Recombinant vector-induced HIV/SIV-specific CD4 T lymphocyte responses in rhesus monkeys

The recently reported modest success of the RV144 Thai trial vaccine regimen in preventing HIV-1 acquisition has focused interest on the potential contribution to that protection of vaccine-elicited CD4⁺ T cell responses. We evaluated the induction of virus-specific CD4⁺ T cell responses in rhesus monkeys using a series of diverse vaccine vectors. We assessed both the magnitudes and functional profiles of the antigen-specific CD4⁺ T cells by measuring cytokine production, memory differentiation, and the expression of mucosal homing molecules. We found that DNA prime/recombinant MVA boost immunizations induced particularly high-frequency virus-specific CD4⁺ T cell responses with polyfunctional repertoires, and these responses were partially preserved following SHIV-89.6P challenge. The majority of the vaccine-elicited CD4⁺ T cells were CD28⁺ memory T cells that expressed low levels of β7. Neither the magnitudes nor the functional profiles of the virus-specific CD4⁺ T cells generated by vaccination were associated with a preservation of CD4⁺ T cells or control of viral replication following SHIV-89.6P challenge. Interestingly, monkeys primed with recombinant Ad5 immunogens showed a dramatic expansion of both the magnitude and polyfunctionality of the vaccine-elicited CD4⁺ T cell responses following envelope protein boost. These results demonstrate that vaccine strategies that include recombinant MVA or recombinant Ad5 vectors can elicit robust CD4⁺ T cell responses.     

Limited dissemination of pathogenic SIV after vaginal challenge of rhesus monkeys immunized with a live, attenuated lentivirus

In non-human primate models of AIDS, attenuated lentiviruses provide the most reliable protection from challenge with pathogenic virus but the extent to which the vaccine virus replicates after challenge is unclear. At 7 and 14 days after vaginal challenge with pathogenic SIVmac239, plasma SIVenv RNA levels were significantly lower in female macaques immunized 6 months earlier with live, attenuated SHIV89.6 compared to unimmunized control animals. In 2 SHIV-immunized, unprotected macaques SIV replication produced moderate-level plasma viremia with dissemination of challenge virus to all tissues on day 14 after challenge. In protected, SHIV-immunized monkeys, SIV replication was controlled in all tissues, from the day of challenge through 14 days post-challenge. Further, in CD8⁺ T cell-depleted SHIV-immunized animals, SIV replication and dissemination were more rapid than in control animals. These findings suggest that replication of a pathogenic AIDS virus can be controlled at the site of mucosal inoculation by live-attenuated lentivirus immunization.     

Resistance to collagen-induced arthritis in rats and rhesus monkeys after immunization with attenuated type II collagen

Immunization of susceptible rodent or primate species with type II collagen (b-CII) from bovine origin induces type II collagen-induced arthritis (CIA). The disease is characterized as a systemic polyarthritis associated with humoral and cellular autoimmunity to CII and shares similarity with human arthritic diseases. The objective of this study was to develop a procedure for induction of resistance to CIA in animals, which possess a certain major histocompatibility complex phenotype that makes them prone to develop CIA (susceptible). It is shown that by immunization with an attenuated form of CII, in which arthritogenic epitopes have been destroyed by heat denaturation, disease resistance is induced in a susceptible inbred rat strain (RT-1^U) and in an outbred population of susceptible rhesus monkeys (lacking the Mamu-A26 allele). In both species the disease resistance is connected with modulation of anti-CII autoantibodies of the IgM isotype. This protocol may provide a basis for effective and safe methods to induce protection to autoimmune arthritis in those subjects that are genetically prone to develop such a disease.     

