We have compared interferon-gamma (IFN-gamma) with saponin and interleukin-1 (IL-1) as adjuvants ... more We have compared interferon-gamma (IFN-gamma) with saponin and interleukin-1 (IL-1) as adjuvants for a blood-stage malaria vaccine in mice with various immunological abnormalities. IFN-gamma was particularly effective in Biozzi low antibody responder mice, mice selectively bred to produce antibody of low affinity, and mice depleted of CD4+ T cells. IFN-gamma and other cytokines may be safe adjuvants for use in human immunodeficiency states.
Tropical medicine and parasitology : official organ of Deutsche Tropenmedizinische Gesellschaft and of Deutsche Gesellschaft für Technische Zusammenarbeit (GTZ), 1990
Interferon gamma (IFN-gamma) is an effective immunological adjuvant when mixed with vaccines prio... more Interferon gamma (IFN-gamma) is an effective immunological adjuvant when mixed with vaccines prior to injection, but the way in which it exerts this effect has been unclear. Because some adjuvants have been shown to affect lymphocyte traffic, and interferons have been shown to have effects on lymphocyte homing molecules, we examine in this study the effects of IFN-gamma and the potent adjuvant saponin on lymphocyte traffic, and show that both of these adjuvants increase lymphocyte homing to an injection site in mice. We have then compared effects on lymphocyte traffic and on MHC class II expression with adjuvant effects in different mouse strains. Effects on lymphocyte traffic were the inverse of adjuvant effects in different strains, however both materials enhanced MHC class II expression, and this enhancement corresponded with adjuvanticity in different strains of mice. A possible explanation for the negative effect of lymphocyte homing may be that the vast majority of T cells homing to the injection sites in response to both IFN-gamma and saponin were of the CD8+ (suppressor/cytotoxic) phenotype.
Cytokines are potentially useful in vaccination as adjuvants or modulators of the type of respons... more Cytokines are potentially useful in vaccination as adjuvants or modulators of the type of response induced. The work below describes the expression of a cloned cytokine gene for murine interleukin-4 (mIL-4) by a live vaccine vector, an attenuated aroA strain (SL7207) of Salmonella typhimurium, in a murine model system. SL7207 was used as a carrier for two different high-level expression vectors. Both resulting strains, designated SL7207(pOmpAmIL-4) and SL7207(pKKmIL-4), expressed the cloned gene product as monitored by both immunological and biological assays. However, SL7207(pOmpAmIL-4) produced mIL-4 at higher levels and was more stable in vitro than SL7207(pKKmIL-4). When SL7207(pOmpAmIL-4) was used as a live vaccine in BALB/c mice, this strain grew and survived at higher levels than the parental attenuated strain or empty plasmid-carrying strain in spleens, livers, and intestines. This difference in growth and survival did not appear to be caused by alterations in specific lymphocyte-mediated anti-Salmonella immune responses such as delayed-type hypersensitivity or serum antibody as measured by enzyme-linked immunosorbent assay; such alterations have been induced by IL-4 administration in other in vivo systems, and the lack of effect here may reflect the fact that IL-4 is not secreted from the bacteria in large quantities, most of the cytokine being in the cytoplasmic-membrane-bound fraction. Conversely, the ability of mouse macrophages to kill the bacteria in vitro was inhibited by bacterial production of mIL-4. This reduction in macrophage killing activity suggests that bacterial production of mIL-4 may be detrimental to host defense against Salmonella infection and may explain the enhanced bacterial growth and survival in vivo.
