Congenic strain confirms putative quantitative trait locus for body weight in the rat

Quantitative trait loci (QTLs) affecting body weight were investigated in the backcross population derived from non-diabetic BB/OK and spontaneously hypertensive rat (SHR) strains. The F1 hybrids were backcrossed onto SHR rats, and QTL analysis was performed separately with the resulting backcross populations for each sex on Chromosomes (Chrs) 1, 3, 4, 10, 13, and 18. The body weight was determined at the age of 14 weeks, and the statistical analysis was performed with MAPMAKER/QTL 1.1b computer program. According to the stringent threshold for a lod score of 3.0, markers on Chr 1 were found to be linked with body weight. The QTL with a peak lod score (5.1) on Chr 1 for a male population was located within markers Igf2 and D1Mgh12. In contrast, in the female population the body weight affecting QTL (lod = 5.7) on Chr 1 was located between the D1Mit3 and Lsn markers. The existence of QTLs on Chr 1 affecting body weight in the male population was confirmed by congenic BB.Sa rats, carrying chromosomal region of SHR (Sa-Igf2) on the genetic background of BB rat.     

Identification of quantitative trait loci influencing wood specific gravity in an outbred pedigree of loblolly pine

We report the identification of quantitative trait loci (QTL) influencing wood specific gravity (WSG) in an outbred pedigree of loblolly pine (Pinus taeda L.). QTL mapping in an outcrossing species is complicated by the presence of multiple alleles (> 2) at QTL and marker loci. Multiple alleles at QTL allow the examination of interaction among alleles at QTL (deviation from additive gene action). Restriction fragment length polymorphism (RFLP) marker genotypes and wood specific gravity phenotypes were determined for 177 progeny. Two RFLP linkage maps were constructed, representing maternal and paternal parent gamete segregations as inferred from diploid progeny RFLP genotypes. RFLP loci segregating for multiple alleles were vital for aligning the two maps. Each RFLP locus was assayed for cosegregation with WSG QTL using analysis of variance (ANOVA). Five regions of the genome contained one or more RFLP loci showing differences in mean WSG at or below the P = 0.05 level for progeny as grouped by RFLP genotype. One region contained a marker locus (S6a) whose QTL-associated effects were highly significant (P > 0.0002). Marker S6a segregated for multiple alleles, a prerequisite for determining the number of alleles segregating at the linked QTL and analyzing the interactions among QTL alleles. The QTL associated with marker S6a appeared to be segregating for multiple alleles which interacted with each other and with environments. No evidence for digenic epistasis was found among the five QTL.     

Computation of the full likelihood function for estimating variance at a quantitative trait locus

The proportion of alleles identical by descent (IBD) determines the genetic covariance between relatives, and thus is crucial in estimating genetic variances of quantitative trait loci (QTL). However, IBD proportions at QTL are unobservable and must be inferred from marker information. The conventional method of QTL variance analysis maximizes the likelihood function by replacing the missing IBDs by their conditional expectations (the expectation method), while in fact the full likelihood function should take into account the conditional distribution of IBDs (the distribution method). The distribution method for families of more than two sibs has not been obvious because there are n(n - 1)/2 IBD variables in a family of size n, forming an n x n symmetrical matrix. In this paper, I use four binary variables, where each indicates the event that an allele from one of the four grandparents has passed to the individual. The IBD proportion between any two sibs is then expressed as a function of the indicators. Subsequently, the joint distribution of the IBD matrix is derived from the distribution of the indicator variables. Given the joint distribution of the unknown IBDs, a method to compute the full likelihood function is developed for families of arbitrary sizes.     

Multiple-trait mapping of quantitative trait loci after selective genotyping using logistic regression

Experiments to map QTL usually measure several traits, and not uncommonly genotype only those animals that are extreme for some trait(s). Analysis of selectively genotyped, multiple-trait data presents special problems, and most simple methods lead to biased estimates of the QTL effects. The use of logistic regression to estimate QTL effects is described, where the genotype is treated as the dependent variable and the phenotype as the independent variable. In this way selection on phenotype does not bias the results. If normally distributed errors are assumed, the logistic-regression analysis is almost equivalent to a maximum-likelihood analysis, but can be carried out with standard statistical packages. Analysis of a simulated half-sib experiment shows that logistic regression can estimate the effect and position of a QTL without bias and confirms the increased power achieved by multiple-trait analysis.     

