Role of Prolactin Receptors in Lymphangioleiomyomatosis

Pulmonary lymphangioleiomyomatosis (LAM) is a rare lung disease caused by mutations in the tumor suppressor genes encoding Tuberous Sclerosis Complex (TSC) 1 and TSC2. The protein product of the TSC2 gene is a well-known suppressor of the mTOR pathway. Emerging evidence suggests that the pituitary hormone prolactin (Prl) has both endocrine and paracrine modes of action. Here, we have investigated components of the Prl system in models for LAM. In a TSC2 (+/-) mouse sarcoma cell line, down-regulation of TSC2 using siRNA resulted in increased levels of the Prl receptor. In human LAM cells, the Prl receptor is detectable by immunohistochemistry, and the expression of Prl in these cells stimulates STAT3 and Erk phosphorylation, as well as proliferation. A high affinity Prl receptor antagonist consisting of Prl with four amino acid substitutions reduced phosphorylation of STAT3 and Erk. Antagonist treatment further reduced the proliferative and invasive properties of LAM cells. In histological sections from LAM patients, Prl receptor immuno reactivity was observed. We conclude that the Prl receptor is expressed in LAM, and that loss of TSC2 increases Prl receptor levels. It is proposed that Prl exerts growth-stimulatory effects on LAM cells, and that antagonizing the Prl receptor can block such effects.     

Molecular mechanisms of oxidative stress-related neonatal jaundice

Oxidative stress is a pathological condition characterized by an overload of oxidant products, named free radicals, which are not well counteracted by antioxidant systems. Free radicals induce oxidative damage to many body organs and systems. In neonatal red blood cells, free-radical mediated-oxidative stress leads to eryptosis, a suicidal death process of erythrocytes consequent to alteration of cell integrity. Neonatal red blood cells are targets and at the same time generators of free radicals through the Fenton and Haber-Weiss reactions. Enhanced eryptosis in case of oxidative stress damage may cause anemia if the increased loss of erythrocytes is not enough compensated by enhanced new erythrocytes synthesis. The oxidative disruption of the red cells may cause unconjugated idiopathic hyperbilirubinemia in neonates. High levels of bilirubin are recognized to be dangerous for the central nervous system in newborns, however, many studies have highlighted the antioxidant function of bilirubin. Recently, it has been suggested that physiologic concentration of bilirubin correlates with higher antioxidant status while high pathological bilirubin levels are associated with pro-oxidants effects. The aim of this educational review is to provide an updated understanding of the molecular mechanisms underlying erythrocyte oxidant injury and its reversal in neonatal idiopathic hyperbilirubinemia.     

Experimental models of maternal–fetal interface and their potential use for nanotechnology applications

During pregnancy, the placenta regulates the transfer of oxygen, nutrients, and residual products between the maternal and fetal bloodstreams and is a key determinant of fetal exposure to xenobiotics from the mother. To study the disposition of substances through the placenta, various experimental models are used, especially the perfused placenta, placental villi explants, and cell lineage models. In this context, nanotechnology, an area of study that is on the rise, enables the creation of particles on nanometric scales capable of releasing drugs aimed at specific tissues. An important reason for furthering the studies on transplacental transfer is to explore the potential of nanoparticles (NPs), in new delivery strategies for drugs that are specifically aimed at the mother, the placenta, or the fetus and that involve less toxicity. Due to the fact that the placental barrier is essential for the interaction between the maternal and fetal organisms as well as the possibility of NPs being used in the treatment of various pathologies, the aim of this review is to present the main experimental models used in studying the maternal–fetal interaction and the action of NPs in the placental environment.     

Exercise training in childhood-onset systemic lupus erythematosus: a controlled randomized trial

Introduction

Exercise training has emerged as a promising therapeutic strategy to counteract physical dysfunction in adult systemic lupus erythematosus. However, no longitudinal studies have evaluated the effects of an exercise training program in childhood-onset systemic lupus erythematosus (C-SLE) patients. The objective was to evaluate the safety and the efficacy of a supervised aerobic training program in improving the cardiorespiratory capacity in C-SLE patients.

Methods

Nineteen physically inactive C-SLE patients were randomly assigned into two groups: trained (TR, n = 10, supervised moderate-intensity aerobic exercise program) and non-trained (NT, n = 9). Gender-, body mass index (BMI)- and age-matched healthy children were recruited as controls (C, n = 10) for baseline (PRE) measurements only. C-SLE patients were assessed at PRE and after 12 weeks of training (POST). Main measurements included exercise tolerance and cardiorespiratory measurements in response to a maximal exercise (that is, peak VO₂, chronotropic reserve (CR), and the heart rate recovery (ΔHRR) (that is, the difference between HR at peak exercise and at both the first (ΔHRR1) and second (ΔHRR2) minutes of recovery after exercise).

Results

The C-SLE NT patients did not present changes in any of the cardiorespiratory parameters at POST (P > 0.05). In contrast, the exercise training program was effective in promoting significant increases in time-to-exhaustion (P = 0.01; ES = 1.07), peak speed (P = 0.01; ES = 1.08), peak VO₂ (P = 0.04; ES = 0.86), CR (P = 0.06; ES = 0.83), and in ΔHRR1 and ΔHRR2 (P = 0.003; ES = 1.29 and P = 0.0008; ES = 1.36, respectively) in the C-SLE TR when compared with the NT group. Moreover, cardiorespiratory parameters were comparable between C-SLE TR patients and C subjects after the exercise training intervention, as evidenced by the ANOVA analysis (P > 0.05, TR vs. C). SLEDAI-2K scores remained stable throughout the study.

