[Foundation of a Veneta Regional Association of Diabetological Health Personnel].
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Diabetes Mellitus↗
Biomedical subjects
S Marchetto
Publications and source records attributed to S Marchetto.
Accessory molecules and T cell activation. II. Antibody binding to L3T4a inhibits Ia-independent mouse T cell proliferation.
Monoclonal antibody (mAb) blocking assays using L3T4- and lymphocyte function-associated antigen-1 (LFA-1)-specific reagents were used as an approach to investigate the involvement of these accessory molecules in various T cell activation pathways. As previously reported, rat mAb to L3T4a and LFA-1A functional epitopes efficiently blocked antigen-driven T helper cell proliferation. In contrast, antigen- and Ia-independent T cell triggering induced by appropriate mAb to the Thy-1 or the T cell receptor molecules were found to be inhibitable by L3T4a- but not LFA-1A-specific mAb, although the extent of inhibition varied, depending on the cell type and the activating signal examined. These results provide further evidence that the inhibiting effects of L3T4-specific mAb on T cell responses may be due, in addition to an impairment of L3T4-class II major histocompatibility complex molecular interaction, to a down regulatory signal possibly transmitted by the L3T4 molecule itself.
Animals↗
Evidence that murine hematopoietic cell subset marker J11d is attached to a glycosyl-phosphatidylinositol membrane anchor.
Glycosyl-phosphatidylinositol (G-PI) has been shown to serve as membrane anchor for cell surface molecules such as Thy-1, Ly-6-controlled ThB and Qa antigens. Here, we present several lines of evidence indicating that the hematopoietic cell lineage (i.e. thymocytes, B cell subset and red blood cells) marker defined by the rat monoclonal antibody J11d is also a G-PI-linked structure. First, surface expression of the J11d-defined molecules, and that of the related antigen B2A2, was found to be specifically reduced by treatment of thymocytes and B lymphoma or hybridoma cells with excess of Staphylococcus aureus PI-specific phospholipase C; this enzyme also solubilizes a 35-40-kDa material from erythrocyte microsomal membranes corresponding to the predominant J11d-reactive red cell surface molecules. Second, Thy-1- mutants of the BW5147, T1M1, S1A or S49 murine T lymphoma cells of the complementary classes A, B, C and E (i.e. shown to be defective in the enzymatic machinery that posttranslationally modify Thy-1 molecules) also lack J11d, or express it at a very low level. Although directed at a G-PI-linked structure, the J11d monoclonal antibody, unlike other reagents to Thy-1 or Ly-6-controlled antigens, failed to induce thymocyte proliferation even in the presence of phorbol myristate acetate and cross-linker monoclonal antibody.
Animals↗
A Thy-1.1-specific monoclonal alloantibody activates both mouse and rat T cells.
In an attempt to further evaluate the role of Thy-1 in the antigen-independent triggering of mouse T cells, we have examined the activating properties of two Thy-1.1-specific mouse monoclonal antibodies (mAb). These reagents were established from an (A.TH X A.TL)F1 hybrid mouse (Thy-1b) immunized with IL-2 producing (BALB/c (Thy-1b) X BW5147 (Thy-1a)) T hybridoma cells. Although both mAb recognized the same Thy-1.1 determinant, one mAb of the gamma 3,kappa class (H171-146) was found to induce several T hybridoma cells to produce IL-2, and AKR thymocytes or cloned helper T cells to proliferate, whereas another mAb of the gamma 1,kappa class (H171-112) failed to do so even in the presence of phorbol myristic acetate (PMA). Increased IL-2 responses of T hybridoma cells were observed when the cell bound Thy-1.1-specific mAb were crosslinked by goat anti-mouse Ig (GaMIg) antibodies. Both a T-cell activating rat anti-Thy-1.2 mAb and the anti-Thy-1.1 mAb H171-146, although directed at distinct cell surface molecules, synergistically stimulated IL-2 production by T hybridoma cells. In addition, the mouse mAb H171-146 was found to stimulate LOU/M rat thymocytes to proliferate in the presence of exogenous IL-2. These data demonstrate that T cells can use Thy-1 as a signal-transducing molecule in both mouse and rat species, and support the notion that the activating properties of Thy-1.1-specific mAb are influenced by their heavy chain isotypes.
