Phenotypic characterization of Lith genes that determine susceptibility to cholesterol cholelithiasis in inbred mice: soluble pronucleating proteins in gallbladder and hepatic biles.

BACKGROUND/AIMS: Gallstone susceptibility is high in C57L inbred mice (males > females) and low in AKR mice, related to variant lithogenic (Lith) genes. We examined the relationship between biliary crystallization-promoting proteins and gallstone susceptibility. METHODS: Biliary protein and lipid concentrations were determined at 0, 7,14, 21, 28 and 56 days on a lithogenic diet. RESULTS: Protein and soluble mucin concentrations in gallbladder biles increased markedly in males, but remained low in females of both strains and correlated with the cholesterol saturation index (CSI). In all groups, IgA and IgM concentrations decreased initially, but increased at later stages. There were no consistent changes in IgG concentrations, but aminopeptidase-N levels were higher in AKR than in C57L. During the lithogenic diet period, the CSI was > or = 2 in C57L males, approximately 1.5 in AKR males, and 1 in females of both strains. Taurodeoxycholate and taurochenodeoxycholate rose sharply in C57L, but remained low in AKR. CONCLUSIONS: Hydrophobic bile salts, cholesterol supersaturation, and possibly, high mucin concentrations are associated with gallstone formation. In vitro crystallization-promoting immunoglobulins and aminopeptidase-N do not appear to be major factors in murine gallstone pathogenesis, in line with the observation that genes encoding these proteins do not co-localize with any known Lith locus.     

Lith1, a major gene affecting cholesterol gallstone formation among inbred strains of mice.

The prevalence of cholesterol gallstones differs among inbred strains of mice fed a diet containing 15% (wt/wt) dairy fat, 1% (wt/wt) cholesterol, and 0.5% (wt/wt) cholic acid. Strains C57L, SWR, and A were notable for a high prevalence of cholelithiasis; strains C57BL/6, C3H, and SJL had an intermediate prevalence; and strains SM, AKR, and DBA/2 exhibited no cholelithiasis after consuming the diet for 18 weeks. Genetic analysis of the difference in gallstone prevalence rates between strains AKR and C57L was carried out by using the AKXL recombinant inbred strain set and (AKR x C57L)F1 x AKR backcross mice. Susceptibility to gallstone formation was found to be a dominant trait determined by at least two genes. A major gene, named Lith1, mapped to mouse chromosome 2. When examined after 6 weeks on the lithogenic diet, the activity of hepatic 3-hydroxy-3-methylglutaryl-CoA reductase (EC 1.1.1.88) was downregulated as expected in the gallstone-resistant strains, AKR and SJL, but this enzyme failed to downregulate in C57L and SWR, the gallstone-susceptible strains. This suggests that regulation of the rate-limiting enzyme in cholesterol biosynthesis may be pivotal in determining the occurrence and severity of cholesterol hypersecretion and hence lithogenicity of gallbladder bile. These studies indicate that genetic factors are critical in determining gallstone formation and that the genetic resources of the mouse model may permit these factors to be identified.     

Molecular genetics of cholesterol cholelithiasis: identification of human and murine gallstone genes

Cholesterol cholelithiasis is one of the most common gastroenterological diseases in Western countries. It is a polygenic disease resulting from disturbed biliary cholesterol homeostasis. Association studies identified six human gallstone candidate genes. Polymorphisms in the genes encoding the apolipoproteins B and E, phospholipid flippase ( ABCB4), cholesterol ester transfer protein ( CETP), cholesterol-7alpha-hydroxylase ( CYP7A1) and ileal bile acid transporter ( SLC10A2) are correlated with gallstone prevalence. Quantitative Trait Locus (QTL) analysis localises additional unknown gallstone genes in inbred mice. Based on the natural variation of cholesterol gallstone susceptibility among different inbred strains, 5 lithogenic ( Lith) loci have been identified. Hepatobiliary transporters (e. g. bile salt export pump Abcb11) and key proteins of the lipoprotein metabolism (e. g. hepatic lipase Lipc) could be established as creedal candidate genes for Lith loci. The rapid progress of mouse and human genome projects provides the basis for the analysis of orthologous human LITH genes in gallstone patients, which might offer new prospects for individual risk assessment and molecular targets for stone prevention.     

