Physician-diagnosed eczema is an independent risk factor for incident mouse skin test sensitization in adults.

BACKGROUND: The disrupted skin barrier in eczema has been associated with an increased risk of immunoglobulin E (IgE) sensitization in childhood. However, it is unclear whether eczema, independent of atopy, is a risk factor for the development of allergic sensitization in adulthood. OBJECTIVE: To determine if skin barrier dysfunction, independent of atopy, is a risk factor for incident sensitization in adult workers at a mouse production and research facility. METHODS: New employees at The Jackson Laboratory enrolled in a cohort study and underwent skin-prick testing (SPT) at baseline and every 6 months to mouse and to a panel of aeroallergens (net wheal ≥3 mm indicated a positive SPT result). Mouse allergen exposure was measured every 6 months by using personal air monitors. Physician-diagnosed eczema was defined as self-reported physician-diagnosed eczema. Cox proportional hazard modeling was used to examine the association between baseline physician-diagnosed eczema and incident mouse skin test sensitization and adjusted for potential confounders. RESULTS: The participants (N = 394) were followed up for a median of 24 months. Fifty-four percent were women, 89% were white, and 64% handled mice. At baseline, 7% of the participants reported physician-diagnosed eczema and 9% reported current asthma; 61% had at least one positive skin test result. At 30 months, 36% of those with eczema versus 14% of those without eczema had developed a positive mouse skin test result (p = 0.02, log-rank test). After adjusting for age, race, sex, smoking status (current, former, never), current asthma, hay fever, the number of positive SPT results at baseline, and mouse allergen exposure, physician-diagnosed eczema was an independent risk factor for incident mouse SPT sensitization (hazard ratio 5.6 [95% confidence interval, 2.1-15.2]; p = 0.001). CONCLUSION: Among adult workers at a mouse production and research facility, physician-diagnosed eczema was a risk factor for incident mouse sensitization, independent of atopy, which indicated that a defect in skin barrier alone may increase the risk of skin sensitization, not just in childhood, but throughout life.

Differences in health status affect susceptibility and mapping of genetic loci for atherosclerosis (fatty streak) in inbred mice.

OBJECTIVE: We observed differences in atherosclerosis susceptibility in mouse inbred strains over the years as the health status of our animal rooms increased. Therefore, we investigated the effect of animal room health status on atherosclerosis susceptibility in different strains. As these data can also be used for genome-wide association mapping, we performed a mapping study and compared our results with previously found quantitative trait loci for atherosclerosis in mouse and humans. METHODS AND RESULTS: Males and females from 48 inbred strains were housed in 2 animal rooms with different health status and given an atherogenic diet. We compared atherosclerosis susceptibility between animal rooms and between sexes and found that susceptibility is dependent on both health status and sex. Subsequently, the data were used for associations with loci on the mouse genome using 63 222 single nucleotide polymorphism. Three loci in males and 4 loci in females were identified using the data from the low-health status room. No significant associations were identified using the data from the high-health status room. CONCLUSIONS: Health status influences susceptibility to atherosclerosis and suggests that microbiological pressure plays an important role in the development of atherosclerosis in many strains. As we were only able to map susceptibility loci using the data from the lower health status room, we argue that susceptibility under these conditions is determined by a few key loci, whereas in the higher health status room different mechanisms might play a role in the differences in atherosclerosis susceptibility between strains and we did not have enough power to map the loci that are involved.

A customized and versatile high-density genotyping array for the mouse.

We designed a high-density mouse genotyping array containing 623,124 single-nucleotide polymorphisms that captures the known genetic variation present in the laboratory mouse. The array also contains 916,269 invariant genomic probes targeted to functional elements and regions known to harbor segmental duplications. The array opens the door to the characterization of genetic diversity, copy-number variation, allele-specific gene expression and DNA methylation, and will extend the successes of human genome-wide association studies to the mouse.

Effects of FXR in foam-cell formation and atherosclerosis development.

