Research progress on the intervention effects of Chinese medicine on microRNA regulating the signaling pathway of ulcerative colitis / 中国药房

Ulcerative colitis （UC） is a chronic non-specific inflammatory disease characterized by the damage of the epithelial barrier of the colon and the destruction of immune homeostasis. It has a long course， no recovery and high recurrence rate， and is recognized as a difficult digestive disease. MicroRNA （miRNA） has been confirmed to be specifically or differentially expressed in both UC patients and UC animal models， so miRNA can be used as markers for UC diagnosis or reference for treatment evaluation. TCM therapy has a definite therapeutic effect， a wide range of effects， and minimal side effects in the treatment of UC， so this article takes miRNA as the starting point and systematically elaborates on the mechanism of TCM regulating UC related signaling pathways by regulating the expression of miRNA. The results show that chlorogenic acid， Anchang decoction， and Fuyang huoxue jiedu formula can regulate the expressions of miR-155， miR-146a and miR-31-5p， etc.， thereby inhibiting signal transducer and activator of transcription （STAT） signal pathway transduction to improve UC. Limonin， ginsenoside Rh2， artesunate， etc. can inhibit nuclear factor-κB （NF-κB） signaling pathway conduction to improve UC by regulating the expressions of miR-214， miR- 155 and miR-19a， etc. Nitidine chloride， berberine， resveratrol， etc. can regulate the expressions of miR-31， miR-146a， miR- 146b， etc.， thereby inhibiting the Toll-like receptor 4 （TLR4） signaling pathway to improve UC. Mango polyphenolics， Compound qinbai granules， and Astragalus membranaceus polysaccharides can regulate the expressions of miR-126 and miR-193a-3p， thereby inhibiting the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin （PI3K/AKT/mTOR） signaling pathway to improve UC.

Correlation and consistency of three methods for tear evaluation in the diagnosis of dry eye / 国际眼科杂志(Guoji Yanke Zazhi)

AIM:To explore the correlation and consistency of three tear assessment methods in the diagnosis of dry eye, which include the Schirmer Ⅰ test(SⅠt), tear meniscus height(TMH)measurement and a new generation of tear detection technology-Strip meniscometry tube(SMTube).METHODS: A diagnostic test study. A total of 183 dry eye outpatients(183 right eyes)in the Dry Eye Center of Henan Eye Hospital were enrolled from May to June 2021. The SⅠt, TMH and SMTube were performed on all patients, and the correlation and consistency of the measurement results were analyzed.RESULTS:The ocular surface disease index(OSDI)of all included patients was 43.75(31.25, 58.33), noninvasive breakup time(NIBUT)was 7.26(4.97, 9.37)s, and the results of SⅠt, TMH and SMTube were 6(2, 12)mm/5min, 0.18(0.14, 0.22)mm and 5(3,8)mm/5s, respectively; The results of correlation analysis: TMH was positively correlated with SMTube(rs=0.751, P<0.001), however, SⅠt had no correlation with TMH(rs=0.139, P=0.060)and SMTube(rs=0.019, P=0.799). The results of consistency analysis: TMH showed good consistency with SMTube(Kappa=0.794, P=0.044), however, SⅠt showed poor consistency with TMH(Kappa=0.271, P=0.074)and SMTube(Kappa=0.193, P=0.070)respectively.CONCLUSION:SMTube is a new, simple and convenient tool for evaluating tear volume. It has a good correlation and consistency with TMH measurement results. Therefore, it can replace TMH measurement, but cannot replace SⅠt in clinical application.

Effect of Jinshui Xiangsheng Prescription on Clinical Outcome of Advanced Prostate Cancer Based on Propensity Score Matching / 中国实验方剂学杂志

ObjectiveTo explore the effect of Jinshui Xiangsheng prescription on the five-year clinical survival outcome of patients with advanced prostate cancer. MethodFrom May 1, 2014 to May 1, 2016, patients with advanced prostate cancer from Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, Shanghai University of Traditional Chinese Medicine and the Urology Department of the Second Affiliated Hospital of Nanjing University of Chinese Medicine were collected and treated with Jinshui Xiangsheng prescription (155 cases in the observation group). According to age and Gleason score, the patients without Jinshui Xiangsheng prescription were matched in a ratio of 1∶1 (155 cases in the control group). The androgen resistance rate, survival rate, median survival time, and median progress free survival time in 1, 3, 5 years were observed. The prognostic factors of advanced prostate cancer were analyzed and screened out by Chi-square test, t test, Kaplan-Meier and Cox survival analysis. ResultThe androgen resistance rates in the observation group in 1， 3， 5 years were 9.0% （14/155）， 72.3% （112/155）， and 92.9% （144/155）， respectively， and those in the control group were 20.6% （32/155）， 87.7% （136/155）， and 97.4% （151/155）. The 1-year （χ2=8.271，P<0.01）and 3-year （χ2=11.613，P<0.01） androgen resistance rates in the observation group were significantly lower than those in the control group. The median survival time and median progress free survival time in the observation group were （26.35±9.01） months and （11.02±4.40） months， respectively， and in the control group were （22.31±9.21） months and （9.87±5.12） months， respectively. The median survival time and median progress free survival time in the observation group were significantly longer than those in the control group （P<0.05， P<0.01）. The cumulative survival rates in 1， 3， 5 years in the observation group were 96.1% （149/155）， 80.6% （125/155）， and 39.4% （61/155）， respectively， and those in the control group were 94.2% （146/155）， 60.0% （93/155）， and 22.6% （35/155）， respectively. The 3-year （χ2=15.828，P<0.01） and 5-year （χ2=10.201，P<0.01） cumulative survival rates in the observation group were significantly higher than those in the control group. The monofactor analysis showed that the prognostic factors involved in Gleason score， initial prostate specific antigen （PSA）， tumor location， tumor stage， castration regimen， radiotherapy， chemotherapy， complete androgen blockade （CAB）， and Jinshui Xiangsheng prescription （P<0.05， P<0.01）. The multivariate analysis showed that initial PSA， tumor location， and tumor stage were the risk factors affecting the survival time of patients with advanced prostate cancer， whereas Jinshui Xiangsheng prescription， castration regimen， chemotherapy， radiotherapy， and CAB were the independent protective factors affecting the prognosis of advanced prostate cancer. ConclusionJinshui Xiangsheng prescription has a protective effect on the survival of patients with advanced prostate cancer， which reduces the androgen resistance rate and death risk of advanced prostate cancer， thus benefiting the survival of patients. Therefore， it deserves further promotion.