Enhanced in vitro stimulation of rhesus macaque dendritic cells for activation of SIV-specific T cell responses

The macaque-simian immunodeficiency virus (SIV) system is one of the best animal models available to study the role of dendritic cells (DCs) in transmission and pathogenesis of HIV, as well as to test DC-based vaccine and therapeutic strategies. To better define and optimize this system, the responsiveness of macaque monocyte-derived DCs to a variety of maturation stimuli was examined. Characteristic immunophenotypic and functional DC maturation induced by standard monocyte conditioned medium (MCM) was compared to the activation induced by a panel of stimuli including soluble CD40L, LPS, Poly I:C, PGE₂/TNF, and a cocktail mixture of PGE₂/TNF/IL-1β/IL-6. Immunophenotypic analysis confirmed that all stimuli induced stable up-regulation of CD25, CD40, CD80, CD83, CD86, HLA-DR, DC-LAMP (CD208), and DEC-205 (CD205). In general, macaque DCs exhibited weaker responses to LPS and Poly I:C than human DCs, and soluble CD40L stimulation induced variable expression of CD25. Interestingly, while the endocytic capacity of CD40L-matured cells was down-modulated comparably to DCs matured with MCM or the cocktail, the T cell stimulatory activity was not enhanced to the same extent. The particularly reproducible and potent T cell stimulatory capacity of cocktail-treated DCs correlated with a more homogenous mature DC phenotype, consistently high levels of IL-12 production, and better viability upon reculture compared to DCs activated by other stimuli. Furthermore, cocktail-matured DCs efficiently captured and presented inactivated SIV to SIV-primed T cells in vitro. Thus, the cocktail represents a particularly potent and useful stimulus for the generation of efficacious immunostimulatory macaque DCs.     

Immunogenicity of wild and attenuated varicella-zoster virus strains in rhesus monkeys

Thirty susceptible rhesus monkeys were inoculated with cell-free varicella-zoster virus strain OKA or strain KMcC. Both wild and attenuated strains were used. No clinical signs characteristic of human varicella were seen in any of the animals. Virus was not isolated from throat swabs, blood, or cerebrospinal fluid. Antibodies were measured by an enhanced plaque neutralization test. The wild and attenuated OKA strains produced comparable levels of antibodies for 3 months after inoculation. Attenuated KMcC strains produced lower titers than the wild strain. On rechallenge 3 months after primary inoculation animals boostered with the attenuated OKA strain developed significantly higher antibody titers than animals receiving the wild strain. Animals primed and challenged with the attenuated KMcC strains showed significantly lower antibody titers than animals which received the wild strain. The results indicate that the immunogenicity of attenuated OKA and KMcC strains in rhesus monkeys parallels the experience obtained with these strains in humans.     

Endocrine cells in the female genital tract

Endocrine cells are normal inhabitants of the para-urethral, Bartholin's and endocervical glands and of mesonephric rests. All these cells were characterized as serotonin-storing cells. In the para-urethral and Bartholin's glands, serotonin-containing cells were most often found in the transitional epithelium of excretory ducts. Endocrine cells participated in some pathological conditions. Abundant argentaffin cells were observed among the terminal ductules in chronic bartholinitis and serotonin-storing cells were identified in a peculiar ectocervical epithelium. Numerous serotonin-storing cells were detected in a well-differentiated adenocarcinoma of cervix occurring in a patient with the Peutz-Jeghers syndrome. Argyrophilic cells were present in cases of endometrial carcinomas; a striking feature was the demonstration of gut peptide hormones in an unusual type of endometrial adenocarcinoma. Finally, serotonin-storing cells were a constituent of Brenner tumours. It is suggested that a similar endocrine pattern may be shared by tissues originating from both Müllerian ducts and the urogenital sinus.     

灭活自体T细胞增强小鼠免疫功能作用机制的研究

接种灭活自身反应性T细胞用于治疗自身免疫性疾病已获初步成效。本研究采用辐射方法灭活ConA活化自体T细胞,经皮下和腹腔免疫正常的C57BL/6J小鼠,结果发现,该免疫方案可明显上调T细胞功能,表现为:免疫小鼠体内活化T细胞增多且Th1型细胞因子分泌增加,淋巴细胞体外增殖能力增强且凋亡细胞明显减少。进一步研究结果提示,接种灭活自体T细胞可能通过上调Fas、GADD45β及下调FasL基因表达而促进T细胞的活化和存活,并增加其生物学功能。这些结果表明,自体T细胞免疫具有增强机体的细胞免疫功能,为其抗肿瘤免疫作用提供了一定的实验依据。     

Localization of SIV in the genital tract of chronically infected female rhesus macaques.

