Activity rhythms in inbred mice. I. Genetic analysis with recombinant inbred strains

Rhythm of activity of inbred mice was recorded automatically by a set of actographs. This rhythm was characterized by six indices in the temporal and frequency domains. Two methods of genetic analysis were applied to these indices using parental strains BALB/c and C57BL/6, the reciprocal F₁'s, and the seven recombinant inbred strains (RI). Findings on the F₁'s show no maternal effect but indicate dominance and heterosis. The RI method successfully rejects the hypothesis of a monogentic correlate for all measures. In line with F₁ data, it demonstrates the presence of a polygenetic correlate: at least one other locus is involved in each of the six outcome parameters.This study was supported by CNRS (URA 1294, affilieé Inserm), MEN (UFR Biomédicale des Saints-Pères, Paris V), Foundation pour la Recherche Medicale.     

Genetic control of mouse cytomegalovirus-induced myocarditis.

Mouse cytomegalovirus (MCMV) infection of mice induced myocarditis, characterized by a mononuclear cell infiltrate with associated necrosis of myofibres. Myocarditis was observed in parallel with viral inclusion-bearing cells in the heart during the acute phase of the infection. Myocarditis also persisted after the acute phase when viral antigens were no longer detectable by immunoperoxidase histochemistry and infectious virus could not be cultivated from various organs. The influence of host genetic factors on the development of cytomegalovirus-induced myocarditis was investigated using H-2 congenic and recombinant inbred mouse strains. Analysis of congenic variants with C57BL/10 and BALB/c backgrounds and the A/J strain revealed that genes linked to the H-2 complex influenced susceptibility to peak levels of MCMV-induced myocarditis seen 7 and 10 days post-infection. In addition, non-H-2 genes of the BALB/c background were important in determining the severity of myocarditis. Analysis of the strain distribution pattern of the CXB recombinant inbred series did not disclose the identity of the BALB/c non-H-2-linked allele conferring susceptibility to MCMV-induced myocarditis. The level of myocarditis seen in the F1 hybrid between the high-responder BALB/c and low-responder C57BL/6 strains suggested dominant inheritance. The amount of viral replication in the major target organs did not correlate with the severity of myocarditis. In conclusion, at least two genes, one mapping to the H-2 complex and another non-H-2-linked gene, influenced the development of myocarditis in MCMV-infected mice.     

Genetic determinants of sleep regulation in inbred mice

Genetic variation in the expression and regulation of sleep was assessed in six inbred mice strains (AK, C, B6, BR, D2, 129). The amount, distribution, and fragmentation of the behavioral states wakefulness (W), slow-wave sleep (SWS), and paradoxical sleep (PS), as well as EEG delta power in SWS, were determined and compared among strains and between baseline and recovery from a 6-hour sleep deprivation (SD) starting at lights-on. In baseline, the most striking strain differences concerned sleep amount, the onset and duration of the main rest period, and SWS fragmentation. The time course of delta power in SWS during the main rest period was similar between strains. Immediately following the SD, high delta power values were reached (higher for AK than for 129). However, the relative increase in delta power, compared to the first 6 hours of the baseline rest period, was not strain-specific. Over the first 6 hours of recovery, W was decreased and PS increased in AK, B6, BR, and 129. In C and D2, time spent in any of the states was not affected by the SD. In contrast, in the recovery dark period, SWS and PS were invariably increased. In recovery, SWS fragmentation was strongly reduced for D2, resulting in the disappearance of the strain differences observed in baseline. Since these inbred strains are fully homozygous and thus can be considered genetic clones, the sleep-related strain differences reported here can be attributed to differences in genotype. Therefore, this study provides a basis for the identification of genetic factors underlying sleep and its regulation.     

Genetic determination of tuberculin hypersensitivity in chicken inbred lines

A dominant inheritance of the ability of CB line chickens to react to tuberculin over the inability of the WB line to develop this type of immune reaction was observed. The tuberculin reactivity in this line seems to be determined by one locus and is associated with the B locus. The involvement of the B locus in the regulation of tuberculin reactivity in other inbred lines of chickens is supported by the reactivity of the CA. IB (N6F2) birds in which the B allele of the IB line was introduced into the CA genotype. The CA. IB (N6F2) chickens manifest strong tuberculin sensitivity, corresponding to the type of reaction of the IB line, not to the weaker type of response of the CA line. The intensity of the tuberculin reaction of the CA and CC lines resembles that of the CB line, but the proportion of nonreactive birds is higher in these two lines than in CB chickens.     