Mutual Information Entropy Analysis of Compatibility of Traditional Chinese Patent Medicines Against Liver Diseases in Chinese Pharmacopoeia（2020 Edition） / 中国实验方剂学杂志

ObjectiveTo investigate the compatibility rule of traditional Chinese patent medicines （TCPMs） against liver diseases through network analysis. MethodWith “liver” as the search term， TCPMs against liver diseases were retrieved from volume Ⅰ of Chinese Pharmacopoeia （2020 edition）， and the basic information of them was collected. TCPMs with same Chinese medicinal materials （CMMs）， usage， and indications， but different dosage forms， were unified as one formula. Mutual information entropy （MIE） of CMM couples was calculated to quantify the relationship between them， and the top 25% CMM pairs in MIE were used to construct the compatibility network， with CMM as node and the relationship between CMM pairs as the edge. Key CMM and frequently used CMM combinations were identified based on node centrality and cluster analysis， respectively. The indications of TCPMs related to the CMMs in clusters were recorded. Cytoscape 3.6.1 was employed for visualization and topology analysis of the compatibility network. ResultA total of 179 TCPMs， involving 428 CMMs， were retrieved. Angelicae Sinensis Radix， Paeoniae Radix Alba， and Glycyrrhizae Radix et Rhizoma were identified as key CMMs with high frequency， and Cuscutae Semen-Lycii Fructus， Citri Reticulatae Pericarpium-Cyperi Rhizoma， and Ecliptae Herba-Ligustri Lucidi Fructus combinations had high MIE. Furthermore， the CMMs were clustered into ten groups corresponding to different diseases which， however， all belonged to digestive diseases. ConclusionThis study unveils potential CMM pairs and common CMM combinations against liver diseases， which can serve as a reference for revealing compatibility rules of CMMs and research and development of Chinese medicine.

Mechanism of Biejiajian Wan Against EMT of Hepatocellular Carcinoma Cells Through NF-κB Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect of Biejiajian Wan （BJJW） on transforming growth factor-β1 （TGF-β1）-induced epithelial-mesenchymal transition （EMT） of HepG2 cells， and explore its mechanism against EMT of hepatocellular carcinoma cells. MethodHepG2 cells were randomly divided into a blank group， a TGF-β1 model group （10 μg·L-1 TGF-β1）， a low-dose BJJW group （10 μg·L-1 TGF-β1+0.55 g·kg-1 BJJW）， a medium-dose BJJW group （10 μg·L-1 TGF-β1+1.1 g·kg-1 BJJW）， a high-dose BJJW group （10 μg·L-1 TGF-β1+2.2 g·kg-1 BJJW）， and a sorafenib group （10 μg·L-1 TGF-β1+0.03 g·kg-1 sorafenib）. The EMT model was induced by 10 μg·L-1 TGF-β1 in HepG2 cells. After treatment with corresponding medicated serum， cell counting kit -8 （CCK-8） assay was used to detect cell proliferation. Cell migration ability was detected by the Transwell assay and wound healing assay. The protein expression related to EMT and nuclear factor-kappa B （NF-κB） signaling pathway was detected by cell immunofluorescence assay and Western blot. ResultCompared with the blank group 4 days later， the TGF-β1 model group showed fusiform and loose cells with widened gap and antennae reaching out， decreased protein expression of E-cadherin （P<0.05）， and increased protein expression of N-cadherin and vimentin （P<0.05）， which indicated that the EMT model was properly induced in HepG2 cells by TGF-β1 stimulation for 4 days. After 48 hours of treatment with the corresponding medicated serum， each medication group showed inhibited proliferation of HepG2 cells that had undergone EMT， especially the low- and high-dose BJJW groups （P<0.01）， and the medium-dose BJJW group showed increased E-cadherin protein expression （P<0.05） and decreased p-p65， N-cadherin， and vimentin protein expression （P<0.05）， as compared with the TGF-β1 model group. As revealed by the transwell assay and wound healing assay， TGF-β1 enhanced the migration ability of HepG2 cells （P<0.05， P<0.01） compared with the results in the blank group， compared with the TGF-β1 model group， the medication groups showed inhibited migration ability of HepG2 cells （P<0.05， P<0.01）. Compared with the blank group， the TGF-β1 model group promoted the expression of p65 and Snail into the nucleus. Compared with the TGF-β1 model group， the medication groups inhibited the expression of p65 and Snail into the nucleus. ConclusionBJJW may inhibit the EMT， proliferation， and migration of HepG2 cells induced by TGF-β1 by suppressing the NF-κB signaling pathway to exert an anti-hepatocellular carcinoma effect.

