was exerted mainly by Fc-receptor-negative cells. Lymphokine production in response to EBV antigens was also initiated during the acute phase. During the convalescence period the serological and cellular immune parameters adjusted to the pattern of a normal EBV-seropositive person.

Cytogenetic aspects in the differential diagnosis of mesenchymal tumors of the gastrointestinal tract

One patient with infectious mononucleosis (IM) was studied from the probable time of Epstein-Barr virus (EBV) infection (38 days before the onset of clinically overt disease), during the incubation and acute phases, until 6 months after clinical remission. Analysis of spontaneous outgrowth of EBV-carrying lymphoblastoid cells, by limiting dilution on feeder layer cultures, showed that virus containing B lymphocytes are already present early during the incubation period. Also low interferon serum levels were detected early after infection, and only before the onset of clinical disease. All other studied clinical laboratory and virus-associated variables were within normal range during the incubation phase, but changed to a pattern characteristic of IM in parallel to the clinical symptoms. During the acute disease EBV-associated nuclear antigen (EBNA)-positive cells could be directly detected among the lymphocytes, and antibodies to EBV antigens appeared. Lymphocytes stained by monoclonal antibodies, detecting Ia-like determinants (activated cells) and supressor cells, increased dramatically, in parallel to a strong increase of functional suppressor cell activity, measured by inhibition of blastogenesis and PWM-induced immunoglobulin production. During the acute phase there was also a decrease of spontaneous cytotoxicity against the NK-sensitive cell line K562, while cytotoxicity (spontaneous) against an autologous EBV-positive lymphoblastoid cell line (LCL) was detected only during this phase. These reactions correlated to the presence of blasts, and the autologous reaction was exerted mainly by Fc-receptor-negative cells. Lymphokine production in response to EBV antigens was also initiated during the acute phase. During the convalescence period the serological and cellular immune parameters adjusted to the pattern of a normal EBV-seropositive person.

Predictive markers for development of strongyloidiasis in patients infected with both Strongyloides stercoralis and HTLV-1.

Severe strongyloidiasis has often been reported to occur in some patients infected with both Strongyloides stercoralis (S. stercoralis) and human T-cell leukaemia virus type 1 (HTLV-1); however, there are few useful predictive markers for the risk of development of strongyloidiasis in these patients. To search for such predictive markers, we examined peripheral blood and stool samples of individuals infected with both S. stercoralis and HTLV-1 in Okinawa, Japan, an area in which both of these are endemic. The HTLV-1 proviral load and antibody titre were examined in relation to the S. stercoralis load as measured by the direct faecal smear method in patients infected with both S. stercoralis and HTLV-1. The Epstein-Barr virus (EBV)-associated nuclear antigen (EBNA) antibody titre was also measured in these patients in order to examine the relationship between host immunity and HTLV-1 proviral load or antibody titre. The direct faecal smear-positive group showed both a higher HTLV-1 proviral load and HTLV-1 antibody titre than the -negative group (P < 0.05). In contrast, inverse correlations of these parameters with the EBNA antibody titre were observed, especially for proviral load (rho = -0.387, P < 0.05). These results suggest that HTLV-1 proviral load and antibody titre influence the S. stercoralis load via disturbance of the host immunity, and that proviral load would be an especially useful predictive marker of the risk of development of strongyloidiasis in patients infected with both S. stercoralis and HTLV-1.

Epstein-Barr Virus Nuclear Antigen (EBNA)-1 Carboxy-Terminal and EBNA-4 Sequence Polymorphisms in Nasal Natural Killer/T-Cell Lymphoma in the United States