Despite the fact that human immunodeficiency virus (HIV) is transmitted primarily by sexual contact, the biology of the sexual transmission of HIV is poorly understood. Simian immunodeficiency virus (SIV) can be transmitted to female rhesus macaques by placing cell-free virus into the vaginal canal, and SIV can be isolated from the vaginal secretions of infected rhesus macaques. The authors examined the genital tracts from 16 chronically infected female rhesus macaques and localized SIV-infected cells using in situ hybridization and immunohistochemistry. SIV-infected cells were found in the genital tract of 13 of the 16 animals examined, and in most cases the SIV-infected cells were located in the submucosa of the cervix and vagina. However, SIV-infected cells were also found in the vaginal epithelium. SIV-infected cells were more common in sites of inflammation than in normal areas. These findings suggest that SIV gains access to genital tract secretions from the cervix and vaginal epithelium.     

Specific antibody in the equine genital tract following systemic and local immunization.

An enzyme-linked immunosorbent assay (ELISA) was adapted for the horse to investigate the production of antibody in response to subcutaneous, intrauterine or intravaginal immunization with dinitro-phenylated human serum albumin. An IgM response was not detected, but antibodies of the IgGab, IgGc, IgGT and IgA isotypes were measured in serum, uterine and vaginal secretions. Local immunization produced antibody titres in serum and secretions, with evidence of significant local production.     

CD57 expression by T cells in the female genital tract of HIV-zx1 infected women

Despite an influx of T cells to the cervix during HIV infection, genital T cells are not associated with control of HIV shedding. CD57 expression by T cells has been associated with enhanced migratory potential and CD57+ T cells have been shown to accumulate in tissues during the late stages of HIV disease. We investigated the impact of HIV-infection and clinical status on the expression of CD57 by T cells from the female genital tract in 13 HIV-infected and 5 uninfected women. We found that cervical and blood-derived T cells expressed similar frequencies of CD57. The frequency of CD57 expression by cervical or blood T cells was not associated with clinical status (CD4 counts). No impairment in IFN-γ production by CD57+ T cells from the genital tract was observed. We conclude that increased T cell senescence does not appear to be a hallmark of genital mucosal HIV-1 infection.     

The development of pigment cells in the eyes of rhesus monkeys

The eyes of rhesus monkeys possess two lines of pigmented cells: those of the pigmented retinal epithelium and those of the uveal tract, also known as stromal pigmented cells. The cells of the pigmented epithelium differentiate first and are fully functional in 65-day old fetuses. By contrast, stromal pigmented cells do not become recognizable until late gestation (140 days). The differences in development and structure of these cells may reflect a different and perhaps distinct function.     

Aging affects the direction selectivity of MT cells in rhesus monkeys

The ability to accurately perceive the direction and speed of moving objects declines during normal aging. This is likely due to functional degradation of cortical neurons. Most neurons in the primate middle temporal area (MT) are direction-selective and their activity is closely linked to the perception of coherent motion. We investigated the mechanisms that underlie this age-related decline by comparing the proportions of direction-selective MT cells in old and young macaque monkeys, using in vivo single-cell recording techniques. Our results showed that the proportion of such cells was lower in old than in young monkeys. Moreover, one type of direction-sensitive cells, pattern cells, was especially sensitive to aging and was affected more severely than another class, component cells. We also found that direction selectivity was affected more severely in MT than in V1 of senescent monkeys. Thus, the functional degradation of MT and V1 cells may mediate perceptual decline in visual motion tasks in old primates.     

T and B cell populations in the peripheral blood of rhesus monkeys.

