[Performance evaluation of a precision dosing service for vancomycin in a tertiary level pediatric hospital]. / Una evaluación de desempeño del servicio de dosificación de precisión para vancomicina en un hospital pediátrico de nivel terciario.

Background: Plasma level-based therapeutic drug monitoring of vancomycin is recommended in the treatment of complex pediatric infections in order to increase the probability of achieving safe and effective pharmacotherapy. Objective: To retrospectively evaluate the activities and performance of pharmacotherapeutic optimization based on vancomycin levels at a tertiary pediatric hospital between 2007 and 2020. Métodos: Vancomycin levels of pediatric patients were analyzed, assessing care quality indicators and analytical verifications, as well as aspects related to teaching and research. The predictive performance of vancomycin levels was evaluated after adjustment of the therapeutic regimen using a population pharmacokinetic optimization program (BestDose v1.126) considering the coefficient of determination (R2), the mean absolute percentage error (MAPE), and the root mean square error (RMSE). Results: 13269 vancomycin level determinations were analyzed; 70% were trough levels and 81% belonged to patients in the intensive care units. Forty percent of the trough levels were within the therapeutic range when adjusted without software. Three hundred seventy-four pharmacotherapeutic interventions, of which 97% were accepted by the treating physician; 75% of the post-adjustment trough levels were within the therapeutic range, compared to 40% when the approach was empirical, a difference that was statistically significant (p=0.03). The values associated with predictive performance (n subgroup of patients = 91) were: R2=0.61, MAPE=28.16%, and RMSE=3.3, which all showed to be adequate. Conclusion: The performance of therapeutic vancomycin monitoring and related pharmacokinetic clinical activities showed to be good.

Introducción: La dosificación de precisión a través del monitoreo de vancomicina basado en sus concentraciones plasmáticas (vancocinemia) es una práctica recomendada en el tratamiento de infecciones pediátricas de alta complejidad para aumentar la probabilidad de lograr una farmacoterapia segura y eficaz. Objetivo: Evaluar retrospectivamente las actividades y el desempeño relacionado a la optimización farmacoterapéutica basada en las vancocinemias (período 2007-2020) de un hospital pediátrico terciario. Métodos: Se analizaron las vancocinemias de pacientes pediátricos, estimándose indicadores de calidad asistencial y verificaciones analíticas, así como también aspectos relacionados a docencia e investigación. Se evaluó el desempeño predictivo de las concentraciones de vancomicina cuando se ajustaron los regímenes terapéuticos con un programa de optimización farmacocinética (BestDose v1.126) considerando el coeficiente de determinación (R2), el error porcentual absoluto medio (MAPE) y la raíz del error cuadrático medio (RMSE). Resultados: Se analizaron 13269 vancocinemias. El 70% fueron valles y el 81% pertenecieron a pacientes de Unidades de Cuidados Intensivos. El 40% de los valles se encontró dentro del margen terapéutico al ajustarse sin programa informático. Se realizaron 347 intervenciones farmacoterapéuticas, el 97% de las cuales fueron aceptadas por el médico tratante; el 75% de los valles posteriores al ajuste entraron en el margen terapéutico, valor significativamente mayor respecto al 40% de cuando el abordaje fue empírico (p=0.03). Los valores asociados al desempeño predictivo, (n subgrupo de pacientes = 91) fueron: R2=0.61, MAPE=28.16% y RMSE=3.3, mostrándose todos adecuados. Conclusión: Las actividades de monitoreo y farmacocinética clínica de vancomicina mostraron un buen rendimiento clínico. Resultados: Se analizaron 13269 vancocinemias. El 70% fueron valles y el 81% pertenecieron a pacientes de Unidades de Cuidados Intensivos. El 40% de los valles se encontró dentro del margen terapéutico al ajustarse sin programa informático. Se realizaron 347 intervenciones farmacoterapéuticas, el 97% de las cuales fueron aceptadas por el médico tratante; el 75% de los valles posteriores al ajuste entraron en el margen terapéutico, valor significativamente mayor respecto al 40% de cuando el abordaje fue empírico (p=0.03). Los valores asociados al desempeño predictivo, (n subgrupo de pacientes = 91) fueron: R2=0.61, MAPE=28.16% y RMSE=3.3, mostrándose todos adecuados. Conclusión: Las actividades de monitoreo y farmacocinética clínica de vancomicina mostraron un buen rendimiento clínico.

