Differences in DBA/1J and DBA/2J reveal lipid QTL genes

Recent advances in mouse genomics have revealed considerable variation in the form of single-nucleotide polymorphisms (SNPs) among common inbred strains. This has made it possible to characterize closely related strains and to identify genes that differ; such genes may be causal for quantitative phenotypes. The mouse strains DBA/1J and DBA/2J differ by just 5.6% at the SNP level. These strains exhibit differences in a number of metabolic and lipid phenotypes, such as plasma levels of triglycerides (TGs) and HDL. A cross between these strains revealed multiple quantitative trait loci (QTLs) in 294 progeny. We identified significant TG QTLs on chromosomes (Chrs) 1, 2, 3, 4, 8, 9, 10, 11, 12, 13, 14, 16, and 19, and significant HDL QTLs on Chrs 3, 9, and 16. Some QTLs mapped to chromosomes with limited variability between the two strains, thus facilitating the identification of candidate genes. We suggest that Tshr is the QTL gene for Chr 12 TG and HDL levels and that Ihh may account for the TG QTL on Chr 1. This cross highlights the advantage of crossing closely related strains for subsequent identification of QTL genes.     

Generating Embryonic Stem Cells from the Inbred Mouse Strain DBA/2J,a Model of Glaucoma and Other Complex Diseases

Mouse embryonic stem (ES) cells are derived from the inner cell mass of blastocyst stage embryos and are used primarily for the creation of genetically engineered strains through gene targeting. While some inbred strains of mice are permissive to the derivation of embryonic stem cell lines and are therefore easily engineered, others are nonpermissive or recalcitrant. Genetic engineering of recalcitrant strain backgrounds requires gene targeting in a permissive background followed by extensive backcrossing of the engineered allele into the desired strain background. The inbred mouse strain DBA/2J is a recalcitrant strain that is used as a model of many human diseases, including glaucoma, deafness and schizophrenia. Here, we describe the generation of germ-line competent ES cell lines derived from DBA/2J mice. We also demonstrate the utility of DBA/2J ES cells with the creation of conditional knockout allele for Endothelin-2 (Edn2) directly on the DBA/2J strain background.     

Presphenoidal synchondrosis fusion in DBA/2J mice

Cranial base growth plates are important centers of longitudinal growth in the skull and are responsible for the proper anterior placement of the face and the stimulation of normal cranial vault development. We report that the presphenoidal synchondrosis (PSS), a midline growth plate of the cranial base, closes in the DBA/2J mouse strain but not in other common inbred strains. We investigated the genetics of PSS closure in DBA/2J mice by evaluating F1, F1 backcross, and/or F1 intercross offspring from matings with C57BL/6J and DBA/1J mice, whose PSS remain open. We observed that PSS closure is genetically determined, but not inherited as a simple Mendelian trait. Employing a genome-wide SNP array, we identified a region on chromosome 11 in the C57BL/6J strain that affected the frequency of PSS closure in F1 backcross and F1 intercross offspring. The equivalent region in the DBA/1J strain did not affect PSS closure in F1 intercross offspring. We conclude that PSS closure in the DBA/2J strain is complex and modified by different loci when outcrossed with C57BL/6J and DBA/1J mice.     

DBA/2J and DBA/2Ha lymphocytes differ in their ability to induce graft-vs-host disease

Graft-vs-host disease (GVHD) is a common occurrence after bone marrow transplantation despite the use of MHC-matched donors and recipients. This indicates that non-MHC loci play an important role in the regulation and development of GVHD. Non-MHC loci have been shown to regulate GVHD in a murine model where acute GVHD results from i.v. injection of C57BL/6J spleen cells into B6D2F1/J [C57BL/6J X DBA/2J)F1) recipients while chronic GVHD results from injection of DBA/2J spleen cells. In contrast to the hyperproduction of Ig and auto-antibodies that is characteristic of the chronic GVHD that occurs after injection of DBA/2J cells, injection of DBA/2Ha cells was found to induce CTL and suppressor cells characteristic of the acute GVHD that results from injection of C57BL/6 cells into B6D2F1/J recipients. Genetic analysis indicated that one autosomal locus is responsible for the different GVHD responses of DBA/2J and DBA/2Ha cells and that the DBA/2Ha allele is dominant. Further studies indicate that the different responses by DBA/2J and DBA/2Ha cells is not due to functional differences between the two sets of cells but by a radiosensitive B6D2F1 recipient immune response which discriminates between the DBA/2J and DBA/2Ha spleen cells.     

