Subash Sad - Academia.edu (original) (raw)
Papers by Subash Sad
Bacterial Activation of Type I Interferons, 2014
The Journal of Immunology, 2006
Immunology and cell biology, Jan 8, 2015
Salmonella enterica serovar Typhimurium (ST) is a virulent intracellular bacterium that conceals ... more Salmonella enterica serovar Typhimurium (ST) is a virulent intracellular bacterium that conceals itself in the phagosomes of infected cells. Although CD8(+) T cells promote protection against various intracellular pathogens, the role of CD8(+) T cells against virulent ST has been unclear due to early fatality of susceptible (B6) mice. Herein, we generated MHC I-deficient mice on the resistant (129SvJ) and susceptible (Nramp1 transgenic B6) background to evaluate the role of CD8(+) T cells against virulent ST. Our results indicate that CD8(+) T cells have a critical protective role in host survival during infection with virulent ST. As antigen presentation and CD8(+) T-cell activation against phagosomal antigens are considered to operate through the cross-presentation pathway, we have evaluated CD8(+) T-cell response against ST in Batf3-deficient mice that lack CD8α dendritic cells (DCs). Using a recombinant of ST that expresses antigen (ST-OVA) mainly in the phagosomes of infected c...
The Journal of Immunology, May 1, 2012
Salmonella enterica serovar Typhimurium (S. Typhimurium) is a virulent pathogen that induces rapi... more Salmonella enterica serovar Typhimurium (S. Typhimurium) is a virulent pathogen that induces rapid host death. Here we observed that host survival after infection with S. Typhimurium was enhanced in the absence of type I interferon signaling, with improved survival of mice deficient in the receptor for type I interferons (Ifnar1 −/− mice) that was attributed to macrophages. Although there was no impairment in cytokine expression or inflammasome activation in Ifnar1 −/− macrophages, they were highly resistant to S. Typhimurium-induced cell death. Specific inhibition of the kinase RIP1 or knockdown of the gene encoding the kinase RIP3 prevented the death of wild-type macrophages, which indicated that necroptosis was a mechanism of cell death. Finally, RIP3-deficient macrophages, which cannot undergo necroptosis, had similarly less death and enhanced control of S. Typhimurium in vivo. Thus, we propose that S. Typhimurium induces the production of type I interferon, which drives necroptosis of macrophages and allows them to evade the immune response.
The Journal of Immunology, May 1, 2014
Ciba Foundation symposium
National Research Council Canada. www.nrc-cnrc.gc.ca. ...
Immunology
Immunization of male rats and monkeys with gonadotropin-releasing hormone (GnRH) conjugated to a ... more Immunization of male rats and monkeys with gonadotropin-releasing hormone (GnRH) conjugated to a carrier results in a dramatic atrophy of the prostate. GnRH, linked to either diphtheria toxoid or tetanus toxoid as carrier, is now being evaluated for its use in the immunotherapy of hormonedependent prostatic enlargement in men. This report deals with the phenomenon of carrier-induced, epitope-specific regulation in the GnRH-carrier system. In experiments designed to assess the influence of the carrier on antibody responses to the 'self' hapten GnRH, we show that preimmunization with carriers diphtheria toxoid and tetanus toxoid results in a strain-dependent inhibition of anti-GnRH responses in mice. Results of adoptive transfer experiments indicate that T cells from carrier-presensitized mice are responsible for suppression of anti-haptenic antibodies and that T cells from conjugate-immunized mice, on the other hand, can actually help overcome hyporesponsiveness.
Immunology
A gonadotropin-releasing hormone (GnRH)-based vaccine is being developed as a method for nonsurgi... more A gonadotropin-releasing hormone (GnRH)-based vaccine is being developed as a method for nonsurgical immunotherapy as immunization with this vaccine results in atrophy of the prostate. This vaccine, a conjugate of GnRH and diphtheria toxoid (DT), provides a unique hapten-carrier system for investigating the influence of carrier presensitization on antibody responses to self haptens. In a recent communication' we showed that preimmunization with carriers diphtheria toxoid and tetanus toxoid results in a strain-dependent inhibition of anti-GnRH responses in mice and that T cells from carrier-presensitized mice are responsible for anti-haptenic suppression. In the present report we describe a strategy for bypassing DT-induced epitopic suppression using a T-helper epitope from DT.