Journal of immunology (Baltimore, Md. : 1950), 1993
A rat mAb (NIM-R5) has recently been prepared against a novel murine B cell activation marker. We... more A rat mAb (NIM-R5) has recently been prepared against a novel murine B cell activation marker. We report here isolation of a cDNA (1-19) encoding the B cell-derived protein recognized by NIM-R5 antibody. This cDNA contains an open reading frame that encodes a polypeptide of 304 amino acids with a predicted molecular weight of 34,500. The existence of a 22-amino acid hydrophobic region located 23 amino acids from the amino terminal of the deduced protein, together with four potential N-linked glycosylation sites, characterize the deduced protein encoded by I-19 cDNA as a typical type II transmembrane glycoprotein. Although I-19 cDNA appears to encode a novel murine protein, its nucleotide sequence and deduced amino acid sequence show approximately 70% homology to the previously reported sequence of human CD38, suggesting that I-19 cDNA encodes either the mouse homologue of CD38 or a closely related protein. Northern blot analysis of the expression of this cDNA product in a variety of cell types, together with immunoprecipitation of the recombinant protein expressed in BaF3 cells, indicated that I-19 cDNA encodes not only the epitope recognized by NIM-R5 but also a protein that is indistinguishable biochemically and in terms of distribution from the murine B cell activation marker recognized by NIM-R5 antibody. Chromosomal mapping studies have localized this locus to the proximal region of mouse chromosome 5. We anticipate that the availability of probes for the murine B cell activation marker recognized by NIM-R5, and the recombinant protein itself, will greatly aid efforts to define the role of this molecule in murine B cell development.
Various cytokines have been shown to be effective immunological adjuvants in a variety of model s... more Various cytokines have been shown to be effective immunological adjuvants in a variety of model systems, enhancing protection induced by viral, bacterial and parasitic vaccines, and increasing parameters of immunity in tumour immunization models and in clinical trials. While in most cases cytokine adjuvanticity is not as powerful as that shown by the best experimental adjuvants, such as saponin and Freund's, it can rival that of the adjuvants presently allowed for human use and there are many possible routes to improvement. The use of cytokines may allow for a choice of which immune parameters are enhanced in order to further enhance protective effects and decrease the negative effects of vaccines.
The specificity of antibody (Ab) responses depends on focusing helper T (Th) lymphocyte signals t... more The specificity of antibody (Ab) responses depends on focusing helper T (Th) lymphocyte signals to suitable B lymphocytes capable of binding foreign antigens (Ags), and away from nonspecific or self-reactive B cells. To investigate the molecular mechanisms that prevent the activation of self-reactive B lymphocytes, the activation requirements of B cells specific for the Ag hen egg lysozyme (HEL) obtained from immunoglobulin (Ig)-transgenic mice were compared with those of functionally tolerant B cells isolated from Ig-transgenic mice which also express soluble HEL. To eliminate the need for surface (s)Ig-mediated Ag uptake and presentation and allow the effects of sIg signaling to be studied in isolation, we assessed the ability of allogeneic T cells from bm12 strain mice to provide in vivo help to C57BL/6 strain-transgenic B cells. Interestingly, non-tolerant Ig-transgenic B cells required both allogeneic Th cells and binding of soluble HEL for efficient activation and Ab production. By contrast, tolerant self-reactive B cells from Ig/HEL double transgenic mice responded poorly to the same combination of allogeneic T cells and soluble HEL. The tolerant B cells were nevertheless normally responsive to stimulation with interleukin 4 and anti-CD40 Abs in vitro, suggesting that they retained the capacity to respond to mediators of T cell help. However, the tolerant B cells exhibited a proximal block in the sIg signaling pathway which prevented activation of receptor-associated tyrosine kinases in response to the binding of soluble HEL. The functional significance of this sIg signaling defect was confirmed by using a more potent membrane-bound form of HEL capable of triggering sIg signaling in tolerant B cells, which markedly restored their ability to collaborate with allogeneic Th cells and produce Ab. These findings indicate that Ag-specific B cells require two signals for mounting a T cell-dependent Ab response and identify regulation of sIg signaling as a mechanism for controlling self-reactive B cells.