Divergent postnatal development of the carotid body in DBA/2J and A/J strains of mice

We have previously shown that the adult DBA/2J and A/J strains of mice differ in carotid body volume and morphology. The question has arisen whether these differences develop during the prenatal or postnatal period. Investigating morphological development of the carotid body and contributing genes in these mice can provide further understanding of the appropriate formation of the carotid body. We examined the carotid body of these mice from 1 day to 4 wk old for differences in volume, morphology, and gene expression of Gdnf family, Dlx2, Msx2, and Phox2b. The two strains showed divergent morphology starting at 1 wk old. The volume of the carotid body increased from 1 wk up to 2 wk old to the level of 4 wk old in the DBA/2J mice but not in the A/J mice. This corresponds with immunoreactivity of LC3, an autophagy marker, in A/J tissues at 10 days and 2 wk. The differences in gene expression were examined at 1 wk, 10 days, and 2 wk old, because divergent growth occurred during this period. The DBA/2J's carotid body at 1 wk old showed a greater expression of Msx2 than the A/J's carotid body. No other candidate genes showed consistent differences between the ages and strains. The difference was not seen in sympathetic cervical ganglia of 1 wk old, suggesting that the difference is carotid body specific. The current study indicates the critical postnatal period for developing distinctive morphology of the carotid body in these mice. Further studies are required to further elucidate a role of Msx2 and other uninvestigated genes.     

cTnT^R92Q转基因小鼠肥厚型心肌病模型的建立

目的建立cTnT^R92Q肥厚型心肌病的转基因小鼠模型。方法把cTnT^R92Q基因插入-αMHC启动子下游,构建转基因表达载体,通过显微注射法建立cTnT^R92Q转基因C57BL/6J小鼠。PCR鉴定cTnT^R92Q转基因小鼠的基因表型,RT-PCR检测基因表达,光学显微镜和超声检测cTnT^R92Q转基因小鼠心脏的病理改变。结果建立了3个不同表达水平的cTnT^R92Q转基因小鼠品系。转入的cTnT^R92Q基因在心脏组织的表达水平高于内源性cTnT。组织学分析显示cTnT^R92Q转基因小鼠心脏变大,心室壁肥厚,心腔变小,心肌细胞排列紊乱,心肌间质纤维增多。超声检查显示心室壁变厚,收缩期容积和舒张期容积显著缩小,射血分数、短轴缩短率明显增加。结论cTnT^R92Q转基因小鼠心脏变大,室壁变厚,心腔变小,心肌细胞排列紊乱,间质纤维化以及心肌舒张功能失调,说明成功建立了cTnT^R92Q转基因小鼠肥厚型心肌病模型,为研究肥厚型心肌病发病机制和药物研发提供了有价值的动物模型。     

Strain Differences in Presynaptic Function: PROTEOMICS,ULTRASTRUCTURE, AND PHYSIOLOGY OF HIPPOCAMPAL SYNAPSES IN DBA/2J AND C57Bl/6J MICE

The inbred strains C57BL/6J and DBA/2J (DBA) display striking differences in a number of behavioral tasks depending on hippocampal function, such as contextual memory. Historically, this has been explained through differences in postsynaptic protein expression underlying synaptic transmission and plasticity. We measured the synaptic hippocampal protein content (iTRAQ (Isobaric Tags for Relative and Absolute Quantitation) and mass spectrometry), CA1 synapse ultrastructural morphology, and synaptic functioning in adult C57BL/6J and DBA mice. DBA mice showed a prominent decrease in the Ras-GAP calcium-sensing protein RASAL1. Furthermore, expression of several presynaptic markers involved in exocytosis, such as syntaxin (Stx1b), Ras-related proteins (Rab3a/c), and rabphilin (Rph3a), was reduced. Ultrastructural analysis of CA1 hippocampal synapses showed a significantly lower number of synaptic vesicles and presynaptic cluster size in DBA mice, without changes in postsynaptic density or active zone. In line with this compromised presynaptic morphological and molecular phenotype in DBA mice, we found significantly lower paired-pulse facilitation and enhanced short term depression of glutamatergic synapses, indicating a difference in transmitter release and/or refilling mechanisms. Taken together, our data suggest that in addition to strain-specific postsynaptic differences, the change in dynamic properties of presynaptic transmitter release may underlie compromised synaptic processing related to cognitive functioning in DBA mice.     

