Recovery from chronic rotavirus infection in mice with severe combined immunodeficiency: virus clearance mediated by adoptive transfer of immune CD8+ T lymphocytes.

Severe combined immunodeficient (SCID) mice lack both functional T and B cells. These mice develop chronic rotavirus infection following an oral inoculation with the epizootic diarrhea of infant mice (EDIM) rotavirus. Reconstitution of rotavirus-infected SCID mice with T lymphocytes from immunocompetent mice allows an evaluation of a role of T-cell-mediated immunity in clearing chronic rotavirus infection. Complete rotavirus clearance was demonstrated in C.B-17/scid mice 7 to 9 days after the transfer of immune CD8+ splenic T lymphocytes from histocompatible BALB/c mice previously immunized intraperitoneally with the EDIM-w strain of murine rotavirus. The virus clearance mediated by T-cell transfer was restricted to H-2d-bearing T cells and occurred in the absence of rotavirus-specific antibody as determined by enzyme-linked immunosorbent assay, neutralization, immunohistochemistry, and radioimmunoprecipitation. Temporary clearance of rotavirus was observed after the transfer of immune CD8+ T cells isolated from the intestinal mucosa (intraepithelial lymphocytes [IELs]) or the spleens of BALB/c mice previously infected with EDIM by the oral route. Chronic virus shedding was transiently eliminated 7 to 11 days after spleen cell transfer and 11 to 12 days after IEL transfer. However, recurrence of rotavirus infection was detected 1 to 8 days later in all but one SCID recipient receiving cells from orally immunized donors. The viral clearance was mediated by IELs that were both Thy1+ and CD8+. These data demonstrated that the clearance of chronic rotavirus infection in SCID mice can be mediated by immune CD8+ T lymphocytes and that this clearance can occur in the absence of virus-specific antibodies.     

Human peripheral blood leukocyte engraftment into SCID mice: critical role of CD4(+) T cells

We examined the influence of donor T lymphocytes on human peripheral blood leukocytes (PBL) engraftment into severe combined immune deficient (SCID) mice. Mice were injected with unfractionated or subset-depleted human PBL, and treated at various times with OKT3, a cytotoxic monoclonal antibody against human CD3(+) T lymphocytes. PBL engraftment, high levels of human Ig, and high incidence of lymphoproliferative disease (lpd) were found in mice transplanted with unfractionated PBL and CD8- or CD14-depleted PBL, and in mice treated with OKT3 at distance from PBL transfer. Animals xenografted with CD3- or CD4-depleted PBL, or treated at transplantation time with OKT3, had very low levels of human Ig and did not develop lpd. PBL engraftment was minimal or absent in these animals, as determined by immunohistochemistry, dot-blot, and RT-PCR analyses. These results demonstrate that the presence of donor CD4(+) T lymphocytes at transplantation time is necessary for observing human PBL engraftment into SCID mice, an essential condition for human Ig production and lpd development.     

Herpes simplex virus-induced stromal keratitis: role of T-lymphocyte subsets in immunopathology.

Herpetic stromal keratitis (SK), a frequent cause of visual impairment, is considered to represent an immune-mediated inflammatory response to persistent herpes simplex virus virions or subcomponents within the corneal stroma. The experimental disease in mice involves the essential participation of T lymphocytes, but the role of T-lymphocyte subsets in either mediating or controlling the disease is uncertain. In this report, rat monoclonal antibodies were used to selectively deplete mice in vivo of CD4+ (helper-inducer) and CD8+ (cytotoxic-suppressor) T-cell populations and the effect on herpetic SK was evaluated. As measured by flow cytometry, mice treated with anti-CD4 monoclonal antibody (GK 1.5) were greater than 95% depleted of CD4+ T lymphocytes and mice treated with anti-CD8 monoclonal antibody (2.43) were 90% depleted of CD8+ T lymphocytes. Depleted and nonspecific mouse ascites-treated control mice were infected topically on the corneas with herpes simplex virus type 1, and the induction of various immune parameters during the acute infection was evaluated. CD4+-depleted mice failed to produce either a significant antiviral antibody or delayed-type hypersensitivity response but were capable of producing normal cytotoxic T-lymphocyte responses. In contrast, CD8+-depleted mice produced antiviral antibody and delayed-type hypersensitivity responses comparable with those in control animals, but cytotoxic T-lymphocyte responses were markedly reduced. Clinical observations of the corneas revealed that SK in CD4+-depleted mice was significantly reduced, whereas in CD8+-depleted mice SK developed more rapidly, was more severe, and involved a greater percentage of mice. These observations implicate the CD4+ T-lymphocyte subset as the principal mediators of SK and CD8+ T lymphocytes as possible regulators that control the severity of SK.     