Animals↗
Identification of 5 topographic domains of the mouse LFA-1 molecule: subunit assignment and functional involvement in lymphoid cell interactions.
We have evaluated the serologic and T cell function inhibiting properties of 10 rat mAb reactive with the mouse LFA-1 molecule. Binding inhibition studies revealed that these mAb identified five topographic domains on LFA-1, including an immunodominant epitope region (A) defined by 6 mAb (H35-89, H68-96, H85-326, H129-37, H154-595, and H155-141) and four other spatially separate epitopes each defined by a single mAb (i.e., B, H154-266; C, H129-296; D, H154-163; and E, H155-78). Immunoprecipitation studies carried out with T cell hybridoma detergent lysate containing native or dissociated alpha and beta LFA-1 subunits permitted assignment of the epitopes A, C, and D to the alpha-chain, while expression of the epitopes B and E required homologous pairing of the alpha and beta LFA-1 subunits. These anti-LFA-1 mAb did not bind to the Mac-1 positive P388D1 cells. All the six mAb directed at epitope A inhibited, in the range of 50 to 95%, the proliferative responses of alloantigen- or soluble-antigen GAT-specific T cell clones and the cytolytic activity of I-Ak-specific CTL clones. MAb reactive with the epitopes C and D also blocked these T cell responses, although to a lesser extent. No inhibition was observed with mAb specific to epitope B, whereas the epitope E-specific mAb H155-78 potentiated control T cell responses by 20 to 40%. Suboptimal amounts of anti-L3T4 mAb H129-19 were found to synergistically enhance the T cell function inhibiting properties of mAb to LFA-1 epitopes A, C, and D. These studies reveal an unexpected diversification of LFA-1 between mouse and rat species and further the functional dissection of this molecule.
Animals↗
Accessory molecules and T cell activation. I. Antigen receptor avidity differentially influences T cell sensitivity to inhibition by monoclonal antibodies to LFA-1 and L3T4.
A series of BALB/c-derived T hybridoma cells, capable of producing interleukin 2 (IL 2) in response to poly(Glu60, Ala30, Tyr10) (GAT) presented by syngeneic B lymphoma cells in the context of Ad restriction determinants, was used as a model system to evaluate the roles of LFA-1 and L3T4 accessory molecules in antigen-specific T cell activation. Examination of the antigen requirement for optimal IL 2 responses revealed marked differences in the apparent avidity of these cells for GAT/Ad complexes. A relationship was observed between this parameter and the susceptibility of T hybridoma cells to inhibition by monoclonal antibodies directed at 5 distinct epitopes of LFA-1, and at A beta d allodeterminants. In contrast, L3T4a-specific monoclonal antibodies were found to block in a similar fashion the antigen-specific IL 2 responses of T hybridoma cells, regardless of the apparent avidity of their antigen receptors. It was also shown that both L3T4+ and L3T4- T hybridoma cells were capable of recognizing GAT plus Ad with high avidity. Thus, the quality of T cell antigen recognition appears to critically influence the involvement of LFA-1, and only to a marginal extent that of L3T4, in antigen-specific T cell activation. The implications of these findings are discussed in the context of recent data indicating that L3T4 may not only be an Ia-binding protein.
Animals↗
[Organization of an antidiabetic center].
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Ambulatory Care Facilities↗
Analysis of the Thy-1 pathway of T cell hybridoma activation using 17 rat monoclonal antibodies reactive with distinct Thy-1 epitopes.