Epstein-Barr virus (EBV) polymorphisms were examined in 12 cases of nasal natural killer (NK)/T-cell lymphoma diagnosed in the United States (U.S.-NL) with respect to the EBV-associated nuclear antigen (EBNA)-1 carboxy (C)-terminal region and the EBNA-4 region. A single dominant EBV strain was found in all cases. EBNA-1 sequences were remarkably homogeneous, showing either a P-ala (2/12) or P-ala variant (9/12) sequence. Other EBNA-1 subtypes known to be common in U.S.-reactive samples, such as P-thr or V-leu, were not identified. The final case had a base deletion with frame shift and premature stop codon. EBNA-1 C-terminal amino acid substitutions were common at codons 499 (10/12 cases), 502 (7/12), 524 (9/12), and 528 (6/12), all previously reported “hot spots.” However, unlike previous reports of other EBV-associated neoplastic and reactive tissues, mutations were absent at residues 487 and 492. Mutations within HLA-A11–restricted immunogenic EBNA-4 epitopes 399–408 and 416–424 occurred in 3 of 12 cases but were not associated with HLA-A11 status. In summary, the exclusive finding of P-ala variant or P-ala EBNA-1 sequences in U.S.-NL cases differs from that reported in U.S.-reactive and non-U.S.-NL cases. Although the significance of this difference is not known for certain, it may be related to geographic and/or site-specific variations, rather than oncogenicity per se.

An animal model for Epstein-Barr virus (EBV)-associated lymphomagenesis in the human: malignant lymphoma induction of rabbits by EBV-related herpesvirus from cynomolgus.

It is very important to develop and analyze animal models of Epstein-Barr virus (EBV)-associated tumors in the human. However, only a few reports on the animal models of EBV infection have been reported. Here we review those previous models and describe the details on our newly developed rabbit model of malignant lymphoma induced by EBV-related virus from cynomolgus. In brief, Si-IIA-EBV or Cyno-EBV induced T-cell lymphomas in rabbits inoculated intravenously (77-90%), orally (82-89%), subcutaneously (3/3) and intraperitoneally (2/3) about 2-5 months later. EBV-DNA was detected in peripheral blood by polymerase chain reaction 2 days after oral inoculation of Cyno-EBV while antiviral capsid antigen immunoglobulin G (IgG) was raised 3 weeks after the inoculation. Rabbit lymphomas and their cell lines contained EBV-DNA and expressed EBV-encoded small RNA-1 and EBV-associated nuclear antigen. Rabbit lymphoma cell lines, some of which have specific chromosomal abnormality, showed tumorigenicity in nude mice. The significance and further research subjects of this animal model will be discussed. We believe that the present rabbit model of lymphoma with specific chromosomal abnormalities is very useful for clarifying the role of EBV in human EBV-associated lymphoma and provides a means for studying prophylactic and therapeutic regimens.

Pyothorax-associated lymphoma: relationship with Epstein-Barr virus, human herpes virus-8 and body cavity-based high grade lymphomas

Pyothorax-associated lymphoma (PAL) is a newly-described entity developing several decades after artificial pneumothorax treatment for pulmonary or pleural tuberculosis. It is known to be associated with Epstein-Barr virus (EBV) with constant expression of the two latent membrane proteins: latent membrane protein (LMP)-1 and EBV-associated nuclear antigen (EBNA)-2. We are reporting three new cases of PAL. All of the tumours were of B-cell lineage and classified as large-cell diffuse lymphomas according to the International Working Formulation for the Classification of Lymphomas. The EBV genome was detected in two of the cases with LMP-1 and EBNA-2 expression. No EBV could be detected in the third case suggesting that different mechanisms may be involved in the pathogenesis of the disease. Body cavity-based high grade lymphomas (BCBL) represent a new disease, developing mainly in human immunodeficiency virus (HIV) infected patients: the tumoural cells often contain both human herpes virus (HHV)-8 (or Kaposi's sarcoma herpes virus) and EBV genomes, suggesting that these viruses might co-operate in the pathogenesis of the disease. The pleural location and the association of EBV have led to speculation that PAL could also be related to HHV-8 infection. However, no HHV-8 genome could be detected in any of the 14 tested cases already reported in the literature nor in the two cases we studied (one EBV-positive and one EBV-negative), suggesting that PAL and BCBL are two different entities.