Rotavirus-specific cytotoxic T lymphocytes appear at the intestinal mucosal surface after rotavirus infection.

The gastrointestinal tract is constantly exposed to a variety of potentially invasive bacteria and viruses. The first line of defense of the host against these pathogens is the intestinal mucosal surface, which consists of epithelial cells, intraepithelial lymphocytes (IELs), mucus, and secretory immunoglobulins. Little is known about the function, memory, or trafficking of IELs after intestinal infection. We found that IELs obtained 6 days after oral inoculation of mice with the intestinal pathogen rotavirus (simian strain RRV) lysed rotavirus-infected target cells; cytotoxic T lymphocytes (CTLs) were responsible for rotavirus-specific cytotoxic activity. Rotavirus-specific cytotoxic activity by IELs was (i) eliminated by treatment with Thy 1.2-specific immunoglobulin M plus complement, (ii) restricted by proteins encoded at the major histocompatibility complex, and (iii) absent in mock-infected animals. Oral inoculation of mice with RRV also induced rotavirus-specific CTLs in splenic and intestinal lymphocytes (mesenteric lymph nodes, Peyer's patch). Parenteral inoculation induced rotavirus-specific CTLs in splenic, intestinal (IELs, mesenteric lymph nodes, Peyer's patch), and nonintestinal lymphocytes (inguinal nodes). Therefore, presentation of rotavirus to the intestinal mucosal surface was not necessary to induce IELs with virus-specific cytotoxic activity. At 4 weeks after oral or parenteral inoculation of mice with RRV, rotavirus-specific CTL precursors appeared among splenic, Peyer's patch, inguinal, and mesenteric node lymphocytes, but not among IELs. IELs with rotavirus-specific cytotoxic activity may be generated from precursors at a site other than the intestinal mucosal surface. Part of the response of the host to enteric infection may include surveillance and lysis of virus-infected villus epithelial cells by IELs.     

Immunogene therapy of tumors with vaccine based on xenogeneic epidermal growth factor receptor

The breaking of immune tolerance against self epidermal growth factor receptor (EGFr) should be a useful approach for the treatment of receptor-positive tumors with active immunization. To test this concept, we constructed a plasmid DNA encoding extracellular domain of xenogeneic (human) EGFr (hEe-p) or corresponding control mouse EGFr (mEe-p) and empty vector (c-p). Mice immunized with hEe-p showed both protective and therapeutic antitumor activity against EGFr-positive tumor. Sera isolated from the hEe-p-immunized mice exhibited positive staining for EGFr-positive tumor cells in flow cytometric analysis and recognized a single 170-kDa band in Western blot analysis. Ig subclasses responded to rEGFr proteins were elevated in IgG1, Ig2a, and Ig2b. There was the deposition of IgG on the tumor cells. Adoptive transfer of the purified Igs showed the antitumor activity. The increased killing activity of CTL against EGFr-positive tumor cells could be blocked by anti-CD8 or anti-MHC class I mAb. In vivo depletion of CD4(+) T lymphocytes could completely abrogate the antitumor activity, whereas the depletion of CD8(+) cells showed partial abrogation. The adoptive transfer of CD4-depleted (CD8(+)) or CD8-depleted (CD4(+)) T lymphocytes isolated from mice immunized with hEe-p vaccine showed the antitumor activity. In addition, the increase in level of both IFN-gamma and IL-4 was found. Taken together, these findings may provide a new vaccine strategy for the treatment of EGFr-positive tumors through the induction of the autoimmune response against EGFr in a cross-reaction between the xenogeneic homologous and self EGFr.     

Rotavirus-specific cytotoxic T lymphocytes cross-react with target cells infected with different rotavirus serotypes.

