Association of a polymorphism in the indoleamine- 2,3-dioxygenase gene and interferon-α-induced depression in patients with chronic hepatitis C.

Interferon (IFN)-α treatment for infectious diseases and cancer is associated with significant depressive symptoms that can limit therapeutic efficacy. Multiple mechanisms have been implicated in IFN-α-induced depression including immune, neuroendocrine and neurotransmitter pathways. To further explore mechanisms of IFN-α-induced depression and establish associated genetic risk factors, single nucleotide polymorphisms in genes encoding proteins previously implicated in IFN-α-induced depression were explored in two self-reported ethnic groups, Caucasians (n=800) and African Americans (n=232), participating in a clinical trial on the impact of three pegylated IFN-α treatment regimens on sustained viral response in patients with chronic hepatitis C. Before treatment, all subjects were free of psychotropic medications and had a score ≤20 on the Center for Epidemiologic Studies Depression Scale (CES-D), which was used to assess depressive symptom severity throughout the study. In Caucasians, a polymorphism (rs9657182) in the promoter region of the gene encoding indoleamine-2,3-dioxygenase (IDO1) was found to be associated with moderate or severe IFN-α-induced depressive symptoms (CES-D>20) at 12 weeks of IFN-α treatment (P=0.0012, P<0.05 corrected). Similar results were obtained for treatment weeks 24, 36 and 48. In subjects homozygous for the risk allele (CC, n=150), the odds ratio for developing moderate or severe depressive symptoms at treatment week 12 was 2.91 (confidence interval: 1.48-5.73) compared with TT homozygotes (n=270). rs9657182 did not predict depression in African Americans, who exhibited a markedly lower frequency of the risk allele at this locus. The findings in Caucasians further support the notion that IDO has an important role in cytokine-induced behavioral changes.

Associations of progesterone receptor polymorphisms with age at menarche and menstrual cycle length.

BACKGROUND: Age at menarche and menstrual cycle characteristics are indicators of endocrine function and may be risk factors for diseases such as reproductive cancers. The progesterone receptor gene (PGR) has been identified as a candidate gene for age at menarche and menstrual function. METHODS: Women office workers ages 19-41 self-reported age at menarche and participated in a prospective study of menstrual function and fertility. First-morning urine was used as the DNA source. 444 women were genotyped for a functional variant in PGR, rs1042838 (Val660Leu), and 264 women were also genotyped for 29 other SNPs across the extended gene region. RESULTS: Genetic variation across PGR was associated with age at menarche using a global score statistic (p = 0.03 among non-Hispanic whites). Women carrying two copies of the Val660Leu variant experienced menarche 1 year later than women carrying one or no copies of the variant (13.6 ± 0.5 vs. 12.6 ± 0.1; p = 0.03). The Val660Leu variant was also associated with decreased odds of short menstrual cycles (17-24 days) (OR, 95% CI: 0.54 [0.36, 0.80]; p = 0.002). CONCLUSION: Genetic variation in PGR was associated with age at menarche and menstrual cycle length in this population. Further investigation of these associations in a replication dataset is warranted.

Examination of reproductive aging milestones among women who carry the FMR1 premutation.

BACKGROUND: The fragile X premutation is characterized by a large CGG repeat track (55-199 repeats) in the 5' UTR of the FMR1 gene. This X-linked mutation leads to an increased risk for premature ovarian failure; interestingly, the association of repeat size with risk is non-linear. We hypothesize that the premutation-associated ovarian insufficiency is due to a diminished oocyte pool and examined reproductive aging milestones by repeat size group to determine if the same non-linear association is observed. METHODS: We analyzed cross-sectional reproductive history questionnaire data from 948 women with a wide range of repeat sizes. RESULTS: We have confirmed the non-linear relationship among premutation carriers for ovarian insufficiency. The mid-range repeat size group (80-100 repeats), not the highest group, had an increased risk for: altered cycle traits (shortened cycle length, irregular cycles and skipped cycles), subfertility and dizygotic twinning. Smoking, a modifiable risk, decreased the reproductive lifespan of women with the premutation by about 1 year, similar to its effect on non-carriers. As expected, premutation carriers were found to be at an increased risk for osteoporosis. CONCLUSIONS: Possible molecular mechanisms to explain the non-linear repeat size risk for ovarian insufficiency are discussed.

Simple methods for assessing haplotype-environment interactions in case-only and case-control studies.