EBV-NK Cells Interactions and Lymphoproliferative Disorders

Epstein-Barr virus (EBV) usually infects epithelial cells in the oropharynx and B lymphocytes asymptomatically. Occasionally, however, EBV infects T-cells and natural killer (NK) cells, and infection of these cells has been associated with the development of leukemias and lymphomas. EBV-positive lymphoproliferative disorders of NK cells have been reported with increasing frequency, but the interactions between EBV and NK cells are not fully understood, in part because NK cells are not usually infected with EBV in vitro. The lymphoma-derived EBV-positive NK cell line, YT, has been useful in the study of EBV infection of NK cells. YT cells express the EBV-associated nuclear antigen (EBNA)-1, the latent membrane protein (LMP)-1, and LMP-2A, but not EBNA-2 and LMP-2B genes. This pattern of latent gene expression is compatible with a type II latency program, normally associated with nasopharyngeal carcinoma, Hodgkin's disease, and T-cell lymphoma. In this report, we summarize recent information on EBV-NK cell interactions and EBV-positive lymphoproliferative disorders of NK cells.

Malignant lymphoma induction in rabbits by intravenous inoculation of Epstein-Barr-virus-related herpesvirus from HTLV-II-transformed cynomolgus leukocyte cell line (Si-IIA).

Malignant lymphomas, which were usually of T-cell type, were induced in 10 of 13 (77%) male rabbits (Japanese white, 8/10; New Zealand white, 2/3) inoculated i.v. with HTLV-II-transformed simian (Cynomolgus) leukocyte cell line (Si-IIA) cells. Of 7 rabbits injected with cell-free pellets from Si-IIA cultures, 5 also developed malignant lymphoma (15-28 days). Lymphoma development was completely inhibited by inactivation of cell-free pellets from Si-IIA culture with ethyl ether and was almost suppressed by neutralization of the cell-free pellets with anti-Si-IIA sera. Herpesvirus particles were discovered very rarely in Si-IIA cells, in addition to C-type virus particles, by electron microscopy. Si-IIA cells were positive for Epstein-Barr-virus (EBV)-associated nuclear antigen (EBNA) by immunofluorescence (IF) test. Antibody response to viral capsid antigen of EBV was also detected in sera from rabbits inoculated with Si-IIA. EBV-encoded RNA-1 (EBER-1) was demonstrated in Si-IIA, the tumor tissues and all rabbit tumor cell lines by in situ hybridization. EBV DNA was also detected in Si-IIA and rabbit lymphoma cell lines by polymerase chain reaction (PCR) and Southern blotting. However, EBV DNA was amplified only by some primers complementary to human EBV sequence (B95-8), but not by other primers. Integration of HTLV-II provirus genome could not be detected in Si-IIA-induced rabbit tumor cells. Moreover, no lymphoma was induced by inoculation of HTLV-IIC and MOT (other HTLV-II-producing human cell lines), B95-8(EBV-producing cell line) or TALL-1 and peripheral leukocytes from normal Cynomolgus (controls). Neither Herpesvirus saimiri nor H. ateles (simian oncogenic viruses) were detected in Si-IIA cells by IF test. These data suggest that the high rate of lymphoma induction in rabbits may not be caused by HTLV-II, human EBV (B95-8) or well-known simian oncogenic viruses, but by EBV-related herpesvirus derived from Si-IIA cells or HTLV-IIA cells, with which Si-IIA was established. The availability of this animal model promises to clarify the role of EBV in human lymphoma and provides a means of studying prophylactic and therapeutic regimens.

A positive correlation between the precursor frequency of cytotoxic lymphocytes to autologous Epstein-Barr virus-transformed B cells and antibody titer level against Epstein-Barr virus-associated nuclear antigen in healthy seropositive individuals.

A limiting dilution analysis was established to determine the precursor frequency (PF) of cytotoxic lymphocytes against autologous B cells transformed with the Epstein-Barr virus (EBV). This method was found to detect mainly self-restricted T-cell activity and little non-self-restricted cytotoxicity. The mean PF in 21 healthy EBV-seropositive persons was 1.4 x 10(-3) (range: 0.03 x 10(-3) to 8.7 x 10(-3)) for peripheral blood mononuclear cells, whereas 4 samples of mononuclear cells obtained from umbilical cord blood had PFs below 0.007 x 10(-3). A positive correlation was observed between the PF and serum antibody titers against EBV-associated nuclear antigen among the seropositive persons.