Splenocytes from adult C57BL6 (H-2b) mice orally inoculated with nonmurine rotaviruses lysed syngeneic rotavirus-infected target cells. Cytotoxic T lymphocytes (CTLs) were responsible for this cytotoxic activity. Cytotoxic activity was (i) detected 7 days after primary oral inoculation; (ii) not detected in uninoculated animals; (iii) specific for rotavirus-infected target cells; (iv) eliminated by treatment with Thy 1.2-specific immunoglobulin M and complement; and (v) restricted at H-2Db. In addition, rotavirus-specific CTLs cross-reacted with target cells infected with different human or animal rotavirus serotypes. Heterotypic protection against rotavirus challenge may be mediated by cross-reactive CTLs.     

IFN-gamma and CD8+ T cells restore host defenses against Pneumocystis carinii in mice depleted of CD4+ T cells

Host defenses against infection are profoundly compromised in HIV-infected hosts due to progressive depletion of CD4+ T lymphocytes and defective cell-mediated immunity. Although recent advances in antiretroviral therapy can dramatically lower HIV viral load, blood CD4+ T lymphocytes are not restored to normal levels. Therefore, we investigated mechanisms of host defense other than those involving CD4+ T lymphocytes against a common HIV-related opportunistic infection, Pneumocystis carinii (PC) pneumonia. Using CD4-depleted mice, which are permissive for chronic PC infection, we show that up-regulation of murine IFN-gamma by gene transfer into the lung tissue results in clearance of PC from the lungs in the absence of CD4+ lymphocytes. This resolution of infection was associated with a >4-fold increase in recruited CD8+ T lymphocytes and NK cells into the lungs. The role of CD8+ T cells as effector cells in this model was further confirmed by a lack of an effect of IFN-gamma gene transfer in scid mice or mice depleted of both CD4+ and CD8+ T cells. Cytokine mRNA analysis revealed that recruited, lung-derived CD8+ T cells had greater expression of IFN-gamma message in animals treated with the IFN-gamma gene. These results indicate that CD8+ T cells are capable of clearing PC pneumonia in the absence of CD4+ T cells and that this host defense function of CD8+ T cells, as well as their cytokine repertoire, can be up-regulated through cytokine gene transfer.     

T hybridoma alpha/beta gene transfected in a murine T cell hybridoma: role of CD4 molecule in vitro and in vivo--engraftment in SCID mice induces T cell maturation.

In the present work, we tested in SCID and Balb/c mice the activity of T hybridoma transfected with T cell receptor (TCR) alpha/beta chain genes. A T cell hybridoma denoted D011107 was used as recipient for transfection of cytotoxic KB5C20 TCR alpha/beta heterodimer genes by protoplast fusion or electroporation. After transfection, the parental D011107 T cell line reexpressed CD5 and CD4 surface molecules. In vitro, we noted strong proliferation and unusual cytotoxic reactivities against H-2k target cells although the transfected cell line does not express the CD8 molecule. The fate of parental and transfected cells was examined in severe combined immunodeficient (SCID) and Balb/c mice at Day 16 after intravenous injection. Cells from bone marrow, thymus, and spleen tissues were analyzed by immunofluorescence. The transfected T cell hybridoma was CD3+ Desire 1+ CD4+ Thy1.2. The SCID mice grafted with the transfected T cell hybridoma presented a high percentage of CD3+ (15%), CD4+ (27%), Thy1.2+ (27.52%), and Desire 1+ (8.74%) cells in the spleen. The percentages of CD3+ (6.2%) and Thy1.2+ (5.06%) cells in the spleen from SCID mice grafted with parental T cell D011107 and from untreated SCID were similar and lower (CD3+, 3.52%; Thy1.2+, 4.34%). It seems that transfected T cells hybridoma grafted in the SCID mice induce significant expression of CD4+ Thy1.2+ Desire 1- cells (17%) in the spleen. These results indicate that transfected T cells graft may allow T cell differentiation. In Balb/c mice, the percentage of different T cell subsets in bone marrow, thymus, or spleen cells in mice injected with transfected T cells was similar to that in untreated mice. We did not observe any cytotoxic or significant allogeneic proliferation in vitro.     

