The Nef-mediated AIDS-like disease of CD4C/human immunodeficiency virus transgenic mice is associated with increased Fas/FasL expression on T cells and T-cell death but is not prevented in Fas-, FasL-, tumor necrosis factor receptor 1-, or interleukin-1beta-converting enzyme-deficient or Bcl2-expressing transgenic mice

CD4(+)- and CD8(+)-T-cell death is a frequent immunological dysfunction associated with the development of human AIDS. We studied a murine model of AIDS, the CD4C/HIV transgenic (Tg) mouse model, to assess the importance of the apoptotic pathway in human immunodeficiency virus type 1 (HIV-1) pathogenesis. In these Tg mice, Nef is the major determinant of the disease and is expressed in immature and mature CD4(+) T cells and in cells of the macrophage/myeloid lineage. We report here a novel AIDS-like phenotype: enhanced death, most likely by apoptosis (as assessed by 7-aminoactinomycin D and annexin V/propidium iodide staining), of Tg thymic and peripheral CD4(+) and CD8(+) T cells. The Tg CD4(+) and CD8(+) T cells were also more susceptible to cell death after activation in vitro in mixed lymph node (LN) cultures. However, activation-induced cell death was not higher in Tg than in non-Tg-purified CD4(+) T cells. In addition, expression of Fas and FasL, assessed by flow cytometry, was increased in CD4(+) and CD8(+) T cells from Tg mice compared to that of non-Tg littermates. Despite the enhanced expression of Fas and FasL on Tg CD4(+) and CD8(+) T cells, Fas (lpr/lpr) and FasL (gld/gld) mutant CD4C/HIV Tg mice developed an AIDS-like disease indistinguishable from lpr/+ and gld/+ CD4C/HIV Tg mice, including loss of CD4(+) T cells. Similarly, CD4C/HIV Tg mice homozygous for mutations of two other genes implicated in cell death (interleukin-1beta-converting enzyme [ICE], tumor necrosis factor receptor 1 [TNFR-1]) developed similar AIDS-like disease as their respective heterozygous controls. Moreover, the double-Tg mice from a cross between the Bcl2/Wehi25 and CD4C/HIV Tg mice showed no major protection against disease. These results represent genetic evidence for the dispensable role of Fas, FasL, ICE, and TNFR-1 on the development of both T-cell loss and organ disease of these Tg mice. They also provide compelling evidence on the lack of protection by Bcl2 against Tg CD4(+)-T-cell death. In view of the high resemblance between numerous phenotypes observed in the CD4C/HIV Tg mice and in human AIDS, our findings are likely to be relevant for the human disease.     

Structurally distinct ligand-binding or ligand-independent Notch1 mutants are leukemogenic but affect thymocyte development, apoptosis, and metastasis differently

We previously found that provirus insertion in T cell tumors of mouse mammary tumor virus/c-myc transgenic (Tg) mice induced two forms of Notch1 mutations. Type I mutations generated two truncated molecules, one intracellular (IC) (Notch1(IC)) and one extracellular (Notch1(EC)), while in type II mutations Notch1 was deleted of its C terminus (Notch1(DeltaCT)). We expressed these mutants in Tg mice using the CD4 promoter. Both Notch1(IC) and Notch1(DeltaCT), but not Notch1(EC), Tg mice developed double-positive (DP) thymomas. These disseminated more frequently in Notch1(DeltaCT) Tg mice. Double (Notch1(IC) x myc) or (Notch1(DeltaCT) x myc) Tg mice developed thymoma with a much shorter latency than single Tg mice, providing genetic evidence of a collaboration between these two oncogenes. FACS analysis of preleukemic thymocytes did not reveal major T cell differentiation anomalies, except for a higher number of DP cells and an accumulation of TCR(high)CD2(high)CD25(high) DP cells in Notch1(IC), and less so in Notch1(DeltaCT) Tg mice. This was associated with enhanced in vivo thymocyte proliferation. However, Notch1(IC), but not Notch1(DeltaCT), DP thymocytes were protected against apoptosis induced in vivo by dexamethasone and anti-CD3 and in vitro by anti-CD3/CD28 Abs. This indicates that the C terminus of Notch1 and/or the conserved regulation by its ligands have a significant impact on the induced T cell phenotype. Therefore, Notch1(IC) and Notch1(DeltaCT) behave as oncogenes for T cells. Because these two Notch1 mutations are very similar to those described in some forms of human T cell leukemia, these Tg mice may represent relevant models of these human leukemias.     

Overexpression of a single Leishmania major gene enhances parasite infectivity in vivo and in vitro

We identified a Leishmania major‐specific gene that can partly compensate for the loss of virulence observed for L. major HSP100 null mutants. The gene, encoding a 46 kD protein of unknown function and lineage, also enhances the virulence of wild type L. major upon overexpression. Surprisingly, the approximately sixfold overexpression of this protein also extends the host range of L. major to normally resistant C57BL/6 mice, causing persisting lesions in this strain, even while eliciting a strong cellular immune response. This enhanced virulence in vivo is mirrored in vitro by increased parasite burden inside bone marrow‐derived macrophages. The localization of the protein in the macrophage cytoplasm suggests that it may modulate the macrophage effector mechanisms. In summary, our data show that even minor changes of gene expression in L. major may alter the outcome of an infection, regardless of the host's genetic predisposition.     

