Experiment
Asthma is a common disease of the airways that is characterized by episodes of reversible airway obstruction, airway hyperresponsiveness, and inflammation. Airway hyperresponsiveness has been associated with asthma susceptibility in both humans and murine models. In particular, the inbred strain of A/J mice is known to have hyperresponsive airways, while the C57BL/6 strain is known to be relatively refractory to broncho-constricting stimuli.
The objective of this study was to replicate the identification of loci that have been previously identified in A/J x C57BL/6 crosses, and in several cases, have not yet been replicated, as well as to detect additional chromosomal regions associated with airway responsiveness.
Recombinant congenic mouse strains AcB/BcA were generated from methacholine-challenge-resistant C57BL/6 (B) mice and methacholine-challenge-susceptible A/J (A) mice. Fifteen distinct AcB strains containing app 13.25% of the B genome on an A/J background (app 84.4% of the A/J genome) and 22 distinct BcA strains containing app 13.24% of the A genome on a B background (app 85.37% of the B genome) were generated. All RC strains were previously genotyped using a panel of 625 informative SSLP markers; all markers in the panel that mapped to a single position in the Mouse Genome Database (Bult, et al. 2008) were selected for further use in the analysis. Six hundred and thirty markers covering the entire genome with an average spacing of 2.7 cM were ultimately used.
Airway responsiveness was assessed using an unrestrained whole-body plethysmorgraph. Ten male mice from each parental strain and 4-12 male mice from each of the 11 AcB and 22 BcA strains used in the study were phenotyped at 8-12 weeks of age. Comparison of the mean phenotype of the AcB strains with the mean phenotype of the BcA strains was performed using a two-tailed unpaired t test. Analysis of the phenotypes across all strains was performed with a one-way ANOVA followed by Bonferroni-corrected pairwise comparisons of every AcB strain with the A strain and every BcA strain with the B strain. Permutation
analysis consisted of 45,000 permutations performed by shuffling the phenotypes across the genotypes; significance was set at p</=0.05.
A strong strain effect was identified in two AcB strains, AcB64 and AcB51, and in two BcA strains, BcA85 and BcA86. Direct measurements of airway resistance were performed in the A/J, C57BL/6, AcB64, AcB51, BcA85 and BcA86 strains as well as several other RC strains. The test for association of alleles that affect airway responsiveness was performed at each of 603 SSLP markers for which the genotypes of the RCS panel were available. The analysis led to the identification of 50 markers that were significantly associated with airway responsiveness and which delimited 16 chromosomal regions.
The text reports that of the 16 regions identified as significant that 8 replicate QTL or regions that have previously been implicated in the control of airway responsiveness in crosses involving the A/J strain; and the other 8 regions have never been reported to be associated with airway responsiveness, Table1 1.
Curator Note: Because the 8 chromosomal regions identified here as replicating QTL, or as regions that were previously implicated in the control of airway responsiveness, were not originally mapped using a mapping population of AcB/BcA RC strains of mice we consider the QTL mapped in this study unique and have assigned unique nomenclature to the 8 QTL cited as previously identified as well as to the 8 QTL identified here as novel on Chrs 1, 5, 7, 8, 10 (2 QTL) and 12 (2 QTL).
QTL Airrq1, airway responsiveness QTL 1, mapped to Chromosome 1 with significant markers D1Mit113 and D1Mit206 in an interval between 172.1-179.6 Mb with a peak p value of 0.000076.
QTL Airrq2, airway responsiveness QTL 2, mapped to Chromosome 2 with significant markers D2Mit80, D2Mit416, D2Mit81, D2Mit293, D2Mit235 and D2Mit367 in an interval between 9.3-40.6 Mb with a peak p value of 0.000032. QTL Airrq2 overlaps with previously mapped QTL Abhr1 (J:66641).
QTL Airrq3, airway responsiveness QTL 3, mapped to Chromosome 2 with significant markers D2Mit133 and D2Mit164 in an interval between 117.9-148.7 Mb with a peak p value of
0.000412. Airrq3 overlaps with previously mapped QTL Bhr1 (J:29231 and J:97472).
QTL Airrq4, airway responsiveness QTL 4, mapped to Chromosome 5 with significant markers D5Mit97, D5Mit31, D5Mit99, D5Mit101 and D5Mit287 in an interval between 132.6 Mb and the telomere with a peak p value of 0.0000545.
