Ting Yang

Correlation between Nutrition and Symptoms: Nutritional Survey of Children with Autism Spectrum Disorder in Chongqing, China.

Restricted diets and inadequate nutrient intake of children with autism spectrum disorder (ASD) have been reported. This study examined the nutritional statuses of children with ASD and the relationships between their behaviors and nutritional intake. A total of 154 children with ASD (age = 5.21 ± 1.83 years) and 73 typically-developing (TD) children (age = 4.83 ± 0.84 years) from Chongqing, China, were enrolled. The severity of ASD was evaluated using the Childhood Autism Rating Scale (CARS). The serum ferritin, folate, vitamin B12, 25(OH) vitamin D, and vitamin A concentrations in the children with ASD were determined. All participants underwent anthropometric examinations, dietary assessments, and questionnaire assessments about their feeding behaviors, and gastrointestinal symptoms. The ZHA, ZWA, and ZBMIA were found to be significantly lower in the children with ASD compared with those without ASD. In addition, the percentages of children exhibiting severe picky eating and severe resistance to new foods, as well as those with a reported general impression of severe eating problems and constipation, were higher among the children with ASD. These children consumed significantly fewer macronutrients compared with the children without ASD. In addition, the children with ASD had the highest rate of vitamin A deficiency, followed by iron deficiency. After adjusting for sex, the vitamin A concentration was found to be negatively correlated with the CARS score (rs = −0.222, p = 0.021). No correlation between the ferritin, folate, vitamin D, or vitamin B12 concentration and the CARS score was found. These results suggest that reduced macronutrient intakes, severe feeding behavior issues, constipation, and vitamin A deficiency are quite common among children with ASD. Further, a low serum vitamin A level may be a risk factor for symptoms of ASD. However, the underlying mechanism should be further studied.

Retinoic Acid Facilitates Toll-Like Receptor 4 Expression to Improve Intestinal Barrier Function through Retinoic Acid Receptor Beta.

Background/Aims: Vitamin A (VA) protects the intestinal epithelial barrier by improving cell migration and proliferation. Our previous studies demonstrated that VA deficiency (VAD) during pregnancy suppresses the systemic and mucosal immune responses in the intestines of offspring and that VA supplementation (VAS) during early life can increase immune cell counts. However, little is known about the mechanisms by which VA regulates intestinal epithelial barrier function. Methods: Caco-2 cells were treated with all-trans retinoic acid (ATRA) for 24 hours to determine the optimum ATRA concentration to which the cells in question respond. Caco-2 cells were infected with recombinant adenoviruses carrying retinoic acid receptor beta (Ad-RARβ) and small interfering RARβ(siRARβ) to assess the effects of RARβ signalling on the expression of specific proteins. A siTLR4 lentivirus was used to knockdown Toll-like receptor 4 (TLR4) in Caco-2 cells to determine its role in the protective effects of VA on the intestinal epithelial barrier, and experiments involving TLR4-knock-out mice were performed to verify the effect of TLR4. VA normal (VAN), VAD and VAS rat models were established to confirm that changes in RARβ, TLR4 and ZO-2 expression levels that occurred following decreases or increases in retinol concentrations in vivo, and the permeability of the Caco-2 cell monolayer, as well as that of the epithelial barrier of the rat intestine was detected by measuring transepithelial resistance (TER) or performing enzyme-linked immunosorbent assay (ELISA). Retinoic acid receptor (RAR), toll like receptor (TLR) and tight junction (TJ) mRNA and protein expression levels in Caco-2 cells and the colon monolayers in the rat and mouse models were measured by PCR and western blotting, respectively. Co-immunoprecipitation (co-IP) and immunofluorescence staining were performed to assess the interactions among RARβ, TLR4 and zonula occluden-2 (ZO-2) in Caco-2 cells, and chromatin immunoprecipitation (ChIP) assay was performed to assess the binding between RARβ and the TLR4 promoter sequence in Caco-2 cells. Results: In the present study, ATRA treatment not only increased the TER of the Caco-2 monolayer but also up-regulated the expression levels of RARβ, TLR4 and ZO-2 in Caco-2 cells. The expression levels of these three proteins were significantly decreased in the colonic epithelial monolayers of VAD rats compared with those of VAN rats and were significantly increased following VAS in the corresponding group compared with the control group. Furthermore, the above changes in TLR4 and ZO-2 expression levels were augmented or attenuated by Ad-RARβ or siRARβ infection, respectively, in Caco-2 cells. Interestingly, siTLR4 down-regulated ZO-2 expression but did not affect RARβ expression in Caco-2 cells, and in VAD mice the lack of TLR4 did not affect ZO-2 expression. We noted direct interactions between RARβ and TLR4, TLR4 and ZO-2 in Caco-2 cells, and ChIP assay showed that RARβ could bind to the TLR4 promoter but not the ZO-2 promoter in Caco-2 cells. Conclusion: Taken together, our results indicate that RARβ enhanced ZO-2 expression by regulating TLR4 to improve intestinal epithelial barrier function in Caco-2 cells, as well as in rat and mouse models, but not in humans.