The Journal of Immunology
After antigenic stimulation, naive CD8+ T cells differentiate into cytotoxic Tc1 cells secreting ... more After antigenic stimulation, naive CD8+ T cells differentiate into cytotoxic Tc1 cells secreting the cytokines IL-2, IFN-gamma, and TNF, which aid their proliferation and effector functions. We have previously shown that IL-4 acts directly on differentiated Tc1 cells to impair subsequent Con A-induced IL-2 production. As IL-4 may be produced in the vicinity of Tc1 cells during normal immune responses, we have further analyzed the short and long term functions of IL-4-treated Tc1 cells. We now show that these cells also have a defect in the synthesis of IFN-gamma, TNF, and IL-10 in response to antigenic stimulation. IL-2 synthesis was the most sensitive, as stimulation of IL-4-treated Tc1 cells with higher numbers of APCs partially restored IFN-gamma, TNF, and IL-10, but not IL-2, synthesis. Injection of allo-specific Tc1 cells into mice expressing the target Ag revealed reduced cytokine synthesis in vivo by IL-4-treated Tc1 cells. Loss of cytokine synthesis did not impair the short term effector functions of Tc1 cells, as they induced adoptively transferred delayed type hypersensitivity in recipient mice and retained both perforin- and Fas-dependent cytolytic mechanisms in vitro. Long term coculture of tumor targets and tumor-specific Tc1 cells indicated that normal Tc1 cells proliferated and killed tumor cells, whereas IL-4-treated Tc1 cells failed to proliferate and hence were unable to curtail the proliferation of tumor cells. These results suggest that IL-4 synthesis in vivo would not affect immediate effector functions of differentiated Tc1 cells, but would compromise immunity by reducing their long term functional capability.
mBio, 2015
Nontyphoidal Salmonella enterica (NTS) infections are a major burden to global public health, as ... more Nontyphoidal Salmonella enterica (NTS) infections are a major burden to global public health, as they lead to diseases ranging from gastroenteritis to systemic infections and there is currently no vaccine available. Here, we describe a highly effective component vaccine against S. enterica serovar Typhimurium in both gastroenteritis and systemic murine infection models. We devised an approach to generate supernatants of S. enterica serovar Typhimurium, an organism that is highly abundant in virulence factors. Immunization of mice with this supernatant resulted in dramatic protection against a challenge with serovar Typhimurium, showing increased survival in the systemic model and decreased intestinal pathology in the gastrointestinal model. Protection correlated with specific IgA and IgG levels in the serum and specific secretory IgA levels in the feces of immunized mice. Initial characterization of the protective antigens in the bacterial culture supernatants revealed a subset of a...
The Tc1 and Tc2 subsets of CD8+ T effector cells secrete different patterns of cytokines, but hav... more The Tc1 and Tc2 subsets of CD8+ T effector cells secrete different patterns of cytokines, but have similar functions, including perforin- and Fas-dependent cytotoxicity, and induction of delayed type hypersensitivity (DTH) reactions involving oedema and granulocytic infiltration. The characteristic cytokines of Tc1 (gamma-interferon) and Tc2 (interleukins 4 and 5) are expressed in vivo during the DTH reaction. Tc1 cells that are deficient in cytokine synthesis also induce similar levels of DTH, supporting the lack of correlation between CD8+ T cell cytokine patterns and DTH. CD8+ T cells often produce lower cytokine levels than CD4 cells because the CD8 cells kill their antigen-presenting cells before full stimulation can occur. This effect can be counteracted by increasing the frequency of stimulation, or using perforin-deficient T cells. A multiparameter analysis of cytokine effects on CD8+ T cell differentiation has been initiated, on the basis of the principle that normal immune responses involve complex cytokine mixtures. All combinations of seven cytokines were tested. In some combinations, the combined effect could not have been predicted from individual cytokine functions. Conditions were identified in which each of interleukins 4, 10 and 12 could have opposite effects on CD8+ T cell differentiation.
