Number of colony-forming unit-Hill colonies among children and teenagers with obesity, dyslipidemia and breastfeeding history.

BACKGROUND AND AIMS: The number of colony-forming unit (CFU)-Hill colonies has been proposed as a biomarker of vascular function and cardiovascular risk in adults but information about its role in children is scarce. This study evaluates the associations between obesity, cardiovascular risk factors and breastfeeding history with the numbers of CFU-Hill colonies in a sample of young people. METHODS AND RESULTS: We selected 49 children and teenagers between ages 10 and 17 (65.3% boys) from Mexican Health Care system. Physical activity and Anthropometric measures data were registered. CFU-Hill colonies were cultured from mononuclear cells obtained from venous blood. We detected inverse associations between the formation of CFU-Hill colonies and body mass index (BMI; ß = -1.53; 95% confidence interval [CI], -1.92, -1.13), triglycerides (ß = -0.26; 95%CI = -0.34, -0.18), total cholesterol (ß = -0.13; 95%CI = -0.17, -0.08), Low Density Lipoprotein (LDL) (ß = -0.20; 95%CI = -0.31, -0.09) and glucose (ß = -0.37; 95%CI = -0.55, -0.18) using multivariate models. Breastfeeding duration showed a 1.46-colony increase for each month of breastfeeding (95%CI = 0.73, 2.18). CONCLUSIONS: CFU-Hill colony-forming capacity in children and teenagers was inversely associated with obesity, dyslipidemia and high blood levels of glucose. In contrast a longer breastfeeding duration was directly associated with an increased number of CFU-Hill colonies. However these results must be confirmed with further studies. Our findings support the importance of promoting breastfeeding and monitoring nutritional and metabolic status at an early age to prevent chronic disease development.

Nestin-expressing cells in the gut give rise to enteric neurons and glial cells.

BACKGROUND: Neuronal stem cells (NSCs) are promising for neurointestinal disease therapy. Although NSCs have been isolated from intestinal musclularis, their presence in mucosa has not been well described. Mucosa-derived NSCs are accessible endoscopically and could be used autologously. Brain-derived Nestin-positive NSCs are important in endogenous repair and plasticity. The aim was to isolate and characterize mucosa-derived NSCs, determine their relationship to Nestin-expressing cells and to demonstrate their capacity to produce neuroglial networks in vitro and in vivo. METHODS: Neurospheres were generated from periventricular brain, colonic muscularis (Musc), and mucosa-submucosa (MSM) of mice expressing green fluorescent protein (GFP) controlled by the Nestin promoter (Nestin-GFP). Neuronal stem cells were also grown as adherent colonies from intestinal mucosal organoids. Their differentiation potential was assessed using immunohistochemistry using glial and neuronal markers. Brain and gut-derived neurospheres were transplanted into explants of chick embryonic aneural hindgut to determine their fate. KEY RESULTS: Musc- and MSM-derived neurospheres expressed Nestin and gave rise to cells of neuronal, glial, and mesenchymal lineage. Although Nestin expression in tissue was mostly limited to glia co-labelled with glial fibrillary acid protein (GFAP), neurosphere-derived neurons and glia both expressed Nestin in vitro, suggesting that Nestin+/GFAP+ glial cells may give rise to new neurons. Moreover, following transplantation into aneural colon, brain- and gut-derived NSCs were able to differentiate into neurons. CONCLUSIONS & INFERENCES: Nestin-expressing intestinal NSCs cells give rise to neurospheres, differentiate into neuronal, glial, and mesenchymal lineages in vitro, generate neurons in vivo and can be isolated from mucosa. Further studies are needed for exploring their potential for treating neuropathies.

Phosphorylated cyclic-AMP-response element-binding protein and thyroid hormone receptor have independent response elements in the rat thyrotropin-releasing hormone promoter: an analysis in hypothalamic cells.

BACKGROUND: Thyrotropin-releasing hormone (TRH) from the hypothalamic paraventricular nucleus (PVN) controls the activity of the hypothalamus-pituitary-thyroid axis. TRH is expressed in other hypothalamic nuclei but is downregulated by 3,3',5-L-triiodothyronine (T(3)) exclusively in the PVN. Thyroid hormone receptors (TRs) bind TRH promoter at Site-4 (-59/-52), also proposed to bind phosphorylated cAMP response element-binding protein (pCREB). However, nuclear extracts from 8Br-cAMP-stimulated hypothalamic cells showed no binding to Site-4 and instead to cAMP response element (CRE)-2 (-101/-94). METHODS: We characterized, by DNA footprinting and chromatin immunoprecipitation, the sites in the rat (-242/+34) TRH promoter that bind to nuclear factors of hypothalamic primary cultures incubated with 8Br-cAMP and/or T(3). RESULTS: In primary cultures of fetal hypothalamic cells, TRH mRNA levels rapidly diminished with 10 nM T(3) while they increased by 1 mM 8Br-cAMP (+/- T(3)). Site-4 was protected from DNase I digestion with nuclear extracts from T(3)-incubated cells but not from controls or from those incubated with 8Br-cAMP, which protected CRE-2; T(3) + 8Br-cAMP coincubation caused no interference. The region protected by nuclear extracts from cAMP-stimulated cells included sequences adjacent to CRE-2-containing response elements of the SP/Krüppel family. A TRbeta2 antibody immunoprecipitated chromatin containing Site-4 but not CRE-2, from cells incubated with T(3). A pCREB antibody immunoprecipitated CRE-2 containing chromatin in controls and more in 8Br-cAMP-stimulated cells but none when cells were incubated only with T(3). Recruitment of the 2 transcription factors was preserved in cells simultaneously exposed to 8Br-cAMP and T(3). DISCUSSION: These results show that pCREB binds to a response element in the TRH promoter (CRE-2) that is independent of Site-4 where TRbeta2 is bound; pCREB and TR do not present mutual interference on their binding sites.