Farnesoid X receptor (FXR), a bile-acid-activated member of the nuclear receptor superfamily, is essential in regulating bile-acid, cholesterol, and triglyceride homeostasis. Disruption of the FXR gene in mice results in a proatherosclerotic lipid profile with increased serum cholesterols and triglycerides. However, the role of FXR in foam-cell formation and atherosclerosis development remains unclear. The current study showed that the peritoneal macrophages isolated from FXR-null mice took up less oxidized LDL-cholesterol (oxLDL-C), which was accompanied by a marked reduction in CD36 expression in these cells. This result appears to be FXR-independent, as FXR was not detected in the peritoneal macrophages. To assess to what extent FXR modulates atherosclerosis development, FXR/ApoE double-null mice were generated. Female mice were used for atherosclerosis analysis. Compared to ApoE-null mice, the FXR/ApoE double-null mice were found to have less atherosclerotic lesion area in the aorta, despite a further increase in the serum cholesterols and triglycerides. Our results indicate that disruption of the FXR gene could attenuate atherosclerosis development, most likely resulting from reduced oxLDL-C uptake by macrophages. Our study cautions the use of serum lipid levels as a surrogate marker to determine the efficiency of FXR modulators in treating hyperlipidemia.

Hepatic lipase expression in macrophages contributes to atherosclerosis in apoE-deficient and LCAT-transgenic mice.

Hepatic lipase (HL) has a well-established role in lipoprotein metabolism. However, its role in atherosclerosis is poorly understood. Here we demonstrate that HL deficiency raises the proatherogenic apoB-containing lipoprotein levels in plasma but reduces atherosclerosis in lecithin cholesterol acyltransferase (LCAT) transgenic (Tg) mice, similar to results previously observed with HL-deficient apoE-KO mice. These findings suggest that HL has functions that modify atherogenic risk that are separate from its role in lipoprotein metabolism. We used bone marrow transplantation (BMT) to generate apoE-KO and apoE-KO x HL-KO mice, as well as LCAT-Tg and LCAT-Tg x HL-KO mice, chimeric for macrophage HL gene expression. Using in situ RNA hybridization, we demonstrated localized production of HL by donor macrophages in the artery wall. We found that expression of HL by macrophages enhances early aortic lesion formation in both apoE-KO and LCAT-Tg mice, without changing the plasma lipid profile, lipoprotein lipid composition, or HL and lipoprotein lipase activities. HL does, however, enhance oxidized LDL uptake by peritoneal macrophages. These combined data demonstrate that macrophage-derived HL significantly contributes to early aortic lesion formation in two independent mouse models and identify a novel mechanism, separable from the role of HL in plasma lipoprotein metabolism, by which HL modulates atherogenic risk in vivo.

Atopy as a Modifier of the Relationships Between Endotoxin Exposure and Symptoms Among Laboratory Animal Workers.

BACKGROUND: Exposure to endotoxin is known to trigger airway inflammation and symptoms, and atopy may modify the relationship between endotoxin exposure and symptom development. OBJECTIVE: To test the a priori hypothesis that atopic status modifies the relationship between endotoxin exposure and respiratory symptom development. METHODS: A prospective study of laboratory workers at The Jackson Laboratories was conducted. Allergy skin testing was performed and population demographic and clinical information was obtained at baseline. Personal exposure assessments for airborne endotoxin and surveys of self-reported symptoms were performed every 6 months. Cox proportional hazards models were used to examine the relationship between endotoxin exposure and development of mouse-associated symptoms and multivariate regression was used to test for interaction. RESULTS: Overall, 16 (9%) of 174 worker-participants developed mouse-associated rhinoconjunctivitis symptoms by 24 months and 8 (5%) developed mouse-associated lower respiratory symptoms by 24 months. Among workers with endotoxin exposure above the median (≥2.4 EU m-3), 5 (6% of 80) atopics reported mouse-associated rhinoconjunctivitis symptoms at 24 months as compared to 3 (3% of 94) non-atopics. Among workers below the median endotoxin exposure (<2.4 EU m-3), 1 (1% of 80) atopic reported mouse-associated rhinoconjunctivitis symptoms at 24 months as compared to 7 (7% of 94) non-atopics. For the combination of symptoms, the adjusted hazard ratio was 6.8 (95% confidence interval: 0.7-67.2) for atopics and 0.07 (95% confidence interval: 0.01-0.5) for non-atopics. CONCLUSION: In this occupational cohort, atopic workers may be more susceptible to, and non-atopic workers protected from, endotoxin-associated upper and lower respiratory symptoms.

