Targeting GATA1 and p2x7r Locus Binding in Spinal Astrocytes Suppresses Chronic Visceral Pain by Promoting DNA Demethylation / 神经科学通报·英文版

Irritable bowel syndrome is a gastrointestinal disorder of unknown etiology characterized by widespread, chronic abdominal pain associated with altered bowel movements. Increasing amounts of evidence indicate that injury and inflammation during the neonatal period have long-term effects on tissue structure and function in the adult that may predispose to gastrointestinal diseases. In this study we aimed to investigate how the epigenetic regulation of DNA demethylation of the p2x7r locus guided by the transcription factor GATA binding protein 1 (GATA1) in spinal astrocytes affects chronic visceral pain in adult rats with neonatal colonic inflammation (NCI). The spinal GATA1 targeting to DNA demethylation of p2x7r locus in these rats was assessed by assessing GATA1 function with luciferase assay, chromatin immunoprecipitation, patch clamp, and interference in vitro and in vivo. In addition, a decoy oligodeoxynucleotide was designed and applied to determine the influence of GATA1 on the DNA methylation of a p2x7r CpG island. We showed that NCI caused the induction of GATA1, Ten-eleven translocation 3 (TET3), and purinergic receptors (P2X7Rs) in astrocytes of the spinal dorsal horn, and demonstrated that inhibiting these molecules markedly increased the pain threshold, inhibited the activation of astrocytes, and decreased the spinal sEPSC frequency. NCI also markedly demethylated the p2x7r locus in a manner dependent on the enhancement of both a GATA1-TET3 physical interaction and GATA1 binding at the p2x7r promoter. Importantly, we showed that demethylation of the p2x7r locus (and the attendant increase in P2X7R expression) was reversed upon knockdown of GATA1 or TET3 expression, and demonstrated that a decoy oligodeoxynucleotide that selectively blocked the GATA1 binding site increased the methylation of a CpG island in the p2x7r promoter. These results demonstrate that chronic visceral pain is mediated synergistically by GATA1 and TET3 via a DNA-demethylation mechanism that controls p2x7r transcription in spinal dorsal horn astrocytes, and provide a potential therapeutic strategy by targeting GATA1 and p2x7r locus binding.

Effect of different cult ure conditions on differentiation of Treg and Th17 in PB-MC / 中国免疫学杂志

Objectiv e:To investigate the effect of different culture conditions on the differentiation of Treg and Th17 to lay a foundation for exploring the methods to reverse the immune tolerance induced by tumor microenvironment.Methods:The IL-6 gene was cloned and stablely transferred into the tumor cell line expressing TGF-β.The conditioned mediums ( CM) were prepared by collecting the culture supernatants of tumor cell lines with or without IL-6 expression and used in the in vitro culture of peripheral blood mononuclear cells ( PBMC ) .The changes of Treg and Th17 in PBMC treated with different CM were detected with flow cytometry ( FCM) .Results:The expression of TGF-βin BEL-7402 was higher than that in HepG2.Thus the BEL-7402 was selected for preparation of cell line stablely transfected with IL -6 gene.ELISA detection confirmed the effective expression of IL -6 by the identified cell lines.It was showed that the Treg increased in PBMC treated with culture supernatants of tumor cells .However,the presence of IL-6 reversed the increase of Treg and promoted the differentiation of Th 17.Conclusion: The culture supernatants of tumor cells increases the proportion of Treg.However,the presence of IL-6 in this CM can reverse the increase of Treg and raise the proportion of Th 17.

