Inhibition of cell-associated herpes simplex virus type 2 glycoproteins by delta 9-tetrahydrocannabinol

This study was conducted to define the effect of micromolar concentrations of delta 9-tetrahydrocannabinol (delta 9-THC) on the biosynthesis and expression of herpes simplex virus type 2 (HSV2)-specified glycoproteins. Dose-related reductions in all species of virus glycoproteins were recorded by one-dimensional SDS-polyacrylamide gel electrophoresis (SDS-PAGE) and autoradiography of [14C]glucosamine-labeled infected Vero cells treated with 10(-7) to 10(-5) M delta 9-THC. A drug dose-related depletion of the mature HSV2 major envelope glycoprotein complex (119-kDa average molecular weight), accompanied by accumulation of immature unglycosylated species, was demonstrated by two-dimensional SDS-PAGE in concert with Western immunoblotting or autoradiography. Light and electron microscopy immunoperoxidase staining revealed that delta 9-THC effected depletion of 119-kDa determinants from the infected cell surface. This depletion occurred concomitantly with accumulation of 119-kDa components at the perinucleus. However, the expression of 119-kDa glycoproteins on the virion envelope was not affected. These results indicate that delta 9-THC inhibits the synthesis, maturation, and cellular transport of HSV2-specified glycoproteins. Decreased expression of virus glycoproteins on the infected cell surface may affect host immune responsiveness to HSV2.     

Thymidine kinase-deficient herpes simplex virus type 2 genital infection in guinea pigs.

In guinea pigs, thymidine kinase-producing strains of herpes simplex virus type 2 replicated to high titer in the vagina and spinal cord, and animals developed severe clinical disease. Infection with thymidine kinase-deficient virus resulted in similar vaginal virus titers; however, animals exhibited little or no clinical illness and only low titers of virus were detected in spinal cord homogenate cultures. Neural and extraneural latent infection as well as recurrent infection were noted in animals inoculated with either thymidine kinase-producing or -deficient viruses. These data suggest that neural pathways are important in the pathogenesis of genital herpes and that virus-coded thymidine kinase may influence virulence but is not required for latency.     

Suppressive effect of delta-9-tetrahydrocannabinol on herpes simplex virus infectivity in vitro.

Delta-9-Tetrahydrocannabinol (THC) was found to reduce the infectivity of herpes simplex virus and was without effect against adenovirus type 2 or poliovirus. The effective THC concentration resulting in an 80% decrement in virus viability was dependent upon the presence or absence of serum in the incubation mixture, as a 5% serum concentration decreased the drug activity by approximately 50-fold. THC-mediated inactivation of herpes simplex virus was both time and dose dependent and did not result in virion disassembly or clumping. The THC-related effect was not influenced by the pH of the suspending medium, suggesting that the mechanism of inactivation differed from that associated with the thermal inactivation of the virus. Thus, the data suggest that THC preferentially reduces the infectivity of the enveloped herpes simplex virus, and that this activity is modulated by the presence of serum proteins.     

The role of dendritic cells in immune responses against vaginal infection by herpes simplex virus type 2

Herpes simplex virus type 2 is a leading cause of genital ulcers that affects more women than men worldwide. Recent evidence indicates that protective immunity can be generated by specialized dendritic cells in the female genital mucosa. This article aims to provide an overview of the effector immunity required for protection from genital herpes, and to discuss the mechanism by which specific subsets of dendritic cells mediate induction of adaptive immunity following genital infection with herpes simplex virus type 2 in vivo.     

Delta 9-tetrahydrocannabinol decreases cytotoxic T lymphocyte activity to herpes simplex virus type 1-infected cells.

The purpose of this study was to examine the effect of delta 9-tetrahydrocannabinol (delta 9-THC), the major psychoactive component of marijuana, on T lymphocyte functional competence against herpes simplex virus Type 1 (HSV1) infection. Spleen cells from C3H/HeJ (H-2k) mice primed with HSV1 and exposed to delta 9-THC were examined for anti-HSV1 cytolytic T lymphocyte (CTL) activity. Flow cytometry was used to determine whether delta 9-THC altered T cytotoxic (Lyt-2+) and T helper (L3T4+) lymphocyte numbers or cell ratios. Nomarski optics microscopy was used to determine whether effector lymphocytes from drug-treated mice were able to bind to virally infected L929 (H-2k) target cells. Cytotoxicity assays demonstrated that CTL from mice exposed to delta 9-THC were deficient in anti-HSV1 cytolytic activity. delta 9-THC in vivo treatment had little effect on the number of T lymphocytes expressing the Lyt-2 or L3T4 antigens. Nomarski optics microscopy revealed that the CTL from the drug-treated mice were able to bind specifically to the HSV1-infected targets. However, delta 9-THC in vivo exposure affected CTL cytoplasmic polarization toward the virus-infected target cell. CTL granule reorientation toward the effector cell-target cell interface following cell conjugation occurred at a lower frequency in co-cultures containing CTL from drug-treated mice. These results suggest that delta 9-THC elicits dysfunction in CTL by altering effector cell-target cell postconjugation events.     