Effect of vitamin A supplementation on gut microbiota in children with autism spectrum disorders - a pilot study

BackgroundDysbiosis of gut microbiota are commonly reported in autism spectrum disorder (ASD) and may contribute to behavioral impairment. Vitamin A (VA) plays a role in regulation of gut microbiota. This study was performed to investigate the role of VA in the changes of gut microbiota and changes of autism functions in children with ASD.ResultsSixty four, aged 1 to 8xa0years old children with ASD completed a 6-month follow-up study with VA intervention. High-performance liquid chromatography was used to assess plasma retinol levels. The Autism Behaviour Checklist (ABC), Childhood Autism Rating Scale (CARS) and Social Responsiveness Scale (SRS) were used to assess autism symptoms. CD38 and acid-related orphan receptor alpha (RORA) mRNA levels were used to assess autism-related biochemical indicators’ changes. Evaluations of plasma retinol, ABC, CARS, SRS, CD38 and RORA mRNA levels were performed before and after 6xa0months of intervention in the 64 children. Illumina MiSeq for 16S rRNA genes was used to compare the differences in gut microbiota before and after 6xa0months of treatment in the subset 20 of the 64 children. After 6xa0months of intervention, plasma retinol, CD38 and RORA mRNA levels significantly increased (all Pxa0<xa00.05); the scores of ABC, CARS and SRS scales showed no significant differences (all Pxa0>xa00.05) in the 64 children. Meanwhile, the proportion of Bacteroidetes/Bacteroidales significantly increased and the proportion of Bifidobacterium significantly decreased in the subgroup of 20 (all false discovery rate (FDR) qxa0<xa00.05).ConclusionsBacteroidetes/Bacteroidales were the key taxa related to VA. Moreover, VA played a role in the changes in autism biomarkers. It remains unclear whether the VA concentration is associated with autism symptoms.Trial registrationThe study protocol was peer reviewed and approved by the institutional review board of Children’s Hospital, Chongqing Medical University in 2013 and retrospectively registered in Chinese Clinical Trial Registry (ChiCTR) on November 6, 2014 (TRN: ChiCTR-ROC-14005442).

TLR2/NFκB signalling regulates endogenous IL-6 release from marrow-derived mesenchymal stromal cells to suppress the apoptosis of PC12 cells injured by oxygen and glucose deprivation.

Two previous studies published by our group identified that mesenchymal stromal cells (MSCs) conferred neuroprotection in a rat model of hypoxic-ischaemic brain damage (HIBD), and that MSCs secreted abundant interleukin-6 (IL‑6) when co‑cultured with oxygen and glucose deprivation (OGD)‑injured PC12xa0cells. The present study has further investigated the role of IL‑6, and explored potential signalling pathways inxa0vitro. Inxa0vitro models were established by co‑culturing OGD‑injured PC12xa0cells with MSCs. Subsequently, the expression levels of the signalling molecules, Toll‑like receptorxa02 (TLR2)/nuclear factorxa0κB (NFκB), and IL‑6 were altered separately in this inxa0vitro model by treatment with an agonist, antagonist, siRNA or overexpression adenovirus. The expression levels of Bxa0cell lymphoma‑associated X (Bax), TLR2, NFκB and IL‑6 were detected by western blot analysis, real‑time polymerase chain reaction or ELISA. The resting membrane potential (RMP) of the PC12xa0cells was analysed by whole‑cell patch‑clamp recordings. Compared with controls or the PC12 co‑culture group, the MSC co‑cultured group induced less expression of Bax, but more IL‑6 secretion. Up- or down-regulation of the TLR2/NFκB signalling pathway resulted in a corresponding increase or decrease in the IL‑6 expression level in the MSCs. Co‑culture with siIL‑6‑MSCs increased the expression levels of Bax and increased the RMP in the OGD PC12xa0cells. In conclusion, the release of IL‑6 from MSCs was regulated via the TLR2/NFκB signalling pathway. Endogenous IL‑6 reduced apoptosis and protected OGD‑injured PC12xa0cells when they were co‑cultured with MSCs. The present study has reported a novel immunomodulatory effect of the microenvironment of neural damage during MSC cytotherapy.

