Biomedical subjects

A Coutinho

Publications and source records attributed to A Coutinho.

At least 127 records · Page 7Linked to original sources

PubMed1991

Normal serum immunoglobulins influence the numbers of bone marrow pre-B and B cells.

The homeostatic mechanisms controlling B lymphocyte output from bone marrow are not well understood. The present experiments evaluated putative influences of circulating immunoglobulins (Ig) on bone marrow (BM) pre-B and B cell populations. Injections into normal mice of Ig isolated from normal mouse serum, resulted in a dose-dependent and reversible reduction in numbers of BM B lineage cells, in particular of small B220+ surface IgM- cells. Maximal effects were observed upon injection of isologous polyclonal Ig and were independent of mature T cells. These results suggest a feedback modulation of peripheral Ig on cellular activities in BM B lineage compartments, mediated by mechanisms that seem to involve the variable regions of the Ig molecule.

Animals↗

PubMed1991

Biased VH gene expression in murine CD5 B cells results from age-dependent cellular selection.

Flow cytometry-purified, peritoneal and splenic CD5+ and CD5- B cells from neonatal and adult C57BL/6 mice were studied for expression of VH and Vx gene families in RNA colony blot assays, and for frequencies of clones secreting antibodies to bromelain-treated mouse red blood cells (BrMRBC), single-stranded DNA, trimethyl ammonium and bovine gamma-globulin, by limiting dilution. The results show few overall differences between the two B cell subsets, which both manifest ontogenic D-proximal VH preferences that are lost with age. Biased VH11 expression in CD5 B cells is high in adult peritoneum and spleen but absent in newborns. It only partly correlates with the selection of anti-BrMRBC reactivity, which is considerably higher in peritoneum than in spleen. No particular Vx bias was observed in any of the populations studied with the possible exception of Vx22 in peritoneal CD5+ B cells. We conclude that the antibody repertoire expressed by peritoneal CD5+ B cells of adult mice is not the result of a genetic program, but rather the consequence of local, age-dependent cellular selection mechanisms.

Aging↗

PubMed1991

Clonal persistence of B lymphocytes in normal mice is determined by variable region-dependent selection.

Many adult splenic B cells die within 1 week in the spleen of adoptive adult recipient mice; in contrast, the cellular environment of newborn recipients allows for their expansion and persistence for several weeks. In the present study, we show that the local environment of adult peritoneal cavity also allows adult splenic B cells to persist for over 2 weeks after intraperitoneal transfer. In order to determine whether the persistence of donor B cells in newborn hosts and in the peritoneum of adult recipients results from a selection process involving the clonal specificities expressed, the variation in time of VH gene family repertoires of donor B cells was analyzed in the hosts. At different times after the transfer of splenic cells from lipopolysaccharide (LPS)-reactive mice into LPS-non responder histocompatible recipients, mRNA colony blot assays were performed. The results show that among the donor adult LPS-reactive B cells, the VH genes are differently used by the expanding or persisting B cells, in both kinds of recipients. Thus, cells expressing J558 or VH11 gene families are, in particular, positively selected, while those expressing D-proximal or J606 and 36-30 VH gene families are less selected. These findings demonstrate that the propensity of B cells to persist and expand is determined by their selection through their immunoglobulin variable regions, rather than by genetic properties linked to particular B cell subsets.

Animals↗

PubMed1991

Stimulation of B and T cells by in vivo high dose immunoglobulin administration in normal mice.

Adult BALB/c mice were injected intravenously with a preparation of pooled normal murine IgG (400 mg/kg/day, on five consecutive days) and studied 8, 15, and 60 days later. High dose IgG administration increased the total numbers of splenic activated B and CD4+ (but not CD8+) T cells, as well as the numbers of splenic Ig-secreting cells, particularly in the IgG isotypes. Reactivities to some autoantigens, but not to bacterial or other heteroantigens, were selectively amplified amongst IgM-secreting cells. IgG administration did not alter the specific primary immune response to heterologous erythrocytes or bacterial dextran. No cellular alterations were detected in the lymph nodes or peritoneal cavity of treated animals. Most of these effects subsided with time, but some autoantibody reactivities remained elevated 60 days later. The present results suggest that the therapeutic effects of high dose IgG administration which have been reported in human diseases might be associated with the immunostimulatory activities of such treatment.

Adjuvants, Immunologic↗

PubMed1991

Second generation immune networks.

Network approaches have had little impact on immunology because they have addressed the wrong questions. They have concentrated on the regulation of clonal immune responses rather than on the supraclonal properties of the immune system that emerge from its network organization, such as natural tolerance and memory. Theoretical advances, observations in unimmunized mice and humans, and the success of novel therapeutics in autoimmune diseases have recently promoted a new burst of research on the structure, temporal dynamics and metadynamical plasticity of immune networks.