Cold-adapted reassortants of A/Ann A r b o r / 6 / 6 0 × A / A l a s k a / 6 / 7 7 viruses made i... more Cold-adapted reassortants of A/Ann A r b o r / 6 / 6 0 × A / A l a s k a / 6 / 7 7 viruses made in MDCK cells have recently been assessed genotypically and for temperature-sensitive and cold-adapted phenotypes. These reassortants were used to infect ferrets and hamsters and to inoculate organ cultures of hamster tracheal rings, in order to assess their degree of virulence. Virulence in the three model systems corresponded quite well, and a correlation between loss of virulence and particular A / A A / 6 / 6 0 genes present in the reassortants was noted. Two different reassortants containing either RNA 2 or RNA 5 (NA gene) alone from A / A A / 6 / 6 0 showed little attenuation from the wild-type parent. A reassortant containing both RNA 2 and the NA gene from A / A A / 6 / 6 0 and all remaining wild-type genes showed some small decrease in virulence compared to the wild-type virus. However a reassortant containing these two A / A A / 6 / 6 0 genes and I~NA 3 as an additional gene from this parent, had a level of attenuation comparable to that of the cold-adapted virus.
Journal of immunology (Baltimore, Md. : 1950), 1996
B cell antigen receptor (BCR)-induced apoptosis in the WEHI-231 B lymphoma cell line can be preve... more B cell antigen receptor (BCR)-induced apoptosis in the WEHI-231 B lymphoma cell line can be prevented by engaging CD40. We have used this cell line to investigate the role of mitogen-activated protein (MAP) kinases in integrating BCR and CD40 signaling. Each of the three types of M A P kinases, the extracellular signal-regulated kinases (ERKs), the c-Jun N-terminal kinases (JNKs), and p38, phosphorylates a distinct set of transcription factors. Thus, activating different combinations of M A P kinases could lead to distinct biological responses. We found that BCR engagement in WEHI-231 cells caused a 15to 20-fold activation of ERK2 and a 2-to 3-fold stimulation of ERKl. CD40 did not activate either of these kinases, nor did it affect BCR-induced ERK activation. In contrast, CD40 engagement caused a 50-to 70-fold increase in JNK activity. BCR cross-linking caused a modest (4-to 8-fold) increase in JNK activity by itself and also potentiated CD40-induced JNK activation. Finally, CD40 caused strong activation of the p38 kinase as well as MAPKAP kinase-2, a downstream target of p38. BCR engagement caused only weak activation of the p38 pathway. In summary, the BCR strongly activates ERK2 and weakly activates ERK1, JNK, and p38, while CD40 markedly stimulates the JNK and p38 kinases. Thus, activation of only ERK2 correlates with apoptosis in WEHI-231 cells, whereas full activation of all three M A P kinase pathways correlates with cell survival. The role of M A P kinases in regulating these responses remains to be tested.
Transforming growth factor-b 1 (TGF-b 1) produced by in ltrating macrophage s plays a role in bro... more Transforming growth factor-b 1 (TGF-b 1) produced by in ltrating macrophage s plays a role in brotic disorders through the induction of myobroblasts. To explore possible mechanisms by which TGF-b 1 may act in this context, we investigated effects of TGF-b 1 on macrophage-like (HS-P) and myo broblastic (MT-9) cells, two novel cell lines developed by us. Immunocytochemicall y, the addition of TGF-b 1 (0, 0.1, 0.5, and 1.0 ng/ml) dose-dependentl y suppressed the expressions of antigens recognized by macrophage /histiocyte-speci c antibodies (ED1 and ED2) in HS-P cells, whereas the addition concomitantly increased the number of anti-a -smooth muscle actin antibody-positive myo broblastic cells, suggesting a possible phenotypica l modulation of macrophages into myo broblasts in the brotic lesions. By contrast, MT-9 cells did not show such immunophenotypica l changes following TGF-b 1 addition. DNA synthesis, measured by tritiated thymidine-incorporation , was inhibited in a dose-dependen t manner in MT-9 cells by TGF-b 1 addition (0, 0.1, 0.2, 0.5, 1.0, 5, and 10 ng/ml), but that in HS-P cells was unchanged . Northern blot analysis revealed that expressions of cell cycle-related early genes, c-jun and c-myc, were increased in HS-P cells after TGF-b 1 (1 ng/ml) addition, with c-jun showing peak expression prior to c-myc. By contrast, the peak expressions of c-jun and c-myc were delayed in TGF-b 1 (1 ng/ml)-added MT-9 cells, and their levels were less in MT-9 cells than in HS-P cells. Furthermore, TGF-b 1 (1 and 10 ng/ml) induced DNA laddering in MT-9 cells, but did not in HS-P cells. Based on these ndings, it was speculated that TGF-b 1 could have induced G1 arrest in cell cycle and apoptosis in MT-9 cells. The present study showed that there were signi cant differences in the effects of TGF-b 1 between macrophage-like HS-P cells and myo broblastic MT-9 cells, presumably depending on divergent susceptibilities to TGF-b 1 between both cell types. Because such cell types are key cells in the brogenesis, HS-P and MT-9 might be useful models for investigating the pathogenesi s of brosis in vitro.
