Wafa Hanna Koury Cabrera

Innate resistance to infection by intracellular bacterial pathogens differs in mice selected for maximal or minimal acute inflammatory response

The intensity of nonspecific immune reaction and the host resistance to facultative intracellular pathogens are found to be associated in lines of mice selected for maximal (AIRmax) or minimal (AIRmin) acute inflammatory reactivity. AIRmax are more resistant than AIRmin mice to Salmonella typhimurium and Listeria monocytogenes infection, the differences between lines in LD50 being > 1000 and 100 times, respectively. This difference was shown to be related to the initial bacterial containment at the infectious focus, and to the control of bacterial multiplication in the spleen during the 1st week after s.  c. inoculation of the bacteria. Specific immune responses were not deeply affected by the selective process: antibody production and delayed‐type hypersensitivity were both of similar intensity in AIRmax and AIRmin mice. The differential susceptibility to infection seems independent of the Nramp‐1 locus polymorphism; therefore, these two lines represent a powerful model for investigating the role of other genetic loci regulating the nonspecific immunity effectors in the course of infectious diseases.

Pulmonary adenoma susceptibility 1 ( Pas1 ) locus affects inflammatory response

Two outbred mouse lines, phenotypically selected for differential subcutaneous (s.c.) acute inflammatory response (AIR), were analysed for urethane-induced lung inflammatory response and susceptibility to lung tumorigenesis. AIRmin mice, which show a low response to s.c. acute inflammation, developed a persistent subacute lung inflammatory response and a 40-fold higher lung tumor multiplicity than did AIRmax mice, which are selected for high response to s.c. acute inflammation and showed a transient lung inflammatory response. A highly significant linkage disequilibrium pattern was observed in AIRmax and AIRmin mice at marker alleles located within a 452-kb pulmonary adenoma susceptibility 1 (Pas1) locus region, thus defining the location of gene candidacy for inflammatory response and for the biological effects of Pas1 in this region. AIRmin and AIRmax mice segregated by descent the Pas1s and Pas1r alleles, respectively, providing evidence for the involvement of the Pas1 locus in the inflammatory response. The 452-kb region contains Kras2 and four additional genes, including the lymphoid-restricted membrane protein (Lrmp) gene, whose Pro→Leu nonconservative variation was linked with inflammatory response and Pas1 allelotype. These results provide a model to explore the mechanism underlying inherited predisposition to lung cancer in the context of a link to inflammation.

Convergent alteration of granulopoiesis, chemotactic activity, and neutrophil apoptosis during mouse selection for high acute inflammatory response

Neutrophil homeostasis was investigated in two mouse lines, AIRmax and AIRmin, genetically selected for high or low acute inflammatory response (AIR) and compared with unselected BALB/c mice. Mature neutrophil phenotype and functions appeared similar in the three mouse lines. However, an unprecedented phenotype was revealed in AIRmax animals characterized by a high neutrophil production in bone marrow (BM), a high number of neutrophils in blood, a high concentration of chemotactic agents in acrylamide‐induced inflammatory exudates, and an increased resistance of locally infiltrated neutrophils to spontaneous apoptosis. In vitro, BM production of neutrophils and eosinophils was accompanied by an unusual high up‐regulation of cytokine receptors as assessed by antibodies to CD131, which bind the common β chain of receptors to interleukin (IL)‐3, IL‐5, and granulocyte macrophage‐colony stimulating factor. An accelerated neutrophil maturation was also observed in response to all‐trans retinoic acid. Several candidate genes can be proposed to explain this phenotype. Yet, more importantly, the results underline that genetic selection, based on the degree of AIR and starting from a founding population resulting from the intercross of eight inbred mouse lines, which display a continuous range of inflammatory responses, can lead to the convergent selection of alleles affecting neutrophil homeostasis. Similar gene combinations may occur in the human with important consequences in the susceptibility to inflammatory or infectious diseases and cancer.

Pristane-induced arthritis in mice selected for maximal or minimal acute inflammatory reaction.