Phencyclidine reduces inherited neocortical spindling in DBA/2J mice

The role of phencyclidine (PCP) in the control of the spike-and-wave spindling episodes (S&W) which can be spontaneously recorded in the electrocorticogram (ECoG) of DBA/2J mice was investigated. PCP (0.1–0.5-1.0–5.0 mg/kg/i.p.) dose dependently reduced both S&W number and duration of DBA/2J mice. PCP reduction is significant 30–60 min after drug administration and lasts for the whole duration of the recording period (240 min). These results suggest that PCP may play an important regulatory role on brain excitability.     

The genetics of aryl hydrocarbon hydroxylase induction in mice: A single gene difference between C57BL/6J and DBA/2J

Twenty-one inbred strains of mice were surveyed for inducibility of hepatic aryl hydrocarbon hydroxylase (AHH) activity by the carcinogen 3-methylcholanthrene (MC). In 11 strains given MC, AHH activity increased 1.3- to 5-fold (inducible), whereas ten strains responded with a less than 0.5-fold increase (noninducible). Neither the inducible nor the noninducible class was homogeneous, and in each considerable variation was found in both the basal activity of AHH and the response to MC. Strains DBA/2J and C57BL/6J were chosen to represent the noninducible and inducible classes, respectively. In the crosses (C57BL/6 × DBA/2)F₁ × DBA/2 and (C57BL/6 × DBA/2)F₂, inducibility segregated as a single autosomal dominant gene. The gene symbols Ahh ⁱ and Ahh ⁿ are proposed for the alleles present in C57BL/6J and DBA/2J, respectively. No genetic linkage was found between the Ahh locus and the following loci: b, d, Es-1, Es-3, Gpd-1, Hbb, Id-1, Pgm-1, and sex. Some implications of this work in the study of mammalian enzyme induction and chemically induced carcinogenesis are discussed. There is a positive correlation between AHH inducibility and the development of an inflammatory response to the topical application of the carcinogen 7,12-dimethylbenzanthracene.     

Retinal Ganglion Cell Dendritic Atrophy in DBA/2J Glaucoma

Glaucoma is a complex disease affecting an estimated 70 million people worldwide, characterised by the progressive degeneration of retinal ganglion cells and accompanying visual field loss. The common site of damage to retinal ganglion cells is thought to be at the optic nerve head, however evidence from other optic neuropathies and neurodegenerative disorders suggests that dendritic structures undergo a prolonged period of atrophy that may accompany or even precede soma loss and neuronal cell death. Using the DBA/2J mouse model of glaucoma this investigation aims to elucidate the impact of increasing intraocular pressure on retinal ganglion cell dendrites using DBA/2J mice that express YFP throughout the retinal ganglion cells driven by Thy1 (DBA/2J.Thy1(YFP)) and DiOlistically labelled retinal ganglion cells in DBA/2J mice. Here we show retinal ganglion cell dendritic degeneration in DiOlistically labelled DBA/2J retinal ganglion cells but not in the DBA/2J.Thy1(YFP) retinal ganglion cells suggesting that a potential downregulation of Thy1 allows only ‘healthy’ retinal ganglion cells to express YFP. These data may highlight alternative pathways to retinal ganglion cell loss in DBA/2J glaucoma.     

Structural and functional differences of the carotid body between DBA/2J and A/J strains of mice.

In a previous study, DBA/2J and A/J inbred mice showed extremely different hypoxic ventilatory responses, suggesting variations in their carotid bodies. We have assessed the morphological and functional differences of the carotid bodies in these mice. Histological examination revealed a clearly delineated carotid body only in the DBA/2J mice. Many typical glomus cells and glomeruli appeared in the DBA/2J but not in the A/J mice. The size of the carotid body in the DBA/2J and A/J mice was 6.3 +/- 0.5 x 10(6) and 1.5 +/- 0.3 x 10(6) micro m(3), respectively. The area immunostained for tyrosine hydroxylase, an estimation of the glomus cell quantity, was four times larger in the DBA/2J mice than in the A/J mice. The individual data points in the DBA/2J mice segregated from those in the A/J mice. ACh increased intracellular Ca(2+) in most clusters (81%) of cultured carotid body cells from the DBA/2J mice, but only in 18% of clusters in the A/J mice. These data suggest that genetic determinants account for the strain differences in the structure and function of the carotid body.     

