The microenvironment of ovarian follicles in fertile dairy cows is associated with high oocyte quality.

We hypothesised that heifers and cows with positive genetic merit for fertility would have a follicular microenvironment that resulted in better quality oocytes. To test this, we compared cumulus cell-oocyte complexes (COC) and follicular fluid from preovulatory follicles of 36 Holstein-Friesian nulliparous heifers and 50 primiparous lactating cows with either positive (POS, +5%) or negative (NEG, -5%) fertility breeding values (FertBV). Established gene markers of oocyte quality were measured in individual cumulus cell masses and oocytes, and concentrations of amino acids, steroids, and metabolites were quantified in corresponding follicular fluid and plasma. The timing of visually detectable oestrus in NEG FertBV heifers was inconsistent with their stage of COC maturation. Retrospective analyses of oestrous activity data indicated that NEG FertBV heifers were sampled earlier. Their recovered COC were morphologically less mature and exhibited differential expression of genes that are associated with follicular maturation (lower levels of BMPR2) and protein processing (higher levels of HSP90B1). Despite consistent sampling times being achieved in the lactating cows, lower concentrations of serine, proline, methionine, isoleucine, and non-esterified fatty acids were present in follicular fluid from POS FertBV cows. This was associated with higher expression of gene biomarkers of good oocyte quality (VCAN, PDE8A) in COC recovered from POS FertBV cows. This study supports our hypothesis that the follicular microenvironment in lactating dairy cows with high genetic merit leads to COC with higher metabolic rates and oocytes of superior quality. Moreover, an additional stressor such as lactation is required for this difference to be pronounced.

Orchestration of protein acetylation as a toggle for cellular defense and virus replication.

Emerging evidence highlights protein acetylation, a prevalent lysine posttranslational modification, as a regulatory mechanism and promising therapeutic target in human viral infections. However, how infections dynamically alter global cellular acetylation or whether viral proteins are acetylated remains virtually unexplored. Here, we establish acetylation as a highly-regulated molecular toggle of protein function integral to the herpesvirus human cytomegalovirus (HCMV) replication. We offer temporal resolution of cellular and viral acetylations. By interrogating dynamic protein acetylation with both protein abundance and subcellular localization, we discover finely tuned spatial acetylations across infection time. We determine that lamin acetylation at the nuclear periphery protects against virus production by inhibiting capsid nuclear egress. Further studies within infectious viral particles identify numerous acetylations, including on the viral transcriptional activator pUL26, which we show represses virus production. Altogether, this study provides specific insights into functions of cellular and viral protein acetylations and a valuable resource of dynamic acetylation events.

NeuroAiD® (MLC601) and amyloid precursor protein processing.

BACKGROUND: Amyloid precursor protein (APP) undergoes cleavage under physiological conditions, predominantly by α- and γ-secretases, to form the nonpathogenic sAPPα and p3 fragments. By contrast, amyloid-beta (Aß) is produced via proteolytic cleavage by ß- and γ-secretases. In Alzheimer's disease (AD), APP is preferentially processed via the amyloidogenic pathway, producing large amounts of Aß that form the major constituent of senile plaques and tau-containing neurofibrillary tangles. Similarly, stroke patients have a higher level of Aß around the area of infarct, suggesting that Aß may mediate at least some of the secondary neurotoxicity observed in stroke patients. METHODS: To investigate the effects of MLC601 (NeuroAiD(®)) on regulation of APP processing, the human neuroblastoma cell line SH-SY5Y was used for all experiments. Stocks of MLC601 were prepared at a final concentration of 50 mg/ml. Cells were treated with different concentrations of MLC601 before assessing changes in the levels of released lactate dehydrogenase (LDH), full-length APP and secreted sAPPα. RESULTS: Concentrations of MLC601 between 1 and 1,000 µg/ml significantly lowered the levels of LDH released into the media when compared to control cells. In contrast, MLC601 concentrations at 5,000 and 10,000 µg/ml resulted in a significant increase in the LDH release. Treatment with 100, 500 and 1,000 µg/ml of MLC601 significantly increases the levels of sAPPα secreted by SH-SY5Y into the media. Treatment with 1,000 µg/ml of MLC601 significantly decreased the levels of full-length APP. CONCLUSION: MLC601 is a possible modulator of APP processing and has implications as a putative therapeutic strategy for the treatment of poststroke dementia and AD.