Ribonucleotide reductase of herpes simplex virus type 2 resembles that of herpes simplex virus type 1.

The ribonucleotide reductase (ribonucleoside-diphosphate reductase; EC 1.17.4.1) induced by herpes simplex virus type 2 infection of serum-starved BHK-21 cells was purified to provide a preparation practically free of both eucaryotic ribonucleotide reductase and contaminating enzymes that could significantly deplete the substrates. Certain key properties of the herpes simplex virus type 2 ribonucleotide reductase were examined to define the extent to which it resembled the herpes simplex virus type 1 ribonucleotide reductase. The herpes simplex virus type 2 ribonucleotide reductase was inhibited by ATP and MgCl2 but only weakly inhibited by the ATP X Mg complex. Deoxynucleoside triphosphates were at best only weak inhibitors of this enzyme. ADP was a competitive inhibitor (K'i, 11 microM) of CDP reduction (K'm, 0.5 microM), and CDP was a competitive inhibitor (K'i, 0.4 microM) of ADP reduction (K'm, 8 microM). These key properties closely resemble those observed for similarly purified herpes simplex virus type 1 ribonucleotide reductase and serve to distinguish these virally induced enzymes from other ribonucleotide reductases.     

Pathogenesis of herpes simplex virus type 2 experimental genital infection in pregnant mice

The progression of herpes simplex-2 genital infection in pregnant mice was studied by detection of viral antigens using immunoperoxidase in tissue sections, electron microscopy and virus isolation. The majority of mice (66.66%) died at 8-9 days post-inoculation. Abortions were observed in 69.23% of the infected mice along with impairment of labor and delivery. Herpes antigens were detected in most of the autonomic nerves of the uterus, including those surrounding small arterioles in the myometrium and the Auerbach and Meissner plexa of the large bowel, but not in the abortions or placentas. The infection of uterine autonomic fibers and myometrial cells could explain the delivery impairment and could have provoked a decrease in blood flow leading to abortions.     

Regulation by recombinant interleukin-2 of protective immunity against recurrent herpes simplex virus type 2 genital infection in guinea pigs.

The goal of our study was to determine whether recombinant interleukin-2 (rIL-2) could modify the recurrence pattern of chronic herpes simplex virus type 2 (HSV-2) genital infection in guinea pigs. Animals that developed symptomatic acute HSV-2 infection were distributed at 14 days after viral inoculation into several treatment groups, which were similar with respect to the severity of acute disease. Three rIL-2 dosages administered for 4 weeks in daily subcutaneous injections were tested in this study: 5 X 10(3), 5 X 10(4), and 2.5 X 10(5) U. Daily observations of the animals showed a significant decrease of the incidence of new recurrent lesions with the use of 5 X 10(4) U of rIL-2 (rate of recurrence, 0.08, compared with 0.21 in untreated controls), whereas the other rIL-2 regimens did not affect the overall rate of recurrence. Weekly analysis of recurrences showed that treatment with 5 X 10(4) U of rIL-2 was effective only during the first 3 weeks of use and that 2.5 X 10(5) U of rIL-2 markedly decreased the rate of recurrence in the first week of treatment but not in subsequent weeks. The loss of clinical protection in both groups coincided with the production of neutralizing antibodies to rIL-2. The immune mechanisms possibly involved in the protective effect of rIL-2 in chronic HSV-2 disease were further investigated. Production of gamma interferon correlated well with clinical protection, and circulating levels dropped at the time when neutralizing antibodies to rIL-2 developed. Nonspecific cytotoxicity represented by natural killer cell and lymphokine-activated killer cell activities was also increased in the treated guinea pigs. Antibody titers and lymphocyte proliferation to herpes simplex antigen were similar in rIL-2 and placebo recipients. Finally, we found that the rIL-2-induced immune stimulation was as protective against recurrent HSV-2 disease in guinea pigs as the viral suppression achieved with acyclovir. However, the biological activity of both drugs was not additive when they were coadministered.     

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.     