Animals↗

PubMed1991

Endogenous VH gene family expression in immunoglobulin-transgenic mice: evidence for selection of antibody repertoires.

VH gene family expression in single cells of the emergent, available and actual B cell repertoires of C57BL/6 mice was compared to that of two immunoglobulin (Ig)-transgenic B6 lines (B6-Sp6 and M54). We found that less than 5% of bone marrow cells of transgenic mice express endogenous VH genes and that the vast majority (95%) of the peripheral, mature B cell repertoire in these animals is composed of cells expressing the VHJ558 transgenic family. Unimmunized transgenic mice, however, diversify VH gene family usage by 'background' Ig-secreting cells in the spleen, greater than 50% of which express endogenous VH genes. The pattern of endogenous VH gene family expression in the actual repertoire of B6-Sp6 mice is indistinguishable from that of normal B6 mice. In contrast, actual repertoires of M54 mice differ by a 4- to 5-fold higher representation of the VHQ52 family. These results demonstrate a powerful positive selection of B cells into the secretory compartments of unimmunized animals, show that actual and available repertoires differ very markedly, and suggest that V region interactions participate in the selection of 'natural antibody' repertoires.

Animals↗

PubMed1991

Clonal analysis of B lymphocyte responses to Plasmodium chabaudi infection of normal and immunoprotected mice.

Parasite infection causes marked perturbations in the host immune system, as shown by hypergammaglobulinemia, autoimmunity and immune depression, but there is little information on the number, specificities and performance of B cell clones activated in the course of infection. We have addressed these questions in a model of murine malaria induced by Plasmodium chabaudi, where primary infection results in very marked B cell responses that shift in Ig isotype pattern in immunoprotected animals, and where immunity can be transferred to naive recipients by injection of serum from late, but not early, infection. We have quantitated B cells responding to infection in two distinct functional compartments, namely blast cells and Ig-secreting cells, and compared normal with immune animals. We have also determined the frequencies of clonal specificities towards several autoantigens (DNA, myosin, transferrin and red cells), non-self protein or polysaccharide antigens (KLH, levan and dextran), and parasite antigens in both compartments, by measuring blast cell reactivities in limiting dilution analyses and Ig secretion in ELISASPOT assays. This experimental design allowed us to assess the specificity of the B cell responses, to compare the clonal composition of these two B cell compartments, and to evaluate putative specific response regulation at the step of terminal differentiation. Our results show that, in this particular experimental system: (i) B cell responses in primary infection are truly non-specific while immune animals show a greater ability to control the massive non-specific response; (ii) parasite specific B cells, particularly those committed to IgG production, are selectively stimulated in immune individuals; (iii) autoreactive B cells are not selectively stimulated, but increased autoantibody production may result from perturbation in the control of terminal differentiation in the respective clones; (iv) clones with specificity to some non-self antigens (e.g. KLH and dextran) are selectively engaged and regulated, which might have implications for the immunosuppression following infection.

Animals↗

PubMed1991

Role of B and T lymphocytes in the specific immunosuppression induced by a protein released by porcine monocytes infected with African swine fever virus.

Some immunobiological aspects of host responses to an immunosuppressive protein (p36) released by porcine monocytes upon infection with African swine fever virus were analysed in a murine system. Treatment of normal, adult C57BL/6 mice with p36 (i) significantly delayed allogenic skin graft rejection; (ii) suppressed the specific plaque-forming cell response to immunization with heterologous erythrocytes; but (iii) induced marked increases in the numbers of 'background' splenic Ig-secreting plaque-forming cells. Cytofluorometric analysis of spleen cells revealed that a considerable fraction of all B cells, as well as CD4 and CD8 T lymphocytes, undergo blast transformation after p36 treatment. The immunosuppressive effects do not seem to result from 'antigenic competition', for they cannot be induced by even higher doses of pig albumin or by culture products of non-infected pig monocytes. Suppression of specific antibody responses and stimulation of 'background' plaque-forming cells are both T cell-dependent, since they are markedly reduced in thymectomized mice and in animals treated with anti-CD4 or anti-CD8 antibodies. This suggests the relationship between non-specific stimulation and specific suppression of 'unrelated' immune responses and reinforces the notion that viral-associated immunosuppression may be due to overstimulation. The present murine experimental model may prove valuable in the study of immunosuppression associated with infection, even for microorganisms which do not infect mice.

African Swine Fever↗

PubMed1991

Xid immunodeficiency imparts increased parasite clearance and resistance to pathology in experimental Chagas' disease.