Comparison of three kinds of free flaps used in patients with oral and oropharyngeal tumors / 中华耳鼻咽喉头颈外科杂志

Objective: To compare the recovery and quality of life of patients with oral and oropharyngeal tumors treated with three kinds of free soft tissue flaps. Methods: The clinical data of 103 patients, including 66 males and 37 females, aged 26-74 years, who underwent primary repair of defects after resection of oral and oropharyngeal tumors in Sichuan Tumor Hospital from July 2014 to August 2020 were analyzed. Anterolateral thigh flap (ALTF) was used in 43 patients, radial forearm free flap (RFFF) in 45 patients, and lateral arm free flap (LAFF) in 15 patients. Postoperative qualities of life of patients were evaluated by the university of Washington quality of life questionnaire and oral health impact scale (HIP-14 Chinese edition). SPSS 23.0 software was used for statistical analysis. Results: The T staging of RFFF or LAFF group was significantly lower than that of ALTF group (P<0.05). There was no significant difference in mean flap areas between ALTF group ((55.87±27.38) cm2) and LAFF group ((49.93±19.44) cm2), while RFFF group had smaller mean flap area ((33.18±6.05) cm2) than ALTF group (t=5.311, P<0.001) and LAFF group (t=3.284, P=0.005). In terms of oral functions including swallowing, mastication, taste and spitmouth, there were no significant differences between LAFF group and RFFF group (P>0.05), but both groups had better oral functions than ALTF group (P<0.05). There was no significant difference in appearance scores between LAFF group (75(75, 75)) and ALTF group (75(75,75) vs.75(75,75),Z=-1.532, P=0.126), and both groups had higher scores than RFFF group (50(50, 75),Z values were -3.447 and -3.005 respectively, P<0.05). RFFF group had higher speech score (100(67, 100)) than LAFF group (67(50, 76),Z=-2.480, P<0.05) and ALTF group (67(33, 67),Z=-5.414, P<0.05). ALTF group had lower mean score of quality of life than RFFF group [72(56,77) vs.79(69, 89),Z=-3.070, P<0.05), but there was no statistical difference in the mean scores of qualities of life between ALTF group and LAFF group (Z=1.754, P=0.079). According to the evaluation of oral health impact scale (HIP-14 Chinese version) 1 year after surgery, individual item scores and the average score of all items in ALTF group were lower than those in RFFF and LAFF groups (P<0.05), with no significant difference between RFFF group and LAFF group (P>0.05). Conclusions: RFFF has unique advantages for small tissue defects, while ALTF is suitable for large tissue defects, such as buccal penetrating defect, whole tongue and near whole tongue defect, and LAFF is a compromise choice between ALTF and RFFF. ALTF is inferior to RFFF and LAFF in oral functional reconstruction, including swallowing, chewing, taste and spittle. ALTF and LAFF are superior to RFFF in postoperative appearance.

Establishment of ultra performance liquid chromatography-tandem mass spectrometry method for measuring plasma caffeine concentration / 中华检验医学杂志

Objective:To establish an ultra-high-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method for plasma caffeine concentration detection, and to explore the clinical value of caffeine therapeutic drug monitoring (TDM) in the treatment of premature infants with respiratory distress syndrome (RDS).Method:Take the plasma sample in a centrifuge tube, add the caffeine deuterated isotope internal standard, then add the protein precipitant, vortex the mixture thoroughly, and centrifuge the supernatant to enter the mass spectrometry analysis. The mobile phase were methanol and water, gradient elution; the column temperature was 45 ℃, the method was established using Shimadzu LC-30AD-CL liquid system and AB SCIEX 4500 QTRAP mass spectrometer, and the sensitivity, specificity, linearity, accuracy imprecision, matrix effect, and carry-over of the method were evaluated. Sample from 30 patients diagnosed with neonatal RSD were collected in the Department of Neonatology of Renmin Hospital of Wuhan University from February to April 2021, then detected the trough concentration of caffeine in premature infants with RDS after taking the same dose of caffeine to assess the impact of individual variation on caffeine drug concentration.Results:The detection limit of caffeine was 0.02 μg/ml, and the lowest limit of quantification was 0.05 μg/ml. It showed good linearity ( R2=0.9986, R>0.99) in the concentration range from 1.0 to 100.0 μg/ml, specificity (recovery rate of 85.52%-114.12%), accuracy (recovery rate 85.97%-114.53%), intra-day and inter-day imprecision ( CV 6.01%-11.28%), matrix effects and carryover pollution were negligible. The trough concentration of 30 preterm infants with RSD after taking the same dose of caffeine (10 mg/kg) was (25.45±11.61) μg/ml, and the coefficient of variation was 44.88%. Conclusion:This study established an accurate and reliable UPLC-MS/MS method with low sample consumption to monitor the blood concentration of caffeine; caffeine TDM has certain clinical application value, which can be used to assist RDS diagnosis and treatment and improve the efficacy of caffeine.