Worsening of memory deficit induced by energy-dense diet in a rat model of early-Alzheimer's disease is associated to neurotoxic Aß species and independent of neuroinflammation.

Diet is a modifiable risk factor for Alzheimer's disease (AD), but the mechanisms linking alterations in peripheral metabolism and cognition remain unclear. Since it is especially difficult to study long-term effects of high-energy diet in individuals at risk for AD, we addressed this question by using the McGill-R-Thy1-APP transgenic rat model (Tg(+/-)) that mimics presymptomatic AD. Wild-type and Tg(+/-) rats were exposed during 6months to a standard diet or a Western diet (WD), high in saturated fat and sugar. Results from peripheral and hippocampal biochemical analysis and in situ respirometry showed that WD induced a metabolic syndrome and decreased presynaptic bioenergetic parameters without alterations in hippocampal insulin signaling or lipid composition. Cognitive tests, ELISA multiplex, Western blot, immunohistochemistry and RT-qPCR indicated that WD worsened cognition in Tg(+/-) rats, increased hippocampal levels of monomeric Aß isoforms and oligomeric species, promoted deposits of N-Terminal pyroglutamate-Aß (AßN3(pE)) in CA1 pyramidal neurons and interneurons, decreased transcript levels of genes involved in neuroprotective pathways such as Sirtuin-1 and increased nitrated proteins. Our results support the concept that in the presence of early Aß pathology, diet-induced metabolic dysfunctions may contribute as a "second hit" to impair cognition. Noteworthy, such effect is not mediated by higher microglia activation or disruption of blood brain barrier. However, it may be attributed to increased amyloidogenic processing of amyloid precursor protein, generation of AßN3(pE) and dysregulation of pathways governed by Sirtuin-1. This evidence reinforces the implementation of prophylactic interventions in individuals at risk for AD.

White Adipose Tissue and Circadian Rhythm Dysfunctions in Obesity: Pathogenesis and Available Therapies.

A combined neuroendocrine, metabolic, and chronobiological view can help to better understand the multiple and complex mechanisms involved in obesity development and maintenance, as well as to provide new effective approaches for its control and treatment. Indeed, we have currently updated data on the whole adipogenic process involved in white adipose tissue (WAT) mass expansion, namely due to a mechanism whereby WAT cells become hypertrophic, thus inducing a serious local (WAT) inflammatory condition that in turn, will impair not only the cross-talk between the hypothalamus and the WAT, but also favoring the development of deep and widespread neuroendocrine-metabolic dysfunction. Moreover, we also have revisited the circadian clock genes involved in dysfunctional WAT mass expansion and the mechanisms that may lead to obesity development, including early metabolic dysfunctions, enhanced oxidative stress and distorted energy homeostasis. The epigenetic changes of clock genes driving metabolic disease and obesity development have also been included in this review. Finally, we have also underlined the relevance of metabolic homeostasis regulation by central and peripheral organ clocks, sleep disturbances, nutrients, and feeding time, as key factors in obesity development as well as both, classical and chronotherapeutic approaches for its prevention and treatment.

Melatonin normalizes clinical and biochemical parameters of mild inflammation in diet-induced metabolic syndrome in rats.

