Evaluation of R‐ (−) and S‐ (+) Clenbuterol enantiomers during a doping cycle or continuous ingestion of contaminated meat using chiral liquid chromatography by LC‐TQ‐MS

Clenbuterol is known to improve competition resistance and muscular growth in athletes. Although it is an illegal drug, its use by farmers is widely spread to induce growth of their cattle. Thus, when clenbuterol is found in the urine of an athlete, there is doubt whether it was consumed with doping purposes or if it is due to the consumption of meat from a clenbuterol‐fed animal. Previous studies suggest that enantiomeric relationship of clenbuterol may be different according to the intake source. However, the enantiomeric relationship throughout a doping cycle or a continuous intake of contaminated meat has not yet been explored. In this first approximation, our aim was the development and validation of a sensitive and rapid method for the determination of S‐ (+) and R‐ (─) clenbuterol enantiomers to be used in a controlled study in rats fed for one week with contaminated meat or simulating a doping cycle. Enantiomers were measured using liquid chromatography coupled to mass spectrometry with a triple quadrupole analyzer (LC‐TQ‐MS) and were separated on an AGP Chiralpak column. The method was fully validated following the VICH (Veterinary International Conference on Harmonization guidelines) and was linear in the range of 12.5–800 pg/mL with a correlation coefficient of ≥0.98 for each enantiomer, and with a limit of quantitation and detection (LOQ and LOD) of 12.5 pg/mL and 6.5 pg/mL, respectively, for both enantiomers. The application of this method pointed out the shift of the enantiomeric relationship in urine from rats during the first five days of the doping cycle compared to those fed with contaminated meat. This finding can be of substantial importance in further doping studies.     

An Automated Telephone Nutrition Support System for Spanish-Speaking Patients With Diabetes

In the United States, Spanish-speaking patients with diabetes often receive inadequate dietary counseling. Providing language and culture-concordant dietary counseling on an ongoing basis is critical to diabetes self-care. To determine if automated telephone nutrition support (ATNS) counseling could help patients improve glycemic control by duplicating a successful pilot in Mexico in a Spanish-speaking population in Oakland, California. A prospective randomized open-label trial with blinded endpoint assessment (PROBE) was performed. The participants were seventy-five adult patients with diabetes receiving care at a federally qualified health center in Oakland, California. ATNS, a computerized system that dialed patients on their phones, prompted them in Spanish to enter (via keypad) portions consumed in the prior 24 hours of various cultural-specific dietary items, and then provided dietary feedback based on proportion of high versus low glycemic index foods consumed. The control group received the same ATNS phone calls 14 weeks after enrollment. The primary outcome was hemoglobin A1c % (A1c) 12 weeks following enrollment. Participants had no significant improvement in A1c (–0.3% in the control arm, –0.1% in the intervention arm, P = .41 for any difference) or any secondary parameters. In our study, an ATNS system did not improve diabetes control in a Spanish-speaking population in Oakland.     

Dynamics of axonal regeneration in adult and aging zebrafish reveal the promoting effect of a first lesion