Expression of GFAP, β-tubulin Ⅲ and synaptophsin and intervention effects of chondroitin sulfate in brain of rats with chronic fluorosis / 中华地方病学杂志

Objective:To observe the expression levels of glial fibrillary acidic protein (GFAP), β-tubulin Ⅲ and synaptophsin, and explore the role of tripartite synapse in the mechanism of central nervous system (CNS) injury and the neuroprotective effect of chondroitin sulfate (CS).Methods:One month old clean grade, 48 female Sparague-Dawley rats and 48 male Sparague-Dawley rats, were randomly divided into 8 groups according to body weight (90 - 120 g) by random number table method, with 12 rats in each group, half male and half female. These rats were fed with water containing different concentrations of sodium fluoride (NaF) [ < 0.5 mg/L (control, CN), 10.0 mg/L (low dose fluoride, LF) and 50.0 mg/L (high dose fluoride, HF)]. Some rats were fed directly for 185 days (CN, LF and HF groups). In addition, rats of CN + normal saline (NS), LF + NS, HF + NS groups and LF + CS, HF + CS groups, were intraperitoneally injected with NS or 0.66 mg/kg CS for 5 consecutive days after 180 days of feeding. After the experiment, the pathological changes of hippocampal CA4 of brain tissue in each group were observed by hematoxylin eosin staining under light microscope, and the expression and distribution of GFAP, β-tubulin Ⅲ and synaptophsin in hippocampal CA4 of rats were detected by immunohistochemistry, the expression of GFAP, β-tubulin Ⅲ and synaptophsin at protein level in hippocampus of rats were detected by Western blotting.Results:Under light microscope, eosinophilic change, loss and irregular arrangement of neuron in the hippocampal CA4 were observed in LF, HF, LF + NS and HF + NS groups. The morphology of LF + CS and HF + CS groups was not significantly changed compared with CN group, but was significant changed compared with LF, HF, LF + NS and HF + NS groups. Immunohistochemical results showed that the rates of positive area of GFAP, β-tubulin Ⅲ and synaptophsin in female and male rats in LF and HF groups were significantly decreased than those in CN group ( P < 0.05); the positive area rates of female and male rats in LF + CS and HF + CS groups were higher than those in LF and HF groups, respectively ( P < 0.05). Western blotting results showed that the proten expression levels of GFAP, β-tubulin Ⅲ and synaptophsin of female and male rats in LF and HF groups (LF group: 0.90 ± 0.09, 0.82 ± 0.08, 1.43 ± 0.14, 0.92 ± 0.02, 1.21 ± 0.15, 0.87 ± 0.02, HF group: 0.58 ± 0.14, 0.73 ± 0.03, 0.63 ± 0.06, 0.67 ± 0.03, 0.87 ± 0.04, 0.70 ± 0.05) were lower than those in CN group (1.24 ± 0.08, 1.09 ± 0.10, 2.64 ± 0.30, 1.54 ± 0.09, 1.72 ± 0.10, 1.13 ± 0.06, P < 0.05). Conclusions:The tripartite synapse and extracellular matrix may take part in pathogenesis of the damages of CNS results from chronic fluorosis; CS may reduce the injury to a certain extent.

Clinical features and prognostic factors of extrathyroidal extension in differentiated thyroid carcinoma / 中华内分泌外科杂志

Objective:To investigate the risk factors related to extrathyroidal extension (ETE) of differentiated thyroid carcinoma (DTC) and the specific effects on the prognosis.Methods:The clinical data of 592 patients with newly diagnosed DTC admitted to our hospital from Jun. 2015 to Jun. 2016 were retrospectively analyzed. The data including the maximum tumor diameter and lymph node metastasis were collected, and the survival data were followed up. Chi-square test and Logistic regression were used to analyze the risk factors associated with ETE. Kaplan-Meier method and Cox proportional risk model were used to analyze the effect of ETE on disease-free survival.Results:There were 100 ETE of 592 DTC patients (16.9%) . Univariate analysis showed that the risk factors for ETE were the largest tumor diameter ≥2 cm, multiple lesions, and lymph node metastasis in the lateral cervical region ( P<0.01) . The follow-up time was 29 to 64 months, and the median follow-up time was 60 months. The 3-year and 5-year DFS rates of patients with DTC and ETE were 86.9% and 83.2% respectively, which were significantly lower than those of patients without ETE ( P<0.001) . Univariate analysis showed that ETE ( P<0.001) was a risk factor for postoperative recurrence in patients with DTC. Cox multivariate regression showed that ETE (HR: 10.564, 95% CI 3.712-30.063, P<0.001) was an independent risk factor for postoperative recurrence in patients with DTC. Conclusions:DTC accompanied by ETE is associated with risk factors such as large tumor diameter and multiple lesions, which has a lower 5-year DFS. Patients with the above characteristics should consider the possibility of ETE before surgery, and focus on long-term follow-up after surgery.