Infection of several mouse strains with Trypanosoma cruzi stimulates high levels of T and B lymphocyte activities which persist during the chronic phase and is followed by specific immunosuppression and severe autoimmune pathology. Infected BALB.Xid mice carrying an X-linked mutation and lacking CD5 B cells, display poor B cell responses to T. cruzi infection, accompanied by low levels of specific and non-specific immunoglobulins in the serum. However, these animals control parasitemia, do not show the wasting observed in BALB/c mice, and develop almost no pathology early in the chronic phase. The infection of (BALB.Xid female x BALB/c male) F1 animals shows that immunodefective males behave like Xid animals in contrast to females which respond as normal BALB/c mice. These results indicate that the Xid locus controls lymphocyte responses, parasite clearance and pathology in experimental Chagas' disease.

Animals↗

PubMed1991

Selective expansion of idiotype sharing T and B cells in cyclosporin A-mediated autoimmunity.

CBA/N mice submitted to autologous bone marrow reconstitution after lethal irradiation and simultaneous Cyclosporin A (CsA) treatment develop a chronic graft-versus-host disease with autoimmune characteristics. When compared to normal controls, diseased mice show an overrepresentation of V beta 8-expressing T cells (65-80% of all CD3+ lymphocytes), together with a marked increase in the titres of serum Ig that specifically bind to F(ab')2 fragments of anti-V beta 8 F23.1 antibodies. Such 'V beta 8-like' Ig V regions are abundantly represented among the IgG2b and mAbs of an unselected collection of hybridomas derived from these mice. These mAbs are not multireactive Ig as they fail to bind to a panel of various antigens and antibodies, but often show simultaneous reactivity with anti-idiotypic mAbs to F23.1 and auto-binding. These molecules may provide the structural basis of V-region specific complementarities, driving the expansion of restricted T and B cell repertoires associated with pathological autoimmunity.

Animals↗

PubMed1991

Proliferative T cell anergy to MIs-1a does not correlate with in vivo tolerance.

Intravenous or intraperitoneal priming of MIs-1b mice with cells from MIs-1a donors drastically reduces secondary in vitro proliferative responses to specific stimulation. We show here that: (i) priming leads to blast transformation of essentially all CD4+ T cells bearing V beta 6 receptors in spleen and lymph nodes, and to their marked clonal expansion; (ii) secondary in vivo (or in vitro) challenges have no effect on the state of activation and numbers of V beta 6 CD4 T cells, which, however, migrate to the site of antigenic exposure; (iii) priming results in the differentiation of specific V beta 6 CD4 T cells to effector helper activities, manifested in vivo by marked increases in the numbers of splenic plasma cells, which include terminally differentiated donor MIs-1a B cells; (iv) primed mice show accelerated 'second set' rejection of antigenic cells; and (v) MIs-1b mice, thymectomized as adults before exposure to MIs-1a cells, show immune responses that are equivalent to those of control animals. We conclude that, in this experimental system, proliferative 'anergy' does not correlate with tolerance but with memory, and relates to the determination of class in immune responses.

Animals↗

PubMed1991

Role of the humoral immune response in resistance to Theiler's virus infection.

Theiler's virus, a murine picornavirus, persists in the central nervous system of susceptible strains of mice, causing chronic inflammation and demyelination in the white matter of the spinal cord. Resistant strains, however, clear the virus and do not develop late disease. In this study, we compared the characteristics of T and B lymphocytes in C57BL/6 (resistant) and SJL/J (susceptible) mice 1 week after intracerebral infection. We detected a marked increase of the number of immunoglobulin M (IgM)-secreting cells in the spleens of C57BL/6 detected a marked increase of the number of immunoglobulin M (IgM)-secreting cells in the spleens of C57BL/6 mice (but not in those of SJL/J mice), which correlated with higher levels of serum IgM antiviral antibodies. The role of the humoral response in virus clearance and resistance was demonstrated by a marked decrease in the number of infected spinal cord cells in SJL/J mice after passive transfer of serum from infected C57BL/6 donors. The B-cell response was found to be partly T cell independent. These results suggest an important role of the early humoral immune response in resistance to Theiler's virus-induced disease.

Animals↗

PubMed1990

V beta 17 gene polymorphism in wild-derived mouse strains: two amino acid substitutions in the V beta 17 region greatly alter T cell receptor specificity.

Of 41 wild-derived mouse strains analyzed, 14 contained T cells bearing V beta 17 receptors in spite of the concomitant expression of I-E antigens. Reciprocal F1 and F2 hybrids of one of these strains, PWK, with laboratory strains revealed different patterns of V beta 17 T cell deletions from those observed with V beta 17 T cells from SJL, implying that the two V beta 17 regions are associated with recognition of distinct superantigens. The structures of the V beta 17 alleles differ by two amino acid substitutions, which lie together in an area distant from the predicted site of T cell receptor interaction with peptide-MHC complexes but overlapping with that implicated in V beta 8.2 recognition of Mls-1 superantigen. This demonstrates that the self-superantigen leading to V beta 17 T cell deletion varies with the allele of the receptor gene and confirms that T cell deletions by such ligands involve interactions with a region of the V beta domain that is distinct from the conventional combining site.