Protective cytotoxic T-cell responses induced by venezuelan equine encephalitis virus replicons expressing Ebola virus proteins

Infection with Ebola virus causes a severe disease accompanied by high mortality rates, and there are no licensed vaccines or therapies available for human use. Filovirus vaccine research efforts still need to determine the roles of humoral and cell-mediated immune responses in protection from Ebola virus infection. Previous studies indicated that exposure to Ebola virus proteins expressed from packaged Venezuelan equine encephalitis virus replicons elicited protective immunity in mice and that antibody-mediated protection could only be demonstrated after vaccination against the glycoprotein. In this study, the murine CD8(+) T-cell responses to six Ebola virus proteins were examined. CD8(+) T cells specific for Ebola virus glycoprotein, nucleoprotein, and viral proteins (VP24, VP30, VP35, and VP40) were identified by intracellular cytokine assays using splenocytes from vaccinated mice. The cells were expanded by restimulation with peptides and demonstrated cytolytic activity. Adoptive transfer of the CD8(+) cytotoxic T cells protected filovirus na?ve mice from challenge with Ebola virus. These data support a role for CD8(+) cytotoxic T cells as part of a protective mechanism induced by vaccination against six Ebola virus proteins and provide additional evidence that cytotoxic T-cell responses can contribute to protection from filovirus infections.     

Induction of humoral and CD8+ T cell responses are required for protection against lethal Ebola virus infection

Ebola virus (EBOV)-like particles (eVLP), composed of the EBOV glycoprotein and matrix viral protein (VP)40 with a lipid membrane, are a highly efficacious method of immunization against EBOV infection. The exact requirements for immunity against EBOV infection are poorly defined at this time. The goal of this work was to determine the requirements for EBOV immunity following eVLP vaccination. Vaccination of BALB/c or C57BL/6 mice with eVLPs in conjunction with QS-21 adjuvant resulted in mixed IgG subclass responses, a Th1-like memory cytokine response, and protection from lethal EBOV challenge. Further, this vaccination schedule led to the generation of both CD4(+) and CD8(+) IFN-gamma(+) T cells recognizing specific peptides within glycoprotein and VP40. The transfer of both serum and splenocytes, but not serum or splenocytes alone, from eVLP-vaccinated mice conferred protection against lethal EBOV infection in these studies. B cells were required for eVLP-mediated immunity to EBOV because B cell-deficient mice vaccinated with eVLPs were not protected from lethal EBOV challenge. We also found that CD8(+), but not CD4(+), T cells are absolutely required for eVLP-mediated protection against EBOV infection. Further, eVLP-induced protective mechanisms were perforin-independent, but IFN-gamma-dependent. Taken together, both EBOV-specific humoral and cytotoxic CD8(+) T cell responses are critical to mediate protection against filoviruses following eVLP vaccination.     

Anti-glycoprotein D monoclonal antibody protects against herpes simplex virus type 1-induced diseases in mice functionally depleted of selected T-cell subsets or asialo GM1+ cells.

Passive transfer of a monoclonal antibody (MAb) specific for glycoprotein D (gD) is highly effective in preventing the development of herpes simplex virus type 1-induced stromal keratitis. In the present study, we investigated whether animals which had been functionally depleted of T-cell subsets or asialo GM1+ cells would continue to be responsive to MAb therapy. BALB/c mice were depleted of CD4+, CD8+, or asialo GM1+ cells by treatment with anti-L3T4, anti-Lyt 2.2, or anti-asialo GM1 antibodies, respectively. Functional depletion of CD4+ cells was documented by the loss of delayed-type hypersensitivity responsiveness, while CD8+ cell depletion was accompanied by abrogation of cytotoxic lymphocyte activity. Anti-asialo GM1 treatment led to the loss of natural killer cell lytic activity. Mice depleted of the desired cell population and infected on the scarified cornea with herpes simplex virus type 1 uniformly developed necrotizing stromal keratitis by 3 weeks postinfection. A single inoculation of anti-gD MAb (55 micrograms) given intraperitoneally 24 h postinfection strongly protected hosts depleted of CD4+ cells against stromal keratitis. Likewise, antibody treatment in CD8+ or asialo GM1+ cell-depleted hosts was as therapeutically effective as that seen in non-cell-depleted mice. We also observed that in cell-depleted mice, the virus spread into the central nervous system and caused encephalitis. The CD4+ cell-depleted mice were the most severely affected, as 100% developed fatal disease. Anti-gD MAb treatment successfully protected all (32 of 32) CD4+-, CD8+-, or asialo GM1(+)-depleted hosts against encephalitis. We therefore conclude that antibody-mediated prevention of stromal keratitis and encephalitis does not require the obligatory participation of CD4+, CD8+, or asialo GM1+ cells. However, when mice were simultaneously depleted of both CD4+ and CD8+ T-cell subsets, antibody treatment could not prevent fatal encephalitis. Thus, antibody can compensate for the functional loss of one but not two T-lymphocyte subpopulations.     