Analysis of the Genome Sequence of Strain GiC-126 of Gloeostereum incarnatum with Genetic Linkage Map

Gloeostereum incarnatum has edible and medicinal value and was first cultivated and domesticated in China. We sequenced the G. incarnatum monokaryotic strain GiC-126 on an Illumina HiSeq X Ten system and obtained a 34.52-Mb genome assembly sequence that encoded 16,895 predicted genes. We combined the GiC-126 genome with the published genome of G. incarnatum strain CCMJ2665 to construct a genetic linkage map (GiC-126 genome) that had 10 linkage groups (LGs), and the 15 assembly sequences of CCMJ2665 were integrated into 8 LGs. We identified 1912 simple sequence repeat (SSR) loci and detected 700 genes containing 768 SSRs in the genome; 65 and 100 of them were annotated with gene ontology (GO) terms and KEGG pathways, respectively. Carbohydrate-active enzymes (CAZymes) were identified in 20 fungal genomes and annotated; among them, 144 CAZymes were annotated in the GiC-126 genome. The A mating-type locus (MAT-A) of G. incarnatum was located on scaffold885 at 38.9 cM of LG1 and was flanked by two homeodomain (HD1) genes, mip and beta-fg. Fourteen segregation distortion markers were detected in the genetic linkage map, all of which were skewed toward the parent GiC-126. They formed three segregation distortion regions (SDR1–SDR3), and 22 predictive genes were found in scaffold1920 where three segregation distortion markers were located in SDR1. In this study, we corrected and updated the genomic information of G. incarnatum. Our results will provide a theoretical basis for fine gene mapping, functional gene cloning, and genetic breeding the follow-up of G. incarnatum.

Analysis of the Genome Sequence of Strain GiC-126 of Gloeostereum incarnatum with Genetic Linkage Map

Gloeostereum incarnatum has edible and medicinal value and was first cultivated and domesticated in China. We sequenced the G. incarnatum monokaryotic strain GiC-126 on an Illumina HiSeq X Ten system and obtained a 34.52-Mb genome assembly sequence that encoded 16,895 predicted genes. We combined the GiC-126 genome with the published genome of G. incarnatum strain CCMJ2665 to construct a genetic linkage map (GiC-126 genome) that had 10 linkage groups (LGs), and the 15 assembly sequences of CCMJ2665 were integrated into 8 LGs. We identified 1912 simple sequence repeat (SSR) loci and detected 700 genes containing 768 SSRs in the genome; 65 and 100 of them were annotated with gene ontology (GO) terms and KEGG pathways, respectively. Carbohydrate-active enzymes (CAZymes) were identified in 20 fungal genomes and annotated; among them, 144 CAZymes were annotated in the GiC-126 genome. The A mating-type locus (MAT-A) of G. incarnatum was located on scaffold885 at 38.9 cM of LG1 and was flanked by two homeodomain (HD1) genes, mip and beta-fg. Fourteen segregation distortion markers were detected in the genetic linkage map, all of which were skewed toward the parent GiC-126. They formed three segregation distortion regions (SDR1–SDR3), and 22 predictive genes were found in scaffold1920 where three segregation distortion markers were located in SDR1. In this study, we corrected and updated the genomic information of G. incarnatum. Our results will provide a theoretical basis for fine gene mapping, functional gene cloning, and genetic breeding the follow-up of G. incarnatum.

Effect of electrothermal acupuncture on moderate to severe cancer pain with / 中国针灸

OBJECTIVE@#To observe the effectiveness and safety of electrothermal acupuncture therapy for patients of moderate to severe cancer pain with @*METHODS@#A total of 60 patients of moderate to severe cancer pain with @*RESULTS@#The variation of NRS scores in the observation group were larger than the control group 3, 5 days into treatment (@*CONCLUSION@#On the basis of the conventional western medication for analgesia, electrothermal acupuncture could relieve pain, reduce the dose of opioid painkillers and improve the quality of life in patients of moderate to severe cancer pain with

Expression, purification and characterization of anti-α2δ1/NanoLuc fusion protein / 生物工程学报

The existence of cancer stem cells is regarded as the major cause for therapeutic resistance and relapse of a variety of cancer types including hepatocellular carcinoma (HCC). However, the tracing of such a subpopulation in vivo has been challenging. We have previously demonstrated that the isoform 5 of the voltage-gated calcium channel α2δ1 subunit, which can be recognized specifically by a monoclonal antibody 1B50-1, is a bona fide surface marker for HCC stem cells. Here we developed a strategy for optical imaging of α2δ1-positive cells by using a fusion protein containing the single chain variable fragment (scFv) of Mab1B50-1 and the luciferase NanoLuc which was tagged with Flag in the C-terminal. The scFv of Mab1B50-1 was fused to the N-terminal of NanoLucFlag using overlap PCR, and the recombinant fragment, which was named as 1B50-1scFv-NanoLucFlag, was subsequently cloned into a eukaryotic expression vector. The resulting construct was transfected into FreeStyle 293F cells in suspension using PEI reagent. The expression of the fusion protein was identified as a protein with molecular weight about 50 kDa by Western blotting. After purification by ANTI-FLAG® M2 affinity chromatography, 1B50-1scFv-NanoLucFlag was demonstrated to bind to α2δ1 positive cells specifically with a Kd value of (18.62±1.84) nmol/L. Furthermore, a strong luciferase activity of 1B50-1scFv-NanoLucFlag was detected in α2δ1 positive cells following incubation with the fusion protein, indicating that the presence of α2δ1 could be quantified using this fusion protein. Hence, 1B50-1scFv-NanoLucFlag provides a potential tool for optical imaging of α2δ1 positive cancer stem cells both in vitro and in vivo.