Characterization of age-related changes in natural killer cells during primary influenza infection in mice

The current investigation examined the importance of natural killer (NK) cells during the innate immune response to primary influenza infection in young and aged mice. Young (6-8 weeks) and aged (22 months) C57BL/6 mice were infected intranasally with influenza A virus, and NK cell-mediated cytotoxicity was determined in lung and spleen during the first 4 days of infection. Aged mice demonstrated both a decrease in influenza-inducible NK activity and a reduction in the percentage and number of NK1.1+ cells in response to primary influenza infection, relative to young mice. In order to further establish a role for NK cells in controlling influenza infection, young mice were depleted of NK cells in vivo by injecting rabbit anit-NK1.1 antibody 2 days and 1 day prior to influenza infection. Young mice depleted of NK cells exhibited increased weight loss and lung virus titers during the course of infection, compared to young mice infected with influenza virus. These data indicate that NK cell function is impaired in response to primary influenza infection in aged mice. More importantly, these results underscore the essential role of NK cells in controlling virus titers in lung during the early course of influenza infection, regardless of age.     

Genetic determination of the immune response in mice to cells of different histological type.

The intensity of the cellular immune response to strong transplantation antigens of the H-2 complex is related to the histological type of cells taken for immunization. Splenic cells from C3H mice immunized BALB/c mice better than cells of non-lymphoid origin from the same donor. The reverse was observed when, instead of BALB/c mice, C57B1/6 and A/He were used in immunization experiments. Analysis of the inheritance of this immunization pattern studied in mice of the first generation from various crosses between strains (BALB/c, C57B1/6 and A/He) and in mice from backcrosses has demonstrated that this immunization pattern is not inherited as a simple Mendelian character. Therefore, it could be supposed that efficiency of immunological control of cells having different histological types, may be genetically determined.     

Genetic susceptibility to respiratory syncytial virus infection in inbred mice

Differences in the severity of respiratory syncytial virus (RSV)-induced lower respiratory disease in infants have been attributed to multiple environmental and genetic factors. To identify the genetic factor(s) influencing RSV susceptibility, we examined RSV infection in eight inbred mouse strains. Lung RSV titers differed significantly between mouse strains: the RSV titers were 15-fold higher in AKR/J (permissive) mice compared with C57BL/6J (resistant) mice at 4 days after inoculation. This strain-specific difference in RSV titers suggested that susceptibility to RSV infection was attributable to genetic differences between strains. To examine the mode of inheritance of RSV susceptibility, F1 and backcross (F1 x AKR/J) progeny were infected and RSV titers determined. RSV titers in the F1 progeny were similar to those found in the resistant (C57BL/6J) parent, suggesting resistance was inherited as a dominant trait. The distribution of RSV titers in backcross progeny were discordant with that predicted for a single gene effect, suggesting susceptibility was influenced by more than one gene. These data suggest that RSV susceptibility is a multigenic trait that should be amenable to resolution by genomic analysis.     

Cyclic synthesis of human cytomegalovirus-induced proteins in infected cells.

Synthesis of most human embryo lung cellular proteins is more sensitive to inhibition by a hypertonic condition than is synthesis of cytomegalovirus (CMV)-induced proteins. Results using the selective suppression of host cell protein synthesis by the hypertonic condition indicate that virus-induced proteins are synthesized in a cyclic manner.     

Characterization of human cardiac infiltrating cells post transplantation. 1. Phenotypic and functional alloreactivity

Sequential cardiac biopsies from patients post transplantation were studied for histological evidence of grades of rejection, the immunophenotype of the mononuclear cell infiltrate if present and, in addition, aliquots of the same biopsy were cultured in vitro with medium containing interleukin-2. The exuding mononuclear cells were expanded and bulk cultures and T cell lines resulting from this were evaluated phenotypically and functionally for donor specific alloreactivity. Results of these studies demonstrate: (1) No strict correlation of histological rejection grades with immunophenotype or degree of mononuclear cell infiltrate. (2) The mononuclear cell cultured from 102 biopsies yielded 47 relatively pure CD4+ T cell cultures and 12 cultures enriched for CD4+ T cells, 16 relatively pure CD8+ T cell cultures and 15 cultures enriched for CD8+ T cells, 7 cultures which consisted of dual marked CD4+ CD8+ T cells and 5 which were negative for both CD4 and CD8 but expressed CD3. (3) Of the 59 total CD4+ T cells, 6 demonstrated donor specific CTL reactivity and of the 31 CD8+ T cells, 26 demonstrated donor specific CTL activity. (4) None of the CD4+ CD8+ T cell cultures demonstrated CTL reactivity and all 5 of the CD3+ CD4- CD8- T cell cultures demonstrated potent donor specific CTL activity. (5) Of the 102 cultures, 89 showed donor specific PLT function. (6) High MHC-Class I expression by cardiac myocytes correlated with high frequency of CTL cultures. These data provide a summary of the phenotype and functional analysis of T cells in cardiac biopsy specimens and provide valuable reagents for further studies on the mechanisms involved in human cardiac allograft rejection.     