Toll like receptor-2 modulates both innate and adaptive immune responses during chronic fungal asthma in mice.

OBJECTIVE AND DESIGN: We investigated the effect of TLR2 gene deletion in a murine model of chronic fungal asthma. METHODS: TLR2 wildtype (TLR2(+/+)) and TLR2 deficient (TLR2(-/-)) mice were sensitized to soluble A. fumigatus antigens and challenged with live A. fumigatus conidia, and the extent of allergic airways disease was analyzed in both groups of mice at 3, 7, 14, and 30 days after conidia. RESULTS: At day 7 post-conidia, TLR2(-/-) mice exhibited significantly lower airway hyperresponsiveness, airway inflammation, and whole lung Th2 cytokine levels compared with the TLR2(+ / +) group. TLR2 deletion also significantly reduced mucus cell metaplasia and peribronchial fibrosis at day 30 after conidia. However, fungal material persisted in the TLR2(-/-) group, and at day 30 after conidia TLR2(-/-) mice exhibited enhanced airway neutrophil recruitment and airway hyperresponsiveness. CONCLUSION: Thus, during chronic fungal asthma in mice, TLR2 is a major contributor to the maintenance of the adaptive Th2-cytokine driven and anti-fungal innate responses.

Role of chemokines in severe asthma.

The severe asthma phenotype is exhibited by a subset of asthma patients whose asthma symptom is poorly controlled by current therapies. Severe asthma represents a high unmet medical need and warrants research into the mechanisms driving the underlying pathophysiology. It is hypothesized that the underlying pathology associated with severe asthma is driving the symptoms experienced by these patients, which may share common features with mild to moderate asthma or may represent a unique pathological phenotype. For the purpose of this review, the pathophysiology associated with asthma in general are described and extended to incorporate severe asthma. Chemokines may contribute towards multiple features of asthma pathophysiology and this current review focuses on the biology of chemokines pertaining to asthma pathophysiology. Chemokines are important recruiters and activators of inflammatory cells and these infiltrating cells interact with resident cells, such as fibroblasts and it is through these pathways that chemokines appear to exert multiple biological actions. Clinical trials are underway with therapeutics targeting chemokine pathways for other inflammatory diseases. It is hoped that the information generated from these studies will contribute towards furthering our understanding of chemokine biology and be applied towards targeting severe asthma.

Prescription drug coverage and spending for Medicare beneficiaries.

Outpatient prescription drug coverage is not a Medicare covered benefit. Debate continues in Congress and elsewhere on modernizing the Medicare benefit package, including proposals that would help the Nation's seniors pay for prescription drugs. Very little is known about which persons within the Medicare population have drug coverage from other sources. Using 1995 data from the Medicare Current Beneficiary Survey (MCBS), the authors present information on who has coverage by various sociodemographic categories. The data indicate higher-than-average levels of coverage for minority persons, beneficiaries eligible for Medicare because of disability, and those with higher incomes.

Abnormal cardiac structure and function in mice expressing nonphosphorylatable cardiac regulatory myosin light chain 2.

A role for myosin phosphorylation in modulating normal cardiac function has long been suspected, and we hypothesized that changing the phosphorylation status of a cardiac myosin light chain might alter cardiac function in the whole animal. To test this directly, transgenic mice were created in which three potentially phosphorylatable serines in the ventricular isoform of the regulatory myosin light chain were mutated to alanines. Lines were obtained in which replacement of the endogenous species in the ventricle with the nonphosphorylatable, transgenically encoded protein was essentially complete. The mice show a spectrum of cardiovascular changes. As previously observed in skeletal muscle, Ca(2+) sensitivity of force development was dependent upon the phosphorylation status of the regulatory light chain. Structural abnormalities were detected by both gross histology and transmission electron microscopic analyses. Mature animals showed both atrial hypertrophy and dilatation. Echocardiographic analysis revealed that as a result of chamber enlargement, severe tricuspid valve insufficiency resulted in a detectable regurgitation jet. We conclude that regulated phosphorylation of the regulatory myosin light chains appears to play an important role in maintaining normal cardiac function over the lifetime of the animal.