CD38 is a 42 kDa membrane-associated ectoenzyme expressed by a large proportion of human and mous... more CD38 is a 42 kDa membrane-associated ectoenzyme expressed by a large proportion of human and mouse lymphocytes. Agonistic antibodies to CD38 induce a strong proliferative response in lymphocytes additionally co-stimulated with other growth co-factors such as IL-4, IL-2 plus accessory cells or sub-mitogenic doses of endotoxin. We show here that B lymphocytes from unstimulated X-linked immunodeficient (xid) mice are unresponsive to CD38 stimulation, both in terms of proliferative response and surface antigen modulation. This CD38 unresponsiveness is evident in the presence of excess quantities of, and normal responses to, the accessory growth co-stimulants required for this response. CD38 molecules expressed on xid B cells are normal in terms of expression levels, size and enzymatic activity, suggesting that CD38 unresponsiveness reflects a down-stream signaling defect. In light of the recent proposal that the xid gene encodes a tyrosine kinase called Bruton's tyrosine kinase (btk), these data suggest that btk is either an integral component or an indirect regulator of the CD38-induced signal transduction pathway.
We have compared interferon-gamma (IFN-gamma) with saponin and interleukin-1 (IL-1) as adjuvants ... more We have compared interferon-gamma (IFN-gamma) with saponin and interleukin-1 (IL-1) as adjuvants for a blood-stage malaria vaccine in mice with various immunological abnormalities. IFN-gamma was particularly effective in Biozzi low antibody responder mice, mice selectively bred to produce antibody of low affinity, and mice depleted of CD4+ T cells. IFN-gamma and other cytokines may be safe adjuvants for use in human immunodeficiency states.
Tropical medicine and parasitology : official organ of Deutsche Tropenmedizinische Gesellschaft and of Deutsche Gesellschaft für Technische Zusammenarbeit (GTZ), 1990
Interferon gamma (IFN-gamma) is an effective immunological adjuvant when mixed with vaccines prio... more Interferon gamma (IFN-gamma) is an effective immunological adjuvant when mixed with vaccines prior to injection, but the way in which it exerts this effect has been unclear. Because some adjuvants have been shown to affect lymphocyte traffic, and interferons have been shown to have effects on lymphocyte homing molecules, we examine in this study the effects of IFN-gamma and the potent adjuvant saponin on lymphocyte traffic, and show that both of these adjuvants increase lymphocyte homing to an injection site in mice. We have then compared effects on lymphocyte traffic and on MHC class II expression with adjuvant effects in different mouse strains. Effects on lymphocyte traffic were the inverse of adjuvant effects in different strains, however both materials enhanced MHC class II expression, and this enhancement corresponded with adjuvanticity in different strains of mice. A possible explanation for the negative effect of lymphocyte homing may be that the vast majority of T cells homing to the injection sites in response to both IFN-gamma and saponin were of the CD8+ (suppressor/cytotoxic) phenotype.