Genetic complementation of defects in vaccine-induced immunity against Schistosoma mansoni in P- and A-strain inbred mice.

Inbred P- and A-strain mice are deficient in their capacity to develop resistance to challenge infection in response to vaccination with irradiated cercariae of Schistosoma mansoni and are also defective in their cell-mediated response as assessed by the activity of antigen-elicited macrophages in killing schistosome larvae in vitro. In contrast, vaccinated (P x A)F1 mice displayed high levels of both immunity to challenge and macrophage larvicidal activity, indicating that the P- and A-strain defects in the vaccine-induced response are controlled by distinct genetic loci.     

Genetic and sex-linked factors influencing hbs antigen clearance 1. nonimmune clearance in inbred strains of mice

Differences in the clearance values of polyvinylpyrrolidonc (K_PVP) and HBs antigen (K_HBs) were seen in several strains of inbred mice. In addition, in all strains studied K_HBs was greater in females than in males, while K_PVP was not significantly different in the two sexes. In the strain with the lowest K_HBs values, the incidence of HBs antibody formation was lower than in the other strains. These data indicate that genetic and sex-linked factors influence nonimmune clearance of HBs antigen in mice. If similar factors exist in man they may explain some of the variation in clinical manifestation of HBV infection.     

Genetic control of B-cell function. I. Two genes determine the magnitude of antibody responses of inbred mice to red blood cells

In this paper we analyze the details of the genetic control of high and low responsiveness to limiting doses of RBC in inbred mice. We find that high and low responsiveness is expressed in several types of antibody responses, is a quantitative phenomenon, and apparently applies to all red blood cell antigens. Using the primary PFC response to SRBC in vitro as test system, we find that inbred mouse strains of independent genetic origin can be classified as high or as low responders. Breeding experiments revealed segregation ratios in F2 and backcross progeny that clearly show a control by 2 independently segregating genes, one of them linked to the Igh gene complex. An analysis of a variety of congenic strains revealed that a high responder Igh gene complex is necessary but insufficient to maintain high responsiveness and that the additional gene required is neither linked to H-2 nor to Lyt-23 (k-chain genes). An analysis of the CXB RI strains confirmed the important role of Igh and suggested that the additional genetic information may be associated with the minor histocompatibility loci H-21 or H-30.In the following paper of this series we show that high and low responsiveness to RBC is exclusively determined by the B cell. Because of the lack of determinant specificity of the response polymorphism we consider the possibility that the role of Igh in the genetic control may not be restricted to encoding specific Ig receptors for RBC. As an alternative, we consider its role as one of two genes with additive or complementing effects in specifying the ability of B cells to become activated in a T-helper-cell-dependent response. Taken together, we think that we have a system at hand to study the mechanism of B-cell activation by a genetic approach.     

Assortative mating in mice. III. Genetic determination of female mating preference

Females from the inbred strains BALB/Ibg and DBA/Ibg and females from the reciprocal crosses between these strains, which were fostered by BALB/Ibg and DBA/Ibg males, were tested for mating preference between BALB/Ibg and DBA/Ibg males. The tests included the use of both the standard and the tubetechnique methods. In these strains, it was found that the main determinant of female mating preference was the genotype of the foster father. Females reared by DBA/Ibg males had a tendency for negative mating preference, whereas females reared by BALB/Ibg did not. The genotype of the mothers had some effect on the degree of the preference. Females reared by BALB/Ibg mothers had somewhat lower preference for BDA/Ibg males than did daughters' of BDA/Ibg females. The effect of the female's own genotype on mating preference was found to be only minor in a comparison done between these strains and their F ₁ crosses which had been raised by foster parents of the same genotype.This work was supported by NIGMS grant GM- 14547.     