Vitamin A improves the symptoms of autism spectrum disorders and decreases 5-hydroxytryptamine (5-HT): A pilot study

Autism spectrum disorders (ASD) are complicated neurodevelopmental disorders. Many studies have demonstrated that children with autism have multiple nutritional deficiencies and increased serum 5-hydroxytryptamine (5-HT) levels. In our previous study, 77.9% of autistic children were found to have vitamin A deficiency, and the concentration of vitamin A was negatively associated with the CARS score. In the present study, we sought to test whether vitamin A supplementation could improve autistic symptoms and decrease serum 5-HT levels. The DSM-V criteria and CARS score were used for symptom description and symptom assessment of the patients, respectively, before and after vitamin A supplementation (VAS). Serum retinol and 5-HT levels, mRNA levels of RAR α, β, and γ and TpH 1 expression were detected in autistic children before and after VAS and in normal children. Serum retinol levels in children with ASD were significantly lower than in control children. Serum 5-HT levels in children with ASD were higher than in control children, which were correlated with symptom severity of children with autism. After VA supplementation, the children with ASD exhibited significant improvement in autism symptoms. Serum retinol concentrations of children with ASD were significantly increased, and serum 5-HT levels were decreased. Moreover, statistically significant changes were observed in mRNA expression levels of RAR α, RAR γ and TpH 1 after VAS compared to baseline. This study suggested that VA supplementation may improve symptoms and reduce 5-HT levels in children with ASD, indicating that VA supplementation is a reasonable therapy at least for a subset of children with autism.

Interleukin-10 Release from Astrocytes Suppresses Neuronal Apoptosis via the TLR2/NFκB Pathway in a Neonatal Rat Model of Hypoxic-Ischemic Brain Damage

The biological function of interleukin‐10 (IL‐10) and the relationship between IL‐10 secretion and the Toll‐like receptor 2 (TLR2) expression levels in the central nervous system following hypoxic‐ischemic brain damage (HIBD) are poorly understood. Here, we intend to elucidate the biological function and mechanism of IL‐10 secretion following HIBD. In this study, we used a neonatal rat model of HIBD and found that rats injected with adeno‐associated virus‐IL‐10‐shRNA (short hairpin RNA) exhibited partially impaired learning and memory function compared to rats administered adeno‐associated virus‐control‐shRNA. In vitro oxygen‐glucose deprivation (OGD) induced IL‐10 release from astrocytes but not from neurons. Pretreatment with exogenous recombinant IL‐10 alleviated OGD‐mediated apoptosis of neurons but not astrocytes. In addition, we also observed that hypoxic injury induced a marked increase in IL‐10 expression in astrocytes as a result of activation of the TLR2/phosphorylated nuclear factor kappa B (p‐NFκB) p65 signaling cascade; furthermore, this effect disappeared upon small interfering RNA targeting rat TLR2 gene (siTLR2) treatment. Pyrrolidinedithiocarbamate, an inhibitor of NFκB activation, reduced the IL‐10 expression levels in both OGD‐injured astrocytes in vitro and the hippocampi of HIBD rats in vivo but did not significantly affect TLR2 expression. Furthermore, a luciferase assay revealed that p‐NFκB p65 could bind the −1700/−1000 bp proximal region of the IL‐10 gene promoter to regulate IL‐10 secretion from astrocytes and that this interaction could be controlled by OGD treatment. These data suggest that HIBD induces IL‐10 secretion from astrocytes to exert a paracrine‐induced anti‐apoptotic effect on injured neurons via the TLR2/NFκB signaling pathway, which may improve learning and memory dysfunction after ischemic injury.