The objective of this study was to evaluate the efficacy of melatonin to affect mild inflammation in the metabolic syndrome (MS) induced by a high-fat diet in rats. Adult Wistar male rats were divided into four groups (n = 16/group): (i) control diet (3% fat); (ii) high-fat (35%) diet; (iii) high-fat diet + melatonin; and (iv) melatonin. Rats had free access to high-fat or control chow and one of the following drinking solutions for 10 wk: (a) tap water; (b) 25 µg/mL of melatonin. Plasma interleukin (IL)-1ß, IL-4, IL-6, IL-10, tumor necrosis factor (TNF)-α, interferon (IFN)-γ, and C-reactive protein (CRP) were measured at two time intervals, that is, the middle of daylight period and the middle of the scotophase. In addition, a number of somatic and metabolic components employed clinically to monitor the MS were measured. Melatonin decreased the augmented circulating levels of IL-1ß, IL-6, TNF-α, IFN-γ, and CRP seen in obese rats and restored the depressed levels of IL-4 and IL-10. Rats fed with the high-fat diet showed significantly higher body weights and augmented systolic blood pressure from the third and fourth week onwards, respectively, melatonin effectively preventing these changes. In high-fat-fed rats, circulating low-density lipoprotein-cholesterol, total cholesterol, and triglyceride concentration augmented significantly, melatonin being effective to counteract these changes. Melatonin-treated rats showed a decreased insulin resistance, the highest values of plasma high-density lipoprotein-cholesterol, and the lowest values of plasma uric acid. The results indicate that melatonin is able to normalize the altered biochemical pro-inflammatory profile seen in rats fed with a high-fat diet.

Melatonin and mitochondrial dysfunction in the central nervous system.

Cell death and survival are critical events for neurodegeneration, mitochondria being increasingly seen as important determinants of both. Mitochondrial dysfunction is considered a major causative factor in Alzheimer's disease (AD), Parkinson's disease (PD) and Huntington's disease (HD). Increased free radical generation, enhanced mitochondrial inducible nitric oxide (NO) synthase activity and NO production, and disrupted electron transport system and mitochondrial permeability transition, have all been involved in impaired mitochondrial function. Melatonin, the major secretory product of the pineal gland, is an antioxidant and an effective protector of mitochondrial bioenergetic function. Both in vitro and in vivo, melatonin was effective to prevent oxidative stress/nitrosative stress-induced mitochondrial dysfunction seen in experimental models of AD, PD and HD. These effects are seen at doses 2-3 orders of magnitude higher than those required to affect sleep and circadian rhythms, both conspicuous targets of melatonin action. Melatonin is selectively taken up by mitochondria, a function not shared by other antioxidants. A limited number of clinical studies indicate that melatonin can improve sleep and circadian rhythm disruption in PD and AD patients. More recently, attention has been focused on the development of potent melatonin analogs with prolonged effects which were employed in clinical trials in sleep-disturbed or depressed patients in doses considerably higher than those employed for melatonin. In view that the relative potencies of the analogs are higher than that of the natural compound, clinical trials employing melatonin in the range of 50-100mg/day are needed to assess its therapeutic validity in neurodegenerative disorders.

Melatonin supplementation decreases prolactin synthesis and release in rat adenohypophysis: correlation with anterior pituitary redox state and circadian clock mechanisms.