B and T cell populations in the peripheral blood of rhesus monkeys are comparable to those of normal human beings. B cells of monkeys, like those of human beings, also have receptors for IgM, IgG and IgA.     

Induction of antibody response to Chlamydia trachomatis in the genital tract by oral immunization.

Groups of BALB/c mice were orally immunized with Chlamydia trachomatis serovar L2/434/Bu in order to characterize the nature and kinetics of the chlamydial antibody response in the cervix and other mucosal sites. These animals were subsequently challenged intravaginally to determine whether oral immunization offers protection against chlamydial antigen shedding in the genital tract. Following oral immunization, immunoglobulin A antibody activity was detected in the genital tract as well as other mucosal sites. Subsequent intravaginal challenges exhibited booster effects on preexisting antibody activity in the genital tract. Significant protection against challenge infection in the genital tract was observed by oral immunization. This was indicated by the absence of any chlamydial antigen shedding in cervical secretions. On the other hand, passively administered chlamydial-specific serum immunoglobulin G antibody did not significantly influence the course of cervical shedding of the organism and did not confer any protection against a subsequent intravaginal challenge. It is concluded that prior oral immunization can induce a secretory antibody response in the genital tract and provide protection against subsequent infection.     

Fine structure of adrenocortical cells in adult male rhesus monkeys

Adrenals of ten mature male rhesus monkeys were studied by electron microscopy. Mitochondria had lamelliform cristae in the zona glomerulosa and tubular cristae in the zona fasciculata and zona reticularis. Agranular reticulum of tubular form was scarce in the zona glomerulosa but present in great abundance in the fasciculata and reticularis. The Golgi complex was prominent in the cells of the zona glomerulosa but poorly developed in the cells of the inner cortical zones. Zona fasciculata cells contained large, highly ordered “stacks” of granular endoplasmic reticulum which showed multiple connections with the random network of tubular agranular reticulum. In the reticularis, granular endoplasmic reticulum was present in more disordered, whorl-like arrays. Lipid droplets in the inner cortical regions were enveloped by tubules of agranular endoplasmic reticulum. Occasional sections showed these tubules ending blindly at or near the surface of the lipid droplets. This association was strongly suggestive of a functional relationship. Dense bodies varied in size and complexity of structure throughout the cortex. They were smallest and simplest in form in the zona glomerulosa; those in the zona reticularis were the largest and most complex.     

Experimental infection of the genital tract of female grivet monkeys by Mycoplasma hominis.

Mycoplasma hominis, a common inhabitant of the mucosae of the genitourinary tract of human and nonhuman primates, was inoculated directly into the uterine tubes of five laparotomized grivet monkeys. A self-limiting acute salpingitis and parametritis developed within a few days in all animals. Although there were no clinical signs of overt disease, the gross pathology was characterized by pronounced oedematous swelling and hyperaemia of the tubes and parametria. Microscopically, cellular infiltrations of lymphocytes and some polymorphonuclear leukocytes were found in the acute phase in the subserosa and muscularis of the tubes and in the parametria. Granulation tissue and fat necrosis appeared at a later stage in the parametria. The infection was associated with a marked antibody response and a moderate rise of the erythrocyte sedimentation rate and leukocyte counts. The capability of M. hominis to produce salpingitis and parametritis in a nonhuman primate would seem to add rather significantly to the available evidence suggesting an etiological role of this organism in inflammatory diseases of the internal female genitals of humans.     

Susceptibility to reinfection after a primary chlamydial genital infection is associated with a decrease of antigen-specific T cells in the genital tract.

