Catherine Laporte

Genetic basis of murine lupus

Systemic lupus erythematosus (SLE) is an autoimmune disorder characterized by the formation of a variety of autoantibodies and subsequent development of severe glomerulonephritis. Etiology of SLE remains unknown even if it is now well established that SLE is under polygenic control as well as the contribution of hormonal and environmental factors. The availability of several murine strains that spontaneously develop an autoimmune syndrome resembling human SLE, such as New Zealand, MRL and BXSB mice has provided useful tools for the genetic dissection of susceptibility to SLE. Moreover, development of various transgenic and mutant mice has made it possible to identify a number of susceptibility genes such as those involved in the regulation of apoptosis or B cell receptor signaling that can trigger lupus-like phenotypes. Obviously, further identification of the genetic defects present in lupus-prone mice is of paramount importance for understanding the immunopathogenesis of SLE.

Dysregulated expression of the Cd22 gene as a result of a short interspersed nucleotide element insertion in Cd22a lupus-prone mice

The Cd22 gene encodes a B cell-specific adhesion molecule that modulates B cell Ag receptor-mediated signal transduction, and is allelic to a lupus-susceptibility locus in New Zealand White (NZW) mice. In this study, we show that, in addition to the wild-type transcripts, NZW (Cd22a) mice synthesize aberrant CD22 mRNAs that contain ∼20–120 nucleotide insertions upstream of the coding region between exons 2 and 3, and/or ∼100–190 nucleotide deletions of exon 4. Sequence analysis revealed that these aberrant mRNA species arose by alternative splicing due to the presence in the NZW strain of a 794-bp sequence insertion in the second intron, containing a cluster of short interspersed nucleotide elements. Both the presence of sequence insertion and aberrantly spliced mRNAs were specific to mice bearing the Cd22a and Cd22c alleles. Up-regulation of CD22 expression after LPS activation appeared impaired in Cd22a spleen cells (twice lower than in Cd22b B cells). Furthermore, we show that partial CD22 deficiency, i.e., heterozygous level of CD22 expression, markedly promotes the production of IgG anti-DNA autoantibodies in C57BL/6 (Cd22b) mice bearing the Y chromosome-linked autoimmune acceleration gene, Yaa. Taken together, these results suggest that a lower up-regulation of CD22 on activated B cells (resulting from Cd22 gene anomaly in Cd22a mice or from CD22 heterozygosity in mutants obtained by gene targeting) is implicated in autoantibody production, providing support for Cd22a as a possible candidate allele contributing to lupus susceptibility.

Differential role of three major New Zealand Black-derived loci linked with Yaa-induced murine lupus nephritis

By assessing the development of Y-linked autoimmune acceleration (Yaa) gene-induced systemic lupus erythematosus in C57BL/6 (B6) × (New Zealand Black (NZB) × B6.Yaa)F1 backcross male mice, we mapped three major susceptibility loci derived from the NZB strain. These three quantitative trait loci (QTL) on NZB chromosomes 1, 7, and 13 differentially regulated three different autoimmune traits: anti-nuclear autoantibody production, gp70-anti-gp70 immune complex (gp70 IC) formation, and glomerulonephritis. Contributions to the disease traits were further confirmed by generating and analyzing three different B6.Yaa congenic mice, each carrying one individual NZB QTL. The chromosome 1 locus that overlapped with the previously identified Nba2 (NZB autoimmunity 2) locus regulated all three traits. A newly identified chromosome 7 locus, designated Nba5, selectively promoted anti-gp70 autoantibody production, hence the formation of gp70 IC and glomerulonephritis. B6.Yaa mice bearing the NZB chromosome 13 locus displayed increased serum gp70 production, but not gp70 IC formation and glomerulonephritis. This locus, called Sgp3 (serum gp70 production 3), selectively regulated the production of serum gp70, thereby contributing to the formation of nephritogenic gp70 IC and glomerulonephritis, in combination with Nba2 and Nba5 in NZB mice. Among these three loci, a major role of Nba2 was demonstrated, because B6.Yaa Nba2 congenic male mice developed the most severe disease. Finally, our analysis revealed the presence in B6 mice of an H2-linked QTL, which regulated autoantibody production. This locus had no apparent individual effect, but most likely modulated disease severity through interaction with NZB-derived susceptibility loci.

A necrotic cell death model in a protist.