The phenotype and genotype of rheumatoid arthritis in the Democratic Republic of Congo

Introduction

Little is known about rheumatoid arthritis in the black, particularly in Congolese, populations. Our objective was to describe the phenotype and genotype of rheumatoid arthritis (RA) in Congolese.

Methods

All consecutive rheumatoid arthritis (RA) patients attending Kinshasa University Hospital in a three-year time period were included. Demographics, clinical features and tobacco consumption were noted. Disease Activity Score (DAS)-28 based on the erythrocyte sedimentation rate (ESR), Health Assessment Questionnaire (HAQ), anti-citrullinated peptide antibodies (CCP) antibodies and rheumatoid factor (RF) were determined. Radiographs were scored according to Sharp-van der Heijde. On a subset of patients and controls HLA-DRB1 typing was performed.

Results

A total of 114 females and 14 males aged 51.2 ± 14.9 were included. Mean duration of symptoms was four years. Moderate tobacco consumption was reported in a minority of patients. DAS-28 at first visit was >5.1 and HAQ ≥0.5 in all patients. X-rays showed joint erosions and/or joint space narrowing, mostly of a moderate grade in 55.8% of patients. Anti-CCP and/or RF were present in 48.6% of patients with available data (n = 72) and in 3.0% of controls (n = 67). Radiographic changes and nodules were more frequent in RF or anti-CCP positive patients. One copy of the shared epitope was found in 13 patients (35.1%) and 3 controls (12.5%). Two copies were found in one patient (2.7%) and in one control (4.2%).

Conclusion

Congolese patients with RA consult long after disease onset. Despite this delay, the majority presents without major damage and is RF, anti-CCP and SE negative. We put forward the hypothesis that besides different environmental factors there is probably also a particular genetic risk profile in Congolese patients, different from the HLA-DRB1 shared epitope.     

Molecular characterization of <Emphasis Type="Italic">Staphylococcus pseudintermedius</Emphasis> strains isolated from clinical samples of animal origin

The aim of this study was to determine the species distribution among 44 randomly selected clinical isolates (30 mecA-positive and 14 mecA-negative) of animal origin previously identified as Staphylococcus intermedius by phenotypic tests and species-specific PCR amplification of the 16S rRNA gene. For this purpose, we used a multiplex PCR for the detection of the nuc gene and restriction fragment length polymorphism analysis of pta gene amplified by PCR. Both methods allow discrimination of Staphylococcus pseudintermedius from the other closely related members of the S. intermedius group and other coagulase-positive staphylococci isolated from animals. Genetic diversity of S. pseudintermedius strains was analyzed by staphylococcal protein A-encoding gene (spa) typing. Multiplex PCR method was used to identify staphylococcal cassette chromosome mec (SCCmec) type in mecA-positive strains. All isolates previously identified as S. intermedius were shown to belong to S. pseudintermedius. According to PCR-based SCCmec typing, SCCmecIII was the most prevalent type (n = 23), and solely seven isolates were designated as non-typeable. Furthermore, the assessment of spa-typing results revealed that the majority of all strains (n = 27) harbored spa type t02, and 17 strains were classified as non-typeable.     

Structure-function analysis of vitamin D(2) analogs as potential inducers of leukemia differentiation and inhibitors of prostate cancer proliferation

We characterized a structure-function relationships of four analogs of vitamin D(2) with extended and branched side-chains. We tested their ability to induce differentiation of human acute myeloid leukemia (AML) cells both in vitro and ex vivo. Our experiments on five human cell lines revealed substantial differences among tested analogs. Analogs with side-chains extended by one (PRI-1906) or two carbon units (PRI-1907) displayed similar or elevated cell-differentiating activity in comparison to 1,25-dihydroxyvitamin D(3) (1,25D), whereas further extending side-chain resulted in substantially lower biological activity (PRI-1908 and PRI-1909). Similar pattern of cell-differentiating activities to that observed in human cell lines has also been shown in blast cells isolated from patients diagnosed with AML. The ability of the analogs to activate expression of CYP24A1 gene has been studied in HL60 cell line. The analog PRI-1906 activated expression of CYP24A1 similarly to 1,25D, while PRI-1907 weaker than 1,25D. In addition, the analogs PRI-1906 and PRI-1907 were able to moderately inhibit proliferation and significantly activate expression of CYP24A1 mRNA in prostate cancer cells PC-3. Finally, we examined the molecular actions triggered by these analogs and found that their biological activity was related to their ability to induce expression and nuclear translocation of VDR and C/EBPβ.