Serologic and Virologic Epidemiology of Epstein-Barr Virus: Relevance to Chronic Fatigue Syndrome

Patients considered to have chronic fatigue syndrome (CFS) have been reported to exhibit an increased antibody response to Epstein-Barr virus (EBV) early antigen complex and capsid antigen, findings that suggest some relationship between EBV and CFS. However, the serologic findings have not been totally consistent among different study groups, and the antibody patterns in asymptomatic individuals may be similar. Moreover, patients with symptomatology indicative of CFS do not appear to have an abnormal burden of EBV in body fluids and manifest only a variable, mild degree of EBV-specific cell-mediated responses. The evidence is growing that the serologic findings of an enhanced EBV state in individuals with CFS-like manifestations, as well as the subsequent reports of increased antibody titers to other viruses, reflect a generalized underlying immunologic dysfunction in these patients. Future studies with criteria-defined CFS study groups in which determinations are made of antibody responses to newly identified EBV-associated nuclear antigen components and distinct EBV proteins in addition to specific virologic and immunologic analyses of EBV may be worthwhile as a means of clarifying the association between EBV and CFS.

Antibodies to Epstein-Barr Virus in Iridocorneal Endothelial Syndrome

Antibody titers to Epstein-Barr virus were determined in 13 patients with iridocorneal endothelial syndrome and in 13 healthy race-, age-, and sex-matched controls. Both the geometric mean titer of IgG antibodies to the Epstein-Barr virus capsid antigen and the proportion with high titers of IgG antibodies to the Epstein-Barr virus capsid antigen (greater than or equal to 1:640) were significantly higher in 12 seropositive patients with iridocorneal endothelial syndrome than in 12 seropositive controls (1/761:1/202, P = .001; 83.3%:8.3%, P less than .001). Ten of 12 seropositive patients with iridocorneal endothelial syndrome and five of 12 seropositive controls had antibodies to Epstein-Barr virus-induced early antigens (greater than or equal to 1:10) (Fisher's Exact Test, P less than .05), while four seropositive patients with iridocorneal endothelial syndrome and one seropositive control had low to undetectable levels of antibodies to Epstein-Barr virus-associated nuclear antigen (less than or equal to 1:5) (P greater than .1). Antibody levels to cytomegalovirus or measles virus were not different between patients with iridocorneal endothelial syndrome and controls. Additional studies showed no evidence of humoral immune disorder or collagen vascular disease in the patients with iridocorneal endothelial syndrome. The serologic profiles suggest that the patients with iridocorneal endothelial syndrome examined had a cellular immune abnormality sufficient to permit reactivation of latent Epstein-Barr virus infection and imply, but do not establish, a role for Epstein-Barr virus infection in the pathogenesis of some cases of the iridocorneal endothelial syndrome.

Detection of Epstein-Barr virus genome in benign polyclonal proliferative T cells of a young male patient

Epstein-Barr virus (EBV) DNA was detected in polyclonal T cells that proliferated transiently in a 21-year-old male (referred to as H.J.) who underwent an apparently benign lymphocytosis (white blood cells, 31 x 10(6)/microL; lymphocyte, 79%) with fever, tonsillar swelling, lymphadenopathy, and hepatosplenomegaly. The symptoms and signs subsided mostly within a month of hospitalization. The major population of the lymphocytes at admission was positive for CD3, CD8 (4/8 ratio, 0.16), WT31, and DR antigen. Eight percent of the leukocytes were too blastoid to be classified as atypical lymphocytes of infectious mononucleosis (IM). The blastoid lymphocytes and the duration and degree of the lymphocytosis and hypergammaglobulinemia appeared inconsistent with IM, whereas the EBV serology indicated either EBV primary infection or a secondary alteration of normal seropositive EBV immunity. The genomic analysis of T-cell receptor beta chain in the peripheral blood mononuclear cells (PBMC) at admission with a C beta probe did not show a monoclonal rearrangement. EBV genome was detected in these cells, using the BamHI W and K probe, but not in the cells after discharge. Analysis of the EBV terminal repeat junctional sequence, using Xho I fragment of the latent membrane protein (LMP) probe binding with the terminus, did not show monoclonal or oligoclonal populations. EBV-associated nuclear antigen (EBNA) was detected in 36% of the PBMC at admission, but not in the later cells. These EBNA- positive cells were found to form rosette with sheep erythrocytes. The PBMC of six acute IM patients contained neither EBV DNA nor EBNA- positive cells. The observations in this case show a unique type of EBV infection in T cells that has not been previously reported.

