Posttransplant lymphoproliferative disease in primary Epstein-Barr virus infection after liver transplantation: the role of cytomegalovirus disease.

Epstein-Barr virus (EBV) plays a major role in the pathogenesis of posttransplant lymphoproliferative disease (PTLD). Patients who undergo primary EBV infection after transplantation are at greater risk of developing PTLD. In this retrospective study, the incidence of EBV infection and associated PTLD in 40 consecutive adult recipients who were seronegative for EBV at the time of liver transplantation were investigated, and risk factors for PTLD were analyzed. Of 37 patients with available timely posttransplant serum samples, 35 (95%) developed primary EBV infection. Of the 40 patients, 13 (33%) developed PTLD a median of 126 days (range, 48-776) after liver transplantation. The factor significantly associated with the development of PTLD was cytomegalovirus disease (relative risk, 7.3; 95% confidence interval, 2.36-22.6; P = .0006). Cytomegalovirus disease is a predictor for the development of PTLD in primary EBV infection after liver transplantation, and it may be a target for prophylactic intervention.

EBV gene expression, EBNA antibody responses and EBV+ peripheral blood lymphocytes in post-transplant lymphoproliferative disease.

Epstein-Barr virus (EBV) is associated with the development of several B cell malignancies including Burkitt's lymphoma (BL), post-transplant lymphoproliferative disease (PTLD), and AIDS-related lymphomas. The latter two diseases result from EBV-driven B cell proliferation in the absence of normal immunosurveillance and as such, represent a heterogenous family of lymphoproliferative disorders. This article reviews studies on EBV gene expression and antibody development in PTLD and introduces recent information on the levels of EBV+ peripheral blood lymphocytes to discuss possible mechanisms of pathogenesis under varying conditions of immunosuppression.

Increased levels of circulating Epstein-Barr virus (EBV)-infected lymphocytes and decreased EBV nuclear antigen antibody responses are associated with the development of posttransplant lymphoproliferative disease in solid-organ transplant recipients.

Epstein-Barr virus (EBV)-associated posttransplant lymphoproliferative disease (PTLD) is an uncommon but potentially fatal complication of immunosuppression in solid-organ transplant recipients. A semiquantitative DNA polymerase chain reaction assay was developed to amplify a unique 269-bp region of the EBNA-1 gene in peripheral blood lymphocytes (PBL) using the primers described by Telenti et al (J Clin Microbiol 28:2187, 1990). Serial samples were studied from 23 transplant recipients, 12 of whom were diagnosed with PTLD. The majority of transplant recipients who were EBV seropositive at the time of transplant surgery and who did not develop PTLD (5 of 7, 71%) exhibited less than a 10-fold increase in the levels of EBV-infected PBL over the 0.1 to 5 EBV genomes/10(6) PBL observed in immunocompetent EBV seropositive controls. Transplant recipients who were seronegative at the time of transplantation and who underwent a primary EBV infection but did not develop PTLD exhibited a reduced capacity to control viremia because the levels of EBV-infected PBL were up to 400 times greater than the 1.0 to 50 EBV genomes/10(6) PBL observed in individuals undergoing acute infectious mononucleosis (Rocci et al: N Engl J Med 296:132, 1977). However, all transplant recipients who developed PTLD exhibited a marked elevation of EBV-infected PBL independent of their serologic state at the time of transplantation. Six of the 10 transplant recipients with PTLD exhibited > or = 300,000 EBV genomes/10(5) PBL, two exhibited 10,000 to 50,000 EBV-infected genomes/10(5) PBL, and one each exhibited 2,500 and 500 EBV genomes/10(5) PBL. However, the latter two samples were obtained 4 to 5 weeks after the diagnosis of PTLD and may reflect a decrease in viral load resulting from immunomodulation. Marked decreases in the levels of EBV nuclear antigen-1 (EBNA-1), EBNA-2, and EBNA-LP antibodies correlated with the increase in EBV-infected PBL. Hence, a quantitative difference in circulating EBV viral load and EBNA antibody levels is evident between transplant recipients with and without PTLD and may be useful as a noninvasive prognostic marker with which to monitor and/or predict the development of PTLD.

Evidence for restricted Epstein-Barr virus latent gene expression and anti-EBNA antibody response in solid organ transplant recipients with posttransplant lymphoproliferative disorders.