Axonal regeneration is a major issue in the maintenance of adult nervous systems, both after nerve injuries and in neurodegenerative diseases. However, studying this process in vivo is difficult or even impossible in most vertebrates. Here we show that the posterior lateral line (PLL) of zebrafish is a suitable system to study axonal regeneration in vivo because of both the superficial location and reproducible spatial arrangement of neurons and targets, and the possibility of following reinnervation in live fish on a daily basis. Axonal regeneration after nerve cut has been demonstrated in this system during the first few days of life, leading to complete regeneration within 24 h. However, the potential for PLL nerve regeneration has not been tested yet beyond the early larval stage. We explore the regeneration potential and dynamics of the PLL nerve in adult zebrafish and report that regeneration occurs throughout adulthood. We observed that irregularities in the original branching pattern are faithfully reproduced after regeneration, suggesting that regenerating axons follow the path laid down by the original nerve branches. We quantified the extent of target reinnervation after a nerve cut and found that the latency before the nerve regenerates increases with age. This latency is reduced after a second nerve cut at all ages, suggesting that a regeneration-promoting factor induced by the first cut facilitates regeneration on a second cut. We provide evidence that this factor remains present at the site of the first lesion for several days and is intrinsic to the neurons.The potential of adult neurons to regenerate their axons and to reinnervate target organs after injury is not as well understood as early axonogenesis. Whether and how this capability is modified on aging is an even less explored area. In all vertebrates studied so far, neurons of the peripheral nervous system retain the ability to reextend peripheral axons and reestablish functional connections. This ability is thought to involve intrinsic mechanisms of repair as well as extrinsic signals from the local environment, mostly coming from Schwann cells and macrophages (1).The posterior lateral line (PLL) of fish is a convenient yet unexplored sensory system to address the issue of axonal regeneration throughout adulthood. The PLL comprises a set of superficial mechanosensory organs called neuromasts, which are distributed over the body and tail. Neuromasts are composed of a core of mechanosensory hair cells providing information about the local water flow, surrounded by accessory cells. The afferent neurons innervating neuromasts are clustered in a ganglion posterior to the otic vesicle, and their peripheral axons extend toward the tail, right under the skin. This sensory system is involved in a large repertoire of behaviors (2).The juvenile PLL of zebrafish comprises four lines of neuromasts that extend at different dorsoventral levels, totaling about 50 organs (3). This pattern is established around 1 mo postfertilization (mpf) and remains essentially unchanged throughout adulthood, except that each juvenile neuromast gives rise to a number of “accessory” neuromasts through a budding process (4, 5) that depends on innervation (6). Bud-neuromasts remain closely associated and form dorsoventrally arranged linear clusters, or “stitches” (7). Here we concentrate on the most extensive of the four lines, the ventral one. This line lies originally along the horizontal myoseptum of the embryo and comprises only five neuromasts. More neuromasts develop during larval life, such that the line eventually consists of one neuromast on every intersomitic border, or about 30 altogether (8). The line migrates ventrally to reach its final position at the juvenile stage, and an axonal branch follows each neuromast during this migration.Axonal regeneration after nerve cut has been demonstrated in the PLL during the first few days of life, leading to complete reinnervation of neuromasts within 24 h (9, 10). Regenerating axons follow either the Schwann cells that ensheathe the nerve (10) or the interneuromast cells that extend between consecutive neuromasts (11). PLL nerve regeneration has not yet been addressed beyond the early larval stage.Here we examine whether the PLL nerve is able to regenerate in adult and aging zebrafish. Our data clearly demonstrate the effectiveness and fidelity of regeneration at all ages studied, from 1 to 15 mpf, although the onset of reinnervation is increasingly delayed with age, thereby linking neuronal aging with a progressive decline in neuronal reactivity to axonal damage. Whenever a second cut is made after complete regeneration, the latency of reinnervation is reduced at all ages, provided the second cut is immediately distal to the first one. We show that although Schwann cells act as guidance cues to help the axons regrow along their original path, they are not involved in this regeneration-promoting effect. We conclude that the promoting effect of a first lesion is mostly caused by an intrinsic, local change occurring in the injured axons. Altogether, our results reveal that the zebrafish PLL is a convenient system to study axonal regeneration in vivo compared with mammalian systems, in which nerve wiring is more complex and less traceable.     

Do bulk‐fill resins decrease the restorative time in posterior teeth? A systematic review and meta‐analysis of in vitro studies

The aim of the present study was to compare the restorative time for filling cavities in posterior teeth using bulk‐fill and conventional resin composites through a systematic review and meta‐analysis. A search for clinical trials and laboratory studies was performed in PubMed/MEDLINE, Scopus, the Latin American and Caribbean Health Sciences Literature database, the Brazilian Library in Dentistry, Cochrane Library, Clinical Trials, and ReBEC (Registro Brasileiro de Ensaios Clínicos) databases without publication year or language restriction. Two reviewers identified eligible studies according to the inclusion criteria: bulk‐fill compared to conventional resin in class I or II, and the restorative time as an outcome. A meta‐analysis of the restorative time mean difference between composites was performed (inverse variance method, random effects model; Z‐test, P ≤ .05). From the 662 eligible studies, 133 were selected for full‐text analysis; three were included in the systematic review and in the meta‐analysis. Overall, the restorative time was lower when bulk‐fill was used (P = .0007, Z = 3.37), as the subgroup full‐body bulk‐fill (P < .00001, Z = 21.00). There was no difference in restorative time between flowable bulk‐fill and conventional resins (P = .08, Z = 1.76). Moderate‐to‐substantial heterogeneity was detected. Full‐body bulk‐fill composites decrease the restorative time in posterior teeth compared to conventional resins. Full‐body bulk‐fill resins require a shorter restorative time to perform restorations in posterior teeth than conventional resins, but the same is not valid for flowable bulk‐fill resin composites.     

Duke Activity Status Index for Cardiovascular Diseases: Validation of the Portuguese Translation

Background

The Duke Activity Status Index (DASI) assesses the functional capacity of patients with cardiovascular disease (CVD), but there is no Portuguese version validated for CVD.