The role of inflammatory and specific immune responses in pristane‐induced arthritis (PIA) was investigated in mouse lines produced by bi‐directional selective breedings for maximal (AIRmax) or minimal (AIRmin) acute inflammatory reaction, comparing the outcome of PIA and the humoral and cellular response to hsp65. Symptoms of arthritis were detected in 50 % AIRmax mice 120 days after pristane injection, reaching a maximal incidence of 65 %, whereas only 7 % of AIRmin mice developed arthritis within an observation period of 200 days. The production of IgG antibody against hsp65 was found to be similar on both lines, although the IgG1 isotype was predominant in AIRmax, and IgG2a in AIRmin line. In vitro T cell proliferation to hsp65 was similar in the two lines, however, ELISPOT assays carried out soon after pristane treatment, demonstrated higher numbers of IL‐6‐, TNF‐α‐ and IL‐4‐secreting cells in the spleen of AIRmax than in AIRmin mice, while higher numbers of IFN‐γ‐producing cells were found in AIRmin mice. These results suggest a major participation of acute inflammatory mechanisms in the susceptibility to PIA. The genetic background which determines high or low AIR favors a Th2‐like response in susceptible AIRmax and Th1‐like response in resistant AIRmin mice at the initial phase of arthritis induction.

Local inflammatory reaction induced by Bothrops jararaca venom differs in mice selected for acute inflammatory response.

Bothrops jararaca venom (BjV) causes severe systemic and local reactions, characterized by an acute inflammatory reaction with accumulation of leukocytes and release of endogenous mediators. The systemic and local effects of BjV were compared in lines of mice genetically selected for maximal (AIR(max)) or minimal (AIR(min)) acute inflammatory reactivity (AIR). The systemic reaction was evaluated by LD(50) and the local reaction by edema formation, cellular influx, release of PGE(2), NO and H(2)O(2) and the production of the pro-inflammatory cytokines IL-6, Tumor necrosis factor-alpha (TNF-alpha) and IFN-gamma. Both mouse lines were equally susceptible to the lethal effects of the venom showing similar LD(50) but differed significantly in terms of the local inflammatory reaction. Footpad edema and leukocyte influx in the peritoneum after BjV inoculation was higher in AIR(max) compared to AIR(min), BALB/c or outbred Swiss mice. Coincidently, higher levels of the soluble mediators PGE(2), IFN-gamma and TNF-alpha were detected in the inflammatory exudate induced by BjV in AIR(max) mice. Cytokines levels were correlated to in vitro NO and H(2)O(2) production. The results demonstrate that the genetic factors selected in AIR(max) and AIR(min) lines of mice interfere in the control of the acute local reaction triggered by BjV venom.

Slc11a1 (Nramp1) alleles interact with acute inflammation loci to modulate wound-healing traits in mice

Lines of mice were obtained by selective breeding for maximum (AIRmax) or minimum (AIRmin) acute inflammation. They present distinct neutrophil influx and show frequency disequilibrium of the solute carrier family 11a member 1(Slc11a1) alleles. This gene is involved in ion transport at the endosomes within macrophages and neutrophils, interfering in their activation. Homozygous AIRmax and AIRmin sublines for the Slc11a1 gene were produced to examine the interaction of this gene with the acute inflammatory loci. The present work investigated wound-healing traits in AIRmax and AIRmin mice, in F1 and F2 intercrosses, and in Slc11a1 sublines. Two-millimeter ear punches were made in the mice and hole closure was measured during 40 days. AIRmax mice demonstrated significant tissue repair while AIRmin mice did not. Significant differences between the responses of male and female mice were also observed. Wound-healing traits demonstrated a correlation with neutrophil influx in F2 populations. AIRmaxSSshowed higher ear-wound closure than AIRmaxRR mice, suggesting that the Slc11a1S allele favored ear tissue repair. QTL analysis has detected two inflammatory loci modulating ear wound healing on chromosomes 1 and 14. These results suggest the involvement of the acute inflammation modifier QTL in the wound-healing phenotype.