QTL Airrq5, airway responsiveness QTL 5, mapped to Chromosome 6 with significant markers D6Mit264 and D6Mit159 in an interval between 5.3-34.7 Mb with a peak p value of 0.0000441. Airrq5 overlaps with previously identified QTL Bhr5 (J:33778) and Bhr8 (J:97472).
QTL Airrq6, airway responsiveness QTL 6, mapped to Chr 6 with significant marker D6Mit178 in an interval between 92.6-98.5 Mb with a peak p value of 0.000319. Airrq6 maps near previously identified QTL Bhr7 (J:59047) and Bhr5 (J:33778).
QTL Airrq7, airway responsiveness QTL 7, mapped to Chr 6 with significant markers D6Mit290 and D6Mit25 with a peak p value of 0.000248.
QTL Airrq8, airway responsiveness QTL 8, mapped to Chromosome 7 with significant markers D7Mit187 and D7Mit259 in an interval between 135.7 Mb and the telomere with a peak p value of 0.000131.
QTL Airrq9, airway responsiveness QTL 9, mapped to Chromosome 8 with significant markers D8Mit271, D8Mit200 and D8Mit186 in an interval between 111.4-129.1 Mb with a peak p value of 0.000304.
QTL Airrq10, airway responsiveness QTL 10, mapped to Chromosome 10 with significant markers D10Mit80 and D10Mit246 in an interval between 8.5-18.1 Mb with a peak p value of 0.000315.
QTL Airrq11, airway responsiveness QTL 11, mapped to Chromosome 10 with significant markers D10Mit205 and D10Mit103 in an interval between 119.4 Mb and the telomere with a peak p value of 0.000102.
QTL Airrq12, airway responsiveness QTL 12, mapped to Chromosome 12 with significant markers D12Mit36, D12Mit52 and D12Mit14 in an interval between 55.8-83.7 Mb with a peak p value of 0.0000886.
QTL Airrq13, airway responsiveness QTL 13, mapped to Chromosome 12 with significant markers D12Mit79, D12Mit133, D12Mit280, D12Mit196, D12Mit8 and D12Mit134 in an interval between 109.5 Mb and the telomere with a peak p value of 0.0000027.
QTL Airrq14, airway responsiveness QTL 14, mapped to Chromosome 15 with significant markers D15Mit111, D15Mit255, D15Mit100, D15Mit115, D15Mit156 and D15Mit105 in an interval between 17.1-74.7 Mb with a peak p value of 0.0000763. QTL Airrq14 overlapped with QTL Bhr2 (J:29231).
QTL Airrq15, airway responsiveness QTL 15, mapped to Chromosome 17 with significant markers D17Mit113 and D17Mit222a in an interval between 3.9 and 25 Mb with a peak p value of 0.000299. QTL Airrq15 overlapped with QTL Bhr3 (J:29231) and to a suggestive asthma phenotype identified on Chr 17 in J:54405 using a (BP2 x BALB/c)F2 cross.
QTL Airrq16, airway responsiveness QTL 16, mapped to Chromosome 17 with significant markers D17Mit88, D17Mit139, D17Mit7 and D17Mit159 in an interval between 47.9-73.9 Mb with a peak p value of 0.0000122. QTL Airrq16 mapped near a suggestive QTL ((LOD = 1.7 at D17Mit215) identified in an (A/J X C3H/HeJ)F1 X C3H/HeJ backcross population used in J:54405.
Table 3 lists the following likely candidate genes:
Ddr2 for QTL Airrq1 (Chr 1, 171.9 Mb);
Hc for QTL Airrq2 (Chr 2, 34.8 Mb);
Fbn1 for QTL Airrq3 (Chr 2, 125.3 Mb);
Utrn for QTL Airrq10 (Chr 10, 12.6 Mb);
Enpp2 for QTL Airrq14 (Chr 15, 54.7 Mb);
Tsc2 for QTL Airrq16 (Chr 17, 24.7 Mb).
Analysis Tools