Hammondia sp. oocysts shed by a Brazilian fox (Lycalopex vetulus) differ from Hammondia heydorni and Hammondia triffittae.

A Brazilian fox (Lycalopex vetulus) was rescued from a highway, and 16 days after maintained in captivity, the fox shed oocysts with sizes compatible with Hammondia sp. and Neospora caninum. DNA extracted from oocysts were initially tested in two PCRs targeting the internal transcribed spacer 1 (ITS-1) of the rDNA of Hammondia heydorni and the Nc-5 gene of N. caninum. A 270-bp product was visualized in the PCR for H. heydorni. No amplification was observed for N. caninum PCR. Since ITS-1-based PCR is not sufficient to differentiate Hammondia species derived from canids, oocyst DNA was examined using multilocus sequence analysis of five genetic fragments [intron 1 of the alpha tubulin gene (intron 1), internal transcribed spaces 1 and 2 (ITS-1 and ITS-2) of the rDNA, 28S rRNA gene (D2/D3 domain), and heat shock protein 70 (Hsp70)]. The Hammondia sp. oocyst from the Brazilian fox, referred here as H-FOXBR isolate, is closely related to H. heydorni and Hammondia triffittae, but differs from these parasites in three genetic markers (alpha tubulin gene, ITS-2, and 28S rRNA). As reported by other research groups, Hammondia spp. excreted by canids are genetically diverse and may encompass additional species besides H. heydorni and H. triffittae. In this study, we confirmed that H-FOXBR has significant genetic differences in comparison to H. heydorni and H. triffittae and may represent a separate species. Further studies are needed to identify the life cycle of this parasite and to characterize the parasite stages in the intermediate and definitive hosts.

Mater, a maternal effect gene required for early embryonic development in mice.

Maternal effect genes produce mRNA or proteins that accumulate in the egg during oogenesis. We show here that Mater, a mouse oocyte protein dependent on the maternal genome, is essential for embryonic development beyond the two-cell stage. Females lacking the maternal effect gene Mater are sterile. Null males are fertile.

Coffee, ADORA2A, and CYP1A2: the caffeine connection in Parkinson's disease.

BACKGROUND AND PURPOSE: In 1-methyl-4-phenyl 1,2,3,6-tetrahydropyridine animal models of Parkinson's disease (PD), caffeine protects neurons by blocking the adenosine receptor A2A (ADORA2A). Caffeine is primarily metabolized by cytochrome P450 1A2 (CYP1A2). Our objective was to examine whether ADORA2A and CYP1A2 polymorphisms are associated with PD risk or modify the caffeine-PD association. METHODS: Parkinson's Epidemiology and Genetic Associations Studies in the United States (PEGASUS) included five population-based case-control studies. One laboratory genotyped four ADORA2A and three CYP1A2 polymorphisms in 1325 PD cases and 1735 age- and sex-matched controls. Information regarding caffeine (coffee) consumption and other lifestyle factors came from structured in-person or telephone interviews. Odds ratios (OR) and 95% confidence intervals (CI) were estimated using logistic regression. RESULTS: Two ADORA2A polymorphisms were inversely associated with PD risk - rs71651683, a 5' variant (adjusted allelic OR = 0.51, 95% CI 0.33-0.80, permutation-adjusted P = 0.015) and rs5996696, a promoter region variant (adjusted OR for AC and CC genotypes compared with the AA wild-type genotype were 0.76 (95% CI 0.57-1.02) and 0.37 (95% CI 0.13-1.01), respectively (permutation-adjusted P for trend = 0.04). CYP1A2 polymorphisms were not associated with PD risk; however, the coffee-PD association was strongest among subjects homozygous for either variant allele rs762551 (P(interaction) = 0.05) or rs2470890 (P(interaction) = 0.04). CONCLUSION: In this consortium study, two ADORA2A polymorphisms were inversely associated with PD risk, but there was weak evidence of interaction with coffee consumption. In contrast, the coffee-PD association was strongest among slow metabolizers of caffeine who were homozygous carriers of the CYP1A2 polymorphisms.