Epstein-Barr virus (EBV) is associated with the development of two human B-cell malignancies, Burkitt's lymphoma and lymphomas that occur in the immunosuppressed host. The latter category of disease has become important recently as it is seen primarily in organ transplant recipients and individuals with acquired immunodeficiency syndrome. One possible mechanism for lymphoma development involves a reduction in or lack of EBV-specific cytotoxic T-cell recognition. In support of this model are previous observations that the expression of EBV nuclear antigen 2 (EBNA2) and latent membrane protein, two viral antigens associated with major histocompatibility complex class I-restricted T-cell killing, are downregulated in Burkitt's lymphoma and in early passage lymphoblastoid cell lines (LCL) derived from the malignant lesions. To determine whether a similar mechanism could occur in the development of posttransplant lymphoproliferative disorders (PTLD), we compared EBV gene expression among 23 PTLD tumor lesions obtained from 11 solid organ transplant recipients and among LCL derived from 3 of these lesions. In this report, we demonstrate, by Southern blot, Western blot, and immunofluorescence analysis, that (1) the tumor lesions exhibit varying patterns of restricted viral gene expression; (2) LCL derived from these lesions may represent the in vitro selection of cell subpopulations; and (3) immunosuppressed individuals have a markedly reduced antibody response to the latent cycle antigens, EBNA1, EBNA2, and EBNA-LP, but not to the lytic cycle viral capsid antigen when compared with normal immunocompetent controls.

Reactivation of Epstein-Barr virus during early infection with human immunodeficiency virus.

Reactivation of Epstein-Barr virus (EBV) in early human immunodeficiency virus (HIV) infection was investigated in 49 homosexual men who seroconverted to HIV (cases) as compared with 49 matched controls who remained seronegative to HIV during a longitudinal study. EBV infection was reactivated in cases 6 months, but not 12 months, prior to HIV seroconversion as compared with controls and remained reactivated during 18 months of follow-up after HIV seroconversion, as shown by increases in immunoglobulin (Ig) G antibody titers to EBV early antigen. Antibody titers to EBV viral capsid antigen did not differ between cases and controls prior to the time of seroconversion to HIV but were significantly increased among cases by the first seropositive study visit and remained elevated during the 18 months after HIV seroconversion. Total serum IgG levels were increased in cases at the visit of seroconversion, and during 18 months of follow-up, but did not correlate with enhanced IgG production specific for EBV antigens. Significant decreases in numbers of CD4+ cells and increases in numbers of CD8+ cells during this early phase of HIV infection were not associated with changes in patterns of EBV antibody responses. Reactivation of EBV beginning 6 months before HIV seroconversion may have implications regarding the role of this herpesvirus in the pathogenesis of HIV.

Epstein-Barr virus transmission via the donor organs in solid organ transplantation: polymerase chain reaction and restriction fragment length polymorphism analysis of IR2, IR3, and IR4.

Two organ transplant recipients who received organs from a common donor and were diagnosed as having an Epstein-Barr virus (EBV)-associated posttransplant lymphoproliferative disorder were studied to determine the mode of EBV transmission. The results of restriction fragment length polymorphism, polymerase chain reaction, and minisatellite DNA analyses demonstrate that both patients had a common strain of EBV and that this strain was transmitted from the donor's organs to both recipients. Posttransplant lymphoproliferative disorder resulted from the proliferation of EBV-immortalized B lymphocytes of the recipient, not those of the donor.

Current understanding of the natural history of genital herpes simplex infections.

Considerable variations in the disease make an accurate portrayal of genital herpes simplex virus (HSV) infections quite difficult. Primary genital HSV infections classically produce ulcerated lesions lasting a mean of 16-21 days in men and 10-16 days in women. HSV shedding has been documented for a mean of 9.1-11.6 days in men and 8.0-14.7 days in women. Probably many adults have a less severe initial genital infection, however, and it may be so mild as to be misdiagnosed or undetected. Studies of recurrent genital HSV infections are often misleading because they overestimate the frequency and severity of recurrences. By necessity, these studies have recruited and evaluated those adults with the worst and most frequent recurrences. Data from these studies show even greater variations than do data on initial episodes of genital HSV infection. The frequency and clinical importance of asymptomatic genital HSV shedding are being recognized and documented only now. Further studies of this phenomenon and of the natural history of all genital HSV infections are needed urgently.

Characteristics and management of pregnancy in women with genital herpes simplex virus infection.