Brugia pahangi in nude mice: protective immunity to infective larvae is Thy 1.2+ cell dependent and cyclosporin A resistant

Mechanisms of protective immunity to larvae of Brugia pahangi were studied in congenitally athymic nude C3H/HeN mice and their syngeneic heterozygous littermates. An average 11% of subcutaneous larval inocula was recovered from control nudes 28 days after inoculation. No worms were recovered from nude recipients of viable splenic Thy 1.2+ T lymphocytes from heterozygotes which had killed a priming dose of B. pahangi larvae. Primed T lymphocytes, depleted of either Lyt 1.1+ or Lyt 2.1+ cells or incubated with anti-Thy 1.2 monoclonal antibody and complement, failed to protect nude mice against a larval challenge. Nor were primed B lymphocytes depleted by Thy 1.2+ T cell contaminants protective. Treatment with cyclosporin A (CsA) did not increase the numbers of worms recovered from heterozygotes nor did CsA treatment of heterozygous cell donors abolish the ability of primed Thy 1.2+ T lymphocytes to transfer protection to nude mice. IgG but not IgM antibody titres to B. pahangi antigens were depressed in all CsA-treated mice. CsA treatment of nude mice had no direct effect upon development of B. pahangi larvae. These results show that protective immunity to larvae of B. pahangi in mice depends upon small numbers of Thy 1.2+ T cells which are CsA-resistant.     

Role of CD4+ and CD8+ T cells in murine resistance to street rabies virus.

Mice of the SJL/J and BALB/cByJ inbred strains are naturally resistant to street rabies virus (SRV) injected via the intraperitoneal route. To determine the cellular mechanism of resistance, monoclonal antibodies specific for CD4+ or CD8+ subsets of T cells were used to deplete the respective cell population in SRV-infected animals. Elimination of CD4+ T-helper cells abrogated the production of immunoglobulin G (IgG) neutralizing antibodies in response to rabies virus infection and reversed the resistant status of SJL/J and BALB/cByJ mice. In contrast, in vivo depletion of CD8+ cytotoxic T cells had no measurable effect on host resistance to SRV. These results indicate that serum neutralizing antibodies of the IgG class are a primary immunological mechanism of defense against rabies virus infection in this murine model of disease. CD8+ cytotoxic T lymphocytes, which have been shown to transfer protection in other rabies virus systems, appear to have no role in protecting mice against intraperitoneally injected SRV.     

Cellular and humoral immunity against vaccinia virus infection of mice

Despite the widespread use of vaccinia virus (VV) as a vector for other Ags and as the smallpox vaccine, there is little information available about the protective components of the immune response following VV infection. In this study, protection against wild-type VV was evaluated in mice with respect to the relative contributions of CD8(+) T cells vs that of CD4(+) T cells and Ab. C57BL/6 mice primed with the Western Reserve strain of VV mount significant IgM and IgG Ab responses, specific cytotoxic T cell responses, IFN-gamma responses in CD4(+) and CD8(+) T cells, and effectively clear the virus. This protection was abrogated by in vivo depletion of CD4(+) T cells or B cells in IgH(-/-) mice, but was not sensitive to CD8(+) T cell depletion alone. However, a role for CD8(+) T cells in primary protection was demonstrated in MHC class II(-/-) mice, where depleting CD8(+) T cells lead to increase severity of disease. Unlike control MHC class II(-/-) mice, the group depleted of CD8(+) T cells developed skin lesions on the tail and feet and had adrenal necrosis. Adoptive transfer experiments also show CD8(+) T cells can mediate protective memory. These results collectively show that both CD4(+) and CD8(+) T cell-mediated immunity can contribute to protection against VV infection. However, CD4(+) T cell-dependent anti-virus Ab production plays a more important role in clearing virus following acute infection, while in the absence of Ab, CD8(+) T cells can contribute to protection against disease.     