Cytokines are potentially useful in vaccination as adjuvants or modulators of the type of respons... more Cytokines are potentially useful in vaccination as adjuvants or modulators of the type of response induced. The work below describes the expression of a cloned cytokine gene for murine interleukin-4 (mIL-4) by a live vaccine vector, an attenuated aroA strain (SL7207) of Salmonella typhimurium, in a murine model system. SL7207 was used as a carrier for two different high-level expression vectors. Both resulting strains, designated SL7207(pOmpAmIL-4) and SL7207(pKKmIL-4), expressed the cloned gene product as monitored by both immunological and biological assays. However, SL7207(pOmpAmIL-4) produced mIL-4 at higher levels and was more stable in vitro than SL7207(pKKmIL-4). When SL7207(pOmpAmIL-4) was used as a live vaccine in BALB/c mice, this strain grew and survived at higher levels than the parental attenuated strain or empty plasmid-carrying strain in spleens, livers, and intestines. This difference in growth and survival did not appear to be caused by alterations in specific lymphocyte-mediated anti-Salmonella immune responses such as delayed-type hypersensitivity or serum antibody as measured by enzyme-linked immunosorbent assay; such alterations have been induced by IL-4 administration in other in vivo systems, and the lack of effect here may reflect the fact that IL-4 is not secreted from the bacteria in large quantities, most of the cytokine being in the cytoplasmic-membrane-bound fraction. Conversely, the ability of mouse macrophages to kill the bacteria in vitro was inhibited by bacterial production of mIL-4. This reduction in macrophage killing activity suggests that bacterial production of mIL-4 may be detrimental to host defense against Salmonella infection and may explain the enhanced bacterial growth and survival in vivo.
Journal of immunology (Baltimore, Md. : 1950), 1993
A rat mAb (NIM-R5) has recently been prepared against a novel murine B cell activation marker. We... more A rat mAb (NIM-R5) has recently been prepared against a novel murine B cell activation marker. We report here isolation of a cDNA (1-19) encoding the B cell-derived protein recognized by NIM-R5 antibody. This cDNA contains an open reading frame that encodes a polypeptide of 304 amino acids with a predicted molecular weight of 34,500. The existence of a 22-amino acid hydrophobic region located 23 amino acids from the amino terminal of the deduced protein, together with four potential N-linked glycosylation sites, characterize the deduced protein encoded by I-19 cDNA as a typical type II transmembrane glycoprotein. Although I-19 cDNA appears to encode a novel murine protein, its nucleotide sequence and deduced amino acid sequence show approximately 70% homology to the previously reported sequence of human CD38, suggesting that I-19 cDNA encodes either the mouse homologue of CD38 or a closely related protein. Northern blot analysis of the expression of this cDNA product in a variety of cell types, together with immunoprecipitation of the recombinant protein expressed in BaF3 cells, indicated that I-19 cDNA encodes not only the epitope recognized by NIM-R5 but also a protein that is indistinguishable biochemically and in terms of distribution from the murine B cell activation marker recognized by NIM-R5 antibody. Chromosomal mapping studies have localized this locus to the proximal region of mouse chromosome 5. We anticipate that the availability of probes for the murine B cell activation marker recognized by NIM-R5, and the recombinant protein itself, will greatly aid efforts to define the role of this molecule in murine B cell development.
Various cytokines have been shown to be effective immunological adjuvants in a variety of model s... more Various cytokines have been shown to be effective immunological adjuvants in a variety of model systems, enhancing protection induced by viral, bacterial and parasitic vaccines, and increasing parameters of immunity in tumour immunization models and in clinical trials. While in most cases cytokine adjuvanticity is not as powerful as that shown by the best experimental adjuvants, such as saponin and Freund's, it can rival that of the adjuvants presently allowed for human use and there are many possible routes to improvement. The use of cytokines may allow for a choice of which immune parameters are enhanced in order to further enhance protective effects and decrease the negative effects of vaccines.