The natural history of genital herpes simplex virus (HSV) infections was investigated in 83 pregnancies in 78 women, and the information was used to determine the need for cesarean section in these women. We studied 163 recurrent episodes with HSV cultures from the cervix and from vulvar lesions every 1 to 3 days. Cervical HSV cultures were obtained weekly from asymptomatic women beginning at 32 weeks' gestation, and 14/462 (3.03%) of these cultures were positive. Cervical cultures obtained during culture-positive vulvar recurrences demonstrated concomitant cervical HSV shedding in 25/165 (15.2%) cultures. Mean duration of 26 genital HSV recurrences was 4.6 +/- 2.8 days with a range of 1 to 13 days. The mean interval between culture-positive HSV recurrences was 59.2 +/- 42.1 days, but many (14/76 = 18%) intervals were less than 21 days. Viral cultures were already positive in 92.3% of cases after 4 days' incubation, so they could be used effectively to determine route of delivery. Following a cautious set of criteria for vaginal delivery in these women, 69.1% were delivered of their infants vaginally and no neonatal morbidity caused by HSV was encountered. Information about the natural history of genital HSV infections obtained from frequent third-trimester viral cultures can be used to manage pregnancy and will reduce the need for cesarean section while avoiding neonatal HSV morbidity.

Effect of prolonged administration of interferon-alpha on pharmacokinetics, fever, lymphocyte proliferative response, and NK cell activity.

Over a 60-week period, a patient with multiple warts received a total of 281 x 10(6) units of human leukocyte interferon (IFN-alpha) by intramuscular (IM) or intralesional (IL) injections. Circulating IFN was significantly higher following IM administration than after IL administration. These pharmacokinetics did not change. The patient's body temperature was always significantly elevated after administration of IFN. However, hyporeactivity to the febrile response developed when the interval between repeated injections of IFN was less than six days. The lymphocyte count was significantly decreased within 5-7 h after administration of IFN and had returned to normal by 24 hours, whereas total WBC, platelet, and monocyte counts were not altered. There was a depression of specific lymphocyte proliferative response to herpes simplex virus after multiple daily injections, but not after prolonged therapy. Circulating natural killer (NK) levels were not elevated during the first two months of IFN therapy. After the patient had received about 100 x 10(6) units of IFN, however, the NK cell level became elevated and remained elevated upon cessation of treatment. NK activity was stimulated by in vitro incubation of peripheral mononuclear cells with 1000 units of IFN during the initial phase of treatment. A decline of in vitro stimulation of NK activity by interferon developed during two subsequent periods of treatment with mean daily doses of 2.46 and 1.07 x 10(6) units of IFN. Long-term therapy in our patient with an average of 4.7 x 10(6) units of IFN/week was well tolerated, did not irreversibly affect platelet or white cell counts or nonspecific or virus-specific cell mediated immune responses, and enhanced circulating NK levels.

Effects of interferon-alpha on human warts.

Two patients with extensive warts which were stable for two years or more were treated with human interferon-alpha to assess the ability of interferon to affect this benign tumor of viral etiology. Intramuscular administration of 96.6 and 135 million units over 12-15 weeks produced softening and decreased scaling of each patient's warts. Double blind, placebo-controlled intralesional injections resulted in progressive disappearance of interferon treated warts. A dose response relationship was shown in eight warts. The minimum effective dose was 1.2 x 10(6) units injected over 15.5 weeks.

Effect of pyran on latency after herpes simplex virus infections.

The immunomodulator pyran protected mice against both herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) infections. In infections of the lip with HSV-1, prophylactic administration of pyran reduced the severity of the herpetic lesions and enhanced their resolution, but did not decrease the high incidence of development of latent HSV-1 infection of the trigeminal ganglia. In vaginal infections with HSV-2, prophylactic administration of pyran either systemically or locally reduced mortality, reduced the incidence of mice with vaginal HSV-2 infection, and did not alter the low incidence of latent infection of the spinal dorsal root ganglia. Pyran treatment before systemic herpetic infection after intravenous inoculation of HSV-2 also reduced mortality and virus replication, as evidenced by a decreased antibody response in the survivors, and it either reduced latent infection in the spinal dorsal root ganglia or did not predispose mice to latent infection. Treatment with the immunomodulator appeared to inhibit or reduce HSV infection early in viral pathogenesis in all three model systems, producing protection from clinical disease and resulting in less virus to induce a systemic antibody response, with either a reduction in latent virus infection or no enhancement of development of latency. In all of the HSV models, the development of latent herpetic infection was closely correlated with sufficient virus replication early in the infection to induce a systemic neutralizing-antibody response.

Immune responses to vaginal or systemic infection of mice with herpes simplex virus type 2.

