[Male sexuality after external continent urinary diversion type Mitrofanoff]. / Étude de la sexualité masculine après dérivation urinaire externe continente de type Mitrofanoff.

OBJECTIVE: To evaluate the influence of continent external urinary diversion type Mitrofanoff on male sexuality. MATERIAL AND METHODS: Between 1992 and 2011, 140 patients underwent continent urinary diversion type Mitrofanoff at an academic hospital. Among 76 men, 46 were interviewed about their sexuality after this operation. This study was performed using a set of validated questionnaires (IIEF, DAN PSS and Urolife), grouped by the model of the CTMH. Patients were divided according to their marital status: group 1: patients married before surgery (15 cases), group 2: patients married after surgery (7 cases) and group 3: singles (24 cases). RESULTS: In the first group, the functional dimension of sexuality was positive with an overall score of 81%, the sexual discomfort score was assessed at 26 % and the sexual satisfaction score was 77%. In the second group, sexual function was considered conserved in all cases with a satisfaction score estimated at 98%. These patients reported a feeling of well-being following the disappearance of urinary incontinence with integrity of their body images. In contrast, in the last group, relatively impaired sexual function was noted (65%) with a satisfaction score estimated at 59%. These disorders were multifactorial, mainly related to neurological causal pathology. CONCLUSION: To our knowledge, this is the first study about male sexuality in patients with a continent urinary diversion type Mitrofanoff. Marital status has a major role in the sexuality of these patients. A prospective study with pre- and postoperative evaluation will better clarify the factors affecting sexuality in these patients.

Two distinct Notch1 mutant alleles are involved in the induction of T-cell leukemia in c-myc transgenic mice.

We have previously characterized a large panel of provirus insertion Notch1 mutant alleles and their products arising in thymomas of MMTV(D)/myc transgenic mice. Here, we show that these Notch1 mutations represent two clearly distinct classes. In the first class (type I), proviral integrations were clustered just upstream of sequences encoding the transmembrane domain. Type I Notch1 alleles produced two types of mutant Notch1 RNA, one of which encoded the entire Notch1 cytoplasmic domain [N(IC)] and the other of which encoded a soluble ectodomain [N(EC)(Mut)] which, in contrast to the processed wild-type ectodomain [N(EC)(WT)], did not reside at the cell surface and became secreted in a temperature-dependent manner. A second, novel class of mutant Notch1 allele (type II) encoded a Notch1 receptor with the C-terminal PEST motif deleted (DeltaCT). The type II Notch1(DeltaCT) protein was expressed as a normally processed receptor [N(EC)(WT) and N(IC)(DeltaCT)] at the cell surface, and its ectodomain was found to be shed into the extracellular medium in a temperature- and calcium-dependent manner. These data suggest that both type I and type II mutations generate two structurally distinct Notch1 N(EC) and N(IC) proteins that may participate in tumor formation, in collaboration with the c-myc oncogene, through distinct mechanisms. Constitutive type I N(IC) and type II N(IC)(DeltaCT) expression may enhance Notch1 intracellular signaling, while secreted or shed type I N(EC)(Mut) and type II N(EC) proteins may differentially interact in an autocrine or paracrine fashion with ligands of Notch1 and affect their signaling.

Structure-function analysis of Ia molecules: in-phase insertion mutagenesis of the amino-terminal domain of the E beta k polypeptide chain.

To identify which segments of the beta 1 domain of the E beta k polypeptide control T cell recognition of antigen, E beta genes were constructed with in-phase insertion mutations. Five independent mutants, with insertions mapping to positions 24, 50 and 93 of the E beta k polypeptide, were obtained. Cell lines expressing these mutated genes were analysed by microfluorometry using a panel of 20 anti-Ek monoclonal antibodies. None of the tested in-phase insertions has resulted in the loss of antibody binding sites. In striking contrast, mutations at position 93 and at a lesser level 50 were indicative of a crucial role of the corresponding regions in T-cell recognition, because they led to significant or complete loss of antigen-presenting function with all but one of the T hybridomas tested. These data are discussed with regard to a model of the foreign antigen binding site of Ia molecules.