The specificity of antibody (Ab) responses depends on focusing helper T (Th) lymphocyte signals t... more The specificity of antibody (Ab) responses depends on focusing helper T (Th) lymphocyte signals to suitable B lymphocytes capable of binding foreign antigens (Ags), and away from nonspecific or self-reactive B cells. To investigate the molecular mechanisms that prevent the activation of self-reactive B lymphocytes, the activation requirements of B cells specific for the Ag hen egg lysozyme (HEL) obtained from immunoglobulin (Ig)-transgenic mice were compared with those of functionally tolerant B cells isolated from Ig-transgenic mice which also express soluble HEL. To eliminate the need for surface (s)Ig-mediated Ag uptake and presentation and allow the effects of sIg signaling to be studied in isolation, we assessed the ability of allogeneic T cells from bm12 strain mice to provide in vivo help to C57BL/6 strain-transgenic B cells. Interestingly, non-tolerant Ig-transgenic B cells required both allogeneic Th cells and binding of soluble HEL for efficient activation and Ab production. By contrast, tolerant self-reactive B cells from Ig/HEL double transgenic mice responded poorly to the same combination of allogeneic T cells and soluble HEL. The tolerant B cells were nevertheless normally responsive to stimulation with interleukin 4 and anti-CD40 Abs in vitro, suggesting that they retained the capacity to respond to mediators of T cell help. However, the tolerant B cells exhibited a proximal block in the sIg signaling pathway which prevented activation of receptor-associated tyrosine kinases in response to the binding of soluble HEL. The functional significance of this sIg signaling defect was confirmed by using a more potent membrane-bound form of HEL capable of triggering sIg signaling in tolerant B cells, which markedly restored their ability to collaborate with allogeneic Th cells and produce Ab. These findings indicate that Ag-specific B cells require two signals for mounting a T cell-dependent Ab response and identify regulation of sIg signaling as a mechanism for controlling self-reactive B cells.
Cold-adapted reassortants of A/Ann A r b o r / 6 / 6 0 × A / A l a s k a / 6 / 7 7 viruses made i... more Cold-adapted reassortants of A/Ann A r b o r / 6 / 6 0 × A / A l a s k a / 6 / 7 7 viruses made in MDCK cells have recently been assessed genotypically and for temperature-sensitive and cold-adapted phenotypes. These reassortants were used to infect ferrets and hamsters and to inoculate organ cultures of hamster tracheal rings, in order to assess their degree of virulence. Virulence in the three model systems corresponded quite well, and a correlation between loss of virulence and particular A / A A / 6 / 6 0 genes present in the reassortants was noted. Two different reassortants containing either RNA 2 or RNA 5 (NA gene) alone from A / A A / 6 / 6 0 showed little attenuation from the wild-type parent. A reassortant containing both RNA 2 and the NA gene from A / A A / 6 / 6 0 and all remaining wild-type genes showed some small decrease in virulence compared to the wild-type virus. However a reassortant containing these two A / A A / 6 / 6 0 genes and I~NA 3 as an additional gene from this parent, had a level of attenuation comparable to that of the cold-adapted virus.
Journal of immunology (Baltimore, Md. : 1950), 1996
B cell antigen receptor (BCR)-induced apoptosis in the WEHI-231 B lymphoma cell line can be preve... more B cell antigen receptor (BCR)-induced apoptosis in the WEHI-231 B lymphoma cell line can be prevented by engaging CD40. We have used this cell line to investigate the role of mitogen-activated protein (MAP) kinases in integrating BCR and CD40 signaling. Each of the three types of M A P kinases, the extracellular signal-regulated kinases (ERKs), the c-Jun N-terminal kinases (JNKs), and p38, phosphorylates a distinct set of transcription factors. Thus, activating different combinations of M A P kinases could lead to distinct biological responses. We found that BCR engagement in WEHI-231 cells caused a 15to 20-fold activation of ERK2 and a 2-to 3-fold stimulation of ERKl. CD40 did not activate either of these kinases, nor did it affect BCR-induced ERK activation. In contrast, CD40 engagement caused a 50-to 70-fold increase in JNK activity. BCR cross-linking caused a modest (4-to 8-fold) increase in JNK activity by itself and also potentiated CD40-induced JNK activation. Finally, CD40 caused strong activation of the p38 kinase as well as MAPKAP kinase-2, a downstream target of p38. BCR engagement caused only weak activation of the p38 pathway. In summary, the BCR strongly activates ERK2 and weakly activates ERK1, JNK, and p38, while CD40 markedly stimulates the JNK and p38 kinases. Thus, activation of only ERK2 correlates with apoptosis in WEHI-231 cells, whereas full activation of all three M A P kinase pathways correlates with cell survival. The role of M A P kinases in regulating these responses remains to be tested.