Changes in retinal neuronal populations in the DBA/2J mouse

DBA/2J (D2) mice develop a form of progressive pigmentary glaucoma with increasing age. We have compared retinal cell populations of D2 mice with those in control C57BL/6J mice to provide information on retinal histopathology in the D2 mouse. The D2 mouse retina is characterized by a reduction in retinal thickness caused mainly by a thinning of the inner retinal layers. Immunocytochemical staining for specific inner retinal neuronal markers, viz., calbindin for horizontal cells; protein kinase C (PKC) and recoverin for bipolar cells, glycine, -aminobutyric acid (GABA), choline acetyltransferase (ChAT), and nitric oxide synthase (NOS) for amacrine cells, and osteopontin (OPN) for ganglion cells, was performed to detect preferentially affected neurons in the D2 mouse retina. Calbindin, PKC, and recoverin immunoreactivities were not significantly altered. Amacrine cells immunoreactive for GABA, ChAT, and OPN were markedly decreased in number, whereas NOS-immunoreactive amacrine cells increased in number. However, no changes were observed in the population of glycine-immunoreactive amacrine cells. These findings indicate a significant loss of retinal ganglion and some amacrine cells, whereas glycinergic amacrine cells, horizontal, and bipolar cells are almost unaffected in the D2 mouse. The reduction in amacrine cells appears to be attributable to a loss of GABAergic and particularly cholinergic amacrine cells. The increase in nitrergic neurons with the consequent increase in NOS and NO may be important in the changes in the retinal organization that lead to glaucomain D2 mice. Thus, the D2 mouse retina represents a useful model for studying the pathogenesis of glaucoma and mechanisms of retinal neuronal death and for evaluating neuroprotection strategies.Jung-Il Moon and In-Beom Kim contributed equally to this work.This work was supported by a Korea Research Foundation Grant (FP 0005) and by BK 21 in Korea.     

Myeloproliferative sarcoma virus: its effects on erythropoiesis in adult DBA/2J mice

Adult susceptible mice (DBA/2J) infected with MPSV (myeloproliferative sarcoma virus), a defective RNA tumour virus, develop splenomegaly and progressive disruption of the haematologic system culminating in death. The present study was specifically directed toward determining the effects of the virus on erythroid differentiation. Early and late precursor cells (erythroid burst-forming units; BFU-E and colony-forming units; CFU-E, respectively) were evaluated by the ability of bone marrow and spleen cells to form colonies of fully differentiated erythroid cells in vitro. MPSV caused substantial modification of both the BFU-E and CFU-E populations in the bone marrow and spleen of infected animals. Changes were detected in the CFU-E population preceding any significant increase in spleen weight. In the bone marrow, the proportion of CFU-E cells increased almost twofold by days 5-10 after virus infection but decreased by day 15. In the spleen, CFU-E frequency rose 40-fold by days 10-15 and then declined steadily prior to death. At the peak of CFU-E expansion, a small proportion of the population appeared to be erythropoietin (Ep) independent, although there was no evidence of a complete switch to Ep-independence which occurs in Friend virus-induced erythroleukemia. Dose-response curves showed that none of these data could be explained in terms of a changing responsiveness to Ep. However, evidence is presented that indicates that BFU-E from MPSV-infected animals lose or have a reduced requirement for burst-promoting activity (BPA) relative to normal cells although their progeny still need Ep for terminal erythroid differentiation.     