Effect of a novel dietary supplement on pH levels of healthy volunteers: a pilot study / 中西医结合学报

<p><b>OBJECTIVE</b>To examine the effects of a greens alkalizing dietary supplement on urinary pH levels in individuals with lower-than-average pH levels.</p><p><b>METHODS</b>The present study investigated the effects of an alkalizing formula (Reserveage Wholeganic Greens(TM)) on four individuals who had average urinary pH levels below 6.0 for three consecutive days. Following the three-day, baseline period, participants received Reserveage Wholeganic Greens(TM) for four consecutive days and were instructed to continue to measure their urine pH levels. Paired samples t-tests were used to examine pH levels before and after a four-day treatment period with Reserveage Wholeganic Greens(TM).</p><p><b>RESULTS</b>Compared to baseline, mean urine pH levels in all volunteers were significantly higher following the supplementation with Reserveage Wholeganic Greens(TM) (5.89 ± 0.20 vs 5.56 ± 0.23; P<0.01). Participants' pH levels were also significantly higher than baseline on days 5, 6, and 7 of the treatment period (P < 0.05). Noteworthy, on day 7, participants' mean pH levels were significantly higher than at the beginning of the treatment period (6.03 ± 0.15 at day 7 vs 5.65 ± 0.24 at day 4; P < 0.01).</p><p><b>CONCLUSION</b>The findings of this study suggest that supplementation with Reserveage Wholeganic Greens(TM) has an alkalizing effect on the body and can increase the urine pH levels in individuals with lower-than-average pH levels.</p>

The accumulation and maturation status of pulmonary conventional dendritic cells in the early phase of acute lung injury / 中华急诊医学杂志

Objective To investigate the accumulation and maturation status of pulmonary conventional dendritic cells (cDCs) in the early phase of acute lung injury (ALI),and to explore the way of the inflammatory responses and lung injury modulated by cDCs in vivo.MethodsMale C57BL/6 mice were randomly ( random number) divided into the normal control group,6 h-ALI group and 24 h-ALI group.Murine model of ALI was made by intra-tracheal administration of lipopolysaccharide (LPS) and lung specimens were taken 6 h or 24 h later.The accumulation and maturation status of pulmonary cDCs were assessed by flow cytometry.IL-6 and TNF-α were quantified to evaluate the lung inflammation.Transcription factors T-bet/GATA-3 mRNA ratio was determined to estimate the balance between Th1/Th2 responses.IFN-γand IL-4 were quantified to evaluate Thl-specific and Th2-specific cytokine production respectively.Lung injury was estimated by lung wet weight/body weight ratio (LWW/BW) and histopathological assessment.Comparison between groups was performed using one -way ANOVA.ResultsCompared with normal control group,LPS challenge resulted in higher level of IL-6 and TNF-α,increased LWW/BW ratio and significant histopathological changes (P ＜0.01 ).The accumulation and maturation of pulmonary cDCs in 6 h-ALI group were significantly increased after LPS challenge (P ＜0.01 ),while the accumulation and maturation of pulmonary cDCs in 24 h-ALI group were significantly lower than that in 6 h-ALI group ( P ＜0.01 ).Compared with normal control group,the expression of T-bet mRNA in 24 h-ALI group was markedly enhanced ( P ＜ 0.01 ) and the production of IFN-γ was increased as well ( P ＜ 0.01 ).ConclusionsThe accumulation and maturation of pulmonary cDCs peaked within 24 h after LPS challenge,pulmonary cDCs may initiate and amplify acute lung inflammation of ALI by enhancing the Th1 immune response and ensuing cytokine production.

The effect of bifico on rat experimental hepatic encephalopathy induced by thioaetamide / 中国药理学通报

AIM　The effect of bifico on experimental hepatic encephalopathy (HE) induced by thioacetamide(TAA) and the possible mechanism of its protective effect were studied. METHODS　Experimental hepatic encephalopathy was induced by ig thioacetamide 250 mg*kg-1 successively for two days; bifico was administraed ig once per day for one week before HE induction. RESULTS　Compared with control HE rat, Bifico improved rat neuro-reflexes score and hepatic injury grade(P<0.05); thus decreased rat serum ammonium and LPS concentrations, respectively (P<0.05,P<0.01). The preparation slightly increased the ratio of chain amino acid to aromatic amino acid as well as reduced the degree of liver necrosis. CONCLUSION　At the dosage of 840 mg*kg-1 ig for one week before liver intoxication, Bifico significantly protects rats from hepatic encephalopathy induced by TAA.