For investigating haplotype-environment interactions in case-control studies, one can implement statistical methods based either on a retrospective likelihood (modeling the probability of haplotype and environment conditional on disease status) or a prospective likelihood (modeling the probability of disease status conditional on haplotype and environment). Retrospective approaches are generally more powerful than prospective approaches, but require an explicit model of the joint distribution of haplotype and environmental factors in the sample with the latter being particularly unattractive to specify. To resolve this issue, we propose a number of simple retrospective procedures for haplotype-environment interaction analysis that do not require explicit modeling of environmental covariates in the sample. We first consider a cases-only procedure, followed by a simple likelihood for case-control data that is proportional to the full-retrospective likelihood. Finally, we consider a retrospective procedure for inference on haplotype-environment interaction effects in matched or finely-stratified case-control studies. Our methods are based on the assumptions that haplotypes and environmental covariates are independent in the target population and that disease is rare. We illustrate our approaches using case-control data from the Finland-United States Investigation of Non-Insulin Dependent Diabetes Mellitus (FUSION) genetic study and simulated data.

Association of FMR1 repeat size with ovarian dysfunction.

BACKGROUND: Women who carry the FMR1 premutation allele have a significantly increased risk for ovarian dysfunction. We hypothesize that molecular characteristics of the FMR1 gene may explain this increased risk. METHODS: Thus, we examined the effect of FMR1 CGG repeat size and related factors on measures of ovarian dysfunction using data from 507 women with a wide range of repeat sizes. RESULTS AND CONCLUSIONS: We found a significant positive association of repeat size with ovarian dysfunction, but have preliminary evidence that this relationship is non-linear. We suggest that FMR1 repeat size in the lower range (<80 repeats) contributes to the variation in age at menopause; thus, FMR1 could be considered a quantitative trait locus. More importantly, when repeat size exceeds this threshold, the increase in risk for ovarian dysfunction is clinically significant. Intriguingly, this risk appears to plateau, or perhaps decrease, among women with very high repeats (> or =100 repeats).

The Finland-United States investigation of non-insulin-dependent diabetes mellitus genetics (FUSION) study. II. An autosomal genome scan for diabetes-related quantitative-trait loci.

Type 2 diabetes mellitus is a complex disorder encompassing multiple metabolic defects. We report results from an autosomal genome scan for type 2 diabetes-related quantitative traits in 580 Finnish families ascertained for an affected sibling pair and analyzed by the variance components-based quantitative-trait locus (QTL) linkage approach. We analyzed diabetic and nondiabetic subjects separately, because of the possible impact of disease on the traits of interest. In diabetic individuals, our strongest results were observed on chromosomes 3 (fasting C-peptide/glucose: maximum LOD score [MLS] = 3.13 at 53.0 cM) and 13 (body-mass index: MLS = 3.28 at 5.0 cM). In nondiabetic individuals, the strongest results were observed on chromosomes 10 (acute insulin response: MLS = 3.11 at 21.0 cM), 13 (2-h insulin: MLS = 2.86 at 65.5 cM), and 17 (fasting insulin/glucose ratio: MLS = 3.20 at 9.0 cM). In several cases, there was evidence for overlapping signals between diabetic and nondiabetic individuals; therefore we performed joint analyses. In these joint analyses, we observed strong signals for chromosomes 3 (body-mass index: MLS = 3.43 at 59.5 cM), 17 (empirical insulin-resistance index: MLS = 3.61 at 0.0 cM), and 19 (empirical insulin-resistance index: MLS = 2.80 at 74.5 cM). Integrating genome-scan results from the companion article by Ghosh et al., we identify several regions that may harbor susceptibility genes for type 2 diabetes in the Finnish population.

Improved inference of relationship for pairs of individuals.

Linkage analyses of genetic diseases and quantitative traits generally are performed using family data. These studies assume the relationships between individuals within families are known correctly. Misclassification of relationships can lead to reduced or inappropriately increased evidence for linkage. Boehnke and Cox (1997) presented a likelihood-based method to infer the most likely relationship of a pair of putative sibs. Here, we modify this method to consider all possible pairs of individuals in the sample, to test for additional relationships, to allow explicitly for genotyping error, and to include X-linked data. Using autosomal genome scan data, our method has excellent power to differentiate monozygotic twins, full sibs, parent-offspring pairs, second-degree (2 degrees ) relatives, first cousins, and unrelated pairs but is unable to distinguish accurately among the 2 degrees relationships of half sibs, avuncular pairs, and grandparent-grandchild pairs. Inclusion of X-linked data improves our ability to distinguish certain types of 2 degrees relationships. Our method also models genotyping error successfully, to judge by the recovery of MZ twins and parent-offspring pairs that are otherwise misclassified when error exists. We have included these extensions in the latest version of our computer program RELPAIR and have applied the program to data from the Finland-United States Investigation of Non-Insulin-Dependent Diabetes Mellitus (FUSION) study.

Binding of human urokinase type plasminogen activator and plasminogen to Borrelia species.