Marginal Vitamin A Deficiency Exacerbates Memory Deficits Following Aβ1-42 Injection in Rats

Background Although clinical vitamin A deficiency (VAD), which is a public health problem developing throughout the world, has been well controlled, marginal vitamin A deficiency (MVAD) is far more prevalent, especially among pregnant women and preschool children in China. Increasing evidence suggests that VAD is involved in the pathogenesis of Alzheimer’s disease (AD). However, whether MVAD, beginning early in life, increases the risk of developing AD has yet to be determined. Objective The goal of this study was to investigate the long-term effects of MVAD on the pathogenesis of AD in rats. Method An MVAD model was generated from maternal MVAD rats and maintained with an MVAD diet after weaning. The males were bilaterally injected with aggregated amyloid β (Aβ)1–42 into the CA3 area of the hippocampus, and the AD-associated cognitive and neuropathological phenotypes were examined. Results We found that MVAD feeding significantly aggravated Aβ1-42-induced learning and memory deficits in the Morris water maze test. MVAD did not induce the mRNA expression of retinoic acid receptors (RARs), a disintegrin and metalloprotease 10 (ADAM10) or insulin-degrading enzyme (IDE) in Aβ1-42-injected rats. Moreover, RARα and RARγ mRNA were positively correlated with ADAM10 mRNA, whereas RARβ mRNA was positively correlated with IDE mRNA. Conclusion Our study suggests that MVAD beginning from the embryonic period perturbs the AD-associated genes, resulting in an enhanced risk of developing AD.

Vitamin A supplementation improves the intestinal mucosal barrier and facilitates the expression of tight junction proteins in rats with diarrhea

OBJECTIVESnThe aim of this study is to investigate the specific effects of vitamin A (VA) on diarrhea in rats and its potential targets to protect the intestinal mucosa.nnnMETHODSnSpecific pathogen-free Sprague Dawley rats were fed a VA deficient (VAD) or VA normal (VAN) diet for 4 wk. Then, half of the VAN rats were treated with a VAN diet and the other half with a lactose VAN diet. VAD rats were randomly assigned to one of four groups and fed a VAD diet, lactose VAD diet, VAN diet with VA supplementation (VAS) via daily intragastric administration, or a lactose VAN diet with daily VAS. Rat weight and degree of diarrhea were evaluated daily. After 15 d, the serum retinol level was measured by high-performance liquid chromatography, and the serum diamine oxidase (DAO) and zonulin concentrations were analyzed by enzyme-linked immunosorbent assays. The small intestine mucosal pathology was observed by hematoxylin and eosin staining. Western blotting was performed to detect the protein expression levels of occludin and claudin-1 in the intestinal mucosa, and the zonula-occludens 1 expression was assessed using immunohistochemistry.nnnRESULTSnVAD limited weight gain in rats and increased the degree of diarrhea. The serum retinol levels and the level of tight junction (TJ) proteins claudin-1 and occludin and grip strength were affected by the interaction between lactose-induced diarrhea and the VA diet. Diarrhea, independent of VAD, significantly decreased rat weight, increased serum DAO levels, damaged small intestine villi, and impaired zonula-occludens 1 protein expression. VAD significantly increased the concentration of zonulin independently of diarrhea, but VAS increased the serum retinol level, reduced the severity of diarrhea, increased the expression levels of the TJ proteins, facilitated the restoration of the small intestine villi that were damaged by the diarrhea, and decreased the concentrations of serum DAO and zonulin.nnnCONCLUSIONSnVAD may aggravate the degree of diarrhea and intestinal mucosal damage during the duration of diarrhea, and VAS helps relieve diarrhea and improves intestinal damage likely by regulating the expression of TJ proteins. Therefore, VA plays a pivotal role in the protection of the intestinal mucosa during instances of diarrhea.