Epitopic analysis of detergent-solubilized HLA molecules by solid-phase radioimmunoassay.

A solid-phase radioimmunoanalysis of plasma membrane molecules solubilized in the presence of detergent was developed. From a crude cell lysate, membrane molecules were specifically immobilized by solid-phase adsorbed monoclonal antibodies. Analysis of these molecules was easily performed with radiolabeled monoclonal antibodies of different specificities. This technique was used to analyze the HLA-DR and HLA-A,B,C molecules expressed by RAJI cells. It was possible (a) to determine which epitopes are expressed on the same molecules; (b) to define subsets of HLA molecules according to the epitopes which they express, and (c) to perform quantitative analysis of HLA molecules on a crude cell lysate.

Human allospecific T cell proliferative clones: different T cell clones expressing an apparently identical HLA class II specificity are heterogeneous in regard to their proliferative inhibition by a panel of monoclonal antibodies against human Ia-like antigens.

Four independent PLT clones displaying an apparently identical class II specificity (i.e., Dw/DR3) were found to give a heterogeneous pattern of inhibition in relation to 15 anti-class II mAb and an anti-beta 2m mAb used as a control. Some clones were inhibited by all anti-class II mAbs, irrespective of the cluster of molecular subsets with which they reacted. Such clones were also inhibited by the control anti-beta 2m mAb. Other clones were inhibited by only a few of the mAb tested. Within this group of T clones following the addition of a limiting amount of conditioned medium the inhibitory data of all independent clones with an identical specificity were inhibited by the same mAb; under these conditions, it was possible to relate the mAb inhibition patterns with the specificity of the T cell clones and these T cell specificities with the epitopic cluster/molecular subsets defined by these mAbs. A new level of T cell subset heterogeneity within T cell clones with apparent identical proliferative specificities is demonstrated, in relation to the T cell clone "susceptibility" or "resistance" to the effects of anti-class II mAbs directed towards their own Ia-like antigens.

A 55,000 Mr surface antigen on activated human T lymphocytes defined by a monoclonal antibody.

A T cell growth factor-dependent alloreactive human T cell line has been used to generate a monoclonal antibody B1.49.9 that reacts with an antigen present on most if not all mitogen or alloantigen activated T cells but not on resting T cells. The T lymphoblastoid cell line HUT-102 is also strongly reactive with B1.49.9 but all other T and non-T leukemia-lymphoma cell lines tested were negative. The B1.49.9 antigen is a glycoprotein of 55,000 Mr on mitogen or alloantigen activated T cells and 50,000 Mr on the cell line HUT-102. Pulse labeling experiments showed that a 40,000 Mr precursor (at approximately 0.7 h) which does not bind to ricin lectin precedes the appearance of the ricin-binding 55,000 Mr form. Comparisons of the monoclonal antibody anti-Tac, which recognizes the IL-2 receptor, to B1.49.9 suggest that B1.49.9 also recognizes a structure similar or identical to the IL-2 receptor.

A G1 cell cycle arrest induced by ligands of the reovirus type 3 receptor is secondary to inactivation of p21ras and mitogen-activated protein kinase.

The reovirus type 3 S1 gene product (type 3 hemagglutinin; HA3) is the viral protein responsible for binding to a mammalian cell-surface receptor. It has been shown that HA3 binding to its receptor inhibits cell growth, even in the continuous presence of serum mitogens. Here, receptor-mediated signal transduction leading to growth arrest was studied after binding with synthetic or recombinant ligands in the absence of viral infection. Receptor ligation caused rapid inactivation of p21(ras), a decrease in Raf phosphorylation and in mitogen-activated protein kinase (MAPK) enzymatic activity, and G1 cell cycle arrest. Transfection and expression of constitutively active v-Has-ras prevented the G1 arrest, indicating that inactivation of p21(ras) is causative. Interestingly, v-Has-ras expression also decreased the efficiency of reoviridae replication, suggesting that inactivation of p21(ras) signals is required at some step of the viral cycle. This study may define new mechanisms regulating cell growth and support the approach of using viral proteins to identify and study cellular receptors. Synthetic receptor ligands with antiproliferative properties may be useful in drug development with the aim of blocking mitosis.