Borrelia burgdorferi binds human urokinase plasminogen activator (uPA), which cleaves plasminogen (Pgn) to plasmin. The ability of other Borrelia species to bind uPA, Pgn, or both was investigated. Borrelia coriacae, Borrelia garinii, Borrelia parkeri, Borrelia anserina, and Borrelia turicatae were compared with infectious and noninfectious B. burgdorferi isolates. All Borrelia species lacked endogenous proteases capable of digesting casein, but all species bound human uPA and Pgn, generating Pgn-dependent proteolytic activity. There were no significant differences in the amount of plasmin, Pgn, or uPA bound per spirochete of the different species. On unfixed borreliae, fluorochrome-conjugated uPA bound to all species. Early binding was at the terminus of B. burgdorferi, whereas diffuse binding was observed on B. coriacae, B. garinii, B. parkeri, and B. turicatae. These studies demonstrate that binding of human uPA and Pgn to borreliae occurs on multiple species with apparent differences in surface distribution.

Binding of human plasminogen and urokinase-type plasminogen activator to the Lyme disease spirochete, Borrelia burgdorferi.

Many bacteria that spread in the skin produce enzymes that digest extracellular matrix components. Borrelia burgdorferi spreads from a skin inoculation site to form the characteristic erythema migrans skin lesion. It was determined that B. burgdorferi does not produce collagenase, elastase, hyaluronidase, or other enzymes that digest extracellular matrix components. However, B. burgdorferi bound human plasmin, plasminogen (Pgn), and urokinase-type plasminogen activator (uPA). When spirochetes were sequentially incubated with Pgn and uPA, bioactive plasmin was generated on the surface of B. burgdorferi. B. burgdorferi did not produce an endogenous Pgn activator. Fluorochrome-conjugated uPA and Pgn colocalized to the terminus of the spirochete. In a mouse model, uPA-treated B. burgdorferi were more infectious than control spirochetes. Binding of host uPA and Pgn to form a bioactive extracellular matrix protease on B. burgdorferi represents a mechanism that could facilitate dissemination and localization of spirochetes to sites of vascular injury.

Effects of topical ethacrynic acid adducts on intraocular pressure in rabbits and monkeys.

We evaluated the effect of topical ethacrynic acid on rabbit and monkey intraocular pressure. In a preliminary experiment, 100-mmol/L ethacrynic acid applied topically to Dutch-Belted rabbit eyes was associated with an 8-mm Hg lowering of intraocular pressure. However, corneal edema was severe, and the corneal epithelium sloughed off. To try to maintain the pressure-lowering effect but reduce the corneal side effects, we attempted to create an adduct of ethacrynic acid by utilizing ethacrynic acid's sulfhydryl reactivity. Ethacrynic acid was mixed with equimolar cysteine to bind the sulfhydryl-reactive sites on ethacrynic acid. The goal was to expose the cornea to adducted ethacrynic acid, which might then dissociate in the anterior chamber via a retro-Michael reaction. Intraocular pressure decreased 8.9 mm Hg (n = 40) with this treatment, and corneal edema was lessened (32 of 40 eyes had mild to no edema). However, we observed that when the eye was treated before ethacrynic acid-cysteine administration with topical acetylcysteine, the corneal side effects were reduced further and the intraocular pressure effect remained. In living cynomolgus monkeys receiving a single pretreatment drop of 75-mmol/L acetylcysteine followed by two drops of 130-mmol/L ethacrynic acid and 130-mmol/L cysteine, an intraocular pressure lowering of 9.9 mm Hg was observed (n = 7). However, in three of seven eyes corneal edema developed. Pretreatment with two drops of acetylcysteine eliminated the pressure-lowering effect but did not confer any added corneal protection. Our results indicate that topical ethacrynic acid-cysteine is effective in lowering the intraocular pressure of rabbits and cynomolgus monkeys and, when combined with acetylcysteine pretreatment, may offer the potential for a new topical therapeutic regimen for use in glaucoma.

Thiol adducts of ethacrynic acid increase outflow facility in enucleated calf eyes.

Ethacrynic acid (ECA), a sulfhydryl (SH)-reactive diuretic drug, has been shown to increase outflow facility (C) both in living monkey eyes and in the calf eye, in vitro (Epstein et al. 1987). In an attempt to increase the therapeutic index of this drug for potential clinical use in glaucoma, we explored the effect of various thiol adducts of ECA on C in the calf eye in vitro. These adducts might be expected to liberate ECA by a reversible retro-Michael type reaction. Enucleated calf eyes were perfused at 25 degrees C at 15 mm Hg for 5 hours with various ECA-thiol adducts. ECA-cysteine at 0.25 mM (for each) increased outflow facility 104% compared to 38% in sham manipulated eyes (n-10; p less than .005). A dose response effect was demonstrated from 0.01 mM to 0.25 mM. A relative potency table (for increasing C) was established for several ECA-thiol adducts: Cysteine = cysteamine greater than glutathione greater than N-Acetyl cysteine greater than thiosalicylic acid greater than N-Acetyl cysteamine. This study identifies the potential of utilizing various derivatives of ECA as outflow pathway acting agents.