AIM:To explore the mechanism of berberine on breast cancer cells based on network pharmacology and in vitro cell experiments.METH-ODS:Firstly,berberine and breast cancer were tak-en as the research objects,the intersection targets of the two were screened by VEEN diagram,GO function and KEGG enrichment analysis were per-formed by R language,and molecular docking and visualization were carried out by Autodock Vina and Pymol software.Then,berberine treated breast cancer MCF-7 cells for 24 h,and then in vi-tro cell experiments were performed.CCK-8 was used to detect cell viability,Edu and plate cloning were used to detect cell proliferation and cloning,and apoptosis was detected by An-nexin V-FITC/PI double staining and Western blot.Laser confocal and CETSA were used to verify the binding effect of berberine and AKT1 protein.RESULTS:The results of network pharmacology showed that berberine had a good binding to the core targets AKT1,AKT2 and MAPK3.Berberine(20,40,80 μmol/L)signifi-cantly inhibited the proliferation and cloning ability of MCF-7 cells in a concentration-dependent man-ner(P＜0.05,P＜0.01).The results of laser confocal and CETSA experiments showed that berberine and AKT1 had a binding effect,and the stability of the two was enhanced after the combination.CONCLU-SION:Berberine inhibits MCF-7 cell proliferation and induces apoptosis in human breast cancer cells by targeting binding to AKT1 protein.

AIM:To explore the mechanism of berberine on breast cancer cells based on network pharmacology and in vitro cell experiments.METH-ODS:Firstly,berberine and breast cancer were tak-en as the research objects,the intersection targets of the two were screened by VEEN diagram,GO function and KEGG enrichment analysis were per-formed by R language,and molecular docking and visualization were carried out by Autodock Vina and Pymol software.Then,berberine treated breast cancer MCF-7 cells for 24 h,and then in vi-tro cell experiments were performed.CCK-8 was used to detect cell viability,Edu and plate cloning were used to detect cell proliferation and cloning,and apoptosis was detected by An-nexin V-FITC/PI double staining and Western blot.Laser confocal and CETSA were used to verify the binding effect of berberine and AKT1 protein.RESULTS:The results of network pharmacology showed that berberine had a good binding to the core targets AKT1,AKT2 and MAPK3.Berberine(20,40,80 μmol/L)signifi-cantly inhibited the proliferation and cloning ability of MCF-7 cells in a concentration-dependent man-ner(P＜0.05,P＜0.01).The results of laser confocal and CETSA experiments showed that berberine and AKT1 had a binding effect,and the stability of the two was enhanced after the combination.CONCLU-SION:Berberine inhibits MCF-7 cell proliferation and induces apoptosis in human breast cancer cells by targeting binding to AKT1 protein.

ObjectiveTo investigate the effect of Yunkang oral liquid on postpartum kidney deficiency in mice. MethodPostpartum mice were randomized into model and low-dose （6 mL·kg^-1）， medium-dose （9 mL·kg^-1）， and high-dose （12 mL·kg^-1） Yunkang oral liquid groups. The mouse model of postpartum kidney deficiency was established by sleep deprivation combined with forced swimming. Another 9 female ICR mice were selected as the normal control group. The mice were administrated with Yunkang oral liquid during the period of modeling. The levels of estradiol （E₂）， progesterone （P）， follicle-stimulating hormone （FSH）， and luteinizing hormone （LH） in the serum were measured by the enzyme-linked immunosorbent assay. The morphological changes of ovaries and uterus were observed by hematoxylin-eosin （HE） staining， and the expression of transforming growth factor （TGF）-β and Smad2/3 was determined by immunohistochemistry and Western blotting. ResultThe mice in the model group showed prolonged estrous cycle， reduced voluntary activity， dorsal temperature， grip strength， and bone strength， and whitening tongue coating. Compared with the model group， Yunkang oral liquid shortened the estrous cycle， increased the voluntary activity， dorsal temperature， grip strength， and bone strength， and alleviated the whitening of tongue coating. Moreover， it elevated the E₂ and P levels and lowered the FSH and LH levels in the serum， decreased ovarian follicular atresia rate， promoted uterine repair， and down-regulated the expression of TGF-β and Smad2/3 in the ovarian and uterine tissues. ConclusionYunkang oral liquid can ameliorate postpartum kidney deficiency in mice by regulating the TGF-β/Smads signaling pathway.