Structural and genetic analyses of HLA class I molecules using monoclonal xenoantibodies.

The monoclonal antibodies B1.23.2 and B9.12 were developed to characterize human class I major histocompatibility gene products. They detect two different epitopes and define on a given lymphoblastoid B cell line two subsets of beta 2 microglobulin-complexed HLA class I molecules. One subset reacts with B9.12 and B1.23.2 and the other one expresses only the B9.12 epitope. Binding assays were performed on C3H mouse L cells which had been transformed with various single HLA-class I genes and the two detected molecular subsets were shown to be encoded by different genetic loci. Unlike B1.23.2, B9.12 detects all the beta 2m-complexed molecules expressed in human B cell lines and recognizes an epitope different from the one defined by W6/32. In contrast to the vast majority of the other reported anti-class I monoclonal antibodies (including B9.12), B1.23.2 recognizes an epitope expressed on both beta 2m-associated and -free HLA class I heavy chains.

Human cytotoxic T cell structures associated with expression of cytolysis. I. Analysis at the clonal cell level of the cytolysis-inhibiting effect of 7 monoclonal antibodies.

Monoclonal antibodies (mAb) derived from BALB/c mice immunized with human anti-HLA-A2 cloned cytotoxic T lymphocytes (CTL) were screened for their ability to block, in the absence of complement, the cytolytic activity of the immunizing CTL clone. Eight cytolysis-inhibiting mAb have been derived. One of these was directed against a monomorphic determinant expressed on HLA-class I molecules and thus probably inhibited cytolysis via a target cell antigen-masking effect. The 7 other mAb recognized "CTL function-associated structures" and did not have to interfere with target cell antigens in order to inhibit cytolysis. F(ab')2 and Fab fragments of these 7 mAb were also inhibitory. Competitive inhibition of binding and preliminary biochemical analysis suggested that these 7 mAb defined on cloned CTL various epitopes of a structure of 30 kDa disulfide-bonded into several multimeric forms when analyzed without reduction. The inhibitory effect of these 7 mAb has been investigated on a series of short-term (1 month) and long-term (greater than 10 months) expanded cloned CTL lines derived in vitro from an in vivo allosensitized individual exhibiting various specificities. Unexpectedly, only 10% of these CTL clones were inhibited. Flow cytofluorimetric analysis further revealed that noninhibited and inhibited CTL clones expressed similar amounts of the 30-kDa structure. Consequently, the inhibition of CTL was heterogeneous when analyzed at the clonal level and not simply related to the presence or absence of this structure. Furthermore, the ability of CTL clones to be inhibited appeared to be unrelated to their HLA-A, B or C specificity or to their lytic activity. The connections between this mAb-defined structure and CTL function are discussed.

Distinct regulatory elements are required for faithful expression of human CD4 in T cells, macrophages, and dendritic cells of transgenic mice.

To identify the regulatory elements controlling expression of the human CD4 (hCD4) gene in different cell types of the immune system, deletion and chimeric (human/murine) reporter genes were constructed and tested in transgenic (Tg) mice. Regulatory elements required for the proper hCD4 expression in the immature double-positive thymic T cells were identified in the enhancer and in the 3' end of intron 1. Expression of hCD4 in macrophages is controlled by at least 2 sets of regulatory elements: one present in front of exon 1 and the second at the 5' end of intron 1. The hCD4 elements required for expression on both myeloid and lymphoid CD8alpha(+) dendritic cells (DCs) from lymph node and thymus were found to be different from those required for macrophage expression. The results indicate that expression of hCD4 in T cells, macrophages, and DCs is controlled by distinct regulatory elements.

Distinct HLA-DR epitopes and distinct families of HLA-Dr molecules defined by 15 monoclonal antibodies (mAb) either anti-DR or allo-anti-Iak cross-reacting with human DR molecule. I. Cross-inhibition studies of mAb cell surface fixation and differential binding of mAb to detergent-solubilized HLA molecules immobilized to a solid phase by a first mAb.