Failure of clonal deletion in neonatally thymectomized mice: tolerance is preserved through clonal anergy.

Self-tolerance is achieved in part through intrathymic deletion of self-reactive T cells. The necessity of the thymus for this process is suggested by the development of autoimmune diseases in neonatally thymectomized (neoTx) mice and by the failure of clonal deletion in nude mice. Indeed, the present study demonstrates that neonatal thymectomy on day 3 after birth results in the failure of clonal deletion of V beta 11+ T cells in BALB/c mice and V beta 5+ and V beta 6+ T cells in DBA/2 mice. However, these potentially autoreactive cells are nonfunctional as measured by proliferation and lymphokine production after stimulation with appropriate anti-V beta mAbs or stimulator cells. It appears that this induction of nonresponsiveness may have occurred extrathymically: the early neonatal thymus (presumably the source of the peripheral T cells observed in neoTx mice) also contains T cells with self-reactive receptors, but these cells are fully functional. Therefore, neonatal thymectomy aborts deletion of self-reactive T cells, but self-tolerance is maintained through functional inactivation of potentially self-reactive clones.

Autoimmune diseases prior to the diagnosis of multiple sclerosis: a population-based case-control study.

The objective of this study was to determine whether patients with multiple sclerosis (MS) are more likely to have other autoimmune disorders particularly prior to the diagnosis of MS. We conducted a population-based case-control study of patients enrolled in the Northern California Kaiser Permanente Medical Care Program. Electronic clinical records through 2005 were used to ascertain incident and prevalent MS cases and identify the presence and timing of 44 other diagnoses. Controls were matched 5:1 for gender, age, and Kaiser membership characteristics. We identified 5296 MS cases (including 924 diagnosed between 2001 and 2004) and 26,478 matched controls. Prior to MS diagnosis, cases were more likely than controls to have uveitis (OR = 3.2, 95%; CI 1.7-5.7), inflammatory bowel disease (IBD, OR = 1.7; 95%CI 1.2-2.5), and Bell's palsy (OR = 3.2; 95%CI 1.2-8.3). Cases were also more likely to develop Guillain- Barré syndrome (GBS, OR = 5.0; 95%CI 1.6-15.4) and bullous pemphigoid (OR = 6.7; 95%CI 1.5-29.9). Cases were not more likely than controls to have or to develop rheumatoid arthritis, lupus or thyroiditis. MS may share environmental triggers, genetic susceptibilities and/or alterations in immune homeostasis with IBD and uveitis, but not with other autoimmune disorders.

Development and validation of the Axillary Sweating Daily Diary: a patient-reported outcome measure to assess axillary sweating severity.

BACKGROUND: Hyperhidrosis is estimated to affect ~ 4.8% of the US population, and most patients experience a negative psychological impact. Here, we describe development and psychometric evaluation of a patient-reported outcome (PRO) measure to assess severity of axillary hyperhidrosis in clinical trials that meets current U.S. regulatory standards to support product approvals. METHODS: Three rounds of hybrid concept-elicitation/cognitive-debriefing qualitative interviews were conducted in adults with clinician-diagnosed primary axillary hyperhidrosis, followed by similar interviews in children/adolescents. The draft measure included diary items for presence, severity, impact and bothersomeness (basis of the Axillary Sweating Daily Diary [ASDD]), exploratory weekly impact items, and a single-item Patient Global Impression of Change (PGIC). Phase 2 (adults only) and phase 3 (adults and children ≥9 years) clinical trial data were utilized to evaluate measurement properties of the resulting draft measure: floor/ceiling effects, nonresponse bias, test-retest reliability, construct validity, and responsiveness were assessed. The primary concept of interest was axillary sweating severity (ASDD Item 2); however, additional supportive concepts were explored to allow for development of a comprehensive hyperhidrosis measure. RESULTS: Twenty-nine patient interviews were conducted (N = 21 adult and N = 8 children/adolescents), resulting in the ASDD (4 items, patients ≥16y) and child-specific ASDD-C (2 items ≥9y to <16y), as well as 6 Weekly Impact items and the PGIC (patients ≥16y). No floor/ceiling effects or response biases were identified. Consistency between hypothesized and observed correlation patterns between ASDD/ASDD-C items and other efficacy measures supported construct validity. Intraclass correlation coefficients supported test-retest reliability (0.91-0.93; Item 2). Large effect sizes (- 2.2 to - 2.4) demonstrated that the ASDD/ASDD-C Item 2 could detect changes in hyperhidrosis severity, supporting the measure's responsiveness. Patients perceiving a moderate improvement in symptoms on the PGIC experienced an average 3.8-point improvement on ASDD axillary sweating severity (Item 2); thus, a 4-point responder threshold was defined as a clinically meaningful change. CONCLUSIONS: Qualitative and quantitative evidence support the reliability and validity of the ASDD/ASDD-C and its use in the clinical evaluation of axillary hyperhidrosis treatments. Further evaluation of this measure in future research studies is warranted to demonstrate consistent performance across different axillary hyperhidrosis populations and in different study contexts.

