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.

Identification and phylogenetic analysis of five highly pathogenic avian influenza (H5N8) viruses isolated in Urumqi in 2016 / 中华微生物学和免疫学杂志

Objective:To analyze the genetic evolution and molecular characteristics of H5N8 avian influenza viruses (AIVs) isolated from the poultry in a live poultry market (LPM) in Urumqi, Xinjiang.Methods:Oropharyngeal and cloacal swabs of poultry were collected from a LPM in Urumqi in 2016. AIVs were isolated by inoculating swab samples into chicken embryos. Hemagglutination test and RT-PCR were used to identify the AIVs. The genes of isolated AIVs were amplified with the universal primers of AIV and whole-genome sequencing was also performed. Pairwise sequence alignment and analysis of phylogenetic and molecular characteristics were performed using BLAST, Clustal W, MEGA-X and DNAStar software.Results:Five H5N8 AIVs were isolated from poultry. These strains shared a nucleotide identity of 99.70%-100.00%, which indicated that they were from the same source, and were named XJ-H5N8/2016. Phylogenetic analysis based on hemagglutinin( HA), NS and PB2 genes showed that these isolates were clustered together with H5N8 AIVs isolated from the migratory swans in Hubei, Shanxi and Sanmenxia, and the ducks in India during 2016 to 2017. Moreover, they were also clustered together with H5N6 AIVs isolated from minks in China and the first case of human infection in Fujian. The phylogenetic tree of neuraminidase( NA) gene indicated the five isolates clustered together with H5N8 AIVs isolated from ducks in India in 2016, and the phylogenetic trees of PB1, MP, PA and NP genes showed that they were clustered together with H5N8 AIVs isolated from wild birds and poultry in Egypt, Cameroon, Uganda, Congo and other African countries in 2017. The HA cleavage sites of XJ-H5N8/2016 contained five consecutive basic amino acids, indicating high pathogenicity. Multiple mutations in the genes of XJ-H5N8/2016 could enhance its virulence and pathogenicity to mammals. Conclusions:The five strains of H5N8 AIVs isolated from the LPM were highly pathogenic and closely related to the H5N8 AIVs isolated from migratory birds and poultry in Hubei, Shanxi, Sanmenxia area, Africa and India during 2016 to 2017. Meanwhile, some of the viral genes were also closely related to the H5N6 AIVs isolated from the minks and human in China. Multiple mutations could increase the virulence and pathogenicity of AIVs to mammals, which could pose a potential threat to public health.

Distribution Characteristics of Traditional Chinese Medicine Syndromes for Abnormal Uterine Bleeding Associated with Ovulatory Dysfunction: A Clinical and Epidemiological Investigation / 中国实验方剂学杂志

Objective:To explore the distribution characteristics of traditional Chinese medicine （TCM） syndromes of abnormal uterine bleeding associated with ovulatory dysfunction （AUB-O）. Method:A clinical and epidemiological investigation was conducted in 6 688 patients with AUB-O from the provincial， municipal， and county/district hospitals in 29 provinces， autonomous regions， and municipalities across China to identify the distribution characteristics of their TCM syndromes. Result:The AUB-O patients nationwide were mainly differentiated into the kidney Qi deficiency syndrome （17.34%）， the spleen Qi deficiency syndrome （13.25%）， the Qi and blood deficiency syndrome （12.62%）， the Qi deficiency and blood stasis syndrome （8.45%）， and the kidney Yin deficiency syndrome （6.88%）. AUB-O resulted from Qi deficiency， Yin deficiency， and blood deficiency， often involving the kidney， spleen and liver. The analysis of the regional distribution of TCM syndromes in AUB-O patients revealed that kidney Qi deficiency， spleen Qi deficiency， and Qi and blood deficiency were the shared syndromes. However， due to regional discrepancy， the TCM syndrome varied widely from one geographic region to another. The kidney Qi deficiency syndrome was more frequently seen in North China， Northwest China， Southwest China， and East China， but less frequently in central China， Northeast China， and South China. The spleen Qi deficiency syndrome occurred most frequently in central China， while the East China had the highest frequency of Qi and blood deficiency syndrome. The spleen and kidney deficiency syndrome was mostly present in central China， North China， and Southwest China， the Qi deficiency （kidney deficiency） and blood stasis syndrome in Northwest China， South China， and North China， the kidney Yin deficiency syndrome in East China， Northwest China， and Northeast China， the deficiency-heat syndrome in Southwest China and East China， the kidney Yang deficiency syndrome in South China， the liver depression and blood heat syndrome in Northeast China， and the liver depression and spleen deficiency syndrome in central China. The diagnostic criteria for the kidney Qi deficiency， spleen Qi deficiency， Qi and blood deficiency， Qi deficiency and blood stasis， and kidney Yin deficiency syndromes were not significantly different from the previous ones. The distinctive symptoms for the kidney Qi deficiency syndrome were irregular vaginal bleeding， heavy menstrual flow， or shortened menstrual cycle， back soreness and pain， and forgetfulness， while those for the spleen Qi deficiency syndrome mainly included the shortened menstrual cycle， mental fatigue， lack of strength， poor appetite， loose stool， and white tongue coating. The Qi and blood deficiency syndrome were mainly manifested as the shortness of breath， laziness to speak， pale complexion， dizziness， and palpitation. The Qi deficiency and blood stasis syndrome were mainly judged by the scanty menstrual flow frequently or occasionally accompanied by blood clots， mental fatigue， lack of strength， and dark purple tongue. The ovulatory bleeding， dizziness， tinnitus， vexing heat in chest， palms and soles， and night sweat were the characteristic signs for the kidney Yin deficiency syndrome. Conclusion:There exist certain rules in the geographical distribution of TCM syndromes of AUB-O patients， which has provided a reference for the clinical treatment of AUB-O in accordance with the local conditions.