Mesenchymal stem cells-derived IL-6 activates AMPK/mTOR signaling to inhibit the proliferation of reactive astrocytes induced by hypoxic-ischemic brain damage

&NA; Mesenchymal stem cells (MSCs) treatment is an effective strategy for the functional repair of central nervous system (CNS) insults through the production of bioactive molecules. We have previously demonstrated that the interleukin‐6 (IL‐6) secreted by MSCs plays an anti‐apoptotic role in injured astrocytes and partly promotes functional recovery in neonatal rats with hypoxic‐ischemic brain damage (HIBD). However, the mechanisms of IL‐6 underlying the proliferation of injured astrocytes have not been fully elucidated. In this study, we investigated the therapeutic effects of MSCs on astrocyte proliferation in neonatal rats subjected to HIBD. A HIBD model was established in Sprague Dawley (SD) rats, and MSCs were administered by intracerebroventricular injection 5 days after HIBD. Rat primary astrocytes were cultured, subjected to oxygen glucose deprivation (OGD) injury and then immediately co‐cultured with MSCs in vitro. Immunofluorescence staining, Cell Counting Kit (CCK)‐8, flow cytometry, Ca2+ imaging, enzyme‐linked immunosorbent assay (ELISA), western blotting, and co‐immunoprecipitation (Co‐IP) were performed. We found that MSCs transplantation not only promoted the recovery of learning and memory function in HIBD rats but also significantly reduced the number of Ki67+/glial fibrillary acidic protein (GFAP)+ cells in the hippocampi 7–14 days after HIBD. In addition to increasing IL‐6 expression in both the hippocampi of HIBD rats and astrocyte culture medium, MSCs treatment in vitro significantly increased the expression levels of glycoprotein (gp) 130 and phosphorylated AMP‐activated protein kinase &agr; (p‐AMPK&agr;) and decreased the expression levels of p‐mammalian target of rapamycin (mTOR) and its downstream targets. Furthermore, MSCs treatment induced a protein‐protein interaction between gp130 and p‐AMPK&agr;. Suppression of IL‐6 expression in MSCs reversed the above regulatory functions of MSCs in hippocampal astrocytes. The utilization of rapamycin further confirmed that mTOR participated in the proliferation of reactive astrocytes. These findings suggest that endogenous IL‐6 produced by MSCs in the HIBD microenvironment provides therapeutic advantages by activating AMPK/mTOR signaling, thus reducing the proliferation of reactive astrocytes.

Effect of educational interventions on health in childhood: a meta-analysis of randomized controlled trials

OBJECTIVEnThe purpose of this study was to summarize the findings of randomized controlled trials (RCTs) investigating any potential effects of educational interventions on health in childhood.nnnSTUDY DESIGNnMeta-analysis.nnnMETHODSnPubMed, Embase, and the Cochrane Library databases were searched to identify all RCTs that fit our analysis through May 2016. Weighted mean difference (WMD) was used to measure the effect of educational interventions in childhood by using a random effects model.nnnRESULTSnThirty RCTs reporting data on 35,296 children were included in the meta-analysis. The summary WMD indicated that children who received educational interventions had lower levels of body mass index (BMI) (WMD: -0.15; 95% CI: -0.24 to -0.05; Pxa0=xa00.003), BMI z-score (WMD: -0.03; 95% CI: -0.05 to -0.02; Pxa0<xa00.001), waist circumference (WMD: -0.97; 95% CI: -1.95 to -0.00; Pxa0=xa00.050), triceps skinfold (WMD: -1.39; 95% CI: -2.41 to -0.37; Pxa0=xa00.008), systolic blood pressure (WMD: -1.13; 95% CI: -2.20 to -0.07; Pxa0=xa00.037), total cholesterol (WMD: -4.04; 95% CI: -7.18 to -0.90; Pxa0=xa00.012), and triglyceride (WMD: -2.62; 95% CI: -4.33 to -0.90; Pxa0=xa00.003). However, educational interventions were found to have little or no significant impact on the waist-to-hip ratio, diastolic blood pressure, high-density lipoprotein, and low-density lipoprotein.nnnCONCLUSIONSnThe study findings prove the positive effects of educational interventions on BMI, BMI z-score, waist circumference, triceps skinfold, systolic blood pressure, total cholesterol, and triglyceride.