Seventeen monoclonal anti-Thy-1 antibodies (mAb) derived from LOU/M rats immunized with mouse T cell clones were used to study the role of Thy-1 in antigen-independent T cell activation. These mAb identified Thy-1.2 or monomorphic determinants and immunoprecipitated a molecule of 25-28 kDa from detergent-solubilized, 125I-labeled T cell surface proteins. Competitive cross-inhibition binding assays demonstrated that these reagents defined 3 epitope groups including either Thy-1.2 (group A) or Thy-1 monomorphic (groups B and C) determinants. Experiments using high titered culture supernatants revealed that all 6 IgG mAb defining the epitope group C, and one IgG2c mAb directed at a determinant in group A were capable of stimulating the terpolymer-L-glutamic acid60-L-alanine33-Ltyrosine10 (GAT) plus I-Ad-reactive BALB/c T cell hybridoma T14-117.9 to produce interleukin 2 (IL2) in the absence of accessory cells. Cross-linking of cell-bound rat mAb by a BALB/c anti-rat kappa chain mAb, or the presence of B cell lymphomas in the culture resulted in an increase of the Thy-1-mediated IL2 responses of this hybridoma. Some mAb from group B required antibody doses exceeding 80 micrograms/ml in order to activate T cells, while others remained nonstimulatory at any dose tested. Striking synergy in mAb-mediated T cell activation was observed when nonmitogenic doses of mAb group groups A and C were mixed in the same culture. Analysis of a panel of GAT plus I-Ad-specific T cell hybridomas revealed that these cells markedly differed in the magnitude of their IL2 responses induced by a given amount of stimulating anti-Thy-1 mAb. Such reagents also stimulated normal thymocytes to express IL2 receptor on their surface. These studies show that the epitopic specificity and the amount of anti-Thy-1 mAb, and the susceptibility of the T cell examined represent important parameters for the triggering of the Thy-1 pathway of T cell activation.
Animals↗
Preferential expression of VK21E light chains on IdX Ia.7 positive monoclonal anti-I-E antibodies.
We previously characterized major (IdX Ia.7) and minor (IdI) idiotopes in a collection of monoclonal alloantibodies reactive with monomorphic (i.e., Ia.7-like) determinants in the structural domain I of the murine class II I-E molecules. In this report, preliminary structural characterization of this antibody family is presented. First, the contribution of isolated H and L chains of the anti-Ia.7 cluster I mAb 41.A to IdX Ia.7 and IdI 41.A idiotope expression was evaluated by testing the capacity of these chains, either isolated or reassociated in homologous or heterologous hybrid Ig, to inhibit the binding of rat or mouse anti-idiotope mAb to IdX Ia.7+ mAb coated plates. It was found that the IdI 41.A idiotope defined by the mouse anti-idiotopic mAb H90-21.1 required the presence of both 41.A H and L chains for complete expression, while the rat mAb-defined IdX Ia.7 idiotope could be detected on isolated and on reassociated 41.A L chain. To evaluate further the structural correlates of the IdX Ia.7 idiotope, H, L, or both H and L chains of 5 A.BY, 4 A.TH and 1 C3H.SW IdX+ anti-Ia.7 mAb, as well as that of 3 A.TH IdX- anti-I-E or anti-I-A and -I-E mAb were subjected to NH2-terminal amino acid sequencing. These analyses demonstrated a) that different H chains corresponding to different subgroups (at least to the VHII and VHIII) could be expressed without apparent modification of IdX Ia.7 idiotope expression and b) that 9 of 11 IdX+ anti-Ia.7 mAb utilized highly homologous L chains of the VK21E subgroup. The relevance of these findings to the genetic control of the idiotypic markers identified in the Ia.7 system is discussed.
Amino Acid Sequence↗
Efficiency of antigen presentation to T cell clones by (B cell X B cell lymphoma) hybridomas correlates quantitatively with cell surface ia antigen expression.
A series of B cell hybridomas was used as a model system to assess quantitatively the role of Ia molecules in antigen presentation to allo- or soluble antigen-reactive T cell clones. These hybrid cell lines were established by fusion between the HGPRT-BALB/c B cell lymphoma M12.4.1 and LPS-stimulated spleen blasts from B10.BR (H-2k) mice. Quantitative cellular absorption of appropriate anti-Ia monoclonal antibodies and flow cytofluorometric analyses revealed that the B cell hybridomas examined herein expressed constitutively a number of surface I-Ak or I-Ek molecules that varied in an order of magnitude of 1 to 5. Such quantitative differences could be correlated precisely with (a) the capacity of B cell hybridomas to activate T cell clones to proliferate and/or to produce interleukin 2 in response to E beta k allodeterminant or to poly(Glu60Ala30Tyr10) presented in the context of I-Ak restriction element, and (b) the amount of monoclonal anti-I-Ak antibody required to inhibit antigen presentation to T cell clones. The possible implications of these data are discussed in the context of current models of regulation of Ia antigen expression by antigen-presenting cells.