Obstetric interventions and maternal morbidity among women who experience severe postpartum hemorrhage during cesarean delivery.

BACKGROUND: Compared to vaginal delivery, women undergoing cesarean delivery are at increased risk of postpartum hemorrhage. Management approaches may differ between those undergoing prelabor cesarean delivery compared to intrapartum cesarean delivery. We examined surgical interventions, blood component use, and maternal outcomes among those experiencing severe postpartum hemorrhage within the two distinct cesarean delivery cohorts. METHODS: We performed secondary analyses of data from two cohorts who underwent prelabor cesarean delivery or intrapartum cesarean delivery at a tertiary obstetric center in the United States between 2002 and 2012. Severe postpartum hemorrhage was classified as an estimated blood loss ≥1500mL or receipt of a red blood cell transfusion up to 48h post-cesarean delivery. We examined blood component use, medical and surgical interventions and maternal outcomes. RESULTS: The prelabor cohort comprised 269 women and the intrapartum cohort comprised 278 women. In the prelabor cohort, one third of women received red blood cells intraoperatively or postoperatively, respectively. In the intrapartum cohort, 18% women received red blood cells intraoperatively vs. 44% postoperatively (P<0.001). In the prelabor and intrapartum cohorts, methylergonovine was the most common second-line uterotonic (33% and 43%, respectively). Women undergoing prelabor cesarean delivery had the highest rates of morbidity, with 18% requiring hysterectomy and 16% requiring intensive care admission. CONCLUSION: Our findings provide a snapshot of contemporary transfusion and surgical practices for severe postpartum hemorrhage management during cesarean delivery. To determine optimal transfusion and management practices in this setting, large pragmatic studies are needed.

A mouse gene encoding an oocyte antigen associated with autoimmune premature ovarian failure.

Autoimmune premature ovarian failure causes young women to develop menopausal symptoms and infertility. A similar syndrome appears in mice with postthymectomy autoimmune premature ovarian failure. We demonstrate that these mice develop antibodies against a 125-kDa protein located in the oocyte cytoplasm (ooplasm). By screening a mouse ovarian complementary DNA expression library with autoimmune serum, we have identified a novel mouse gene with a 3.75-kb ovarian transcript, the expression of which is restricted to the oocyte. The longest open reading frame (3333 bp) encodes an oocyte-specific protein, designated OP1 (ooplasm-specific protein 1). The protein is composed of 1111 amino acids with a predicted molecular mass of 125,502 Da. Based on its primary structure, it appears to be novel and has no motifs to suggest a localization other than in the cytoplasm. The ability of immune serum from mice with ovarian autoimmunity to react specifically with recombinant OP1 raises the possibility that OP1 as an antigen may play a role in murine autoimmune premature ovarian failure.

Glycoprotein hormone alpha-subunit functions synergistically with progesterone to stimulate differentiation of cultured human endometrial stromal cells to decidualized cells: a novel role for free alpha-subunit in reproduction.