Ciba Foundation symposium, 1995
Our knowledge of the cytokine secretion patterns of T cells and other cells is clearly becoming m... more Our knowledge of the cytokine secretion patterns of T cells and other cells is clearly becoming more complex. The T helper 1 (Th1) and Th2 patterns may represent the extremes of a spectrum of cytokine regulatory patterns controlled by several cell types. CD8+ T cells can also secrete either Th1-like or Th2-like cytokine patterns, and they can contribute to bystander B cell activation. Interactions occur between immune cytokine regulatory networks and other systems, and pregnancy and responses against infection can profoundly influence each other.
Journal of immunology (Baltimore, Md. : 1950), 2007
Typhoid fever and gastroenteritis caused by Salmonella enterica species are increasing globally. ... more Typhoid fever and gastroenteritis caused by Salmonella enterica species are increasing globally. Pregnancy poses a high risk, but it is unclear how maternal immunity to infection is altered. In mice, susceptible strains die of S. enterica serovar typhimurium (ST) infection within 7 days whereas resistant mice (129 x 1/SvJ) develop a chronic infection. We found that virulent ST infection during pregnancy, in normally resistant 129 x 1/SvJ mice, evoked approximately 100% fetal loss and surprisingly >60% host fatality, with a median survival of 6 days. Splenic bacterial load was 1000-fold higher in pregnant mice. This correlated to a diminished splenic recruitment/expansion of innate immune cells: dendritic cells, neutrophils, and NK cells. In particular, the splenic expansion and activation of NK cells postinfection seen in nonpregnant mice was lacking in pregnancy. Most notably, pregnant-infected mice had decreased production of serum IL-12 and increased IL-6 levels. Moreover, ute...
Journal of immunology (Baltimore, Md. : 1950), 2005
During infection with lymphocytic choriomeningitis virus, CD8(+) T cells differentiate rapidly in... more During infection with lymphocytic choriomeningitis virus, CD8(+) T cells differentiate rapidly into effectors (CD62L(low)CD44(high)) that differentiate further into the central memory phenotype (CD62L(high)CD44(high)) gradually. To evaluate whether this CD8(+) T cell differentiation program operates in all infection models, we evaluated CD8(+) T cell differentiation during infection of mice with recombinant intracellular bacteria, Listeria monocytogenes (LM) and Mycobacterium bovis (BCG), expressing OVA. We report that CD8(+) T cells primed during infection with the attenuated pathogen BCG-OVA differentiated primarily into the central subset that correlated to reduced attrition of the primed cells subsequently. CD8(+) T cells induced by LM-OVA also differentiated into central phenotype cells first, but the cells rapidly converted into effectors in contrast to BCG-OVA. Memory CD8(+) T cells induced by both LM-OVA as well as BCG-OVA were functional in that they produced cytokines and ...
Cancer research, Jan 15, 2003
Archaeosome adjuvants formulated as archaeal ether glycerolipid vesicles induce strong CD4(+) as ... more Archaeosome adjuvants formulated as archaeal ether glycerolipid vesicles induce strong CD4(+) as well as CD8(+) CTL responses to entrapped soluble antigens. Immunization of mice with ovalbumin (OVA) entrapped in archaeosomes composed of the total polar lipids of Methanobrevibacter smithii resulted in a potent OVA-specific CD8(+) T-cell response, and subsequently, the mice dramatically resisted solid tumor growth of OVA-expressing EG.7 cells and lung metastasis of B16OVA melanoma cells. Prophylactic protection was antigen-specific because tumor curtailment was not seen in mice injected with antigen-free archaeosomes. Similarly, there was no protection against B16 melanoma cells lacking OVA expression. Furthermore, in vivo depletion of CD8(+) T cells abrogated the protective response, indicating that the antitumor immunity was mediated by CTLs. Depletion of CD4(+) T cells also resulted in partial loss of tumor protection, suggesting a beneficial role for T-helper cells. Interestingly ...