A series of HLA-DR-reactive mouse anti-human B cell or anti-Ia monoclonal antibodies (mAb) have been used to explore the serological complexity of human class II antigens at the determinant level, using two techniques: (a) cross antibody-binding competitor assays using 125I-labeled and-unlabeled mAb were performed to study the topological organization of the corresponding determinants to determine epitopic clusters recognized by this collection of mAb and (b) differential reactivity of mAb to detergent-solobilized solid-phase-immobilized HLA-DR molecules to determine epitopes expressed on identical DR isotypes. The fifteen mAb could be classified according to the first technique as falling into three different epitopic clusters. Using the second technique, we were able to define at least two independent molecular subsets, one co-expressing two of the three epitopic clusters and the second expressing only the third one. We could not formally identify molecular subsets expressing only one of the first two clusters, using the second technique. The precise serological mapping of the determinants recognized by various anti-class II mAb should prove very useful if such mAb were to be introduced in anti-class II-specific T cell clone blocking experiments. We anticipate that some of them should facilitate the correlation at the clonal level between the T cell repertoire and the epitopes or molecular subsets defined by these mAb. However, within mAb belonging apparently to a same cluster, some could mediate different biological effects.

Murine H-2Dd-reactive monoclonal antibodies recognize shared antigenic determinant(s) on human HLA-B7 or HLA-B27 molecules or both.

We have evaluated the serological relationships between the murine H-2Dd and human HLA molecules using four H-2Dd-reactive monoclonal antibodies (mAbs) produced in the A.BY (KbIbDb) anti-A.TL (KsIkDd) combination. In the mouse, these reagents exhibited three distinct reactivity patterns: Dd, Ks, and H-2u (mAb 81.L); Dd, H-2p, and H-2u (mAb 81.R); and Dd, Kd, H-2p, H-2u, and H-2v (mAbs 97.G and 97.H). Sequential immunoprecipitation and cross-competitive mAb binding experiments revealed that these mAbs recognized determinants in two spatially distinct polymorphic domains on the H-2Dd molecule of B10.A(5R) cells (defined by mAbs 81.L and 81.R, 97.H, and 97.G, respectively). MAbs 81.R, 97.G, and 97.H, but not 81.L, also defined an HLA-linked polymorphism in the human, the main characteristics of which can be summarized as follows: (i) on B lymphoblastoid cell lines, mAbs 81.R and 97.H bound to cells expressing the HLA-B7, HL-B27 or Bw40 cross-reacting specificities, (ii) on peripheral blood lymphocyte (PBL) panel mAb 81.R exerted C dependent cytotoxicity to 118 of 400 cells tested, including almost all HLA-B7 or HLA-B27 cells or both (r: 0.952), (iii) the expression of the 81.R cross-reacting determinant segregated in an informative family with the parental haplotype carrying the HLA-B7 allele, and (iv) mAbs 81.R, 97.G, and 97.H recognized topologically related determinants on the same class I molecule(s) of the human B lymphoblastoid cells JY (HLA-A2,2, -B7,7). These data support the view that some, but not all H-2Dd allotopes have been conserved throughout evolution and are associated in the human with the HLA-B7, -B27 cross-reacting specificities.

Nef harbors a major determinant of pathogenicity for an AIDS-like disease induced by HIV-1 in transgenic mice.

Transgenic (Tg) mice expressing the complete coding sequences of HIV-1 in CD4+ T cells and in cells of the macrophage/dendritic lineages develop severe AIDS-like pathologies: failure to thrive/weight loss, diarrhea, wasting, premature death, thymus atrophy, loss of CD4+ T cells, interstitial pneumonitis, and tubulo-interstitial nephritis. The generation of Tg mice expressing selected HIV-1 gene(s) revealed that nef harbors a major disease determinant. The latency and progression (fast/slow) of the disease were strongly correlated with the levels of Tg expression. Nef-expressing Tg thymocytes were activated and alpha-CD3 hyperresponsive with respect to tyrosine phosphorylation of several substrates, including LAT and MAPK. The similarity of this mouse model to human AIDS, particularly pediatric AIDS, suggests that Nef may play a critical role in human AIDS, independently of its role in virus replication.