Detection of Epstein-Barr Virus Genome in Benign Polyclonal Proliferative T Cells of a Young Male Patient

Epstein-Barr virus (EBV) DNA was detected in polyclonal T cells that proliferated transiently in a 21-year-old male (referred to as H.J.) who underwent an apparently benign lymphocytosis (white blood cells, 31 x 10(6)/microL; lymphocyte, 79%) with fever, tonsillar swelling, lymphadenopathy, and hepatosplenomegaly. The symptoms and signs subsided mostly within a month of hospitalization. The major population of the lymphocytes at admission was positive for CD3, CD8 (4/8 ratio, 0.16), WT31, and DR antigen. Eight percent of the leukocytes were too blastoid to be classified as atypical lymphocytes of infectious mononucleosis (IM). The blastoid lymphocytes and the duration and degree of the lymphocytosis and hypergammaglobulinemia appeared inconsistent with IM, whereas the EBV serology indicated either EBV primary infection or a secondary alteration of normal seropositive EBV immunity. The genomic analysis of T-cell receptor beta chain in the peripheral blood mononuclear cells (PBMC) at admission with a C beta probe did not show a monoclonal rearrangement. EBV genome was detected in these cells, using the BamHI W and K probe, but not in the cells after discharge. Analysis of the EBV terminal repeat junctional sequence, using Xho I fragment of the latent membrane protein (LMP) probe binding with the terminus, did not show monoclonal or oligoclonal populations. EBV-associated nuclear antigen (EBNA) was detected in 36% of the PBMC at admission, but not in the later cells. These EBNA-positive cells were found to form rosette with sheep erythrocytes. The PBMC of six acute IM patients contained neither EBV DNA nor EBNA-positive cells. The observations in this case show a unique type of EBV infection in T cells that has not been previously reported.

Effects of d-nicotine on contraction and relaxation induced by I-nicotine in the tracheal and main bronchial preparations of guinea pig and rabbit

An Epstein-Barr virus (EBV) genome-positive epithelial hybrid cell line, NPC-KT, derived from the fusion of primary nasopharyngeal carcinoma cells with a human epithelial cell line of adenoid origin has previously been described (Takimoto, Kamide, and Umeda, 1984a; Takimoto, Ogura, Sato, and Hatano, 1984b; Takimoto, Ogura, Sato, Umeda, and Hatano, 1985). The NPC-KT cells produce virus (NPC virus) with both transforming and lytic properties. An EBV genome-negative human lymphoblastoid cell line, Ramos, was infected with NPC virus. Three months after infection, more than 90 % of the cell population expressed EBV-associated nuclear antigen (EBNA). Approximately 40% of the cells exhibited a brilliant pattern, whereas the remaining 50% of the cells showed a faint granular pattern. The result suggests that there may be heterogeneity in NPC virus.

Immune status towards Epstein-Barr virus in a group of sicilian children

The prevalence of antibodies to Epstein-Barr virus-determined antigens was studied in 17 children with acute infectious mononucleosis (IM) and in 263 children hospitalized for diseases unrelated to EBV infection. Antibodies against Epstein-Barr viral capsid antigens (VCA) were observed in 173 patients of the control group (66%), but 58 of them (33,5%) had not yet developed antibodies against Epstein-Barr virus-associated nuclear antigen (EBNA). IgM-specific antibodies were not found in any of the children of the control group but were present in all of the 17 patients with IM. The rates of positivity for IgA anti-VCA and IgG anti-early antigen (EA) were similar in all age groups. Anti-viral capsid antigen IgG seropositivity increased to 83% by the age of six years, the mean geometric titre being highest between the ages of 2 and 4 years. Our results suggest that in Sicilian children the primary infection occurs prevalently early in life, in parallel with the occurrence of IM.