Mechanism of Biejiajian Wan in Reversing EMT in Rat Hetapic Oval Cells via Wnt/ β-catenin Pathway / 中国实验方剂学杂志

Objective:To study the effect of Biejiajian Wan on the epithelial-mesenchymal transition （EMT） of rat hepatic oval cells induced by transforming growth factor- β1（TGF-β1）， in order to explore its mechanism in reversing EMT. Method:WB-F344 cells were divided into five groups： blank group， TGF-β1 model group （10 μg·L-1TGF-β1）， low-dose group （10 μg·L-1TGF-β1+0.55 g·kg-1Biejiajian Wan）， medium-dose group （10 μg·L-1TGF-β1+1.1 g·kg-1Biejiajian Wan）， high-dose group （10 μg·L-1TGF-β1+2.2 g·kg-1Biejiajian Wan）. Except blank group， TGF-β1 was used to induce WB-F344 cells in all of the remaining groups to construct an EMT model. After the cells were treated with low， medium and high doses of Biejiajian Wan serum， the changes of migration ability of WB-F344 cells were detected by cell scratching test. The expressions of E-cadherin， N-cadherin and Vimentin were detected by Western blot. Real-time PCR was used to detect the changes in the expression of β-catenin mRNA. The expression of β-catenin was detected by cell immunofluorescence assay. Result:Compared with normal WB-F344 cells， the intercellular space of WB-F344 cells became loose from tight， and the morphology changed from cobblestone to fibroblast after TGF-β1 induced WB-F344 cells for 4 days， and the expression of E-cadherin protein decreased， while the expression of N-cadherin protein increased （P<0.01）， indicating that the EMT model of WB-F344 cells was successfully built. Compared with the blank group， the migration ability of WB-F344 cells in TGF-β1 model group was enhanced （P<0.01）， compared with TGF-β1 model group， Biejiajian Wan could significantly inhibit the migration ability of WB-F344 cells； specifically， the low-dose group had no statistical significance， and the medium and high-dose groups had statistical significance （P<0.05）. Western blot results showed that compared with the blank group， the expression of E-cadherin decreased， whereas those of N-cadherin and Vimentin increased in the TGF-β1 model group （P<0.01）， compared with TGF-β1 model group， E-cadherin protein expression was increased in the low， medium and high-dose groups， while the expressions of N-cadherin and Vimentin was decreased； specifically， the low-dose groups had no statistical significance， and the medium and high dose groups had statistical significance （P<0.05，P<0.01）. Real-time PCR results showed that compared with the blank group， the mRNA expression of β-catenin in the TGF-β1 model group was increased （P<0.05）， whereas compared with TGF-β1 model group， the mRNA expression of β-catenin in the low， medium and high-dose groups of Biejiajian Wan was reduced （P<0.01）. The results of cellular immunofluorescence showed that compared with the blank group， the fluorescence expression of β-catenin in the cell nucleus was enhanced in the TGF-β1 model group； and compared with the TGF-β1 model group， the expression of β -catenin in the cell nucleus of the low， medium and high-dose groups of Biejiajian Wan decreased， and the inhibitory effect of Biejiajian Wan on β-catenin in the cell nucleus was positively correlated with its concentration. Conclusion:Biejiajian Wan may reverse the EMT process that TGF-β1 induced WB-F344 cells， and inhibit the migration of WB-F344 cells by inhibiting Wnt/β-catenin signaling pathway.

Advances in Study on Farnesoid X Receptor in Digestive System Tumors / 胃肠病学

Farnesoid X receptor (FXR) is the first discovered bile acid receptor. In recent years, more and more studies have shown that there are disorders of FXR expression in malignant tumors of digestive system. This article reviewed the advances in study on FXR in the digestive system tumors.

Repurposing of clinically approved drugs for treatment of coronavirus disease 2019 in a 2019-novel coronavirus-related coronavirus model / 中华医学杂志(英文版)

BACKGROUND@#Medicines for the treatment of 2019-novel coronavirus (2019-nCoV) infections are urgently needed. However, drug screening using live 2019-nCoV requires high-level biosafety facilities, which imposes an obstacle for those institutions without such facilities or 2019-nCoV. This study aims to repurpose the clinically approved drugs for the treatment of coronavirus disease 2019 (COVID-19) in a 2019-nCoV-related coronavirus model.@*METHODS@#A 2019-nCoV-related pangolin coronavirus GX_P2V/pangolin/2017/Guangxi was described. Whether GX_P2V uses angiotensin-converting enzyme 2 (ACE2) as the cell receptor was investigated by using small interfering RNA (siRNA)-mediated silencing of ACE2. The pangolin coronavirus model was used to identify drug candidates for treating 2019-nCoV infection. Two libraries of 2406 clinically approved drugs were screened for their ability to inhibit cytopathic effects on Vero E6 cells by GX_P2V infection. The anti-viral activities and anti-viral mechanisms of potential drugs were further investigated. Viral yields of RNAs and infectious particles were quantified by quantitative real-time polymerase chain reaction (qRT-PCR) and plaque assay, respectively.@*RESULTS@#The spike protein of coronavirus GX_P2V shares 92.2% amino acid identity with that of 2019-nCoV isolate Wuhan-hu-1, and uses ACE2 as the receptor for infection just like 2019-nCoV. Three drugs, including cepharanthine (CEP), selamectin, and mefloquine hydrochloride, exhibited complete inhibition of cytopathic effects in cell culture at 10 μmol/L. CEP demonstrated the most potent inhibition of GX_P2V infection, with a concentration for 50% of maximal effect [EC50] of 0.98 μmol/L. The viral RNA yield in cells treated with 10 μmol/L CEP was 15,393-fold lower than in cells without CEP treatment ([6.48 ± 0.02] × 10vs. 1.00 ± 0.12, t = 150.38, P < 0.001) at 72 h post-infection (p.i.). Plaque assays found no production of live viruses in media containing 10 μmol/L CEP at 48 h p.i. Furthermore, we found CEP had potent anti-viral activities against both viral entry (0.46 ± 0.12, vs.1.00 ± 0.37, t = 2.42, P < 0.05) and viral replication ([6.18 ± 0.95] × 10vs. 1.00 ± 0.43, t = 3.98, P < 0.05).@*CONCLUSIONS@#Our pangolin coronavirus GX_P2V is a workable model for 2019-nCoV research. CEP, selamectin, and mefloquine hydrochloride are potential drugs for treating 2019-nCoV infection. Our results strongly suggest that CEP is a wide-spectrum inhibitor of pan-betacoronavirus, and further study of CEP for treatment of 2019-nCoV infection is warranted.