Mood congruence and depressive deficits in memory: a forced-recall analysis.

Two experiments examined the contribution of reporting biases to mood-congruent recall patterns and diminished levels of recall frequently associated with depressed mood states. In Experiment 1, participants classified as dysphoric (n = 14) or nondepressed (n = 21) on the basis of scores on the Beck Depression Inventory and the Profile of Mood States made self-referential judgements regarding a series of affectively valenced words. Subsequently they were given an unexpected forced-recall test, which encouraged guessing to meet the output requirement (i.e. 40 responses) of the test. Nondepressed subjects confidently reported more positive words than dysphoric subjects, but the latter produced significantly more correct guesses of words that were positively valenced. Similar findings were obtained in Experiment 2, in which dysphoric (n = 40) and nondepressed subjects (n = 40) performed both self-referent and orthographic judgements of affectively valenced words, followed by either a free- or forced-recall test. The findings suggest that positive and negative trait words were adequately encoded in memory, but, consistent with cognitive theories of depression, their accessibility to retrieval was differentially limited. In addition, however, the results implicate an important contribution of diminished motivation and/or conservative report criterion in the manifestation of depression-related biases and deficits in recall.

Confirmation of identity by a metallic knee prosthesis in a severely burnt body.

Confirmation of identification of the deceased may be obtained in several ways. In this case, identification was confirmed by matching the serial numbers on a prosthetic knee joint and co-existing anatomical abnormalities with the joint serial number and X-ray reports in the hospital notes of the suspected victim.

Ipodate treatment of hyperthyroidism in cats.

OBJECTIVE: To evaluate the efficacy and safety of ipodate in the treatment of hyperthyroidism in cats. DESIGN: Prospective case series. ANIMALS: 12 cats with hyperthyroidism treated at The Animal Medical Center between November 1994 and March 1996. PROCEDURE: Each cat initially received 100 mg of ipodate/d, PO. The drug's effects on clinical signs, body weight, heart rate, and serum triiodothyronine (T3) and thyroxine concentrations were evaluated 2, 4, 6, 10, and 14 weeks after initiation of treatment. A CBC and serum biochemical analyses were performed at each evaluation to monitor potential adverse effects of the drug. Dosage of ipodate was increased to 150 mg/d and then to 200 mg/d at 2-week intervals if a good clinical response was not observed. RESULTS: 8 cats responded to treatment and 4 did not. Among cats that responded, mean body weight increased and mean heart rate and serum T3 concentration decreased during the study period. Among cats that did not respond, mean body weight decreased and mean heart rate and serum T3 concentration were not significantly changed. Serum thyroxine concentration remained high in all cats. Adverse clinical signs or hematologic abnormalities attributable to ipodate treatment were not reported in any of the cats. CLINICAL IMPLICATIONS: Ipodate may be a feasible alternative to methimazole for medical treatment of hyperthyroidism in cats, particularly those that cannot tolerate methimazole and are not candidates for surgery or radiotherapy. Cats with severe hyperthyroidism are less likely to respond to ipodate than are cats with mild or moderate disease, and cats in which serum T3 concentration does not return to the reference range are unlikely to have an adequate improvement in clinical signs.

Anterior chamber depth measurement in clinical practice.

This study compares the validity of anterior chamber depth estimates provided by a slit-lamp technique with those obtained from pachometry and ultrasonography. The technique provides depth estimates which are valid to within +/- 0.33 mm relative to standard pachometry, and to within +/- 0.42 mm relative to ultrasonography. The technique is simple to perform, requires no extra slit-lamp attachments, and may be carried out on any slit-lamp which has a calibrated variable length of slit.

A longitudinal study of changes in body composition and basal metabolism in physically active elderly men.