Bacterial Activation of Type I Interferons, 2014
The Journal of Immunology, 2006
Immunology and cell biology, Jan 8, 2015
Salmonella enterica serovar Typhimurium (ST) is a virulent intracellular bacterium that conceals ... more Salmonella enterica serovar Typhimurium (ST) is a virulent intracellular bacterium that conceals itself in the phagosomes of infected cells. Although CD8(+) T cells promote protection against various intracellular pathogens, the role of CD8(+) T cells against virulent ST has been unclear due to early fatality of susceptible (B6) mice. Herein, we generated MHC I-deficient mice on the resistant (129SvJ) and susceptible (Nramp1 transgenic B6) background to evaluate the role of CD8(+) T cells against virulent ST. Our results indicate that CD8(+) T cells have a critical protective role in host survival during infection with virulent ST. As antigen presentation and CD8(+) T-cell activation against phagosomal antigens are considered to operate through the cross-presentation pathway, we have evaluated CD8(+) T-cell response against ST in Batf3-deficient mice that lack CD8α dendritic cells (DCs). Using a recombinant of ST that expresses antigen (ST-OVA) mainly in the phagosomes of infected c...
The Journal of Immunology, May 1, 2012
Salmonella enterica serovar Typhimurium (S. Typhimurium) is a virulent pathogen that induces rapi... more Salmonella enterica serovar Typhimurium (S. Typhimurium) is a virulent pathogen that induces rapid host death. Here we observed that host survival after infection with S. Typhimurium was enhanced in the absence of type I interferon signaling, with improved survival of mice deficient in the receptor for type I interferons (Ifnar1 −/− mice) that was attributed to macrophages. Although there was no impairment in cytokine expression or inflammasome activation in Ifnar1 −/− macrophages, they were highly resistant to S. Typhimurium-induced cell death. Specific inhibition of the kinase RIP1 or knockdown of the gene encoding the kinase RIP3 prevented the death of wild-type macrophages, which indicated that necroptosis was a mechanism of cell death. Finally, RIP3-deficient macrophages, which cannot undergo necroptosis, had similarly less death and enhanced control of S. Typhimurium in vivo. Thus, we propose that S. Typhimurium induces the production of type I interferon, which drives necroptosis of macrophages and allows them to evade the immune response.
The Journal of Immunology, May 1, 2014
Ciba Foundation symposium
National Research Council Canada. www.nrc-cnrc.gc.ca. ...
Immunology
Immunization of male rats and monkeys with gonadotropin-releasing hormone (GnRH) conjugated to a ... more Immunization of male rats and monkeys with gonadotropin-releasing hormone (GnRH) conjugated to a carrier results in a dramatic atrophy of the prostate. GnRH, linked to either diphtheria toxoid or tetanus toxoid as carrier, is now being evaluated for its use in the immunotherapy of hormonedependent prostatic enlargement in men. This report deals with the phenomenon of carrier-induced, epitope-specific regulation in the GnRH-carrier system. In experiments designed to assess the influence of the carrier on antibody responses to the 'self' hapten GnRH, we show that preimmunization with carriers diphtheria toxoid and tetanus toxoid results in a strain-dependent inhibition of anti-GnRH responses in mice. Results of adoptive transfer experiments indicate that T cells from carrier-presensitized mice are responsible for suppression of anti-haptenic antibodies and that T cells from conjugate-immunized mice, on the other hand, can actually help overcome hyporesponsiveness.
Immunology
A gonadotropin-releasing hormone (GnRH)-based vaccine is being developed as a method for nonsurgi... more A gonadotropin-releasing hormone (GnRH)-based vaccine is being developed as a method for nonsurgical immunotherapy as immunization with this vaccine results in atrophy of the prostate. This vaccine, a conjugate of GnRH and diphtheria toxoid (DT), provides a unique hapten-carrier system for investigating the influence of carrier presensitization on antibody responses to self haptens. In a recent communication' we showed that preimmunization with carriers diphtheria toxoid and tetanus toxoid results in a strain-dependent inhibition of anti-GnRH responses in mice and that T cells from carrier-presensitized mice are responsible for anti-haptenic suppression. In the present report we describe a strategy for bypassing DT-induced epitopic suppression using a T-helper epitope from DT.