Mapping a quantitative trait locus involved in melanotic encapsulation of foreign bodies in the malaria vector, Anopheles gambiae

A Plasmodium-refractory strain of Anopheles gambiae melanotically encapsulates many species of Plasmodium, whereas wild-type mosquitoes are usually susceptible. This encapsulation trait can also be observed by studying the response of refractory and susceptible strains to intrathoracically injected CM-Sephadex beads. We report the results of broad-scale quantitative trait locus (QTL) mapping of the encapsulation trait using the bead model system. Interval mapping using the method of maximum likelihood identified one major QTL, Pen1. The 13.7-cM interval containing Pen1 was defined by marker AGH157 at 8E and AGH46 at 7A on 2R. Pen1 was associated with a maximum LOD score of 9.0 and accounted for 44% of the phenotypic variance in the distribution of phenotypes in the backcross. To test if this QTL is important for encapsulation of Plasmodium berghei, F2 progeny were infected with P. berghei and evaluated for degree of parasite encapsulation. For each of the two markers that define the interval containing Pen1, a significant difference of encapsulation was seen in progeny with at least one refractory allele in contrast with homozygous susceptible progeny. These results suggest that Pen1 is important for melanotic encapsulation of Plasmodium as well as beads.     

The absorption of vitamin B12 after subtotal pancreatectomy in the rat

Subtotal pancreatectomy in the rat was performed by a new method. Except for a small remnant along the splenal hilus, the gland was completely removed. Exocrine pancreatic insufficiency was confirmed by fat and nitrogen malabsorption. A moderate diabetes mellitus was seen. Insulin substitution or diet restriction was unnecessary. The absorption of vitamin B12 after oral administration of 10 ng 57CoB12 was 5.0 +/- 0.2 (ng 57CoB12 +/- S.E.M.) in unoperated rats and 5.1 +/- 0.1 in pancreatectomized rats (p greater than 0.5). Pancreatic extract (PE) (100 mg) increased the absorption of 57CoB12 in unoperated rats (p less than 0.01), but not in pancreatectomized rats (p greater than 0.2). PE (300 mg) reduced the absorption in unoperated rats (p less than 0.01). The results indicate that an optimal amount of pancreatic secretion may stimulate the absorption of vitamin B12.     

Approximately 70% of fucose-labeled glycopeptides from the cell surface and cellular material of rat

This article describes a model for allocating staff within a large psychiatric hospital. The model provides an objective framework within which one can test alternative staff operating policies before making critical decisions concerning the employment of one category of personnel as opposed to another. It is based on objective data describing patient needs and staff functioning patterns, rather than subjective opinions concerning staff deployment. Besides being useful for the short-term deployment of staff and budgetary resources, it can also be used as a long-range planning tool for testing modifications in policy decisions and budget proposals. The algorithm employed, mixed-integer linear programming, is readily available; computer costs and running time are relatively minimal.     

Identification of quantitative trait loci governing arthritis severity and humoral responses in the murine model of Lyme disease

A spectrum of disease severity has been observed in patients with Lyme disease, with approximately 60% of untreated individuals developing arthritis. The murine model of Lyme disease has provided strong evidence that the genetic composition of the host influences the severity of arthritis following infection with Borrelia burgdorferi: infected C3H mice develop severe arthritis while infected C57BL/6N mice develop mild arthritis. Regions of the mouse genome controlling arthritis severity and humoral responses during B. burgdorferi infection were identified in the F2 intercross generation of C3H/HeNCr and C57BL/6NCr mice. Rear ankle swelling measurements identified quantitative trait loci (QTL) on chromosomes 4 and 5, while histopathological scoring identified QTL on a unique region of chromosome 5 and on chromosome 11. The identification of QTL unique for ankle swelling or histopathological severity suggests that processes under distinct genetic control are responsible for these two manifestations of Lyme arthritis. Additional QTL that control the levels of circulating Igs induced by B. burgdorferi infection were identified on chromosomes 6, 9, 11, 12, and 17. Interestingly, the magnitude of the humoral response was not correlated with the severity of arthritis in infected F2 mice. This work defines several genetic loci that regulate either the severity of arthritis or the magnitude of humoral responses to B. burgdorferi infection in mice, with implications toward understanding the host-pathogen interactions involved in disease development.     