Relationships among cell-mediated immune (CMI) responses, neutralizing antibody, and macrophages in host resistance to vaginal or systemic infection with HSV-2 were defined in BALB/c mice. The importance of T-lymphocytes in resistance to either systemic or vaginal infection was demonstrated by increased susceptibility of mice depleted of T-lymphocytes. A role for non-specific resistance against the systemic, but not vaginal, infection was demonstrated by increased susceptibility of mice depleted of macrophages. The temporal relationships among the serum antibody response, and the delayed-type hypersensitivity (DTH) and splenic proliferative CMI responses were defined in individual animals. Vaginal infection was associated exclusively with relatively transient CMI responses which appeared during the acute infection. After systemic infection, CMI responses appeared during the acute infection. After systemic infection, CMI responses appeared during the acute infection and persisted for approximately five weeks, while a humoral response appeared in surviving animals and persisted for at least four months. The results suggest that CMI is the predominant host response to vaginal HSV-2 infection, while both CMI and macrophage-mediated antiviral activity may be involved in recovery from primary systemic infection with HSV-2.

Immune responses to vaginal or systemic infection of BALB/c mice with herpes simplex virus type 2.

The temporal relationships among the humoral and cellular immune responses were defined in BALB/c mice after vaginal or systemic infection with herpes simplex virus type 2 (HSV-2). After vaginal infection, mice showed evidence of clinical vaginitis on days 4 to 6 and HSV-2 replication was detected locally in the vaginal secretions, cervix, vagina, and uterus before the virus subsequently spread to the central nervous system. Death from encephalitis occurred between 7 and 10 days after infection. Vaginal infection was associated with significant delayed type hypersensitivity and splenic proliferative cell-mediated immune responses which appeared during the acute infection and waned by 3 weeks. There was almost no evidence of a systemic neutralizing antibody response at any time after vaginal infection. In contrast to the local vaginal infection, systemic i.v. HSV-2 infection induced a humoral response as well as the two cellular immune responses. Although both cellular immune responses appeared during the acute infection (days 6 to 14) and persisted for approximately 5 weeks, the humoral response appeared in surviving animals and persisted for at least 4 months. Thus, vaginal HSV-2 infection was associated primarily with transient cellular immune responses, whereas i.v. HSV-2 infection induced prolonged systemic humoral and cellular immune responses.

Cellular basis of interferon formation and hyporeactivity after exposure to bacterial lipopolysaccharide.

Endotoxin apparently induces interferon in only a few types of cell, yet it produces strong hyporesponsiveness to a large number of agents. After incubation for 24 hr with endotoxin in vitro, tissue cultures of thymus, spleen, mesenteric lymph nodes, and to a lesser extent, liver and lung produced interferon. Lymphoid tissues of bone marrow-derived (B-) cells (sacculus rotundus, appendix, and Peyer's patches) and kidney did not produce interferon. Adherent spleen cells produced more interferon than nonadherent cells. Purification of spleen cells on a bovine serum albumin gradient showed that light, DNA-synthesizing cells made interferon in response to endotoxin or polyriboinosinic-polyribocytidylic acid. Mouse spleen cells produced a "late" interferon 24-48 hr after exposure to endotoxin, which, in contrast to "early" interferon (produced at 0-24 hr) from spleen and other tissues, is stable at 56 C for 60 min. It is suggested that this late interferon represents a "B-cell" interferon. Hyporeactivity was produced in vitro by endotoxin only in tissues that make endotoxin-induced interferon, a fact consistent with the theory that the interferon-inducing mechanism must be initiated before hyporeactivity results. The fact that endotoxin has been found to act on a fairly large range of cells (i.e., macrophages, thymus-derived cells, and probably B-lymphocytes) explains its ability to produce broad hyporesponsiveness.

Rapid onset of hyporesponsiveness to interferon induction on re-exposure to polyribonucleotide.

Only doses of complexed polyriboinosinic: polyribocytidylic acids sufficient to induce interferon, stimulated hyporesponsiveness to re-induction in rabbit kidney cells. Results of experiments designed to delineate the earliest time of appearance of hyporesponsiveness suggested the time of onset was less than i h after the end of a i h exposure to the inducer. The duration of hyporesponsiveness was about 24 h. Interferon itself did not produce hyporesponsiveness. Hence hyporesponsiveness must be related to an event or a substance synthesized before interferon production. The best candidate is a control protein inhibiting interferon production which is rapidly synthesized following exposure to an inducer.