Glycoprotein hormone-free alpha subunit is secreted by the pituitary throughout the menstrual cycle and by the placenta during pregnancy. We showed previously that free alpha subunit stimulated PRL secretion from term pregnancy decidual cells, suggesting a function for free alpha in pregnancy. However, no role has been ascribed to free alpha in the normal menstrual cycle. Using an in vitro model, we examined the role of alpha subunit in regulating human endometrial stromal cell differentiation (decidualization). PRL and insulin-like growth factor binding protein-1 (IGFBP-1), specific decidual secretory products, were used as markers of decidualization. We found that alpha subunit acted synergistically with progesterone (P) to induce more rapid decidualization with higher output (2- to 6-fold) of PRL and IGFBP-1, compared with P alone (P < 0.01). The effect of alpha was dose dependent, with stimulatory activity starting at 0.05 ng/ml and reaching maximal levels at 1-2 ng/ml. These levels correspond to serum concentrations of free alpha found during the luteal phase of the menstrual cycle when endometrial decidualization occurs in vivo. These findings demonstrate new biological activity for alpha subunit in the regulation of human endometrial decidualization and indicate that free alpha plays a role in human reproduction. Furthermore, demonstration of potential bioactivities of free alpha subunit has important implications for understanding normal endocrine function and various pathological conditions.

The use of human recombinant gonadotropin receptors to search for immunoglobulin G-mediated premature ovarian failure.

Anti-FSH receptor antibodies, detected using animal systems, have been reported in a few patients with premature ovarian failure (POF). However, assays based on animal receptors may be inappropriate for detecting inhibiting antibodies in humans. Accordingly, we tested for interfering antibodies in patients with POF using a recombinant system expressing human (h) FSH and LH receptors. A mouse adrenal cell line transfected with the hFSH receptor (Y1-hFSHR) exhibits a dose-dependent increase in progesterone when exposed to hFSH. An embryonal kidney cell line transfected with the hLH receptor gene (hLHR-293) exhibits a dose-dependent increase in cAMP when exposed to hLH. We isolated immunoglobulins G (IgG) from 38 patients with POF and 14 normal women. We stimulated Y1-hFSHR and hLHR-293 cells with hFSH or hLH in the presence of these IgG and determined the resulting progesterone or cAMP response. The progesterone and cAMP responses obtained in the presence of IgG from patients with POF did not differ significantly from the responses in the presence of IgG from normal women. In contrast, antigonadotropin polyclonal antibodies isolated in the same manner as the above IgGs caused a greater than 90% reduction in the response of the Y1-hFSHR and hLHR-293 cells. We did not detect inhibitory antibodies in any of our 38 patients with POF. Therefore, if blocking antibodies interfering with gonadotropin-receptor interaction are a mechanism for POF, they account for a small minority of cases (< 8%).

A potential novel mechanism for precocious puberty in juvenile hypothyroidism.

Some children with juvenile hypothyroidism exhibit unexplained precocious puberty. Interaction of TSH with the human FSH receptor (hFSH-R) is a possible pathophysiological mechanism for this syndrome that has not been explored due to the lack of hFSH-free TSH preparations and the scarcity of a suitable hFSH-R-based assay system. To devise an in vitro FSH bioassay suitable for exploring this mechanism, we expressed hFSH-R complementary DNA in COS-7 cells and stimulated them with recombinant hTSH (rec-hTSH). Rec-hTSH elicited a dose-dependent cAMP response in the in vitro hFSH-R bioassay; however, the concentration of rec-hTSH required for half-maximal stimulation was several logs greater than that of hFSH. Rec-hTSH acted as a competitive inhibitor of hFSH at the hFSH-R, indicating that hTSH and hFSH are acting through the same receptor, namely the hFSH-R. This provides a potential novel mechanism for the precocious puberty of juvenile hypothyroidism.

Development of luteinized graafian follicles in patients with karyotypically normal spontaneous premature ovarian failure.