Variation of the gene encoding the nuclear bile salt receptor FXR and gallstone susceptibility in mice and humans

BACKGROUND/AIMS: From quantitative trait locus mapping in inbred mice, we identified the Nr1h4 gene encoding the nuclear bile salt receptor FXR as a candidate gene for the cholesterol gallstone susceptibility locus Lith7. Here, we investigated further an association of the gene encoding FXR and gallstone susceptibility in mice and humans. METHODS: The Nr1h4 gene was sequenced in inbred mouse strains with susceptible and resistant Lith7 alleles. Quantitative RT-PCR was employed to determine mRNA expression levels. Gallstone carriers and control subjects of three different populations comprising 1004 individuals were genotyped for polymorphisms of the orthologous human gene detected by sequencing. RESULTS: Expression and sequence analyses in inbred mice were consistent with Nr1h4 underlying Lith7. In the human populations, we identified three frequent haplotypes that accounted for > 95% of all haplotypes observed. In a Mexican population, the most common haplotype NR1H4_1 was associated with gallstone prevalence. In contrast, NR1H4_1 displayed no association with gallstone prevalence in a German population, whereas in a Chilean population we observed a trend towards a protective effect of NR1H4_1. CONCLUSIONS: Our study in an inbred mouse model and in three ethnically distinct populations indicates complex interactions of NR1H4 alleles and other risk factors for the development of cholelithiasis.     

Genome-wide analysis of hepatic fibrosis in inbred mice identifies the susceptibility locus Hfib1 on chromosome 15

BACKGROUND & AIMS: Host genetic factors are likely to contribute to the variable course of hepatic fibrosis in response to chronic liver injury. Similarly, the fibrotic response differs among inbred mouse strains after challenge with CCl(4). Our aim was to identify unknown susceptibility loci for hepatic fibrosis in a cross between fibrosis-susceptible and -resistant inbred mice. METHODS: Seven inbred mouse strains were treated with CCl(4), and hepatic fibrosis was phenotypically characterized by histology, hepatic hydroxyproline levels, and serum surrogate markers. F(1) hybrids of susceptible BALB/cJ and resistant A/J inbred strains were intercrossed to obtain 358 F(2) progeny. Quantitative trait loci (QTL) that determine hepatic fibrosis were identified by genome-wide interval mapping and haplotype analysis. RESULTS: In this model, marked strain differences in fibrosis susceptibility exist, with BALB/c inbred mice being most susceptible. The hydroxyproline levels of F(1) mice resemble the resistant parental strains, indicating that fibrosis susceptibility is a recessive trait. QTL analysis identifies a susceptibility locus on chromosome 15 that significantly affects the stage of fibrosis and hydroxyproline levels. According to standard nomenclature, this locus is called Hfib1 (hepatic fibrogenic gene 1). Hfib1 is defined by genetic markers D15Mit26 and D15Mit122. A suggestive QTL on chromosome 2 colocalizes with the complement factor 5 gene, known to be mutated in the resistant strain A. CONCLUSIONS: The set of inbred strains provides a framework for systematic analysis of fibrogenic genes. QTL mapping is useful to identify genetic susceptibility loci for hepatic fibrosis that might harbor new molecular targets for antifibrotic drug design.     

Major quantitative trait locus on chromosome 2 for glucose tolerance in diabetic SMXA-5 mouse established from non-diabetic SM/J and A/J strains