The Journal of Immunology
After antigenic stimulation, naive CD8+ T cells differentiate into cytotoxic Tc1 cells secreting ... more After antigenic stimulation, naive CD8+ T cells differentiate into cytotoxic Tc1 cells secreting the cytokines IL-2, IFN-gamma, and TNF, which aid their proliferation and effector functions. We have previously shown that IL-4 acts directly on differentiated Tc1 cells to impair subsequent Con A-induced IL-2 production. As IL-4 may be produced in the vicinity of Tc1 cells during normal immune responses, we have further analyzed the short and long term functions of IL-4-treated Tc1 cells. We now show that these cells also have a defect in the synthesis of IFN-gamma, TNF, and IL-10 in response to antigenic stimulation. IL-2 synthesis was the most sensitive, as stimulation of IL-4-treated Tc1 cells with higher numbers of APCs partially restored IFN-gamma, TNF, and IL-10, but not IL-2, synthesis. Injection of allo-specific Tc1 cells into mice expressing the target Ag revealed reduced cytokine synthesis in vivo by IL-4-treated Tc1 cells. Loss of cytokine synthesis did not impair the short term effector functions of Tc1 cells, as they induced adoptively transferred delayed type hypersensitivity in recipient mice and retained both perforin- and Fas-dependent cytolytic mechanisms in vitro. Long term coculture of tumor targets and tumor-specific Tc1 cells indicated that normal Tc1 cells proliferated and killed tumor cells, whereas IL-4-treated Tc1 cells failed to proliferate and hence were unable to curtail the proliferation of tumor cells. These results suggest that IL-4 synthesis in vivo would not affect immediate effector functions of differentiated Tc1 cells, but would compromise immunity by reducing their long term functional capability.
mBio, 2015
Nontyphoidal Salmonella enterica (NTS) infections are a major burden to global public health, as ... more Nontyphoidal Salmonella enterica (NTS) infections are a major burden to global public health, as they lead to diseases ranging from gastroenteritis to systemic infections and there is currently no vaccine available. Here, we describe a highly effective component vaccine against S. enterica serovar Typhimurium in both gastroenteritis and systemic murine infection models. We devised an approach to generate supernatants of S. enterica serovar Typhimurium, an organism that is highly abundant in virulence factors. Immunization of mice with this supernatant resulted in dramatic protection against a challenge with serovar Typhimurium, showing increased survival in the systemic model and decreased intestinal pathology in the gastrointestinal model. Protection correlated with specific IgA and IgG levels in the serum and specific secretory IgA levels in the feces of immunized mice. Initial characterization of the protective antigens in the bacterial culture supernatants revealed a subset of a...
The Tc1 and Tc2 subsets of CD8+ T effector cells secrete different patterns of cytokines, but hav... more The Tc1 and Tc2 subsets of CD8+ T effector cells secrete different patterns of cytokines, but have similar functions, including perforin- and Fas-dependent cytotoxicity, and induction of delayed type hypersensitivity (DTH) reactions involving oedema and granulocytic infiltration. The characteristic cytokines of Tc1 (gamma-interferon) and Tc2 (interleukins 4 and 5) are expressed in vivo during the DTH reaction. Tc1 cells that are deficient in cytokine synthesis also induce similar levels of DTH, supporting the lack of correlation between CD8+ T cell cytokine patterns and DTH. CD8+ T cells often produce lower cytokine levels than CD4 cells because the CD8 cells kill their antigen-presenting cells before full stimulation can occur. This effect can be counteracted by increasing the frequency of stimulation, or using perforin-deficient T cells. A multiparameter analysis of cytokine effects on CD8+ T cell differentiation has been initiated, on the basis of the principle that normal immune responses involve complex cytokine mixtures. All combinations of seven cytokines were tested. In some combinations, the combined effect could not have been predicted from individual cytokine functions. Conditions were identified in which each of interleukins 4, 10 and 12 could have opposite effects on CD8+ T cell differentiation.