Improved plasma cholesterol levels in men after a nutrition education program at the worksite

Eighty management-level male employees participated in a company-sponsored comprehensive physical that included determination of plasma total cholesterol, low-density lipoprotein cholesterol, high-density lipoprotein cholesterol, and triglyceride levels and percentage of body fat. After the lipid screening, each employee met with a registered dietitian who explained the results of the lipid analysis and discussed risk factors for coronary heart disease with an emphasis on diet. Seventy employees had a triglyceride level above 5.17 mmol/L and were invited to participate in a nutrition education program. Thirty-three (mean age = 44 years) chose to participate (intervention group); the other 37 (mean age = 35 years) served as controls (control group). Thus, the design of the study was not random. All subjects completed 3-day dietary records before and after the nutrition education program. Nutrition intervention consisted of (a) individualized instruction about the step 1 diet; (b) group sessions (1 hour every 3 months) on eating out, dietary fiber, and maintaining heart healthy behaviors; and (c) individualized follow-up by telephone (one call per month). The results of the year-long program revealed that men in the intervention group decreased dietary intake of energy (2,546 +/- 162 kcal to 2,246 +/- 125 kcal) and cholesterol (444 +/- 5.3 mg to 304 +/- 1.6 mg) and percentage of energy from total fat (38 +/- 3.4% to 31 +/- 2.6%) and protein (24 +/- 3.5% to 20 +/- 2.2%). Their consumption of carbohydrate and dietary fiber increased (38 +/- 2.1% to 45 +/- 2.5% and 8.0 +/- 2.3 g to 23.0 +/- 3.5 g, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)     

Genetic mapping of two new blood pressure quantitative trait loci in the rat by genotyping endothelin system genes

The endothelin system, consisting of a series of potent vasoconstrictor peptides and their receptors, is potentially important in the control of blood pressure. We found that the gene coding for endothelin-2 (ET2), also known as vasoctive intestine peptide, cosegregated strongly with systolic blood pressure in a F2 population [F2(S x LEW)] derived from a cross of the Dahl salt-sensitive (S) rat and the Lewis (LEW/NCrlBR) (LEW) rat. The ET2 locus was assigned to rat chromosome 5. The testis-specific histone (HITH) locus also strongly cosegregated with blood pressure in the F2(S x LEW) population and was assigned to rat chromosome 17. Genetic maps of the regions containing the quantitative trait loci (QTL) for blood pressure on chromosomes 5 and 17 were constructed and the QTL were localized using the MAPMAKER/QTL program. The rat genes for endothelin-1, endothelin-3, and endothelin receptor A did not cosegregate with blood pressure in several F2 populations tested and were assigned to rat chromosomes 17, 3, and 19, respectively. Endothelin receptor B cosegregated weakly with blood pressure and was provisionally assigned to rat chromosome 15. We conclude that, in the rat, one new blood pressure QTL is located on chromosome 5 marked by the ET2 locus and another new QTL is located on chromosome 17 near the HITH locus.     

Spontaneous mutation for a quantitative trait in Drosophila melanogaster. II. Distribution of mutant effects on the trait and fitness

Exposure of the Chinese hamster ovarian AuxB1 cell line in vitro to fractionated X-irradiation generated sublines designated DXR-10, which proved resistant to multiple drugs and overexpressed P-glycoprotein (Pgp), as judged by Western blotting using the C219 monoclonal antibody. Further characterization of these irradiated DXR-10 sublines has provided evidence for: (i) the expression of cross-resistance to gramacidin D, taxol, puromycin and Navelbine, but not to daunomycin or mitoxantrone; (ii) overexpression of the class I Pgp, as judged by Western blotting using the C494 monoclonal antibody; (iii) decreased accumulation of 3H-vincristine, which could be enhanced by verapamil addition; (iv) unaltered accumulation and subcellular distribution of adriamycin; (v) significantly increased rhodamine 123 accumulation in the presence of verapamil; (vi) plasma-membrane ultrastructural modifications resulting in a significantly increased surface area; (vii) numerous clonal karyotypic alterations, with abnormalities involving the long arm of chromosome 1 being consistently identified; (viii) a lack of overexpression of sorcin; (ix) increased total glutathione levels and overexpression of glutathione S-transferase pi. The fact that only certain of these features are considered characteristic of the 'classic' multidrug-resistant CHRC5 cell line supports our earlier proposal that exposure to fractionated X-irradiation results in the expression of a unique drug-resistance phenotype.     