Enhanced neointima formation following arterial injury in immune deficient Rag-1-/- mice is attenuated by adoptive transfer of CD8 T cells

T cells modulate neointima formation after arterial injury but the specific T cell population that is activated in response to arterial injury remains unknown. The objective of the study was to identify the T cell populations that are activated and modulate neointimal thickening after arterial injury in mice. Arterial injury in wild type C57Bl6 mice resulted in T cell activation characterized by increased CD4(+)CD44(hi) and CD8(+)CD44(hi) T cells in the lymph nodes and spleens. Splenic CD8(+)CD25(+) T cells and CD8(+)CD28(+) T cells, but not CD4(+)CD25(+) and CD4(+)CD28(+) T cells, were also significantly increased. Adoptive cell transfer of CD4(+) or CD8(+) T cells from donor CD8-/- or CD4-/- mice, respectively, to immune-deficient Rag-1-/- mice was performed to determine the T cell subtype that inhibits neointima formation after arterial injury. Rag-1-/- mice that received CD8(+) T cells had significantly reduced neointima formation compared with Rag-1-/- mice without cell transfer. CD4(+) T cell transfer did not reduce neointima formation. CD8(+) T cells from CD4-/- mice had cytotoxic activity against syngeneic smooth muscle cells in vitro. The study shows that although both CD8(+) T cells and CD4(+) T cells are activated in response to arterial injury, adoptive cell transfer identifies CD8(+) T cells as the specific and selective cell type involved in inhibiting neointima formation.     

Partially TAP-independent protection against Listeria monocytogenes by H2-M3-restricted CD8+ T cells

Effective protection against Listeria monocytogenes requires Ag-specific CD8(+) T cells. A substantial proportion of CD8(+) T cells activated during L. monocytogenes infection of C57BL/6 mice are restricted by the MHC class Ib molecule H2-M3. In this study, an H2-M3-restricted CD8(+) T cell clone specific for a known H2-M3 epitope (fMIGWII) was generated from L. monocytogenes-infected mice. The clone was cytotoxic, produced IFN-gamma, and could mediate strong protection against L. monocytogenes when transferred to infected mice. Macrophages pulsed with heat-killed LISTERIAE: presented Ag to the clone in a TAP-independent manner. Both TAP-independent and -dependent processing occurred in vivo, as TAP-deficient mice infected with L. monocytogenes were partially protected by adoptive transfer of the clone. This is the first example of CD8(+) T cell-mediated, TAP-independent protection against a pathogen in vivo, confirming the importance of alternative MHC class I processing pathways in the antibacterial immunity.     

CD8+ T lymphocytes protect SCID mice against Encephalitozoon cuniculi infection

Microsporidia are obligate intracellular parasites that cause opportunistic infections in immunocompromised patients. The role of two main T cell subsets in anti-microsporidial immunity has been studied using an Encephalitozoon cuniculi-severe combined immunodeficient (SCID) mouse model. Whereas SCID mice reconstituted with CD4+ T lymphocyte-depleted naive BALB/c splenocytes resolved the infection, adoptive transfer of CD8+ T cell-depleted splenocytes failed to protect the animals against a lethal E. cuniculi infection. Splenocytes from E. cuniculi-immune mice specifically killed syngeneic infected macrophages in a short-term 51Cr-release assay. These results suggest the crucial role of cytotoxic T lymphocytes in the protection against E. cuniculi infection.     