Epstein-Barr Virus (EBV) Infection by Cocultivation of Fibroblast-Dominant Cell Line and EBV-Transformed Lymphocytes

A long-term cell culture epitheloid cell line (NPC-F/L) was established from the fibroblast-dominant cell line derived from nasopharyngeal carcinoma tissue by cocultivation with an Epstein-Barr virus (EBV)-transformed lymphoblastoid cell line of adenoid origin. The cells were investigated by karyotyping, by immunofluorescent testing for EBV-associated nuclear antigen (EBNA), early antigen (EA), and viral capsid antigen (VCA) and by a light microscopy. The cells were shown to be epithelial not only by morphology but also by immunocytochemical staining with monoclonal antibody to cytokeratins and they expressed EBNA and EA but not VCA. The cells developed histologically undifferentiated squamous cell carcinoma when inoculated subcutaneously into the back of athymic nude BALB/c(nu/nu) mice, which were similar to the human tumors from which the fibroblastoid cell line was established. The presence of EBNA in touch smears prepared both from the biopsy tissue of human and nude mice was demonstrated. Karyotyping analysis indicated an aneuploid chromosome with a model number of 74.

The Human Nasopharynx as a Reservoir for Epstein-Barr Virus

Nineteen human nasopharyngeal tissue specimens biopsied from patients with suspected nasopharyngeal carcinoma (NPC) and 15 adenoid tissue specimens obtained by surgery were successfully grown in culture without bacterial and fungal contamination. Fibroblastic growth developed in all of the cultures. In some cases, epithelial growth was also seen among the fibroblastic growth. The cells became round and floated in the culture fluid between 19th and 57th day of culture in 6 out of 10 NPC tissue cultures, 3 out of 9 tissue cultures of apparently normal nasopharyngeal mucosa of patients referred for exclusion of NPC, and 4 out of 15 adenoid tissue cultures. The floating cells consisted of B-lymphoblastoid cells and were positive Epstein-Barr virus (EBV)-associated nuclear antigen (EBNA). The data suggest that a small number of B-lymphocytes in the human nasopharynx serve as a reservoir for EBV.

A B-cell line having chromosome 14 aberration at break band q11 derived from an adult T-cell leukemia patient.

A B‐cell line having translocations of chromosome 14 at break band q11 (the assigned locus of the α‐chain gene of the T‐cell antigen receptor) and chromosome 3 at break band p25 (the assigned locus of the c‐raf‐1 oncogene) was established from peripheral blood leukocytes of an adult T‐cell leukemia (ATL) patient. The same chromosome 14 aberration at break band q11 and chromosome 3 aberration at break band p25 were also found in fresh T‐cell leukemia cells. The B‐cell line is surface immunoglobulin (sIg)+, immunoglobulin gene rearrangement+, ATL‐specific antigen (ATLA)+, HTLV‐1 proviral genome+, Epstein‐Barr virus (EBV)‐associated nuclear antigen (EBNA)+ and the EBV DNA genome+. The fresh T‐leukemic cells were T‐cell receptor gene rearrangement+, the HTLV‐1 proviral genome+ and EBV DNA genome−.

Establishment and Characterization of a Human Myeloid Cell Line From Philadelphia Chromosome–Negative Myeloblastic Leukemia Arising in a Patient With Myelodysplastic Syndrome

A new hematopoietic cell line derived from a patient with Philadelphia chromosome (Ph1)-negative myeloblastic leukemia arising from a form of myelodysplastic syndrome (MDS) is described. This cell line, designated TMM, consists of immature cells with the morphological characteristics of young myeloblasts and grows in suspension culture with a doubling time of about 30 hours. By cytochemical analysis the cultured cells were positive for acid phosphatase. They were free of the Epstein-Barr virus-associated nuclear antigen as well as terminal deoxynucleotidyl transferase. Further phenotypic analysis revealed the expression of the myelomonocytic-specific antigen Leu-M1 and receptors for the Fc portion of IgG. Partial differentiation of these cells could be induced by dimethyl sulfoxide, tetradecanoyl phorbol acetate, or hypoxanthine and resulted in cells of the myeloid series expressing lysozyme and receptors for the C3b complement protein. The karyotype was 46,XY, lacked the Ph1 chromosome, and displayed no abnormalities at the light microscopic level. No rearrangement of the bcr-c-abl gene complex was found. This cell line should be useful for studying an important type of the heterogeneous population constituting Ph1-negative myeloblastic leukemia, arising in this instance from MDS, as well as for studying differentiation and proliferation of human pluripotent stem cells.