Animals↗
Shared idiotope on monoclonal anti-Ia.7 antibodies reactive with determinants in a structural domain of the I-E molecules.
In previous studies, heterologous anti-idiotypic (anti-Id) antisera against the C3H.SW 14-4-4S or the A.TH 41.A anti-Ia.7 monoclonal antibodies (mAb) were shown to identify an interstrain cross-reactive idiotypic specificity (IdX.Ia.7) expressed on monoclonal or conventional anti-Ia.7 alloantibodies. The objective of the present investigation was to characterize further this IdX at the idiotopic level. To this end, 11 hybridomas producing IgG1, IgG2a, or IgM anti-Id mAb were derived from a rat immunized with a mixture of 10 A.TH or A.BY anti-Ia.7 mAb. The specificity of the latter anti-Id mAb was determined by direct Id binding radioimmunoassay (RIA) with the use of a panel of 52 anti-Ia mAb derived from hybridomas produced in various inbred mouse strains. These rat anti-Id mAb recognized idiotopes expressed on i) all anti-Ia.7 mAb against determinants in the topographic domain I of the I-Ek molecule but not on 18 other anti-I-Ek mAb directed at epitopes in domains II or III; ii) three of 19 anti-I-Ak mAb; and iii) one A.TL-derived anti-I-As mAb. Competitive Id binding assays revealed that among the 14 IdX+ anti-Ia.7 mAb, one (81.B) was bound to a lesser extent by various rat anti-Id mAb, suggesting that heterogeneity probably exists in this antibody family. By contrast, two isologous (B10.S(7R)) anti-Id mAb to the IdX.Ia.7+ mAb 41.A displayed a specificity restricted to 41.A individual idiotopes (IdI). Rat anti-IdX.Ia.7 and mouse anti-41.A IdI mAb inhibited the binding of 125I-labeled mAb 41.A to CBA spleen cells. These two sets of mAb bound in a noncompetitive fashion to mAb 41.A-coated plates, indicating that their corresponding public or private idiotopes were spatially distinct. These data may have implications for in vivo manipulations of anti-Ia immune responses.
Animals↗
Dissection of the Poly(Glu60Ala30Tyr10) (GAT)-specific T-cell repertoire in H-2Ik mice. II. The use of monoclonal antibodies to study the recognition of Ia antigens by GAT-reactive T-cell clones.
Twenty-five allospecific monoclonal antibodies (mAb), produced in the A. TH. A.BY, or B10.S (7R) anti-A.TL combinations, were shown to recognize determinants organized in four spatially distinct polymorphic regions on the same I-Ak-encoded molecule(s). These reagents were used to assess the recognition of the class II major histocompatibility complex (MHC) determinants in a series of GAT-reactive A.TL T-cell clones exhibiting various restriction specificity or alloreactivity patterns. Of the proliferative responses of 13 cloned T cells, 12 responses were found to be inhibited similarly by the same set of mAbs.A hierarchy in the blocking effects of these reagents that could be correlated with the spatial organization of their determinants was observed. (i) All the mAbs defining the epitope region I (i.e., recognizing public Ia.1- or Ia.17-like determinants, presumably expressed on the A beta subunit) and some of those identifying new public determinants in the epitope region II profoundly inhibited these T-cell responses. (ii) Intermediate blocking was observed when mAbs recognizing public determinants in the epitope region III were used. (iii) Finally, among the mAbs that identified the epitope group IV, the Ia.19-specific mAb 39.J was inhibitory, whereas mAbs directed against private Ia.2-like determinants were not. By contrast, the GAT-specific proliferative response of the T-cell clone AT-20.1, which recognized its nominal antigen in an extensively cross-reactive MHC-restricted fashion, could only be inhibited by a subset of the mAbs recognizing epitopes in groups I and II, but not by those recognizing epitopes in groups III and IV. It was also shown that the same subset of I-Ak-and I-Au-reactive mAbs displayed similar blocking effects on the proliferation of two T-cell clones exhibiting dual specificity for I-Ak- and I-Au-restricting and/or I-Ak- and I-Au-alloactivating determinants. Finally, all the cloned T-cell responses examined were found to be inhibited by rat mAbs against the LFA.1 molecule or the murine equivalent of the human OKT4 differentiation antigen. These studies suggest that class II specific mAbs can impair proliferation of cloned T-cells by a mechanism(s) other than the masking of the T-cells' restriction determinants per se.