In the laboratory rat, a number of physiological parameters display seasonal changes even under constant conditions of temperature, lighting, and food availability. Since there is evidence that prolactin (PRL) is, among the endocrine signals, a major mediator of seasonal adaptations, the authors aimed to examine whether melatonin administration in drinking water resembling in length the exposure to a winter photoperiod could affect accordingly the 24-h pattern of PRL synthesis and release and some of their anterior pituitary redox state and circadian clock modulatory mechanisms. Melatonin (3 µg/mL drinking water) or vehicle was given for 1 mo, and rats were euthanized at six time intervals during a 24-h cycle. High concentrations of melatonin (>2000 pg/mL) were detected in melatonin-treated rats from beginning of scotophase (at 21:00 h) to early photophase (at 09:00 h) as compared with a considerably narrower high-melatonin phase observed in controls. By cosinor analysis, melatonin-treated rats had significantly decreased MESOR (24-h time-series average) values of anterior pituitary PRL gene expression and circulating PRL, with acrophases (peak time) located in the middle of the scotophase, as in the control group. Melatonin treatment disrupted the 24-h pattern of anterior pituitary gene expression of nitric oxide synthase (NOS)-1 and -2, heme oxygenase-1 and -2, glutathione peroxidase, glutathione reductase, Cu/Zn- and Mn-superoxide dismutase, and catalase by shifting their acrophases to early/middle scotophase or amplifying the maxima. Only the inhibitory effect of melatonin on pituitary NOS-2 gene expression correlated temporally with inhibition of PRL production. Gene expression of metallothionein-1 and -3 showed maxima at early/middle photophase after melatonin treatment. The 24-h pattern of anterior pituitary lipid peroxidation did not vary after treatment. In vehicle-treated rats, Clock and Bmal1 expression peaked in the anterior pituitary at middle scotophase, whereas that of Per1 and Per2 and of Cry1 and Cry2 peaked at the middle and late photophase, respectively. Treatment with melatonin raised mean expression of anterior pituitary Per2, Cry1, and Cry2. In the case of Per1, decreased MESOR was observed, although the single significant difference found between the experimental groups when analyzed at individual time intervals was increase at early scotophase in the anterior pituitary of melatonin-treated rats. Melatonin significantly phase-delayed expression of Per1, Per2, and Cry1, also phase-delayed the plasma corticosterone circadian rhythm, and increased the amplitude of plasma corticosterone and thyrotropin rhythms. The results indicate that under prolonged duration of a daily melatonin signal, rat anterior pituitary PRL synthesis and release are depressed, together with significant changes in the redox and circadian mechanisms controlling them.

Disrupted chronobiology of sleep and cytoprotection in obesity: possible therapeutic value of melatonin.

From a physiological perspective the sleep-wake cycle can be envisioned as a sequence of three physiological states (wakefulness, non-rapid eye movement, NREM, sleep and REM sleep) which are defined by a particular neuroendocrine-immune profile regulating the metabolic balance, body weight and inflammatory responses. Sleep deprivation and circadian disruption in contemporary "24/7 Society" lead to the predominance of pro-orexic and proinflammatory mechanisms that contribute to a pandemic metabolic syndrome (MS) including obesity, diabetes and atherosclerotic disease. Thus, a successful management of MS may require a drug that besides antagonizing the trigger factors of MS could also correct a disturbed sleep-wake rhythm. This review deals with the analysis of the therapeutic validity of melatonin in MS. Melatonin is an effective chronobiotic agent changing the phase and amplitude of the sleep/wake rhythm and having cytoprotective and immunomodulatory properties useful to prevent a number of MS sequels. Several studies support that melatonin can prevent hyperadiposity in animal models of obesity. Melatonin at a low dose (2-5 mg/day) has been used for improving sleep in patients with insomnia and circadian rhythm sleep disorders. More recently, attention has been focused on the development of potent melatonin analogs with prolonged effects (ramelteon, agomelatine, tasimelteon, TK 301). In clinical trials these analogs were employed in doses considerably higher than those usually employed for melatonin. In view that the relative potencies of the analogs are higher than that of the natural compound, clinical trials employing melatonin doses in the range of 50-100 mg/day are needed to assess its therapeutic value in MS.

Expression of erythroblastic leukemia viral oncogene homolog (erbBS) mRNAs and possible splice variants in 3T3-L1 preadipocytes.

Previously, we studied the erythroblastic leukemia viral oncogene homolog (erbB) family of tyrosine kinase growth factor receptors in terms of protein expression, modulation and activation in the 3T3-L1 cell line. In the present study, the presence of full-length erbB mRNAs, and splice or proteolytic erbB variants, was evaluated using RT-PCR. Epidermal growth factor receptor (EGFR)/erbB1 expression was confirmed. Wild-type (wt) ErbB2, human erbB2 (HER2)-extracellular domain (ECD) and herstatin mRNA expression were analyzed. Restriction analysis confirmed wt expression. 3T3-L1 cells exhibited HER2-ECD and herstatin mRNA expression, although at extremely low levels (compared to the control cell lines). ErbB3 cDNA was amplified using mouse mammary tumors and rat acines as positive controls. ErbB4 was not positively identified in these cells. In conclusion, this study demonstrates that 3T3-L1 cells express EGFR/erbB1, erbB2 and erbB3, and possibly, certain erbB2 splice or proteolytic variants, which are worth pursuing.