Rules of acupoint selection and drug use in clinical treatment of hypertension with acupoint application therapy / 中国针灸

OBJECTIVE@#To explore the rules of acupoint selection and drug use in treatment of hypertension with acupoint application therapy.@*METHODS@#The articles of the clinical research of hypertension treated with acupoint application therapy were retrieved from Chinese journal full-text database (CNKI), VIP database (VIP) and Wanfang databases from the time of establishment to January 20, 2019. The database was set up with Microsoft Excel 2010. Using the cloud platform of the ancient and modern medicine record, the frequency statistical and clustering analyses were conducted.@*RESULTS@#A total of 117 articles were collected, including 191 prescriptions, 60 aucpoints and 236 kinds of herbal drugs. It was found in the frequency statistical analysis that the top 6 acupoints in use frequency were Yongquan (KI 1), Quchi (LI 11), Taichong (LR 3), Shenque (CV 8), Sanyinjiao (SP 6) and Neiguan (PC 6). According to the correlation analysis, corresponding to these top 6 acupoints, the pairs of acupoints were Sanyinjiao (SP 6) and Yongquan (KI 1), Shenque (CV 8) and Yongquan (KI 1), Neiguan (PC 6) and Yongquan (KI 1), Zusanli (ST 36) and Sanyinjiao (SP 6), Sanyinjiao (SP 6) and Neiguan (PC 6) with Yongquan (KI 1), as well as Yongquan (KI 1) and Neiguan (PC 6) with Sanyinjiao (SP 6). The dominant meridians were the kidney meridian, the conception vessel and the bladder meridian. The special acupoints referred to -source point, -connecting point, back- point and front- point. The top 3 herbal drugs in use frequency included , and . The herbs used were mainly warm and slight cold in nature and neutral in property. The frequencies of the drug use were similar in the application for cold and heat purposes. The common flavors of the herbal medicines were pungent, sweat and bitter and the liver, kidney and spleen meridians were generally involved in meridian tropism.@*CONCLUSION@#In treatment of hypertension with acupoint application therapy, the commonly used single acupoint is Yongquan (KI 1), which is generally combined with Sanyinjiao (SP 6), Shenque (CV 8), Neiguan (PC 6) and Zusanli (ST 36). The correlation is emphasized on the application of special acupoints, meridian points and organs. The vesicatory herbal drugs are predominant in the drug use. In generally, this therapy embodies the treatment principles as tonifying for the deficiency and reducing for the excess, as well as balancing of cold and heat.

Repurposing of clinically approved drugs for treatment of coronavirus disease 2019 in a 2019-novel coronavirus (2019-nCoV) related coronavirus model / 中华医学杂志(英文版)

Background@#Medicines for the treatment of 2019-novel coronavirus (2019-nCoV) infections are urgently needed. However, drug screening using live 2019-nCoV requires high-level biosafety facilities, which imposes an obstacle for those without such facilities or 2019-novel coronavirus (2019-nCoV). This study aims to repurpose the clinically approved drugs for the treatment of coronavirus disease 2019 (COVID-19) in a 2019-nCoV related coronavirus model.@*Methods@#A 2019-nCoV related pangolin coronavirus GX_P2V/pangolin/2017/ Guangxi was described. Whether GX_P2X uses angiotensin-converting enzyme 2 (ACE2) as the cell receptor was investigated by using small interfering RNA (siRNA) -mediated silencing of ACE2. The pangolin coronavirus model was used to identify drug candidates for treating 2019-nCoV infection. Two libraries of 2406 clinically approved drugs were screened for their ability to inhibit cytopathic effects on Vero E6 cells by GX_P2X infection. The antiviral activities and antiviral mechanisms of potential drugs were further investigated. Viral yields of RNAs and infectious particles were quantified by quantitative real-time polymerase chain reaction (qRT-PCR) and plaque assay, respectively.@*Results@#The spike protein of coronavirus GX_P2V shares 92.2% amino acid identity with that of 2019-nCoV isolate Wuhan-hu-1, and uses ACE2 as the receptor for infection just like 2019-nCoV. Three drugs-cepharanthine (CEP), selamectin and mefloquine hydrochloride exhibited complete inhibition of cytopathic effects in cell culture at 10 μmol/L. CEP demonstrated the most potent inhibition of GX_P2V infection, with a concentration for 50% of maximal effect [EC50] of 0.98 μmol/L. The viral RNA yield in cells treated with 10 μmol/L CEP was 15,393-fold lower than in cells without CEP treatment ([6.48±0.02]×10-4 vs. 1.00±0.12, t=150.38, P<0.001) at 72 h post-infection (p.i.). Plaque assays found no production of live viruses in media containing 10 μmol/L CEP at 48 h p.i. Furthermore, we found CEP has potent antiviral activities against both viral entry (1.00±0.37 vs. 0.46±0.12, t=2.42, P<0.05) and viral replication (1.00±0.43 vs. [6.18±0.95]×10-4, t=3.98, P<0.05).@*Conclusions@#Our pangolin coronavirus GX_P2V is a workable model for 2019-nCoV research. CEP, selamectin and mefloquine hydrochloride are potential drugs for treating 2019-nCoV infection. Our results strongly suggest that CEP is a wide-spectrum inhibitor of pan-betacoronavirus, and clinical trial of CEP for treatment of 2019-nCoV infection is warranted.