Conclusion

A 3-month aerobic exercise training was safe and capable of ameliorating the cardiorespiratory capacity and the autonomic function in C-SLE patients.

Trial registration

NCT01515163.     

Mercury Concentration in Breast Milk and Infant Exposure Assessment During the First 90 Days of Lactation in a Midwestern Region of Brazil

Breast milk samples collected from 18 nursing mothers between the 15th and 90th day of lactation were digested in nitric acid in a microwave, and total mercury (THg) levels were quantified by atomic fluorescence spectrometry. Participants responded to a 24-h dietary recall questionnaire on the 74th and 76th day of lactation and to a Food Frequency Questionnaire querying the frequency of fish intake over the last 90 days. Usual intake was estimated using the PC-SIDE software package. A meal of fish was offered on the 75th day of lactation. Mothers’ individual mean THg levels ranged from <0.76 to 22.7 ng/mL during the period, and the mean level for all samples (n?=?142) was 6.47?±6.04 ng/mL. The multilevel mixed linear model used showed high heterogeneity of the mercury levels among the mothers, and THg levels did not change significantly over the period under study. However, a significant increase in THg levels was observed after the intervention with the fish meal. Exposure increased for most infants on the 90th day of lactation, with intakes exceeding the THg provisional tolerable weekly intake (PTWI) at least once during the period for 77.8 % of samples. Mothers consumed mostly food from the fat and grain groups, and a significant correlation was detected between consumption of food of these groups and breast milk THg levels (p?=?0.006 and 0.007). A significant correlation was also found between vegetable consumption and carbohydrate intake and THg levels in the samples (p?=?0.015 and 0.045, respectively). No correlation was found between mothers’ daily fish consumption frequency and THg levels. Although this study showed that mercury intake by infants during lactation may exceed the toxicologically safe exposure level (PTWI), we nevertheless believe that the benefits of lactation for both the mother and the infant outweigh the eventual risks that this exposure may represent.     

Development of a 1-microl scale assay for mitogen-activated kinase kinase 7 using 2-D fluorescence intensity distribution analysis anisotropy

This paper describes the development of a robust, miniaturizable, and quantitative fluorescence-based assay for mitogen-activated protein kinase kinase 7 (MKK7). As a first step, the basic steady-state kinetics of the MKK7-catalyzed phosphorylation of c-Jun N-terminal kinases (JNKs) 1, 2, and 3 were defined using standard radiometric methods. Subsequently, the authors found that in addition to the holo JNKs, a series of novel small peptides (based on the region around the JNK phosphorylation site) are also substrates, provided that these were prephosphorylated on the Y residue of the TPY motif. One of these peptide substrates was used in the development of a fluorescence polarization-based assay using an antibody as a sensor. The assay was successfully miniaturized for use with conventional fluorescence polarization (FP) reader technology in 8.5 microl and on the single microl scale using Evotec proprietary 2-dimensional fluorescence intensity distribution analysis (2D-FIDA) anisotropy and liquid handling technology. The steady-state kinetic parameters derived using the FP or 2D-FIDA anisotropy format assays correlated well with those generated using a radiometric assay. Moreover, the quantitative sensitivity to known inhibitors was maintained independent of the format and assay volume. In addition, the authors found that the 2D-FIDA anisotropy assay exhibited superior performance statistics (typical Z' = approximately 0.5) relative to conventional FP (typical Z' = 0.3) and yielded the additional benefit of order-of-magnitude savings in terms of reagent costs. The 2D-FIDA anisotropy assay was used to carry out a successful high-throughput screening in 1-microl final volume against company file compounds.     

Actions of melatonin in the reduction of oxidative stress

Melatonin was discovered to be a direct free radical scavenger less than 10 years ago. Besides its ability to directly neutralize a number of free radicals and reactive oxygen and nitrogen species, it stimulates several antioxidative enzymes which increase its efficiency as an antioxidant. In terms of direct free radical scavenging, melatonin interacts with the highly toxic hydroxyl radical with a rate constant equivalent to that of other highly efficient hydroxyl radical scavengers. Additionally, melatonin reportedly neutralizes hydrogen peroxide, singlet oxygen, peroxynitrite anion, nitric oxide and hypochlorous acid. The following antioxidative enzymes are also stimulated by melatonin: superoxide dismutase, glutathione peroxidase and glutathione reductase. Melatonin has been widely used as a protective agent against a wide variety of processes and agents that damage tissues via free radical mechanisms.     

Optimisation of a quantitative polymerase chain reaction-based strategy for the detection and quantification of human herpesvirus 6 DNA in patients undergoing allogeneic haematopoietic stem cell transplantation

Human herpesvirus 6 (HHV-6) may cause severe complications after haematopoietic stem cell transplantation (HSCT). Monitoring this virus and providing precise, rapid and early diagnosis of related clinical diseases, constitute essential measures to improve outcomes. A prospective survey on the incidence and clinical features of HHV-6 infections after HSCT has not yet been conducted in Brazilian patients and the impact of this infection on HSCT outcome remains unclear. A rapid test based on real-time quantitative polymerase chain reaction (qPCR) has been optimised to screen and quantify clinical samples for HHV-6. The detection step was based on reaction with TaqMan^® hydrolysis probes. A set of previously described primers and probes have been tested to evaluate efficiency, sensitivity and reproducibility. The target efficiency range was 91.4% with linearity ranging from 10-10⁶ copies/reaction and a limit of detection of five copies/reaction or 250 copies/mL of plasma. The qPCR assay developed in the present study was simple, rapid and sensitive, allowing the detection of a wide range of HHV-6 loads. In conclusion, this test may be useful as a practical tool to help elucidate the clinical relevance of HHV-6 infection and reactivation in different scenarios and to determine the need for surveillance.