QTL influencing baseline hematocrit in the C57BL/6J and DBA/2J lineage: age-related effects

Baseline serum hematocrit varies substantially in the population. While additive genetic factors account for a large part of this variability, little is known about the genetic architecture underlying the trait. Because hematocrit levels vary with age, it is plausible that quantitative trait loci (QTL) that influence the phenotype also show an age-specific profile. To investigate this possibility, hematocrit was measured in three different age cohorts of mice (150, 450, and 750 days) of the C57BL/6J (B6) and the DBA2/J (D2) lineage. QTL were searched in the B6D2F₂ intercross and the BXD recombinant inbred (RI) strains. The effects of these QTL were explored across the different age groups. On the phenotypic level, baseline serum hematocrit declines with age in a sex-specific manner. In the B6D2F₂ intercross, suggestive QTL that influence the phenotype were located on Chromosomes (Chr) 1, 2, 7, 11, 13, and 16. With the exception of the QTL on Chr 2, all of these QTL exerted their largest effect at 750 days. The QTL on Chr 1, 2, 7, 11 and 16 were confirmed in the BXD RIs in a sex- and age-specific manner. Linkage analysis in the BXD RIs revealed an additional significant QTL on Chr 19. Baseline serum hematocrit is influenced by several QTL that appear to vary with the age and sex of the animal. These QTL primarily overlap with QTL that have been shown to regulate hematopoietic stem cell phenotypes.     

A genome-wide set of congenic mouse strains derived from DBA/2J on a C57BL/6J background

In the analysis of complex traits, congenic strains are powerful tools because they allow characterization of a single locus in the absence of genetic variation throughout the remainder of the genome. Here, we report the construction and initial characterization of a genome-wide panel of congenic strains derived from the donor strain DBA/2J on the background strain C57BL/6J. For many strains, we have carried out high-density SNP genotyping to precisely map the congenic interval and to identify any contaminating regions. Certain strains exhibit striking variation in litter size and in the ratio of females to males. We illustrate the utility of the set by "Mendelizing" the complex trait of myocardial calcification. These 65 strains cover more than 95% of the autosomal genome and should facilitate the analysis of the many genetic trait differences that have been reported between these parental strains.     

Hypertrophic Cardiomyopathy as an Oligogenic Disease: Transcriptomic Arguments

Molecular Biology - Hypertrophic cardiomyopathy (HCM) is the most common genetically determined heart pathology and is often accompanied by fatal complications. Today, the traditional view of the...     

C57BL/6J and DBA/2J mice vary in lick rate and ingestive microstructure

Fluid licking in mice is an example of a rhythmic behavior thought to be under the control of a central pattern generator. Inbred strains of mice have been shown to differ in mean or modal interlick interval (ILI) duration, suggesting a genetic-based variation. We investigated water licking in the commonly used inbred strains C57BL/6J (B6) and DBA/2J (D2), using a commercially available contact lickometer. Results from 20-min test sessions indicated that D2 mice lick at a faster rate than B6 mice (10.6 licks/s vs. 8.5 licks/s), based on analysis of the distribution of short-duration ILIs (50-160 ms). This strain difference was independent of sex, extent of water deprivation or total number of licks. D2 mice also displayed a faster lick rate when the strains were tested with a series of brief (5 s) trials. However, when ingestion over the entire 20-min session was analyzed, it was evident that D2 mice had an overall slower rate of ingestion than B6 mice. This was because of the tendency for D2 mice to have more very long pauses (>30 s) between sequences of licking bursts. Overall, it appeared that D2 mice licked more efficiently, ingesting more rapidly during excursions to the spout that were fewer and farther between.     

Latent inhibition and extinction of passive avoidance in mice C57BL/6J AND DBA/2J

The study was carried out in mice C57BL/6J and DBA/2J for comparative analysis of two interference processes: latent inhibition and extinction of passive avoidance produced with an unconditioned aversive stimulus of different parameters (0.5 and 0.25 mA). With a strong training to new stimulus, impairment of extinction has been detected only in mice DBA/2J. Reduction in the strength of punishment during training was accompanied by acceleration of extinction in mice C57BL/6J and its appearance in mice DBA/2J. The learning of passive avoidance in strong and weak reinforcement was the same for both strains of mice. Interline differences were found also in the analysis of latent inhibition. With strong and weak training to conditional stimulus, lost of novelty by repeated an 8-fold pre-exposures to the experimental chamber, in DBA/2J mice, in contrast to C57BL/6J, latent inhibition was disrupted. In addition, DBA/2J mice showed impairment of extinction with weak training to non-relevant stimulus.