Fatty acids and neurodevelopment.

Knowledge of the importance of docosahexaenoic acid (DHA), arachidonic acid (AA), and long-chain polyunsaturated fatty acids (LCPUFAs) in neurodevelopment was originally obtained from animal studies. These fatty acids are rapidly accreted in brain during the first postnatal year in animal and human infants, and they are found in high concentrations in breast milk. Reports of enhanced intellectual development in breast-fed children, and reports linking LCPUFA deficiency with neurodevelopmental disorders have stressed the physiological importance of DHA in visual and neural systems. In addition to high concentrations of fatty acids in breast milk, they are also present in fish and algae oil and have recently been added to infant formulas. Esterified poplyunsaturated fatty acids act in cellular membranes, in signal transduction, in neurotransmission, and in the formation of lipid rafts. Nonesterified polyunsaturated fatty acids can modulate gene expression and ion channel activities, thus becoming neuroprotective agents. The conversion of linoleic acid and alpha-linolenic acid into ARA and DHA have led to randomized clinical trials that have studied whether infant formulas supplemented with DHA or both DHA and ARA would enhance visual and cognitive development. This review gives an overview of fatty acids and neurodevelopment, focusing on the findings from these studies.

Dexamethasone represses cAMP rapid upregulation of TRH gene transcription: identification of a composite glucocorticoid response element and a cAMP response element in TRH promoter.

Hypothalamic proTRH mRNA levels are rapidly increased (at 1 h) in vivo by cold exposure or suckling, and in vitro by 8Br-cAMP or glucocorticoids. The aim of this work was to study whether these effects occurred at the transcriptional level. Hypothalamic cells transfected with rat TRH promoter (-776/+85) linked to the luciferase reporter showed increased transcription by protein kinase (PK) A and PKC activators, or by dexamethasone (dex), but co-incubation with dex and 8Br-cAMP decreased their stimulatory effect (as observed for proTRH mRNA levels). These effects were also observed in NIH-3T3-transfected cells supporting a characteristic of TRH promoter and not of hypothalamic cells. Transcriptional regulation by 8Br-cAMP was mimicked by noradrenaline which increased proTRH mRNA levels, but not in the presence of dex. PKA inhibition by H89 avoided 8Br-cAMP or noradrenaline stimulation. TRH promoter sequences, cAMP response element (CRE)-like (-101/-94 and -59/-52) and glucocorticoid response element (GRE) half-site (-210/-205), were analyzed by electrophoretic mobility shift assays with nuclear extracts from hypothalamic or neuroblastoma cultures. PKA stimulation increased binding to CRE (-101/-94) but not to CRE (-59/-52); dex or 12-O-tetradecanoylphorbol-13-acetate (TPA) increased binding to GRE, a composite site flanked by a perfect and an imperfect activator protein (AP-1) site in the complementary strand. Interference was observed in the binding of CRE or GRE with nuclear extracts from cells co-incubated for 3 h with 8Br-cAMP and dex; from cells incubated for 1 h, only the binding to GRE showed interference. Rapid cross-talk of glucocorticoids with PKA signaling pathways regulating TRH transcription constitutes another example of neuroendocrine integration.

Dexamethasone rapidly regulates TRH mRNA levels in hypothalamic cell cultures: interaction with the cAMP pathway.

The biosynthesis of thyrotropin-releasing hormone (TRH) in the hypothalamic paraventricular nucleus (PVN) is subject to neural and hormonal regulations. To identify some of the potential effectors of this modulation, we incubated hypothalamic dispersed cells with dexamethasone for short periods of time (1-3 h) and studied the interaction of this hormone with protein kinase C (PKC) and PKA signaling pathways. TRH mRNA relative changes were determined by the RT-PCR technique. One hour incubation with 10(-10)-10(-4) M dexamethasone produced a concentration-dependent biphasic effect: an inhibition was observed on TRH mRNA levels at 10(-10) M, an increase above control at 10(-8)-10(-6) M and a reduction at higher concentrations (10(-5)- 10(-4) M). The stimulatory effect of 10(-8) M dexamethasone on TRH mRNA was essentially independent of new protein synthesis, as evidenced by cycloheximide pretreatment. Changes in TRH mRNA levels were reflected by enhanced TRH cell content. Incubation with a cAMP analogue (8-bromo-cAMP, 8Br-cAMP) or with a PKC activator (12-O-tetradecanoylphorbol-13-acetate, TPA) increased TRH mRNA levels after 1 and 2 h, respectively. An increase in TRH mRNA expression was observed by in situ hybridization of dexamethasone or 8Br-cAMP-treated cells. The interaction of dexamethasone, PKA and PKC signaling pathways was studied by combined treatment. The stimulatory effect of 10(-7) M TPA on TRH mRNA levels was additive to that of dexamethasone; in contrast, coincubation with 10(-3) M 8-Br-cAMP and dexamethasone diminished the stimulatory effect of both drugs. An inhibition was observed when the cAMP analogue was coincubated with TPA or TPA and dexamethasone. These results demonstrate that dexamethasone can rapidly regulate TRH biosynthesis and suggest a cross talk between cAMP, glucocorticoid receptors and PKC transducing pathways.