AIMS/HYPOTHESIS: The SMXA-5 mouse is one of the SMXA recombinant inbred substrains established from the non-diabetic SM/J and A/J strains, and is a model for polygenic type 2 diabetes, characterised by moderately impaired glucose tolerance and hyperinsulinaemia. These diabetic traits are worsened by feeding a high-fat diet. The aim of this study was to dissect the diabetogenic loci in the A/J regions of the SMXA-5 genome that contribute to diabetes-related traits. MATERIALS AND METHODS: We analysed the quantitative trait loci (QTL) for diabetes-related traits and obesity in (SM/JxSMXA-5)F(2) intercross mice fed a high-fat diet. To verify the function of the responsible locus that was mapped in the present study, we constructed a congenic strain and characterised its diabetes-related traits. RESULTS: A major QTL for glucose tolerance, free-fed blood glucose concentration and BMI was mapped on chromosome 2. This locus existed near D2Mit15, with the highest logarithm of the odds score (12.6) for glucose concentration at 120 min in a glucose tolerance test, and was designated T2dm2sa. The diabetogenic allele of T2dm2sa originated in the A/J strain. SM.A-T2dm2sa, a congenic strain that introgressed the T2dm2sa region of A/J genome into SM/J, exhibited overt impaired glucose tolerance and hyperinsulinaemia. CONCLUSIONS/INTERPRETATION: The development of impaired glucose tolerance in SM.A-T2dm2sa mice confirmed the results of QTL analysis for diabetes-related traits in F(2) intercross mice. The present results suggest that there are latent diabetogenic loci in the genomes of non-diabetic A/J and SM/J mice, and that the coexistence of these loci, including T2dm2sa, causes impaired glucose tolerance in SMXA-5 and SM.A-T2dm2sa mice.     

Cholelithiasis: genetic hypothesis

In inbred mice gallstone susceptibility is determined by Lith (lithogenic) genes which promote cholesterol hypersecretion in bile as a response to a high-fat diet. At least three major classes of proteins can be considered as candidate genes: (a) enzymes involved in cholesterol metabolism regulation; (b) trans-membrane carrier proteins from hepatocyte into bile; (c) cytosolic transfer proteins which regulate intrahepatocyte trafficking. The main candidates are: Spgp, a transmembrane protein which produces bile salt transport. Its gene map in Lith 1 region (Chromosome 2) and its expression is increased in susceptible inbred strains of mice (C57L) of inbred mice. Cmoat is a carrier protein that promotes the secretion of conjugated substances in bile. Its gene map in Lith 2 region (Chromosome 19) and its expression is increased in susceptible inbred strains of mice fed on a lithogenic diet. HGMR is the enzyme that regulates de novo synthesis of cholesterol in the liver. Its activity increases in resistant strains fed on a lithogenic diet and unvaries in susceptible strains. Its gene does not reside in any Lith region, but one Lith gene could be responsible for its activity regulation. To date, the hypothesis of a genetic basis of cholelithiasis in men has only been investigated in one study, in which the association of cholelithiasis and a mutation in the C7AH gene was documented in a group of Mexican patients.     

Quantitative trait loci analysis of tail tendon break time in mice of C57BL/6J and DBA/2J lineage

Tail tendon break time (TTBT), a measure of collagen cross-linking, shown to increase with age differs significantly among inbred strains of mice, indicating underlying genetic influences. This study was aimed to identify quantitative trait loci (QTLs) associated with tail tendon break time at three ages (200, 500, and 800 days of age) for 23 BxD recombinant inbred strains of mice and B6D2F(2) mice derived from C57BL/6J and DBA/2J strains. Heritability estimates were calculated, and QTL analyses were conducted using interval-mapping methods. Mean tail tendon break time values were higher in males and increased nonlinearly with age. Eight total QTLs were nominated in the B6D2F(2) mice at the three measured ages, with the QTL at 800 days confirmed in the recombinant inbred strains. Allelic effect modeling for the identified QTLs suggests differences in gene action between sexes. Candidate genes in the QTL regions include collagen genes and an advanced glycation end-product receptor. The QTLs identified demonstrate influence at some but not all ages.     

Chromosomal organization of candidate genes involved in cholesterol gallstone formation: a murine gallstone map

Epidemiologic and family studies indicate that cholesterol gallstone formation is in part genetically determined. The major contribution to our current understanding of gallstone genes derives from animal studies, particularly cross-breeding experiments in inbred mouse strains that differ in genetic susceptibility to cholesterol gallstone formation (quantitative trait loci mapping). In this review we summarize how the combined use of genomic strategies and phenotypic studies in inbred mice has proven to be a powerful means of dissecting the complex pathophysiology of this common disease. We present a "gallstone map" for the mouse, consisting of all genetic loci that have been identified to confer gallstone susceptibility as well as putative candidate genes. Translation of the genetic loci and genes between mouse and human predicts chromosomal regions in the human genome that are likely to harbor gallstone genes. Both the number and the precise understanding of gallstone genes are expected to further increase with rapid progress of the genome projects, and multiple new targets for early diagnosis and prevention of gallstone disease should become possible.     