Animals↗
Clonal analysis of B- and T-cell responses to Ia antigens. I. Topology of epitope regions on I-Ak and I-Ek molecules analyzed with 35 monoclonal alloantibodies.
Thirty-five Iak-specific monoclonal alloantibodies, derived from hybridomas constructed by fusion between mouse myeloma and spleen cells from A.TH alloimmune mice (Is anti-Ik), have been used to estimate the allotypic polyporphism of the Ik-gene products. Cross-blocking studies using 17 mAb specific for the I-A molecule indicated that six determinants, which were associated with the conventional specificities Ia.2 and Ia.19, were organized in at least three distinct polymorphic areas of the I-Ak molecules. Similarly, another group of six determinants, which did not correspond to previously described conventional Ia specificities, were found to be topologically heterogeneous. By contrast, the five epitopes associated with the Ia. 1 specificity were clustered into a single region of this molecule. In addition the potentiation of binding observed between mAb specific for topologically distinct epitope regions of the I-Ak molecule, suggested that the latter may undergo conformational changes after binding of a given mAb. A similar analysis of 17 mAb specific for the I-Ek molecule indicated that specificity Ia. 7 of the E alpha chain (as defined in this series by eight mAb) was composed of three topologically distinct polymorphic areas, one of which is also spatially related to a complex cluster of eight new determinants of the I-Ek molecule. Finally, one mAb identified a so far undescribed shared determinant of the I-Ak and I-Ek molecules. The present results, which provide a new estimate of the allotypic polymorphism of the Iak antigens, are discussed with regard to their functional, biochemical, and evolutionary implications.
Animals↗
[Lipoprotein profile estimated by nephelometry in peripheral arterial disease (author's transl)].
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Adult↗
[Nurse-patient communication concerning the education of the diabetic patient: revision of a protocol].
The educational process of a diabetic patient is a long life experience. The monitoring experience of the educational activity with diabetic patients is described. Nurses working at the Diabetic Centre in Rovigo perceived difficulties in the communication with IDDM patients (Insulin Dependent Diabetes Mellitus) especially at the first diagnosis of the disease and during the first patients' encounters. A thorough assessment of the nurse patient communication (analysis of the problems perceived and specific problems encountered; videotapes of nurse-patient communication and discussion with the psychologist on the verbal and non verbal communication styles adopted; discussion with the team on the content of the communication) was the occasion for modifying the information protocols. The experience involved also the nurses caring for patients in the general wards, in order to guarantee continuity in care and information between the ward and the Centre. A 'Survival Kit' for newly diagnosed IDDM patients was devised and an agreement was reached not only on contents to be taught during the first encounters but also on skills that the patients is expected to acquire and on the teaching methods.
Adolescent↗
[Group educational intervention in diabetic foot: the objectives, pedagogical strategies, level of knowledge and results].
Detailed contents and aims of an educational intervention for diabetic foot are presented, together with evaluation tools and results at 3, 6 and 12 month interval after the course. Knowledge level and "health" of the foot were assessed at 12 month interval on 2/3 of the 100 diabetic patients originally exposed to the educational intervention. After 12 months 80% of patients showed a good retention of knowledge on practical issues related to foot care, while 77% of patients with persistent foot problems were not independent (because of hypo-mobility or sight problems) in foot care.
Diabetic Angiopathies↗
Lupus-like onset of recurrent Kawasaki disease in an adolescent boy.
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Adolescent↗
[The methodology and planning of an educational intervention in diabetic foot].
Diabetic foot is considered one of the most threatening and disabling complications for a diabetic patient: lesions of the extremities can become so severe that the person may risk the amputation of the toe, foot or leg. Methodology of planning and implementation of an educational intervention for the prevention of the diabetic foot is presented in its various steps: from the identification of priorities and contents to the problems and difficulties encountered in the implementation of the program.
Diabetes Complications↗