A torrid zone on mouse chromosome 1 containing a cluster of recombinational hotspots.

Within the 2.38-Mb Ath1 region of mouse chromosome 1, 42 of 45 genetic crossovers from crosses between C57BL/6J (B6) and either C3H/HeJ (H) or Mus spretus (SPRET) occurred in four zones (A-D); zone A, 100 kb long, contained a cluster of at least four recombination hotspots. F1 sperm assays indicate that within this "torrid zone" the most active hotspot (A3) can initiate recombination on H and SPRET but not B6 chromosomes. The A3 DNA sequence contains a (G/C)TTT repeat, long stretches of A or T, and a cyclic variation in AT content. Recombination was drastically reduced in a cross between B6 and a B6.SPRET Ath1 congenic strain, but was unaffected in a B6 x B6.H Ath1 congenic cross. Similar nonrandom clustering of hotspots has been observed in yeast and the major histocompatibility complexes of human and mouse. To the extent that torrid zones are a general feature of mammalian genomes, they have considerable implications for genetic mapping strategies in both human populations and mouse crosses.

Quantitative trait locus mapping of genes that regulate phospholipid transfer activity in SM/J and NZB/BlNJ inbred mice.

OBJECTIVE: Phospholipid transfer protein (PLTP), an important protein in the transfer of phospholipids between lipoprotein particles and in the remodeling of HDL, is regulated at both the transcriptional and the protein level. We performed quantitative trait locus (QTL) analysis to identify genomic loci regulating PLTP activity in mice. METHODS AND RESULTS: Plasma PLTP activity was measured in 217 male F2 progeny from a SM/J x NZB/B1NJ intercross. Two QTL for plasma PLTP activity in mice fed chow (Pltpq1 and Pltpq2) were found on chromosomes 3 (34 cM, logarithm of odds [LOD] 3.5) and 10 (66 cM, LOD 4.1); two additional QTL in mice fed atherogenic diet (Pltpq3 and Pltpq4) were found on chromosomes 9 (56 cM, LOD 4.5) and 15 (34 cM, LOD 5.0); and one QTL (Pltiq1) for the inducibility of PLTP activity was found on chromosome 4 (70 cM, LOD 3.7). Several candidate genes for these 5 QTL were tested by sequence comparison and expression studies. CONCLUSIONS: We identified five significant loci involved in PLTP activity in the mouse and provided supporting evidence for the candidacy of Nr1h4 and Apof as the genes underlying Pltpq2.

Assessing the risk of transmission of three infectious agents among mice housed in a negatively pressurized caging system.