Distinct phenotypes of obesity-prone AKR/J, DBA2J and C57BL/6J mice compared to control strains

OBJECTIVE: To characterize and compare three obesity-prone inbred strains, AKR/J, DBA/2J and C57BL/6J, to three control strains, C3H/HeJ, BALB/cByJ and C57L/J, selected based on their normal eating patterns and moderate weight gain on high-calorie diets. METHODS AND PROCEDURES: These six strains were examined at 5 weeks of age while still of normal body weight, and they were maintained for 1 day or 3 weeks on different feeding paradigms with macronutrient diets. Measurements were taken of macronutrient intake, body weight and body fat accrual, circulating hormones and metabolites, and the hypothalamic peptide, galanin. RESULTS: The three control strains each selected a balanced diet with 50% carbohydrate and 15-25% fat when given a choice of macronutrients, and they had similar, normal range of scores for the measures of body weight, adiposity, the hormones, insulin and leptin, and the metabolites, glucose and triglycerides. When compared to this control baseline, the obesity-prone strains with similar total caloric intake to controls selected a diet with significantly more fat (30-40%) and less carbohydrate (<40%). They also had greater adiposity, with the largest differences detected for the AKR/J and DBA/2J strains. These two obesity-prone strains compared to control strains had elevated levels of insulin and leptin. They also had higher triglyceride levels and increased expression and levels of galanin in the hypothalamic paraventricular nucleus. A very different pattern was detected in the obesity-prone C57BL/6J strain, which exhibited a stronger preference for protein as well as fat, normal levels of insulin, leptin and triglycerides, hyperglycemia relative to all other strains, and a small increase in galanin. CONCLUSION: These comparisons to control strains revealed a distinct phenotype in the two obesity-prone strains, AKR/J and DBA/2J, which is very similar to that described in obesity-prone, outbred rats. They also identified a clearly different phenotype in the obesity-prone C57BL/6J strain.     

Hyperhomocysteinemia from trimethylation of hepatic phosphatidylethanolamine during cholesterol cholelithogenesis in inbred mice

Because hyperhomocysteinemia can occur in cholesterol gallstone disease, we hypothesized that this may result from trimethylation of phosphatidylethanolamine (PE), which partakes in biliary phosphatidylcholine (PC) hypersecretion during cholesterol cholelithogenesis. We fed murine strains C57L/J, C57BL/6J, SWR/J, AKR/J, PE N-methyltransferase (PEMT) knockout (KO), PEMT heterozygous (HET), and wildtype (WT) mice a cholesterol/cholic acid lithogenic diet (LD) for up to 56 days and documented biliary lipid phase transitions and secretion rates. We quantified plasma total homocysteine (tHcy), folate, and vitamin B12 in plasma and liver, as well as biliary tHcy and cysteine secretion rates. Rate-limiting enzyme activities of PC synthesis, PEMT and cytidine triphosphate: phosphocholine cytidylyltransferase (PCT), S-adenosylmethionine (SAM), and S-adenosylhomocysteine (SAH) were measured in liver homogenates. Other potential sources of plasma tHcy, glycine N-methyltransferase (GNMT) and guanidinoacetate N-methyltransferase (GAMT), were assayed by gene expression. Plasma tHcy and PEMT activities became elevated during cholelithogenesis in gallstone-susceptible C57L, C57BL/6, and SWR mice but not in the gallstone-resistant AKR mice. Persisting in C57L mice, which exhibit the greatest Lith gene burden, these increases were accompanied by elevated hepatic SAM/SAH ratios and augmented biliary tHcy secretion rates. Counter-regulation included remethylation of Hcy to methionine concurrent with decreased folate and vitamin B12 levels and Hcy transsulfuration to cysteine. Concomitantly, methylenetetrahydrofolate reductase (Mthfr), betaine-homocysteine methyltransferase (Bhmt), and cystathionine-β-synthase (Cbs) were up-regulated, but Gnmt and Gamt genes were down-regulated. PEMT KO and HET mice displayed biliary lipid secretion rates and high gallstone prevalence rates similar to WT mice without any elevation in plasma tHcy levels. CONCLUSION: This work implicates up-regulation of PC synthesis by the PEMT pathway as a source of elevated plasma and bile tHcy during cholesterol cholelithogenesis.     