Mechanism of Biejia Jianwan in Suppressing Epithelial-mesenchymal Transition in DEN-induced Rat Hepatocellular Carcinoma via TGF-β/Smad Pathway / 中国实验方剂学杂志

Objective:To study the effect of Biejia Jianwan on expressions of signal molecules and target genes of transforming growth factor-β (TGF-β)/Smad pathway in diethylnitrosamine (DEN)-induced rat hepatocellular carcinoma, and explore the mechanisms of Biejia Jianwan suppressing the invasion of hepatocellular carcinoma. Method:The rats were divided into three group, namely normal group, model group and Biejia Jianwan group (2.2 g·kg-1·d-1). Rats in Biejia Jianwan group and model group received intraperitoneal injections of DEN to induce sequential chronic inflammation, cirrhosis and hepatocellular carcinoma. At the sign of cirrhosis, rats in Biejia Jianwan group began taking Biejia Jianwan by gavage for 6 weeks. Rat blood was collected to measure serum levels of biochemical markers of liver function tests, including alanine aminotransferase(ALT), aspartate aminotransferase(AST), total bilirubin(TBIL), albumin(Alb), γ-glutamyl transpeptadase(GGT), alkaline phosphatase(ALP). Rat livers were fixed in formalin and stained with hematoxylin-eosin (HE)staining， quantitative real-time PCR was used to test the mRNA expressions of TGF-β1， and Western blot was used to test protein expressions of TGF-β1, Smad2/3, p-Smad2/3, N-cadherin, E-cadherin and Vimentin. Result:All of the levels of biochemical markers showed no difference in Biejia Jianwan group and model group. Biejia Jianwan could improve the pathological changes of balloon-like degeneration, edema, and necrosis in liver cancer tissues. Importantly, the treatment dramatically decreased the mRNA expression of TGF-β1(P<0.01), and the protein expressions of TGF-β1, p-Smad2(P<0.01). Besides, the protein expression of N-cadherin and Vimentin were decreased significantly (P<0.01). Conclusion:Biejia Jianwan can inhibit epithelial-mesenchymal transition (EMT) in hepatocellular carcinoma cells activated via TGF-β/Smad pathway by reducing TGF-β1 expression, so as to suppress the metastasis and invasion of hepatocellular carcinoma.

Effects of narrow-band UVB irradiation on Wnt5a and Frizzled2 signaling molecules in patients with psoriasis vulgaris / 中华物理医学与康复杂志

Objective:To observe the effect of narrow band ultraviolet B (NB-UVB) treatment of patients with psoriasis vulgaris on the expression of Wnt5a and Frizzled2 signaling molecules and its relationship with skin lesions so as to explore the mechanism of treating psoriasis vulgaris with NB-UVB.Methods:Thirty-two patients with psoriasis vulgaris and 30 healthy volunteers were selected into a case group and a healthy control group. The case group was treated with NB-UVB irradiation for 8 weeks. The mRNA and protein associated with Wnt5a and Frizzled2 signaling molecules in the skin tissues of both groups were detected before and after the phototherapy using a reverse transcription-polymerase chain reaction and western blotting respectively. Psoriasis area and severity index(PASI) scores were recorded before and after the treatment for the case group, and the correlation between Wnt5a and Frizzled2 protein content and PASI score was analyzed.Results:Before the treatment the expression of Wnt5a and Frizzled2 molecular mRNA and protein in the lesioned area of the case group was significantly higher than in the healthy area of the case group and in the healthy group. It had decreased significantly after the treatment. Before the treatment, the expression of Wnt5a and Frizzled2 protein in the lesioned area was significantly and positively correlated with the severity of the disease. However, after the NB-UVB treatment, the average PASI score was significantly decreased. The expression of Wnt5a and Frizzled2 protein in the lesioned area was still significantly and positively correlated with the severity of the disease.Conclusion:NB-UVB irradiation can significantly reduce the mRNA and protein expression levels of Wnt5a/Frizzled2 signaling molecules in the lesions of patients with psoriasis vulgaris, which may be one of the mechanisms of NB-UVB in treating psoriasis.