Previous studies from our institution have shown that ventilated caging run at negative pressure to a mouse room dramatically reduced exposure of personnel to the major mouse allergen, Mus m 1. The current study was designed to determine whether negative cage ventilation posed an inordinate risk for spread of infectious agents between cages and/or racks. B6;129S-Tnfsf5(tm1Imx)/J (TNF) mice, which were naturally and persistently infected with Pneumocystis carinii, Helicobacter bilis, and Pasteurella pneumotropica, were used as the source of infections. Uninfected C3Smn.CB17-Prkdc(scid)/J (SCID) mice with severe combined immunodeficiency were used to detect transmission. The following methods were used to detect transmission of infections: polymerase chain reaction (PCR) amplification and histological examination of lungs for P. carinii; PCR of fecal specimens or cecal contents for H. bilis; and culture of oropharyngeal, tracheal, or vaginal swabs for P. pneumotropica. We determined whether transmission of the three agents occurred via direct contact (cohabitation), exposure to soiled bedding, and/or by handling naive SCID mice after handling infected TNF mice. During a 12-week period, all three infectious agents were readily transmitted to uninfected mice by cohabitation. Transmission was much less efficient and occurred later among mice exposed to contaminated bedding. Transmission did not occur as a result of handling. We then studied transmission of the three infectious agents among mice housed in individually ventilated cages run at negative pressure in a small, crowded mouse room. Transmission of P. carinii was detected at the end of the 12-month study in the densely populated room, probably because the exhaust from the changing station passed over soiled cages and caused aerosolization of particulates. Caging systems run at negative pressure effectively reduce personnel exposure to allergens and may also inhibit the transmission of infectious diseases.

Genetic variation in TRPS1 may regulate hip geometry as well as bone mineral density.

Trps1 has been proposed as a candidate gene for a mouse bone mineral density (BMD) QTL on Chromosome (Chr) 15, but it remained unclear if this gene was associated with BMD in humans. We used newly available data and advanced bioinformatics techniques to confirm that Trps1 is the most likely candidate gene for the mouse QTL. In short, by combining the raw genetic mapping data from two F2 generation crosses of inbred strains of mice, we narrowed the 95% confidence interval of this QTL down to the Chr 15 region spanning from 6 to 24cM. This region contains 131 annotated genes. Using block haplotyping, all other genes except Trps1 were eliminated as candidates for this QTL. We then examined associations of 208 SNPs within 10kb of TRPS1 with BMD and hip geometry, using human genome-wide association study (GWAS) data from the GEFOS consortium. After correction for multiple testing, six TRPS1 SNPs were significantly associated with femoral neck BMD (P=0.0015-0.0019; adjusted P=0.038-0.048). We also found that three SNPs were highly associated with femoral neck width in women (rs10505257, P=8.6×10(-5), adjusted P=2.15×10(-3); rs7002384, P=5.5×10(-4), adjusted P=01.38×10(-2)). In conclusion, we demonstrated that combining association studies in humans with murine models provides an efficient strategy to identify new candidate genes for bone phenotypes.

Mouse BMD quantitative trait loci show improved concordance with human genome-wide association loci when recalculated on a new, common mouse genetic map.

Bone mineral density (BMD) is a heritable trait, and in mice, over 100 quantitative trait loci (QTLs) have been reported, but candidate genes have been identified for only a small percentage. Persistent errors in the mouse genetic map have negatively affected QTL localization, spurring the development of a new, corrected map. In this study, QTLs for BMD were remapped in 11 archival mouse data sets using this new genetic map. Since these QTLs all were mapped in a comparable way, direct comparisons of QTLs for concordance would be valid. We then compared human genome-wide association study (GWAS) BMD loci with the mouse QTLs. We found that 26 of the 28 human GWAS loci examined were located within the confidence interval of a mouse QTL. Furthermore, 14 of the GWAS loci mapped to within 3 cM of a mouse QTL peak. Lastly, we demonstrated that these newly remapped mouse QTLs can substantiate a candidate gene for a human GWAS locus, for which the peak single-nucleotide polymorphism (SNP) fell in an intergenic region. Specifically, we suggest that MEF2C (human chromosome 5, mouse chromosome 13) should be considered a candidate gene for the genetic regulation of BMD. In conclusion, use of the new mouse genetic map has improved the localization of mouse BMD QTLs, and these remapped QTLs show high concordance with human GWAS loci. We believe that this is an opportune time for a renewed effort by the genetics community to identify the causal variants regulating BMD using a synergistic mouse-human approach.

Genetic influence on electrocardiogram time intervals and heart rate in aging mice.