Genetic dissection of quantitative trait loci specifying sedative/hypnotic sensitivity to ethanol: mapping with interval-specific congenic recombinant lines

BACKGROUND: Linkage studies alone do not produce sufficient resolution to narrow the location of a quantitative trait locus (QTL) to a small-enough chromosomal region for gene identification. One solution to this problem is to use interval-specific congenic recombinant (ISCR) lines to narrow the chromosomal interval known to contain the QTL. In previous work, we mapped four QTLs for differential ethanol sensitivity in the inbred long-sleep (ILS) and inbred short-sleep (ISS) strains and generated reciprocal congenic strains in which each full QTL interval from ILS was bred onto the ISS background and vice versa. METHODS: ISCR lines were derived by identifying mice carrying recombination events in the congenic interval during backcrossing of the ISS.ILS.Lore congenics to ISS. Recombinant mice were backcrossed to ISS, and progeny carrying the ISCR chromosome were identified and tested to determine whether the ISCR region carried the donor Lore QTL. RESULTS: We developed multiple ISCR lines for each Lore QTL, in which the QTL interval was broken into a number of smaller intervals. For all four QTLs, we reduced the size of the interval, in one case to 3.7 cM. CONCLUSIONS: Use of ISCR lines can narrow each Lore candidate region to a few centimorgans. Such an interval size is conducive to brute-force approaches to identify candidate genes, entailing bioinformatics, gene expression, and DNA sequencing strategies.     

Identification of two novel female-specific non-major histocompatibility complex loci regulating collagen-induced arthritis severity and chronicity, and evidence of epistasis

OBJECTIVE: To identify additional sex-specific and epistatic quantitative trait loci (QTL) regulating collagen-induced arthritis (CIA) severity overall, as well as within different stages during the disease course, in an intercross between major histocompatibility complex-identical inbred rat strains DA/Bkl (susceptible) and ACI/Hsd (resistant). METHODS: Arthritic male (DA x ACI)F2 intercross offspring (n = 143) were analyzed separately from the females (n = 184). Phenotypic extremes (maximum arthritis scores [MAS]) were genotyped and used for QTL analysis. All 327 rats were genotyped with the simple sequence-length polymorphism (SSLP) markers closest to the peak of Cia7 and Cia10, the major loci previously identified in this intercross, and with SSLPs covering chromosomes 12 and 18. Phenotypes studied were disease onset, arthritis severity scores on days 14-39, MAS, mean and cumulative arthritis scores, delayed-type hypersensitivity, and antibody responses to rat type II collagen. RESULTS: A new female-specific arthritis-severity recessive locus was identified on rat chromosome 12 (Cia25), with a maximum effect observed on day 28 (logarithm of odds [LOD] 4.7). The homozygous DA genotype at Cia25 was associated with a 45% higher median arthritis score in females. Sequencing analyses of the Cia25 candidate gene Ncf1 revealed polymorphisms between DA and ACI. The previously identified locus, Cia10, was found to be male-specific. A 2-locus interaction model analysis identified a novel recessive chromosome 18 QTL, Cia26, which was dependent on Cia7, with its maximum effect observed at later stages during the disease course (peak LOD score of 3.6 for arthritis scores on day 39). CONCLUSION: This study identified 2 novel female-specific loci, and 1 male-specific locus. Cia25 regulates MAS and disease severity during the mid-to-late stages of the disease course and may be accounted for by Ncf1 polymorphisms. Cia26 is in epistasis with Cia7 and regulates later stages of disease, suggesting an involvement in disease perpetuation and/or chronicity.     