Objectives

To translate and adapt cross-culturally the DASI for the Portuguese-Brazil language, and to verify its psychometric properties in the assessment of functional capacity of patients with CVD.

Methods

The DASI was translated into Portuguese, then checked by back-translation into English and evaluated by an expert committee. The pre-test version was first evaluated in 30 subjects. The psychometric properties and correlation with exercise testing was performed in a second group of 67 subjects. An exploratory factor analyses was performed in all 97 subjects to verify the construct validity of the DASI.

Results

The intraclass correlation coefficient for test-retest reliability was 0.87 and for the inter-rater reliability was 0.84. Cronbach''s α for internal consistency was 0.93. The concurrent validity was verified by significant positive correlations of DASI scores with the VO₂max (r = 0.51, p < 0.001). The factor analysis yielded two factors, which explained 54% of the total variance, with factor 1 accounting for 40% of the variance. Application of the DASI required between one and three and a half minutes per patient.

Conclusions

The Brazilian version of the DASI appears to be a valid, reliable, fast and easy to administer tool to assess functional capacity among patients with CVD.     

Alterations of LXRα and LXRβ expression in the hypothalamus of glucose-intolerant rats

Liver X receptor (LXR) α and β are nuclear receptors that are crucial for the regulation of carbohydrate and lipid metabolism. Activation of LXRs in the brain facilitates cholesterol clearance and improves cognitive deficits, thus they are considered as promising drug targets to treat diseases such as atherosclerosis and Alzheimer's disease. Nevertheless, little is known about the function and localization of LXRs in the brain. Here, we studied the expression of LXR in the brains of rats that received free access to 10% (w/v) fructose group (FG) in their beverages or water control drinks (control group (CG)). After 6 weeks rats in the FG presented with hypertriglyceridemia, hyperinsulinemia, and became glucose intolerant, suggesting a progression toward type 2 diabetes. We found that hypothalamic LXR expression was altered in fructose-fed rats. Rats in the FG presented with a decrease in LXRβ levels while showing an increase in LXRα expression in the hypothalamus but not in the hippocampus, cerebellum, or neocortex. Moreover, both LXRα and β expression correlated negatively with insulin and triglyceride levels. Interestingly, LXRβ showed a negative correlation with the area under the curve during the glucose tolerance test in the CG and a positive correlation in the FG. Immunocytochemistry revealed that the paraventricular and ventromedial nuclei express mainly LXRα whereas the arcuate nucleus expresses LXRβ. Both LXR immunosignals were found in the median preoptic area. This is the first study showing a relationship between glucose and lipid homeostasis and the expression of LXRs in the hypothalamus, suggesting that LXRs may trigger neurochemical and neurophysiological responses for the control of food intake and energy expenditure through these receptors.     

A Mediterranean-style, low-glycemic-load diet decreases atherogenic lipoproteins and reduces lipoprotein (a) and oxidized low-density lipoprotein in women with metabolic syndrome

The objective was to assess the impact of a Mediterranean-style, low-glycemic-load diet (control group, n = 41) and the same diet plus a medical food (MF) containing phytosterols, soy protein, and extracts from hops and Acacia (MF group, n = 42) on lipoprotein atherogenicity in women with metabolic syndrome. Plasma lipids, apolipoproteins (apos), lipoprotein subfractions and particle size, low-density lipoprotein (LDL) oxidation, and lipoprotein (a) were measured at baseline, week 8, and week 12 of the intervention. Three-day dietary records were collected at the same time points to assess compliance. Compared with baseline, women decreased energy intake from carbohydrate (P < .001) and fat (P < .001), whereas they increased energy intake from protein (P < .001). A significant increase in energy from monounsaturated fatty acids was also observed as well as increases in eicosapentaenoic acid and docosahexaenoic acid, whereas trans-fatty acid intake was reduced (P < .00001). The atherogenic lipoproteins, large very low-density lipoprotein (P < .0001) and small LDL (P < .0001), were reduced, whereas the ratio of large high-density lipoprotein to smaller high-density lipoprotein particles was increased (P < .0001). Apolipoprotein B was reduced for all women (P < .0001), with a greater reduction in the MF group (P < .025). Oxidized LDL (P < .05) and lipoprotein (a) (P < .001) were reduced in both groups at the end of the intervention. Consumption of a Mediterranean-style diet reduces the risk for cardiovascular disease by decreasing atherogenic lipoproteins, oxidized LDL, and apo B. Inclusion of an MF may have an additional effect in reducing apo B.