Tumor-secreted vascular endothelial growth factor A increases the pulmonary metastasis from nasopharyngeal carcinoma / 中华耳鼻咽喉头颈外科杂志

Objective@#Vascular endothelial growth factor A (VEGFA) was investigated as the key protein which might promote the specific metastasis progress of nasopharyngeal carcinoma.@*Methods@#Sixteen specimens of pulmonary metastasis carcinoma and counterparts in primary nasopharyngeal carcinoma tissue were collected from patients. The expression of VEGFA through immunohistochemistry was investigated.VEGFA was knocked down by siRNA in two cell lines of nasopharyngeal carcinoma (CNE-1 and 5-8F), MTT and Transwell test were used to explore the role of VEGFA in praxiology. Then shRNA was used to cultivate the stable CNE-1 cell line with down-regulated-expression of VEGFA. The nude mice models were built through tail vein injection of specific nasopharyngeal carcinoma cells, and lungs were collected to perform further metastasis analysis.@*Results@#Previous genetic studies showed that VEGFA had higher expression in metastasis tissue, and the result was validated in the present study using immunohistochemistry. The percentage of positive cells was 84.8% in pulmonary metastasis group, 51.5% in primary tissue group (t=8.639, P<0.05), average optical density was 0.154 in pulmonary metastasis group, 0.061 in primary tissue group (t=18.791, P<0.05). Low expression of VEGFA inhibited cell viability of optical density value of CNE-1 in siRNA gourp was 0.715, 0.902 in control group (t=7.274, P<0.05); 5-8F in siRNA group was 0.715, 0.935 in control group (t=7.751, P<0.05). Number counting suppressed migration of CNE-1 in siRNA group was 52 per high-power lens, 124 per high-power lens in control group (t=29.380, P<0.05), 5-8F in siRNA group was 65 per high-power lens, 155 per high-power lens in control group (t=18.181, P<0.05). Number counting invasion of CNE-1 in siRNA gourp was 38 per high-power lens, 86 per high-power lens in control group (t=27.665, P<0.05), 5-8F in siRNA group was 52 per high-power lens, 116 per high-power lens in control group (t=40.972, P<0.05) in vitro. Furthermore, knock-down of VEGFA in nasopharyngeal carcinoma reduced the pulmonary metastasis in vivo. Number counting of tumor volumes in shRNA group was 2.4, and 11.0 in control group (t=6.143, P<0.05); average optical density of immunohistochemistry in shRNA group was 0.033, and 0.176 in control group (t=15.734, P<0.05).@*Conclusions@#Results above reveal the overexpression of VEGFA in nasopharyngeal carcinoma can facilitate the pulmonary metastasis. Targeting VEGFA may provide a new biomarker of clinical study.

Study on therapeutic mechanisms of Danshen injection in intrahepatic cholestasis of pregnancy / 中国中药杂志

<p><b>OBJECTIVE</b>To study the therapeutic mechanisms of Danshen injection in intrahepatic cholestasis of pregnancy (ICP).</p><p><b>METHOD</b>The experimental serum was collected from 6 patients with ICP before therapy when in hospital. Endothelial cells were isolated from human umbilical veins chosen among the 10 normal pregnancy according to Jaffe's method. Serum of 20% concentration of ICP and different concentration of Danshen injection were cocultured with human vascular endothelial cells (HUVECs) for 24 hours. MTT method was applied to measuring the vitality of HUVECs. The expression value of vascular endothelial growth factor (VEGF) of different groups was detected by immunocytochemistry and enzyme-linked immunosorbent assay (ELISA) respectively.</p><p><b>RESULT</b>Compared with ICP group, the value of MTT and VEGF in the Danshen injection therapeutic groups were increased with the increase of concentration and moreover, 8 g x L(-1) of Danshen injection was the optical concentration. By immunocytochemistry, the expression of VEGF in HUVECs in the Danshen injection therapeutic groups was also enhanced with the increase of concentration.</p><p><b>CONCLUSION</b>Danshen injection can protect HUVECs against the injury of ICP serum and promote the expression of VEGF. Danshen injection improves HUVECs by increasing the value of VEGF.</p>