Deficient anti-listerial immunity in the absence of perforin can be restored by increasing memory CD8+ T cell numbers

Compared with wild-type (WT) mice, Listeria monocytogenes (LM)-vaccinated perforin-deficient (PKO) mice have elevated levels of CD8(+) T cell memory, but exhibit reduced levels of protection against virulent LM. In this study, Ag-specific CD8(+) T cells from LM-vaccinated WT and PKO mice were used in adoptive transfer assays to determine the contribution of perforin-dependent cytolysis in protective immunity to LM. Perforin deficiency resulted in an approximately 5-fold reduction in the per-cell protective capacity of Ag-specific memory CD8(+) T cells that was not caused by differences in memory cell quality as measured by CD62L/CD27 expression, TCR repertoire use, functional avidity, differences in expansion of Ag-specific cells upon infection, or maintenance of memory levels over time. However, perforin-deficient CD8(+) T cells exhibited reduced in vivo cytotoxic function compared to WT CD8(+) T cells. Consistent with the existence of perforin-independent effector pathways, double-vaccinated PKO mice were as resistant to challenge with LM as single-vaccinated WT mice. Thus, increasing the number of memory CD8(+) T cells can overcome diminished per-cell protective immunity in the absence of perforin.     

Antiviral protection by CD8+ versus CD4+ T cells. CD8+ T cells correlating with cytotoxic activity in vitro are more efficient in antivaccinia virus protection than CD4-dependent IL.

T cell-mediated protection against a recombinant vaccinia virus was evaluated in mice with respect to the relative contributions of CTL vs that of T cell-dependent IL and of CD4+ cells. H-2b mice primed with the wildtype of vesicular stomatitis virus serotype Indiana (VSV-IND wt) mount an in vitro measurable cytotoxic response against the nucleoprotein (NP) of VSV-IND and are protected against a challenge infection with a vaccinia-VSV recombinant virus expressing the NP of VSV-IND (vacc-IND-NP). Their protective mechanism was highly susceptible to in vivo depletion of CD8+ T cells, but resistant to CD4+ depletion or treatment with anti-IFN-gamma and anti-TNF-alpha. Surprisingly, also VSV-CTL nonresponder H-2k mice were protected against a challenging infection with vacc-IND-NP when primed with VSV-IND wt. In contrast to the CTL responder H-2b mice, this protection was highly susceptible to CD4+ T cell depletion and to anti-IFN-gamma or anti-TNF-alpha treatment, but resistant to CD8+ T cell depletion. Antibodies were not responsible because they failed to transfer protection; in contrast CD4+ T cells conferred significant protection. VSV-CTL responder H-2b and nonresponder H-2k mice were protected almost equally well against a challenge dose of 10(3) pfu vacc-IND-NP inoculated intracerebrally. However, after intracerebral challenge with 5 x 10(6) pfu vacc-IND-NP, the CTL nonresponder mice died, whereas the CTL responder mice eliminated the virus by day 5. These results collectively show that CD4+ T cell-dependent IL may mediate antiviral protection, but their efficiency is relatively weak compared with CD8-mediated protection correlating with cytotoxic activity in vitro.     

Murine CD8+ cytotoxic T lymphocytes lyse Toxoplasma gondii-infected cells

Studies were performed to determine whether CTL against Toxoplasma gondii-infected cells could be induced in a murine model of T. gondii infection in which CD8+ T lymphocytes have been shown to play a major role in resistance against this parasite. In 51Cr release assays, nylon wool nonadherent spleen cells from BALB/c (H-2d) mice immunized with the temperature-sensitive (ts-4) mutant strain of T. gondii were cytotoxic for T. gondii-infected P815 (H-2d) mastocytoma cells but not for uninfected cells. This cytotoxic activity was remarkably increased after in vitro stimulation with T. gondii-infected syngeneic spleen cells. The effector cells were shown to be CD8+ T lymphocytes, because the cytotoxicity was significantly inhibited by depletion of CD8+ T lymphocytes but not by depletion of CD4+ T lymphocytes. This cytotoxic activity was genetically restricted. Spleen cells from T. gondii-immune BALB/c mice were not cytotoxic for T. gondii-infected EL4 (H-2b) thymoma cells, whereas spleen cells from T. gondii-immune C57B1/6 (H-2b) mice were cytotoxic for T. gondii-infected EL4 cells but not for T. gondii-infected P815 cells. The cytolytic activity of CD8+ T lymphocytes against T. gondii-infected cells might be a mechanism whereby these cells confer resistance against T. gondii.