Cholesterol synthesis inhibition distal to squalene upregulates biliary phospholipid secretion and counteracts cholelithiasis in the genetically prone C57L/J mouse

BACKGROUND AND AIMS: Newly synthesised cholesterol contributes poorly to biliary lipid secretion but may assume greater importance when the rate limiting enzyme 3-hydroxy-3-methylglutaryl-CoA reductase (HMGR) is upregulated. As this occurs in the gall stone susceptible C57L/J inbred mouse, we employed two cholesterol biosynthesis inhibitors, Tu 2208 and Ro 48-8071, potent inhibitors of squalene epoxidase and oxidosqualene-lanosterol cyclase, respectively, to assess their potential in preventing cholesterol cholelithiasis in the C57L/J mouse strain. Mice were fed a lithogenic diet comprising a balanced nutrient intake with 15% dairy fat, 1% cholesterol, and 0.5% cholic acid added. METHODS: We determined gall stone phenotype, HMGR activity, biliary lipid secretion rates, and counterregulatory events in male C57L/J mice and gall stone resistant AKR treated with Tu 2208 (30-60 mg/kg/day) or Ro 48-8071 (30-100 mg/kg/day), while ingesting chow or the lithogenic diet. RESULTS: Both agents reduced the gall stone prevalence rate from 73% to 17% in C57L/J mice, inhibited HMGR activity, and decreased hepatic cholesterol concentrations without appreciably influencing biliary cholesterol secretion. In C57L as well as AKR mice, both agents increased biliary phospholipid (which is mostly phosphatidylcholine) secretion rates and at the highest doses effectively reduced the biliary cholesterol saturation index. CONCLUSIONS: Cholesterol biosynthesis inhibitors acting distally to squalene do not reduce biliary cholesterol secretion rates despite reductions in cholesterol biosynthesis and hepatocellular levels. However, they effectively prevent gall stone formation through stimulation of pathways that lead to enhanced biliary phospholipid secretion.     

Allelic variation in the GABA A receptor gamma2 subunit is associated with genetic susceptibility to ethanol-induced motor incoordination and hypothermia, conditioned taste aversion, and withdrawal in BXD/Ty recombinant inbred mice

BACKGROUND: Behavioral genomics has made dramatic progress toward mapping quantitative trait loci (QTLs) that contain genes responsible for phenotypic differences in a variety of behavioral responses to alcohol (ethanol). We previously identified a QTL on mouse Chromosome 11 that affects genetic predisposition to acute alcohol withdrawal. Among mice derived from the C57BL/6J (B6) and DBA/2J (D2) inbred strains, this QTL (Alcw3) accounts for 12% of the genetic variability in withdrawal liability. Candidate genes within this QTL encode the gamma-aminobutyric acid type A (GABA A) receptor gamma2, alpha1, alpha6, and beta2 subunits. We recently identified a coding sequence polymorphism between the B6 and D2 strains for the GABA A receptor gamma2 subunit gene (Gabrg2). In this study, we expand our analysis to a panel of BXD strains derived from the B6 and D2 progenitor strains. These BXD strains provide 26 fixed recombinant genotypes that can be used to examine genetic correlations, for example, between a phenotype of interest and allelic variation in a candidate gene. METHODS: Gabrg2 was cloned and sequenced from the 26 BXD recombinant inbred strains. We analyzed genetic correlations between allelic variation in Gabrg2 and alcohol phenotypes previously measured in the BXD strain means. RESULTS: Allelic variation in Gabrg2 is correlated genetically with predisposition to acute alcohol withdrawal and may underlie the Alcw3 locus. In addition, Gabrg2 is associated with ethanol-conditioned taste aversion, ethanol-induced motor incoordination, and ethanol-induced hypothermia. A trend is observed for chronic ethanol withdrawal, ethanol-induced loss of righting reflex, and tolerance to ethanol-induced hypothermia and ataxia. CONCLUSIONS: Functionally relevant variation in Gabrg2, or a closely linked gene, is correlated genetically with some, but not all, behavioral responses to alcohol. The alcohol-related phenotypes associated with Gabrg2 generally may be characterized as debilitating or motivationally negative.     

Arthritis in MRL/lpr mice is under the control of multiple gene loci with an allelic combination derived from the original inbred strains