Stereospecific presynaptic inhibitory effect of delta9-tetrahydrocannabinol on cholinergic transmission in the myenteric plexus of the guinea pig.

delta9-Tetrahydrocannabinol (THC) is very lipid soluble, as are many anesthetic agents. The action of anesthetics is nonspecific; isomers are equieffective. THC is optically active; therefore, the effects of its stereoisomers were studied on the electrically and chemically stimulated longitudinal muscle strip of guinea pig ileum. The results demonstrate that both isomers depress the response to electrical stimulation in a dose-related manner. The maximum effect is gradually reached in approximately 20 min. The (-) isomer is 24.6-fold more active than the (+) isomer (ED50 for (-) THC is 1.25 X 10(-7) M, for (+) THC, 3.08 X 10(-6) M) and the site of action appears to be presynaptic because responses to ACh are not significantly depressed. The depressant effects are relatively nonreversible. Membrane concentrations calculated at the ED50 values for the (-) isomer are of the order of 0.5 mM/kg dry membrane, well within the range for anesthesia. Thus THC may be regarded as a partial anesthetic since some of its actions are similar to those of the classical anesthetics, yet it possesses selective action at the neuronal membrane or tissue level.     

Reinfection of guinea pigs with herpes simplex virus type 2: failure to establish a second latent ganglionic infection

Guinea pigs were infected with herpes simplex virus type 2 (HSV 2) and at various times thereafter reinfected at a different site with the same or a different strain of HSV 2. Viruses were isolated from infected tissues either from the site of primary or secondary infection or from regional ganglia. Viral isolates were characterised by DNA fingerprinting using restriction endonucleases capable of differentiating between the virus strains used. The second infection resulted in a second site of peripheral persistence of the virus, but did not establish a second site of ganglionic infection.     

Delta 9-tetrahydrocannabinol decreases alpha/beta interferon response to herpes simplex virus type 2 in the B6C3F1 mouse

This study was undertaken to determine the effect of delta 9-tetrahydrocannabinol (delta 9-THC) on polyinosinic:polycytidylic acid [poly(I):poly(C)]-induced, and on herpes simplex virus type 2 (HSV-2)-induced, alpha/beta interferon in the B6C3F1 mouse. Animals were administered delta 9-THC, or the diluent, intraperitoneally for 4 consecutive days or at various time intervals prior to administration of the interferon inducer. Poly(I):poly(C) or HSV-2 was injected intravenously on Day 4. Animals receiving poly(I):poly(C) and treated with delta 9-THC at doses ranging from 5 to 100 mg/kg exhibited significantly lower titers of interferon than mice given poly(I):poly(C) and the diluent. Diminished interferon titers occurred in HSV-2-infected animals treated with delta 9-THC in doses exceeding 15 mg/kg when compared to virus-infected animals given the diluent. This suppression of early interferon persisted through 24 hr.     

Effect of phosphorothioate homo-oligodeoxynucleotides on herpes simplex virus type 2-induced DNA polymerase

Effects of phosphorothioate oligodeoxynucleotides of different chain length and base composition on herpes simplex virus (HSV) type 2 (strain 333)-induced DNA polymerase have been examined in vitro. The anti-HSV-2 DNA polymerase activity was related to the base composition of the analogs, with the order of potency: deoxycytidine greater than thymidine greater than deoxyadenosine, for compounds with equal chain length. The potency was also related to oligomer chain length, since it was observed that the longer the chain length, the more potent the inhibition exerted. Among all the compounds tested, the phosphorothioate oligodeoxycytidine 28-mer (S-(dC)28) was the most potent inhibitor of HSV-2-induced DNA polymerase. This inhibition was competitive with an activated DNA template with a Ki value of 7 nM. It was also a competitive inhibitor of the DNA polymerase-associated exonuclease activity with a Ki value of 5 nM. In contrast, this compound showed less inhibition of human DNA polymerase alpha, beta, and gamma, as well as HSV-1 (strain KOS) and Epstein-Barr virus-induced DNA polymerase. The possibility that S-oligomers can serve as primers for DNA elongation was also investigated. Poly(dG).S-(dC)28 and poly(dA).S-(T)28 are poor substrates for DNA elongation catalyzed by HSV-2 DNA polymerase. In summary, phosphorothioate oligonucleotides could be anti-template inhibitors of HSV DNA polymerase. This information may lead to the development of a new class of selective anti-HSV agents.     

单纯疱疹病毒2型疫苗的研究进展

单纯疱疹病毒2型(herpes simplex virus type 2,HSV-2)是人类疱疹病毒家族中α家族成员,可引起多年龄段的人群生殖器疱疹及其他疱疹疾病。由于该病毒具有复杂的基因结构及感染病理,其感染至今尚无有效的治疗方法。该病毒可抑制机体免疫系统,在神经细胞潜伏感染,具有重激活特性,因此,其疫苗的研制亦需要复杂的技术,且面临重重的挑战。