Aryl hydrocarbon receptor polymorphism modulates DMBA-induced inflammation and carcinogenesis in phenotypically selected mice

We tested the role of aryl hydrocarbon receptor (Ahr) gene polymorphism in the inflammatory response and in skin and lung tumorigenesis in 2 lines of mice phenotypically selected for maximum or minimum acute inflammatory reaction (AIRmax and AIRmin, respectively). Following 7,12‐dimethylbenz[a]anthracene (DMBA) treatment, AIRmin but not AIRmax mice showed early skin reactions and eventually developed malignant skin tumors and lung adenocarcinomas. In skin tissue, transcript levels of IL1β, Tnf, Il6, Tgfβ1 and Cyp1b1 genes were upregulated in AIRmin but not AIRmax mice, consistent with the inflammatory responses to the carcinogen. These findings appeared to be related to the homozygosity status of the Ahr functional A375V polymorphism, which influences the binding capability of the receptor for DMBA: the 375A allele, encoding the high‐affinity ligand‐binding receptor (Ahrb1), segregated in AIRmin mice, whereas AIRmax mice carried the 375V, corresponding to the low‐affinity binding receptor (Ahrd), to DMBA. The differential segregation of Ahr functional Ahrdversus Ahrb1 alleles in AIRmax and AIRmin suggests a role for the Ahr gene in the control of inflammatory responsiveness and tumor development of these mouse lines.

Met proto-oncogene juxtamembrane rare variations in mouse and humans: differential effects of Arg and Cys alleles on mouse lung tumorigenesis

Analysis of seven candidate genes mapping in the 1-Mb region of the mouse pulmonary adenoma resistance 4 (Par4) locus revealed a single amino-acid change, consisting in a nonconservative Arg968Cys variation in the juxtamembrane domain of the Met proto-oncogene-encoded protein. The BALB/c strain (resistant allele) carried the Arg allele, whereas the SWR/J mouse strain (Par4-susceptible allele) carried the Cys variation, recently proven to functionally modulate tumorigenesis. Seven genetic linkage crosses herein analysed and six crosses reported in the literature pointed to the candidacy of the Met gene for Par4. Analysis of genomic DNA of 126 lung adenocarcinoma patients for the Met juxtamembrane domain revealed the same Arg/Cys variation at the mouse homologous position in one patient; two other patients carried additional variants in the same domain, suggesting a potential role for rare MET juxtamembrane variants in human lung cancer.

Quantitative trait loci in Chromosomes 3, 8, and 9 regulate antibody production against Salmonella flagellar antigens in the mouse.

Two mouse lines were produced by bidirectional selection according to the high (HIII) or low (LIII) antibody responsiveness against Salmonella flagellar antigens (Selection III). In the present work we conducted a genomewide scan to map the quantitative trait loci (QTL) involved in the antibody response regulation in these selected mice. HIII and LIII genomes were screened with microsatellite markers and those found polymorphic between the lines (146) were used for linkage analysis in F2 (HIII × LIII) intercross. Simple interval mapping analysis was performed using Mapmanager QTX software. Three highly significant QTL linked to antibody production against Salmonella flagellar antigens have been demonstrated in Chromosomes 3, 8, and 9. HIII and LIII lines differ in the resistance to several diseases, therefore, the relevance of these QTL with the genetic factors involved in infections, autoimmunity, and neoplastic disease progression is discussed.

Genetic Control of IL-1β Production and Inflammatory Response by the Mouse Irm1 Locus

Genome-wide linkage analysis using single nucleotide polymorphism arrays was carried out in pedigrees of mice differing in the extent of acute inflammatory response (AIRmax or AIRmin). The AIR phenotype was determined by quantifying the number of infiltrating cells in the 24-h exudate induced by Biogel P-100 s.c. injection and by ex vivo IL-1β production by leukocytes stimulated with LPS and ATP. We mapped the major inflammatory response modulator 1 locus on chromosome 7, at the 1-logarithm of odds (LOD) confidence interval from 116.75 to 139.75 Mb, linked to the number of infiltrating cells (LOD = 3.61) through the production of IL-1β (LOD = 9.35). Of several interesting candidate genes mapping to the inflammatory response modulator 1 locus, 28 of these were differentially expressed in the bone marrow of AIRmax and AIRmin mice. These findings represent a step toward the identification of the genes underlying this complex phenotype.