Ciba Foundation symposium, 1995
Our knowledge of the cytokine secretion patterns of T cells and other cells is clearly becoming m... more Our knowledge of the cytokine secretion patterns of T cells and other cells is clearly becoming more complex. The T helper 1 (Th1) and Th2 patterns may represent the extremes of a spectrum of cytokine regulatory patterns controlled by several cell types. CD8+ T cells can also secrete either Th1-like or Th2-like cytokine patterns, and they can contribute to bystander B cell activation. Interactions occur between immune cytokine regulatory networks and other systems, and pregnancy and responses against infection can profoundly influence each other.
Journal of immunology (Baltimore, Md. : 1950), 2007
Typhoid fever and gastroenteritis caused by Salmonella enterica species are increasing globally. ... more Typhoid fever and gastroenteritis caused by Salmonella enterica species are increasing globally. Pregnancy poses a high risk, but it is unclear how maternal immunity to infection is altered. In mice, susceptible strains die of S. enterica serovar typhimurium (ST) infection within 7 days whereas resistant mice (129 x 1/SvJ) develop a chronic infection. We found that virulent ST infection during pregnancy, in normally resistant 129 x 1/SvJ mice, evoked approximately 100% fetal loss and surprisingly >60% host fatality, with a median survival of 6 days. Splenic bacterial load was 1000-fold higher in pregnant mice. This correlated to a diminished splenic recruitment/expansion of innate immune cells: dendritic cells, neutrophils, and NK cells. In particular, the splenic expansion and activation of NK cells postinfection seen in nonpregnant mice was lacking in pregnancy. Most notably, pregnant-infected mice had decreased production of serum IL-12 and increased IL-6 levels. Moreover, ute...
Journal of immunology (Baltimore, Md. : 1950), 2005
During infection with lymphocytic choriomeningitis virus, CD8(+) T cells differentiate rapidly in... more During infection with lymphocytic choriomeningitis virus, CD8(+) T cells differentiate rapidly into effectors (CD62L(low)CD44(high)) that differentiate further into the central memory phenotype (CD62L(high)CD44(high)) gradually. To evaluate whether this CD8(+) T cell differentiation program operates in all infection models, we evaluated CD8(+) T cell differentiation during infection of mice with recombinant intracellular bacteria, Listeria monocytogenes (LM) and Mycobacterium bovis (BCG), expressing OVA. We report that CD8(+) T cells primed during infection with the attenuated pathogen BCG-OVA differentiated primarily into the central subset that correlated to reduced attrition of the primed cells subsequently. CD8(+) T cells induced by LM-OVA also differentiated into central phenotype cells first, but the cells rapidly converted into effectors in contrast to BCG-OVA. Memory CD8(+) T cells induced by both LM-OVA as well as BCG-OVA were functional in that they produced cytokines and ...
Cancer research, Jan 15, 2003
Archaeosome adjuvants formulated as archaeal ether glycerolipid vesicles induce strong CD4(+) as ... more Archaeosome adjuvants formulated as archaeal ether glycerolipid vesicles induce strong CD4(+) as well as CD8(+) CTL responses to entrapped soluble antigens. Immunization of mice with ovalbumin (OVA) entrapped in archaeosomes composed of the total polar lipids of Methanobrevibacter smithii resulted in a potent OVA-specific CD8(+) T-cell response, and subsequently, the mice dramatically resisted solid tumor growth of OVA-expressing EG.7 cells and lung metastasis of B16OVA melanoma cells. Prophylactic protection was antigen-specific because tumor curtailment was not seen in mice injected with antigen-free archaeosomes. Similarly, there was no protection against B16 melanoma cells lacking OVA expression. Furthermore, in vivo depletion of CD8(+) T cells abrogated the protective response, indicating that the antitumor immunity was mediated by CTLs. Depletion of CD4(+) T cells also resulted in partial loss of tumor protection, suggesting a beneficial role for T-helper cells. Interestingly ...