The effect of selective phosphodiesterase inhibitors on plasma insulin concentrations and insulin secretion in vitro in the rat

We have examined in rats the effects of Org 9935 (4,5-dihydro-6-(5,6-dimethoxy-benzo[b]-thien-2-yl)-methyl-1-(2H)-p yridazinone), a selective inhibitor of type 3 phosphodiesterase (phosphodiesterase 3) and Org 30029 (N-hydroxy-5,6-dimethoxy-benzo[b]-thiophene-2-carboximidamide HCl), an inhibitor of phosphodiesterase 3/4 on rat plasma insulin and glucose concentrations in pentobarbitone-anaesthetised rats and on insulin secretion by rat isolated islets. We have also compared their effects on islet phosphodiesterase activity. Org 9935 (0.1 and 1.0 mg kg(-1) i.v. 15 min previously) dose dependently elevated fasting and post-glucose (0.25 g kg(-1) i.v.) plasma insulin concentrations. Org 30029 in a dose of 10 mg kg(-1), but not 1 mg kg(-1), also increased plasma insulin concentrations. Neither drug modified either fasting or post-glucose plasma glucose concentrations. Each drug augmented glucose-induced insulin release by rat isolated islets in a static incubation system, with approximate EC50 values of 1.5 microM for Org 9935 and 20 microM for Org 30029. Phosphodiesterase activity, in both supernatant and pellet fractions of islet homogenates, was inhibited concentration dependently by each drug. Although the shape of the concentration-inhibition curve for Org 30029 precluded estimation of an IC50 value, this drug was clearly much less potent than Org 9935 (IC50 about 50 nM) in inhibiting islet phosphodiesterase activity. We conclude that the increase in plasma insulin produced by each drug is a consequence of augmented insulin secretion, probably secondary to inhibition of phosphodiesterase 3 in the islet beta cell, with a resultant elevation in cAMP. The failure of the drugs to modify plasma glucose may be due to concomitant inhibition of cAMP phosphodiesterase in liver and adipose tissue.     

Correlation of plasma corticosterone levels with running activity in the blinded rat

Blinded female reats were placed in running-wheel cages to monitor the phase of their activity cycle. After approximately a month of adaptation to the wheels, jugular vein blood samples were withdrawn at the beginning and end of the running phase of activity and analyzed for plasma corticosterone. By the time blood samples were obtained, the blined rats' activity cycles had desynchronized from the lighting cycle of the room in an apparently free-running fashion, and were out of phase with each other. The corticosterone levels were high at the beginning of the running phase and low at the end: begin run corticosterone was 34.5 +/- 13, end run corticosteronewas 14.2 +/- 10 (mean +/-SD in mug/100 ml of plasma; t = 7.93, df = 82, P less than 0.001). It was concluded that blinded rats do have an adrenocortical rhythm and that it is in phase with the activity cycle of each individual rat.     

Identification of a complex congenital heart defect susceptibility locus by using DNA pooling and shared segment analysis

The isotope dilution technique of [6-3H]glucose, [U-14C]lactate and [l-14C]propionate was used to evaluate the effect of dietary chromium (Cr) supplementation on whole-body kinetics of glucose, lactate, and propionate in rams. Rams were fed a high grain diet at 2% of body weight with or without 0.5 ppm of supplemental Cr from chelated Cr for the initial 14 days, and then intake was increased to 2.5% at body weight for the last 9 days. Weight gain was enhanced (P < 0.01) with Cr supplementation. Plasma concentrations of glucose, lactate, and propionate were not influenced by Cr supplementation. Turnover rates of glucose and lactate, and their interconversion were also not influenced. Propionate turnover rate tended to increase (P = 0.11) and the conversion of propionate to glucose increased (P < 0.05) with Cr supplementation, leading the increased proportional contribution of propionate to glucose turnover rate (P < 0.05). Chromium supplementation may influence the contribution of each glucogenic substrate for glucose production in rams fed a high grain diet.     

Expression and characterization of a 70-kDa fragment of the insulin receptor that binds insulin. Minimizing ligand binding domain of the insulin receptor

In order to characterize regions of the insulin receptor that are essential for ligand binding and possibly identify a smaller insulin-binding fragment of the receptor, we have used site-directed mutagenesis to construct a series of insulin receptor deletion mutants. From 112 to 246 amino acids were deleted from the alpha-subunit region comprising amino acids 469-729. The receptor constructs were expressed as soluble insulin receptor IgG fusion proteins in baby hamster kidney cells and were characterized in binding assays by immunoblotting and chemical cross-linking with radiolabeled insulin. The shortest receptor fragment identified was a free monomeric alpha-subunit deleted of amino acids 469-703 and 718-729 (exon 11); the mass of this receptor fragment was found by mass spectrometry to be 70 kDa. This small insulin receptor fragment bound insulin with an affinity (Kd) of 4.4 nM, which is similar to what was found for the full-length ectodomain of the insulin receptor (5.0 nM). Cross-linking experiments confirmed that the 70-kDa receptor fragment specifically bound insulin. In summary we have minimized the insulin binding domain of the insulin receptor by identifying a 70-kDa fragment of the ectodomain that retains insulin binding affinity making this an interesting candidate for detailed structural analysis.