While necrotic cell death is attracting considerable interest, its molecular bases are still poorly understood. Investigations in simple biological models, taken for instance outside the animal kingdom, may benefit from less interference from other cell death mechanisms and from better experimental accessibility, while providing phylogenetic information. Can necrotic cell death occur outside the animal kingdom? In the protist Dictyostelium, developmental stimuli induced in an autophagy mutant a stereotyped sequence of events characteristic of necrotic cell death. This sequence included swift mitochondrial uncoupling with mitochondrial 2′,7′-dichlorofluorescein diacetate fluorescence, ATP depletion and increased oxygen consumption. This was followed by perinuclear clustering of dilated mitochondria. Rapid plasma membrane rupture then occurred, which was evidenced by time-lapse videos and quantified by FACS. Of additional interest, developmental stimuli and classical mitochondrial uncouplers triggered a similar sequence of events, and exogenous glucose delayed plasma membrane rupture in a nonglycolytic manner. The occurrence of necrotic cell death in the protist Dictyostelium (1) provides a very favorable model for further study of this type of cell death, and (2) strongly suggests that the mechanism underlying necrotic cell death was present in an ancestor common to the Amoebozoa protists and to animals and has been conserved in evolution.

The Sgp3 Locus on Mouse Chromosome 13 Regulates Nephritogenic gp70 Autoantigen Expression and Predisposes to Autoimmunity

By interval mapping of a backcross progeny between New Zealand White (NZW) and C57BL/6 (B6) mice bearing the Y chromosome-linked autoimmune acceleration gene Yaa, we previously identified a genetic locus on mid-chromosome 13, here designated as Sgp3, showing a major effect on the expression of a nephritogenic autoantigen, gp70. In this study, the NZW-derived Sgp3 region was transferred by backcross procedure and marker-assisted selection on the B6 background to produce three independent congenic strains B6.NZW-Sgp3/1, -Sgp3/2, and -Sgp3/3. We show that NZW homozygosity at a single 3 centiMorgans (∼12 megabases (Mb)) interval between markers D13Mit142 and D13Mit254 mediates increased basal serum levels of gp70 in B6.NZW-Sgp3/1 and B6.NZW-Sgp3/2 mice and with a higher degree in males (∼15 μg/ml) than in females (∼9 μg/ml) as compared with B6 (∼2 μg/ml), revealing a gender effect. However, their gp70 levels are still lower than that of NZW mice (∼60 μg/ml). In addition, B6.NZW-Sgp3/1 and B6.NZW-Sgp3/2 mice showed a moderate 2- to 3-fold increase in serum gp70 in response to LPS, which contrasted with over a 10-fold increase in NZW mice. Although both B6.NZW-Sgp3/1 and B6.NZW-Sgp3/2 mice failed to produce significant amounts of gp70 anti-gp70 immune complexes, unexpectedly, aged B6.NZW-Sgp3/2 congenic males bearing the Yaa gene developed increased titers of IgG autoantibodies to DNA and chromatin. Our data indicate that Sgp3 is involved in a complex process of gp70 production under polygenic control and may provide a significant contribution to lupus susceptibility not only through up-regulation of gp70 autoantigen production but also predisposition to autoimmunity.

Cannabis and Young Users—A Brief Intervention to Reduce Their Consumption (CANABIC): A Cluster Randomized Controlled Trial in Primary Care

PURPOSE Brief intervention to reduce cannabis is a promising technique that could be adapted for use in primary care, but it has not been well studied in this setting. We tested the efficacy of a brief intervention conducted by general practitioners among cannabis users aged 15 to 25 years. METHODS We performed a cluster randomized controlled trial with 77 general practitioners in France. The intervention consisted of an interview designed according to the FRAMES (feedback, responsibility, advice, menu, empathy, self-efficacy) model, while the control condition consisted of routine care. RESULTS The general practitioners screened and followed up 261 young cannabis users. After 1 year, there was no significant difference between the intervention and control groups in the median number of joints smoked per month among all users (17.5 vs 17.5; P = .13), but there was a difference in favor of the intervention among nondaily users (3 vs 10; P = .01). After 6 months, the intervention was associated with a more favorable change from baseline in the number of joints smoked (−33.3% vs 0%, P = .01) and, among users younger than age of 18, smoking of fewer joints per month (12.5 vs 20, P = .04). CONCLUSIONS Our findings suggest that a brief intervention conducted by general practitioners with French young cannabis users does not affect use overall. They do, however, strongly support use of brief intervention for younger users and for moderate users.