Genetic analysis of the innate immune responses in wild-derived inbred strains of mice

The vertebrate immune system has evolved to recognize nucleic acids of bacterial and viral origin. Microbial DNA, as well as synthetic oligonucleotides based on these motifs, activates innate immune pathways mediated by the family of Toll-like receptors (TLR) initiating a cascade of signals in immune cells necessary for responses to pathogens. However, not all of the proteins that participate in TLR-mediated responses have been identified. In studies described herein, we observed significant variation in innate immune responses among selected wild-derived strains of mice. Specifically, we show that mice of MOLF/Ei, Czech/Ei, and MSM/Ms strains are hypo-responsive to polyinosinic-polycytidylic acid (poly(I:C)) because of a mutation in Tlr3. In addition, we discovered a hypo-response to cytosine guanine dinucleotide in MOLF/Ei mice and established that it is not linked to Tlr9, but to another locus. Further inquiry revealed that this hypo-response is transmitted as a monogenic dominant trait that can be mapped and cloned through positional cloning methods. These results suggest the existence of a novel molecule that can alter pro-inflammatory signals or activate additional signal transduction pathways. In addition, they support the wild-derived mouse strain as a forward genetic tool for the identification of novel immunological phenotypes.     

Genetic control of mast cell development in bone marrow cultures. Strain-dependent variation in cultures from inbred mice.

A comparison was made of the capacity of bone marrow cells (BM) from genetically distinct strains of mice to develop into mast cells under defined conditions of in vitro culture. In the presence of conditioned media derived from ConA treated spleen cells from normal or Trichinella spiralis-infected mice, mast cell development occurred readily. After 21 days of culture mast cells comprised more than 90% of the total cell population. BM taken from certain strains of mice (SWR and NIH) produced large numbers of mast cells, total cell numbers increasing between 2 and 5 fold; other strains (C57BL/10 [B10] B10 congenics) produced relatively few mast cells, total cell numbers not increasing above the starting concentration or declining during culture. The genetic factors determining the strain-response phenotype (no. of mast cells in culture) were predominantly associated with the background genome. No significant differences in response were noted between the B10 congenic strains B10 [H-2b], B10.G [H-2q] or B10.BR [H-2k], which differ only at the MHC, whereas major differences were seen between B10.G and the other H-2q strains [SWR and NIH]. Response phenotype was not inherited as a simple dominant trait; F1 progeny of high x low responder strains were intermediate between the parental values. The expression of genetic influences upon mast cell response phenotype appears to be at both the level of mast cell precursor cells, as determined from limiting dilution assays of BM from high, low and F1 (high x low) strains, and at the level of mast cell proliferation, as determined by repeated sub-culture of mast cells from these strains.     

Characterization of oral Yersinia enterocolitica infection in three different strains of inbred mice

Several studies have highlighted differences in the resistances of various mouse strains to intravenous (i.v.) infection with Yersinia enterocolitica. In particular, differences in resistance and immunological response between BALB/c and C57BL/6 mouse strains have been determined. Following i.v infection, C57BL/6 mice are more resistant to Y. enterocolitica than are BALB/c mice. However, because Y. enterocolitica is typically a food-borne pathogen, the oral route of infection more accurately reflects the natural route of infection. Therefore, it was of interest to ascertain if the differences in resistance between mouse strains observed for an i.v. infection can be recapitulated following an oral infection. C57BL/6j, BALB/cj, and 129X1/Svj mouse strains presented no differences in 50% lethal dose (LD(50)) following oral infection with Y. enterocolitica. Subsequent analysis of cytokine levels, bacterial colonization and immune cell populations following oral infection confirmed characteristics previously described following i.v. Y. enterocolitica infection. All tissues analyzed from each mouse strain demonstrated a polarized Th1 cytokine profile and inflammatory cell influx throughout a 7-day course of infection. This immune response was present in all tissues and increased as bacterial colonization progressed. The lack of a differing LD(50) phenotype and common trends in immunological response among the three mouse strains tested suggests that oral infection is a useful model for studying the host response to Y. enterocolitica infection.     

Respiratory syncytial virus infection in inbred mice.

Respiratory syncytial virus infected the nose and lungs of each of 20 strains of inbred mice, with viral titers varying 100-fold from least permissive to most permissive strains. Viral titers appeared to be under genetic control, but did not correlate with the H-2 haplotype.