Understanding the genetic influence on ECG time intervals and heart rate (HR) is important for identifying the genes underlying susceptibility to cardiac arrhythmias. The objective of this study was to determine the genetic influence on ECG parameters and their age-related changes in mice. ECGs were recorded in lead I on 8 males and 8 females from each of 28 inbred strains at the ages of 6, 12, and 18 mo. Significant interstrain differences in the P-R interval, QRS complex duration, and HR were found. Age-related changes in the P-R interval, QRS complex duration, and HR differed among strains. The P-R interval increased with age in 129S1/SvlmJ females. The QRS complex duration decreased with age in C57BR/J males and DBA2/J females but increased in NON/ShiLtJ females. HR decreased in C57L/J females and SM/J and P/J males but increased in BALB/cByJ males. Differences between males and females were found for HR in SJL/J mice and in the P-R interval in 129S1/SvlmJ mice. Broad-sense heritability estimates of ECG time intervals and HR ranged from 0.31 for the QRS complex duration to 0.52 for the P-R interval. Heritability estimates decreased with age for the P-R interval. Our study revealed that genetic factors play a significant role on cardiac conduction activity and age-related changes in ECG time intervals and HR.

The mouse as a model for human biology: a resource guide for complex trait analysis.

The mouse has been a powerful force in elucidating the genetic basis of human physiology and pathophysiology. From its beginnings as the model organism for cancer research and transplantation biology to the present, when dissection of the genetic basis of complex disease is at the forefront of genomics research, an enormous and remarkable mouse resource infrastructure has accumulated. This review summarizes those resources and provides practical guidelines for their use, particularly in the analysis of quantitative traits.

Sex does not seem to influence the formation of aortic lesions in the P-407-induced mouse model of hyperlipidemia and atherosclerosis.

Coronary heart disease secondary to atherosclerosis is the leading cause of death for men in the United States. Using a new, nontransgenic, non-fat-fed mouse model of hyperlipidemia and atherosclerosis developed in our laboratory, we investigated the effect of sex on lipid profiles and subsequent aortic atherosclerotic lesion formation. Female and male C57BL/6 mice, which consumed a low-fat diet, were treated with either normal saline or poloxamer 407 (P-407), a triblock copolymer comprised of poly(oxyethylene) and poly(oxypropylene) units, for 4 months. Blood samples were obtained at 0, 1, 2, 3, and 4 months, whereas hearts and livers were harvested only at 4 months, because this model requires approximately 4 months for significant atheroma formation. P-407-treated mice of either sex demonstrated a profound increase in plasma cholesterol and triglyceride; at 3 and 4 months the plasma lipids were significantly (p < 0.05) higher for male mice compared with female mice. Aortas retrieved from P-407-treated mice of either sex after 4 months demonstrated a significant (p < 0.001) increase in the mean atherosclerotic lesion size compared with their respective saline-treated controls, but there was no significant (p > 0.05) difference between lesion sizes for P-407-treated male mice (1.02 +/- 0.074 x 10(5) microm(2)) compared with P-407-treated female mice (1.14 +/- 0.28 x 10(5) microm(2)). Livers harvested at 4 months from either sex of P-407-treated mice displayed no damage to hepatocytes but increased proliferation of macrophages (Kupffer cells), which contained sequestered lipids. Thus, male C57BL/6 mice form atherosclerotic lesions as extensive as female mice in the P-407 mouse model of atherosclerosis.

The ligand-binding function of hepatic lipase modulates the development of atherosclerosis in transgenic mice.