We tested the hypothesis that the intensity of specific antichlamydial T cell-mediated immunity in the genital tract of female guinea pigs infected intravaginally with the chlamydial agent of guinea pig inclusion conjunctivitis would determine the resistance or susceptibility to reinfection after a primary chlamydial infection. T cell-enriched lymphocytes were isolated by collagenase treatment of genital tract tissues from either infected or control uninfected female guinea pigs at various times after infection. The nylon wool-enriched T lymphocytes were evaluated for expression of antigen-specific T cell-mediated immunity in vitro by using a blast transformation assay. Both uninfected and infected genital tracts contained T cells, as evidenced by reactivity to concanavalin A, although a greater number of T lymphocytes was detected in the genital tracts of infected animals compared with that in controls. Significant antigen-specific T-cell activity could be detected in the genital tract tissue by 7 days after a primary genital tract infection with the chlamydial agent of guinea pig inclusion conjunctivitis. When antigen-specific activity was assessed at different times after infection, the intensity of the response of genital tract-associated T lymphocytes was directly proportional to the degree of resistance of the animals to genital challenge. Thus, susceptibility of animals to reinfection by chlamydiae appears to be associated with the intensity of the local T cell-mediated immune responses in the genital tract of infected animals.     

Activation of a myelin basic protein-specific human T cell clone by antigen-presenting cells from rhesus monkeys

This study addresses the capacity of peripheral blood mononuclearcells (PBMC) from rhesus monkeys (Macaca mulatta) to presentmyelin basic protein (MBP), a candidate auto-antigen for multiplesclerosis, to MBP-speclfic human CD4⁺ T cell clones. MHC-restrictlonof the human T cell clones was determined with HLA-DR-transfectedL cells, and epitope specificity was established with a panelof overlapping 20-mer peptides. The MHC-DR region of the rhesusmonkeys (Mamu) was characterized serologlcally and by sequenceanalysis. We identified one CD4⁺ HLADRB1*0301-restrlcted T_h1-llkehuman T cell clone (ES-BP8) that was activated to proliferationwith human or rhesus monkey MBP, or peptide MBP 29–48presented by PBMC from six different rhesus monkeys expressingthe Mamu-DRB1*0305 or -DRB1*0306 alleles. After transformationto continuous growth with Herpesvirus salmiri, the T cell clonecould still be stimulated by antigen (Ag) and Ag-presentlngcells (APC) from monkeys. Two other T cell clones with the sameHLA-restriction and the same peptide-specificity did not respondto MBP presented by these rhesus monkeys. The exon 2 sequencesHLA-DRB1*0301, Mamu-DRBV^*0305 and -DRB 1* 0306 differ at positions32, 47, 67, 73 and 86. These amino acid differences are notcritical for the binding of MBP 29–48 and do not abrogate-recognitionby the clone ES-BP8, but interfere with the recognition of thetwo other HLA-DRB1^*0301-restricted T cell clones. In conclusion,studying Ag-presentation from rhesus monkey may provide furtherinsight Into the Interaction of antigenic peptide, TCR and MHC.Furthermore, these results could form a useful basis for theIn vivo transfer of human auto-Ag-specific T cells into rhesusmonkey recipients.     

Influence of gonadal hormones and genital afferents on EEG activity of the hypothalamus in adult male rhesus monkeys

Electrodes were stereotaxically implanted in hypothalamic and cerebral cortical regions of 15 adult male gonadally intact rhesus monkeys. The electrical activity of these regions was recorded electroencephalographically before and after genital stimulation. Observations were also taken after stimulation on skin, abdomen and nipples and repeated daily for 4 to 6 days, before and after intravenous injections of testosterone propionate (0.4 mg/kg). The experiment was repeated on the same animal after gonadectomy. The influence of gential afferents appears to produce focal inhibition of electroencephalographic (EEG) activity in ventromedial nucleus (VMN) in monkeys with intact gonads. Exogenous administration of testosterone propionate produces EEG facilitation in VMN with simultaneous inhibition of the anterior hypothalamic area. With continued increase in amounts of circulating hormone, the EEG activity in the posterior hypothalamic area is also inhibited by the incoming genital afferent information. Gonadectomy brings about EEG slowing with genital stimulation in ventromedial nucleus before hormone injection but this was less marked than that observed in EEG records of intact preparation.