Ageing in the elderly is usually characterised by loss of fat-free mass (FFM) and reduction in basal metabolic rate (BMR). These age-related changes probably vary in rate, timing and extent between subjects in response to differences in leisure or occupational physical activity, disease and several other factors. Information on rates of change in BMR and FFM is limited by study design (most published work is cross-sectional rather than longitudinal), and possibly by methodology (use of imprecise and/or biased methods for assessment of changes in body composition). In the present study BMR and body composition were first measured in 22 physically active elderly men (mean age 62) in good health and measurements were repeated 6.5 years later. Changes in BMR, FFM and percentage body fat were small and not statistically significant over the time period (paired t, P > 0.05). The study suggests that physically active elderly men in good health in this age range can show very small age-related declines in BMR and FFM.

Reproducibility of cardiorespiratory measurements during submaximal and maximal running in children.

With the increased use of oxygen consumption measurements in clinical and sporting studies, measurement variability has become more important to both the paediatric clinician and the sports scientist. In this study we assessed the reproducibility of cardiorespiratory measurements during submaximal and maximal running in children. Ten healthy, physically active boys (mean(s.d.) age 10.7(0.71) years) performed two submaximal and two maximal running tests within a 4 week period. The submaximal protocol consisted of three 6 minute runs at 7.2, 8.0 and 8.8 km/h. Every attempt was made to minimize the sources of non-biological variability at each testing session. During submaximal exercise, oxygen consumption (VO2), heart rate (HR) and fractional utilization appeared to be the most reliable measures accounting for over two-thirds of the total variation (coefficients of reliability (CR) of 68%, 94% and 82% respectively). Ventilation (Ve) and respiratory exchange ratio (RER) proved to be only moderately reliable accounting for less than half of the total variation (CR 50% and 45% respectively). At maximal exercise, VO2, Ve and time to exhaustion were most reliable, accounting for approximately two-thirds of the total variation (CR 65%, 63% and 63% respectively). Within this test environment, a two visit submaximal assessment was capable of estimating VO2 with a standard error of +/- 1.25 ml/kg/min. Similarly, for maximal testing a two visit assessment estimated peak VO2 with a standard error of +/- 2.28 ml/kg/min. On the evidence of these results a two visit assessment for submaximal and maximal exercise testing seems adequate to estimate the stability of submaximal cardiorespiratory measures and peak VO2 in healthy, normally active boys.

Prediction of body density from skinfold thickness in elderly subjects: are the existing equations valid?

In 33 healthy elderly subjects, body density was measured using hydrodensitometry and predicted from the sum of four skinfold thicknesses. In the men, mean measured density was 1.045 kg/l and mean predicted density 1.041 kg/l, with bias and limits of agreement 0.004 (+/- 0.016) kg/l and 95% confidence interval (CI) for the difference 0.001-0.008 kg/l. In the women, mean measured density was 1.012 kg/l and mean predicted density 1.020 kg/l, and 95% CI for the difference was -0.016-0.000 kg/l. The bias was therefore in the opposite direction in women than men, and limits of agreement were wider (-0.008 +/- 0.030 kg/l) in the women. Biases in prediction of density lead to systematic error in estimation of body fatness in elderly subjects, and this systematic error is more likely to be of practical significance in women than men.

Measuring the body composition of elderly subjects: a comparison of methods.

There is a paucity of data on differences between methods for the assessment of body composition in elderly subjects. Studies on younger adults suggest that such differences are of some practical significance at the individual level. In the present study the following methods of estimating percentage body fatness (BF%) were compared in healthy elderly men and women (mean age 70 (SD 6) years: densitometry; skinfold thickness; total body water; bioelectrical impedance (BIA) using an age-specific predictive equation and the manufacturers' equation; body mass index (BMI). Though BF% estimates from the various methods tended to be highly correlated with those from densitometry and with each other, differences between methods at the individual level were marked. In particular, the age-specific equations based on BMI and BIA systematically overestimated BF% relative to the other methods. Biases between BF% estimates derived from densitometry, skinfolds, BIA (manufacturers' equation) and total body water were less marked, indicating little evidence of systematic differences between these methods in elderly subjects. Individual differences between methods were slightly greater than those reported in some studies of younger adults, but this may be of little practical significance, and may be considered inevitable in view of variability between and within subjects in the extent to which the underlying assumptions of these two-component methods are met in elderly subjects.