OBJECTIVE: To clarify the mode of inheritance and the genome origins of arthritis in a lupus-prone strain of mice, MRL/MpJ, bearing a Fas deletion mutant gene, lpr (MRL/lpr). METHODS: Using non-lupus-prone strains of mice, C3H/HeJ-lpr/lpr (C3H/lpr), (MRL/lpr x C3H/lpr)F(1) intercross and MRL/lpr x (MRL/lpr x C3H/lpr)F(1) backcross mice were prepared. Arthritis in individual mice was analyzed by histopathologic grading, and the genomic DNA of the backcross mice was examined by simple sequence-length polymorphism analysis to determine the polymorphic microsatellite markers highly associated with arthritis. RESULTS: Arthritis-susceptibility loci with significant linkage were mapped between D15Mit111 and D15Mit18 (map position 17.8-18.7 cM) on chromosome 15 and between D19Mit112 and D19Mit72 (map position 43.0-55.0) on chromosome 19 (logarithm of odds scores 3.5 and 4.3, respectively). Three other loci, one mapped to each of chromosomes 1, 2, and 7, showed suggestive linkage. Loci homozygous for MRL alleles on chromosomes 1 and 19 enhanced arthritis in both sexes, whereas other loci on chromosomes 2 and 15 selectively affected males. A locus homozygous for MRL alleles on chromosome 7 inhibited arthritis in both sexes. Three of these loci were found to originate from an LG/J strain and 1 from an AKR/J strain. Some combinations of these loci showed an additive effect in a hierarchical manner on the development of arthritis. CONCLUSION: Arthritis in MRL/lpr mice is a complex pathologic manifestation resulting from the cumulative effect of multiple gene loci with an allelic combination derived from the original inbred strains.     

von Willebrand disease in the RIIIS/J mouse is caused by a defect outside of the von Willebrand factor gene [published erratum appears in Blood 1995 Sep 15;86(6):2461]

An animal model for human type I von Willebrand disease (vWD) has been previously described in the inbred mouse strain RIIIS/J. Murine vWD is characterized by a prolonged bleeding time, normal von Willebrand factor (vWF) multimer distribution, autosomal dominant inheritance, and proportionately decreased plasma vWF antigen, ristocetin cofactor, and factor VIII (FVIII) activities. To study the molecular genetics of murine vWD, a portion of the vWF gene surrounding exon 28 was cloned, sequenced, and used to develop two informative DNA sequence polymorphisms for rapid genotyping by DNA polymerase chain reaction. RIIIS/J mice were crossed with PWK/Ph mice, an inbred line of Mus musculus musculus, and the F1 progeny backcrossed to the parental PWK/Ph strain. vWF antigen levels in F1 mice were not significantly different from the parental RIIIS/J strain but were markedly decreased compared with the parental PWK/Ph mice. Genetic linkage analysis of 104 backcross progeny showed no correlation between vWF antigen level and vWF genotype. These data indicate that murine vWD is caused by a defect at a novel genetic locus, distinct from the murine vWF gene. The distribution of vWF antigen levels among backcross progeny suggests the presence of one major dominant vWD gene in the RIIIS/J mouse with possible modifying contributions from one or more additional minor loci. These observations may provide new insights into the molecular basis and variable expressivity of human vWD.     

Mapping quantitative trait loci that influence serum insulin-like growth factor binding protein-5 levels in F2 mice (MRL/MpJ X SJL/J)

Recent studies using twins and inbred strains of mice reveal evidence for genetic mechanisms contributing to variation in circulating levels of IGF-I, IGF-II, and IGF binding protein (IGFBP)-3. To examine the hypothesis that serum IGFBP-5 levels have a strong heritable component, we intercrossed two inbred strains of mice, MRL/MpJ and SJL, which exhibit 79% difference in serum IGFBP-5 levels (554 +/- 68 vs. 309 +/- 51 ng/ml respectively, P < 0.001). A genome-wide scan was carried out using 137 polymorphic markers in 633 F2 female mice. Serum IGFBP-5 levels in the F2 progeny showed a normal distribution with an estimated heritability of 74%. Whole genome-wide scans for cosegregation of genetic marker data with high or low serum IGFBP-5 levels revealed six different quantitative trait loci (QTL) in chromosomes 1, 9 (two), 10, and 11 (two), which together explained 24% of F2 variance. Chromosome 11 QTL exhibited the highest LOD score (7.5). Based on the past findings that IGFBP-5 is an important bone formation stimulator, we predicted IGFBP-5 to contribute to bone mineral density variation in F2 mice. Accordingly, we found two of the six IGFBP-5 QTLs (Chrs 1 and 11) identified for serum IGFBP-5 phenotype also showed significant association with total body bone mineral density phenotype (measured by dual energy x-ray absorptiometry) in the F2 mice.