Transforming growth factor-b 1 (TGF-b 1) produced by in ltrating macrophage s plays a role in bro... more Transforming growth factor-b 1 (TGF-b 1) produced by in ltrating macrophage s plays a role in brotic disorders through the induction of myobroblasts. To explore possible mechanisms by which TGF-b 1 may act in this context, we investigated effects of TGF-b 1 on macrophage-like (HS-P) and myo broblastic (MT-9) cells, two novel cell lines developed by us. Immunocytochemicall y, the addition of TGF-b 1 (0, 0.1, 0.5, and 1.0 ng/ml) dose-dependentl y suppressed the expressions of antigens recognized by macrophage /histiocyte-speci c antibodies (ED1 and ED2) in HS-P cells, whereas the addition concomitantly increased the number of anti-a -smooth muscle actin antibody-positive myo broblastic cells, suggesting a possible phenotypica l modulation of macrophages into myo broblasts in the brotic lesions. By contrast, MT-9 cells did not show such immunophenotypica l changes following TGF-b 1 addition. DNA synthesis, measured by tritiated thymidine-incorporation , was inhibited in a dose-dependen t manner in MT-9 cells by TGF-b 1 addition (0, 0.1, 0.2, 0.5, 1.0, 5, and 10 ng/ml), but that in HS-P cells was unchanged . Northern blot analysis revealed that expressions of cell cycle-related early genes, c-jun and c-myc, were increased in HS-P cells after TGF-b 1 (1 ng/ml) addition, with c-jun showing peak expression prior to c-myc. By contrast, the peak expressions of c-jun and c-myc were delayed in TGF-b 1 (1 ng/ml)-added MT-9 cells, and their levels were less in MT-9 cells than in HS-P cells. Furthermore, TGF-b 1 (1 and 10 ng/ml) induced DNA laddering in MT-9 cells, but did not in HS-P cells. Based on these ndings, it was speculated that TGF-b 1 could have induced G1 arrest in cell cycle and apoptosis in MT-9 cells. The present study showed that there were signi cant differences in the effects of TGF-b 1 between macrophage-like HS-P cells and myo broblastic MT-9 cells, presumably depending on divergent susceptibilities to TGF-b 1 between both cell types. Because such cell types are key cells in the brogenesis, HS-P and MT-9 might be useful models for investigating the pathogenesi s of brosis in vitro.
CD38 is a 42 kDa membrane-associated ectoenzyme expressed by a large proportion of human and mous... more CD38 is a 42 kDa membrane-associated ectoenzyme expressed by a large proportion of human and mouse lymphocytes. Agonistic antibodies to CD38 induce a strong proliferative response in lymphocytes additionally co-stimulated with other growth co-factors such as IL-4, IL-2 plus accessory cells or sub-mitogenic doses of endotoxin. We show here that B lymphocytes from unstimulated X-linked immunodeficient (xid) mice are unresponsive to CD38 stimulation, both in terms of proliferative response and surface antigen modulation. This CD38 unresponsiveness is evident in the presence of excess quantities of, and normal responses to, the accessory growth co-stimulants required for this response. CD38 molecules expressed on xid B cells are normal in terms of expression levels, size and enzymatic activity, suggesting that CD38 unresponsiveness reflects a down-stream signaling defect. In light of the recent proposal that the xid gene encodes a tyrosine kinase called Bruton's tyrosine kinase (btk), these data suggest that btk is either an integral component or an indirect regulator of the CD38-induced signal transduction pathway.
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Papers by Andrew Heath