To investigate the separate contributions of the lipolytic versus ligand-binding function of hepatic lipase (HL) to plasma lipoprotein metabolism and atherosclerosis, we compared mice expressing catalytically active wild-type HL (HL-WT) and inactive HL (HL-S145G) with no endogenous expression of mouse apoE or HL (E-KO x HL-KO, where KO is knockout). HL-WT and HL-S145G reduced plasma cholesterol (by 40 and 57%, respectively), non-high density lipoprotein cholesterol (by 48 and 61%, respectively), and apoB (by 36 and 44%, respectively) (p < 0.01), but only HL-WT decreased high density lipoprotein cholesterol (by 67%) and apoA-I (by 54%). Compared with E-KO x HL-KO mice, both active and inactive HL lowered the pro-atherogenic lipoproteins by enhancing the catabolism of autologous (125)I-apoB very low density/intermediate density lipoprotein (VLDL/IDL) (fractional catabolic rates of 2.87 +/- 0.04/day for E-KO x HL-KO, 3.77 +/- 0.03/day for E-KO x HL-WT, and 3.63 +/- 0.09/day for E-KO x HL-S145G mice) and (125)I-apoB-48 low density lipoprotein (LDL) (fractional catabolic rates of 5.67 +/- 0.34/day for E-KO x HL-KO, 18.88 +/- 1.72/day for E-KO x HL-WT, and 9.01 +/- 0.14/day for E-KO x HL-S145G mice). In contrast, the catabolism of apoE-free, (131)I-apoB-100 LDL was not increased by either HL-WT or HL-S145G. Infusion of the receptor-associated protein (RAP), which blocks LDL receptor-related protein function, decreased plasma clearance and hepatic uptake of (131)I-apoB-48 LDL induced by HL-S145G. Despite their similar effects on lowering pro-atherogenic apoB-containing lipoproteins, HL-WT enhanced atherosclerosis by up to 50%, whereas HL-S145G markedly reduced aortic atherosclerosis by up to 96% (p < 0.02) in both male and female E-KO x HL-KO mice. These data identify a major receptor pathway (LDL receptor-related protein) by which the ligand-binding function of HL alters remnant lipoprotein uptake in vivo and delineate the separate contributions of the lipolytic versus ligand-binding function of HL to plasma lipoprotein size and metabolism, identifying an anti-atherogenic role of the ligand-binding function of HL in vivo.

Hepatic ABCG5 and ABCG8 overexpression increases hepatobiliary sterol transport but does not alter aortic atherosclerosis in transgenic mice.

The individual roles of hepatic versus intestinal ABCG5 and ABCG8 in sterol transport have not yet been investigated. To determine the specific contribution of liver ABCG5/G8 to sterol transport and atherosclerosis, we generated transgenic mice that overexpress human ABCG5 and ABCG8 in the liver but not intestine (liver G5/G8-Tg) in three different genetic backgrounds: C57Bl/6, apoE-KO, and low density lipoprotein receptor (LDLr)-KO. Hepatic overexpression of ABCG5/G8 enhanced hepatobiliary secretion of cholesterol and plant sterols by 1.5-2-fold, increased the amount of intestinal cholesterol available for absorption and fecal excretion by up to 27%, and decreased the accumulation of plant sterols in plasma by approximately 25%. However, it did not alter fractional intestinal cholesterol absorption, fecal neutral sterol excretion, hepatic cholesterol concentrations, or hepatic cholesterol synthesis. Consequently, overexpression of ABCG5/G8 in only the liver had no effect on the plasma lipid profile, including cholesterol, HDL-C, and non-HDL-C, or on the development of proximal aortic atherosclerosis in C57Bl/6, apoE-KO, or LDLr-KO mice. Thus, liver ABCG5/G8 facilitate the secretion of liver sterols into bile and serve as an alternative mechanism, independent of intestinal ABCG5/G8, to protect against the accumulation of dietary plant sterols in plasma. However, in the absence of changes in fractional intestinal cholesterol absorption, increased secretion of sterols into bile induced by hepatic overexpression of ABCG5/G8 was not sufficient to alter hepatic cholesterol balance, enhance cholesterol removal from the body or to alter atherogenic risk in liver G5/G8-Tg mice. These findings demonstrate that overexpression of ABCG5/G8 in the liver profoundly alters hepatic but not intestinal sterol transport, identifying distinct roles for liver and intestinal ABCG5/G8 in modulating sterol metabolism.