Despite having amenorrhea and markedly elevated serum gonadotropin levels, some women with karyotypically normal spontaneous premature ovarian failure, nevertheless, have ovarian follicles that function intermittently. Graafian follicles capable of responding to these high FSH levels are faced with high serum LH levels as well, which might induce inappropriate luteinization and prevent normal follicle function. We examined this possibility using weekly blood sampling and sonography in 65 patients. Nearly 50% of our patients demonstrated ovarian follicle function [serum estradiol, > 183 pmol/L (50 pg/mL)] during a median of 4 months of observation (range, 2-6 months). However, during this observation, only 16% achieved an ovulatory serum progesterone level [> 9.5 nmol/L (3.0 ng/mL)]. We imaged an antral follicle by sonography in over 40% of patients (27 of 65), and serum estradiol was significantly greater when an antral follicle was present. The follicles in these patients were not functioning normally, however. In contrast to normal women, patients with ovarian failure had poor correlation between follicle diameter and serum estradiol. We biopsied these antral follicles in 6 patients and found luteinized Graafian follicles in all cases. Therefore, luteinized Graafian follicles account for at least 60% of the antral structures imaged (95% confidence limit). Thus, inappropriate luteinization of Graafian follicles appears to be a major pathophysiological mechanism in patients with karyotypically normal spontaneous premature ovarian failure.

Karyotypically normal spontaneous premature ovarian failure: evaluation of association with the class II major histocompatibility complex.

Patients with premature ovarian failure (POF) have been reported to have an increased frequency of the major histocompatibility class (MHC) class II antigen HLA-DR3. Here we attempt to confirm this association. We performed MHC class II immunophenotyping of HLA-DR antigens 1-10 on 102 North American caucasians with confirmed POF and 102 control caucasian women. All patients had experienced amenorrhea before the age of 40 yr and had elevated serum gonadotropins on repeated study. We found no significant increase in HLA-DR3 frequency in patients with POF when compared to our control group (P = 0.52) or even when compared to a large reference population (n = 1927) that did not differ significantly from our control group (P = 0.47). Our patients did have an increased frequency of HLA DR4 compared to this large reference population (41% vs. 23%; P < 0.001), but we were unable to demonstrate increased HLA DR4 frequency using our control group (31%; P = 0.14). In conclusion, despite a power of 99%, we were unable to confirm a significant increase in MHC class II HLA-DR3 frequency in patients with POF.

Study design in genetic epidemiology: theoretical and practical considerations.

Recent advances in molecular genetics have created new opportunities and challenges for genetic epidemiologists. Here we review some of the issues that arise when designing a study involving the genetic epidemiology of chronic diseases of late onset, such as cancer. We discuss two considerations that influence the choice of design. The first consideration is the study's goals. We describe the goals of identifying new susceptibility genes for a disease, of estimating important characteristics of known genes, and of learning how to prevent the disease in the genetically susceptible. We indicate how these goals affect the choice of design and present some guidelines for choosing designs that effectively address them. The second consideration is the set of practical constraints to successfully conducting the research. These contraints include problems of potential selection bias, reduced response rates, problems particular to family registries, problems particular to the cultures of various ethnic groups, and ethical issues. We indicate how these constraints affect the choice of design and discuss ways to deal with them.

A comparison of dementia in Alzheimer's disease and multiple sclerosis.

We compared results of comprehensive neuropsychological testing in 42 patients with clinically diagnosed Alzheimer's disease (AD) and in an equal number of patients with clinically definite chronic-progressive multiple sclerosis. Age, sex, and education were controlled using demographically corrected T scores based on a large normal sample. Both groups showed significant impairment on the test battery, but the degree of dementia was more severe in the patients with AD. A deviation score analysis, controlling for overall level of cognitive impairment, revealed significant differences between the groups. Alzheimer's disease was associated with relatively greater impairment of learning, memory, and verbal skills, whereas the MS group showed greater relative impairment of attention, incidental memory, and psychomotor functions. These data suggest that both the degree and pattern of mental impairement differ in patients with AD and patients with multiple sclerosis. Our results support a distinction between "gray matter" and "white matter" dementia, and may help clarify the issue of "cortical" vs "subcortical" dementia by demonstrating neuropsychological differences based on secure neuropathologic distinctions.