Amino Acid Sequence↗

PubMed1990

Localization of gamma/delta T cells to the intestinal epithelium is independent of normal microbial colonization.

Using monoclonal antibodies identifying all gamma/delta and alpha/beta T cell receptors in cytofluorometric analysis, we have compared the composition of intestinal intraepithelial lymphocytes (i-IEL) in euthymic and athymic germ-free (GF) and conventional (SPF) mice. The results show a marked influence of microbial colonization in the numbers of single-positive (CD4+ or CD8+) alpha/beta i-IEL, but little effect in the pool size or characteristics of gamma/delta i-IEL. In young athymic mice, virtually no alpha/beta i-IEL are detected, while considerable numbers of gamma/delta i-IEL remain, though reduced in GF animals.

Animals↗

PubMed1990

Dibucaine interaction with phospholipid vesicles. A resonance energy-transfer study.

Resonance energy transfer between a local anaesthetic, dibucaine (donor) and a set of functionalized probes [n-(9-anthroyloxy)stearic acids, n = 2, 3, 6, 7, 9 and 12 and 16-(9-anthroyloxy)palmitic acid] (acceptors) was found to be an efficient process with a critical radius of transfer Ro = 2.1 nm, this interaction being used to locate the drug in a model membrane system, small unilamellar vesicles of dipalmitoylglycerophosphocholine, both above and below the temperature of the gel-to-the-liquid-crystal transition of the phospholipid. From the sequence of relative quenching efficiencies of dibucaine fluorescence upon incorporation of the probes, it was concluded that the drug intercalates in the membrane near the glycerol backbone of the lipid. In addition, it was found that dibucaine location is not significantly affected upon crossing the phase-transition temperature of the phospholipid. Dibucaine photophysics was also studied and the short lifetime of the neutral form of the anaesthetic with respect to that of the monoprotonated species was attributed to an intramolecular charge-transfer interaction. From the study of its partition coefficient between the membrane and the aqueous phase, it was concluded that the only significant species present in the membrane is the charged one.

1,2-Dipalmitoylphosphatidylcholine↗

PubMed1990

Thymic epithelium tolerizes for histocompatibility antigens.

The role of thymic epithelium in the establishment of tissue tolerance was analyzed with a murine chimeric system. All T cells differentiated from birth onward in a thymus comprising allogeneic epithelium and syngeneic hematopoietic cells. Embryonic thymic rudiments that contained no hematopoietic cells from C3H (H-2k) donors were grafted to newborn athymic (nude) BALB/c (H-2d) mice. Chimeras that had normal T cell numbers and function rejected third-party skin grafts, but permanently accepted grafts syngeneic to the thymic epithelium. In vitro functional assays did not always correlate with the state of tolerance in vivo. Thus, pure thymic epithelium induces tolerance to histocompatibility antigens.

Animals↗

PubMed1990

Selective peripheral expansion and activation of B cells expressing endogenous immunoglobulin in mu-transgenic mice.

Two different lines of C57BL/6 mice (IgHb) carrying complete rearranged mu chain genes from BALB/c (IgMa) were analyzed for the expression and secretion of endogenous as well as transgenic immunoglobulins at the level of single cells. Quantitation of B cells expressing endogenous IgMb by cytofluorometry, limiting dilution analyses of clonal precursors and secretory cell assays revealed a marked selective expansion, activation and terminal differentiation of those cells producing endogenous immunoglobulins. Thus, the very infrequent IgMb-bearing B cells produced in bone marrow of transgenic mice accumulate in spleen, where they are activated and account for roughly half of all natural immunoglobulin-secreting cells. These observations indicate that mu-transgenic mice are valuable in studies of the antibody repertoire selection operating in unprimed animals but their use could be misleading in the analyzing "monoclonal" immune system.

Animals↗

PubMed1990

Divergency in the specificity of the induction and maintenance of neonatal suppression.

Neonatal treatment of BALB/c mice with anti-VHT15 antibodies suppresses serum expression of VHT15 immunoglobulins in adult animals (2 months) which remains for over 8 months in half of the cases. Suppressed mice, however, contain control numbers of B cells expressing genes of the S107VH family and producing VHT15 after mitogenic stimulation. Furthermore, immunization with phosphorylcholine (PC) breaks suppression and stimulates the production of VHT15 anti-PC antibodies. These animals, however, contain no detectable B lymphocytes expressing the T15 idiotype and produce no T15 idiotype-positive antibodies in response to PC. These results are discussed in the context of lymphocyte repertoire selection.

Animals↗