Down-modulation of erbB2 activity is necessary but not enough in the differentiation of 3T3-L1 preadipocytes.

The high incidence of obesity-related pathologies, led to the study of the mechanisms involved in preadipose cell proliferation and differentiation. Here, we demonstrate that modulation of erbB2, plays a fundamental role during proliferation and adipogenic induction of preadipocytes. Using 3T3-L1 cells as model, we demonstrate that EGF (10 nM, 5 min) in addition to stimulate receptor tyrosine phosphorylation of both erbB2 and EGFR, is able to induce the heterodimer erbB2-EGFR. We treated proliferating 3T3-L1 cells with two inhibitors, AG 825 (IC(50) 0.35 microM, 54 times more selective for erbB2 than for EGFR, IC(50) 19 microM), and AG 879 (IC(50) of 1 microM for erbB2 versus 500 microM for EGFR). We found that both inhibited the proliferation on a dose-dependent basis, reaching a 30% maximal inhibition at 100 microM (P < 0.001) for AG825, and a 20% maximal inhibition at 10 microM (P < 0.001) for AG 879. These results involve erbB2 in 3T3-L1 proliferation. When studying the differentiation process, we found that the action of MIX-Dexa immediately activates MEK, JNK and p38 kinases. We observed that PD98059 and SP600125 (MEK-ERK and JNK inhibitors, respectively) added 1 h prior to the MIX-Dexa induction produced a decrease in erbB2 expression after 6 h, which is even greater than the one produced by the inducers, MIX-Dexa. This work supports erbB2 as a key factor in 3T3-L1 adipogenesis, acting mostly and not only during the proliferative phase but also during the differentiation through modulation of both its expression and activity.

ErbB2 and EGFR are downmodulated during the differentiation of 3T3-L1 preadipocytes.

The expression of receptors belonging to the epidermal growth factor receptor subfamily has been largely studied these last years in epithelial cells mainly as involved in cell proliferation and malignant progression. Although much work has focused on the role of these growth factor receptors in the differentiation of a variety of tissues, there is little information in regards to normal stromal cells. We investigated erbB2 expression in the murine fibroblast cell line Swiss 3T3L1, which naturally or hormonally induced undergoes adipocyte differentiation. We found that the Swiss 3T3-L1 fibroblasts express erbB2, in addition to EGFR, and in a quantity comparable to or even greater than the breast cancer cell line T47D. Proliferating cells increased erbB2 and EGFR levels when reaching confluence up to 4- and 10-fold, respectively. This expression showed a significant decrease when growth-arrested cells were stimulated to differentiate with dexamethasone and isobutyl-methylxanthine. Differentiated cells presented a decreased expression of both erbB2 and EGFR regardless of whether the cells were hormonally or spontaneously differentiated. EGF stimulation of serum-starved cells increased erbB2 tyrosine phosphorylation and retarded erbB2 migration in SDS-PAGE, suggesting receptor association and activation. Heregulin-alpha1 and -beta1, two EGF related factors, had no effect on erbB2 or EGFR phosphorylation. Although 3T3-L1 cells expressed heregulin, its specific receptors, erbB3 and erbB4, were not found. This is the first time in which erbB2 is reported to be expressed in an adipocytic cell line which does not depend on non EGF family growth factors (thyroid hormone, growth hormone, etc.) to accomplish adipose differentiation. Since erbB2 and EGFR expression were downmodulated as differentiation progressed it is conceivable that a mechanism of switching from a mitogenic to a differentiating signaling pathway may be involved, through regulation of the expression of these growth factor receptors.