Cognitive loss in multiple sclerosis. Case reports and review of the literature.

Neuropsychological and neuroradiologic evidence of cerebral lesions is described for 12 patients with multiple sclerosis in whom cognitive disability was far greater than any other neurologic disability. Cognitive dysfunction resulted in significant functional impairment at work or home in three fourths (9 of 12) of the patients described here, despite mild physical disability (mean Kurtzke Expanded Disability Status Scale score, 3.2). A unique feature of the neurologic examination in these patients was the presence of prominent frontal release signs (gait apraxia and placing response) in the lower extremities. Two new scales, a Cognitive Function Scale and a Frontal Release Scale, were adapted for the investigation of these patients. The extensive literature relating to cognitive dysfunction in multiple sclerosis is reviewed and discussed with regard to its clinical relevance.

Helicobacter pylori, pepsinogen, and risk for gastric adenocarcinoma.

The objective of this project was to determine the association of Helicobacter pylori infection and serum pepsinogen levels on subsequent risk for gastric adenocarcinoma. This nested case-control study was set in a large health maintenance organization. One hundred thirty-six cases of gastric adenocarcinoma and 136 matched controls without adenocarcinoma from a large cohort that had contributed serum in the 1960's were studied. The presence of IgG against H. pylori had previously been determined by enzyme-linked immunosorbent assay. Serum levels of pepsinogens I and II were ascertained by radioimmunoassay. In a sample of subjects, the presence of antiparietal cell antibodies was determined by immunofluorescent antibody assay (Nichols Laboratory). There were 98 cases of adenocarcinoma of the antrum, body, or fundus (distal cancers) and 30 of the cardia or gastroesophageal junction (proximal cancers). By univariate analysis, H. pylori infection [odds ratio (OR), 3.6; P < 0.001] and serum pepsinogen I < 50 ng/ml (OR = 2.9; P = 0.003) were both associated with development of distal cancer. In multivariate analysis, there was interaction between the two variables; H. pylori in the absence of low pepsinogen I was independently associated with cancer (OR, 2.4; P = 0.04) but low pepsinogen I in the absence of H. pylori infection was not associated with cancer (OR, 0.8; P > 0.5). In combination, however, H. pylori infection and a low pepsinogen I were associated with a marked increase in the risk of developing distal malignancy (OR, 10.0; P = 0.08).(ABSTRACT TRUNCATED AT 250 WORDS)

Neurology practice patterns in Colorado.

Eighty-one percent (47/58) of private-sector neurologists in Colorado responded to a mail survey of practice patterns. We evaluated patient load, degree of principal versus consultative care, and use of neurodiagnostic tests. Broad areas of neurologic education that were perceived to have been lacking during residency training were also addressed. Eight-four percent (41/49) of responding neurologists agreed to participate in a more comprehensive prospective study of neurology practice patterns. The prospective data will provide information to determine how many neurologists are needed and the applications for neurologic education.

Clinical course of spontaneous subarachnoid hemorrhage: a population-based study in King County, Washington.

Over 2 years, we identified 171 patients in King County, Washington, experiencing an incident subarachnoid hemorrhage and characterized their clinical course and outcome. Most (65%) were women and most (68%) were under age sixty-five. Only five died without medical attention. The remaining 166 patients were hospitalized and had CTs of the head. Of these, 103 underwent aneurysm surgery, 40 developed acute hydrocephalus, 32 had symptomatic vasospasm, and 30 re-bled. Sixty-eight percent (68%) survived to 1 month after the bleed and 62% to 1 year. Independent predictors of good recovery by 1 month after the bleed included youth, a high score on the admission Glasgow Coma Scale, and absence of blood on the first CT. In this population-based series, at 1 month after the bleed, approximately one-third of patients were dead, one-third had neurologic deficits, and one-third were doing well.