Background Bacteria are the most diverse and widely sourced microorganisms in the indoor air of subway stations, where pathogenic bacteria can spread through the air, leading to increased health risks. Objective To understand the status and distribution characteristics of indoor air bacterial pollution in subway stations and compartments in a city of Central South China, and to provide a scientific basis for formulating intervention measures to address indoor air bacteria pollution in subways. Methods Three subway stations and the compartments of trains parking there in a city in Central South China were selected according to passenger flow for synchronous air sampling and monitoring. Temperature, humidity, wind speed, carbon dioxide (CO₂), fine particulate matter (PM_2.5), and inhalable particulate matter (PM₁₀) were measured by direct reading method. In accordance with the requirements of Examination methods for public places-Part 3: Airborne microorganisms (GB/T 18204.3-2013), air samples were collected at a flow rate of 28.3 L·min⁻¹, and total bacterial count was estimated. Bacterial microbial species were identified with a mass spectrometer and pathogenic bacteria were distinguished from non-pathogenic bacteria according to the Catalogue of pathogenic microorganisms transmitted to human beings issued by National Health Commission. Kruskal-Wallis H test was used to compare the subway hygiene indicators in different regions and time periods, and Bonferroni test was used for pairwise comparison. Spearman correlation test was used to evaluate the correlation between CO₂ concentration and total bacterial count. Results The pass rates were 100.0% for airborne total bacteria count, PM_2.5, and PM₁₀ in the subway stations and train compartments, 94.4% for temperature and wind speed, 98.6% for CO₂, but 0% for humidity. The overall median (P₂₅, P₇₅) total bacteria count was 177 (138,262) CFU·m⁻³. Specifically, the total bacteria count was higher in station halls than in platforms, and higher during morning peak hours than during evening peak hours (P<0.05). A total of 874 strains and 82 species were identified by automatic microbial mass spectrometry. The results of identification were all over 9 points, and the predominant bacteria in the air were Micrococcus luteus (52.2%) and Staphylococcus hominis (9.8%). Three pathogens, Acinetobacter baumannii (0.3%), Corynebacterium striatum (0.1%), and Staphylococcus epidermidis bacilli (2.2%) were detected in 23 samples (2.6%), and the associated locations were mainly distributed in train compartments during evening rush hours. Conclusion The total bacteria count in indoor air varies by monitoring sites of subway stations and time periods, and there is a risk of opportunistic bacterial infection. Attention should be paid to cleaning and disinfection during peak passenger flow hours in all areas.

Objective:A nested-PCR assay is developed to detect and identify the genomic DNA of Brucella vaccine A19 strain. Methods:The whole genomic sequences of Brucella vaccine A19 strain and other Brucella spp. strains were compared and analyzed. The primers were designed by nucleotide difference sites. The nested-PCR assay was established to detect and identify Brucella vaccine A19 strain. The genomic DNA of Brucella vaccine A19 strain was extracted and diluted. The diluted template DNA was tested for sensitivity of using nested-PCR assay. And the specificity of nested-PCR assay was tested for the genomic DNA of other Brucella spp. strains and non- Brucella spp. strains. Results:The minimum detection limit of the nested-PCR assay was 3.43 fg. The nested-PCR assay established for amplification of Brucella vaccine A19 strain showed 246 bp electrophoresis bands, while other Brucella spp. strains showed 314 bp electrophoresis bands, and non- Brucella spp. strains did not produce electrophoresis bands. Conclusions:The nested-PCR assay established has the characteristics of high sensitivity and specificity. It can be detected when there is one copy of Brucella vaccine A19 strain genomic DNA in the reaction system. This method is particularly suitable for the detection and identification of trace genomic DNA of Brucella vaccine A19 strain in sample.

Objective:To establish a quantitative real-time PCR detection system for Brucella S2 vaccine strain. Methods:Based on the differences in the entire genome sequence between Brucella S2 vaccine strain and other reference strains of Brucella, primers and probes were designed to establish a quantitative real-time PCR detection system for Brucella S2 vaccine strain. The DNA of 22 reference strains of Brucella and 8 non- Brucella control strains were obtained from the National Institute for Infectious Disease Control and Prevention of the Chinese Center for Disease Control and Prevention. At the same time, environmental samples were obtained from the brucellosis vaccine manufacturers, and bacterial DNA from environmental samples was extracted using a blood/tissue genomic DNA extraction kit. The obtained DNA was pre-amplified by conventional PCR, and then subjected to quantitative real-time PCR secondary amplification (nested fluorescence quantitative PCR) using the amplified PCR product as a template. The specific fluorescence curve and corresponding number of cycles (Ct value) were observed, and the sensitivity was tested. Results:The quantitative real-time PCR detection system established did not detect specific fluorescence curves (without Ct values) for 21 reference strains of Brucella and 8 non- Brucella control strains, except for S2 vaccine strains. The established detection system had a minimum detection limit of 4.34 fg (genomic DNA) for detecting the DNA of Brucella S2 vaccine strain; DNA of Brucella S2 vaccine strain was detected in 3 of the 14 environmental samples collected. Conclusion:The quantitative real-time PCR detection system established can detect Brucella S2 vaccine strain in samples, with good sensitivity and specificity.

Objective:To analyze the differences in whole blood selenium (Se), zinc (Zn), copper (Cu), magnesium (Mg), calcium (Ca), and iron (Fe) levels of rural older adults among areas with different soil selenium levels, and explore the main factors associated with the six nutrient elements status, so as to provide a basis for further evaluating the health risks of people in areas with different soil selenium levels.Methods:Four administrative villages were randomly selected from the Se-deficient (soil Se content < 0.175 mg/kg), Se-sufficient (soil Se content 0.175 - < 0.400 mg/kg), Se-rich (soil Se content 0.400 - < 3.000 mg/kg) and Se-excessive (soil Se content ≥3.000 mg/kg) areas, respectively, in Enshi Tujia and Miao Autonomous Prefecture (Enshi Prefecture) of Hubei Province in 2017 - 2018. And 100 elderly people aged 60 years or older (half male and half female) were randomly selected as the survey subjects in each servey site. The basic information such as general demography and lifestyle was collected through face-to-face questionnaires. Physical examination was performed and fasting venous blood was collected in the morning. The contents of blood Se, Zn, Cu, Mg, Ca, and Fe were determined by inductively coupled plasma mass spectrometry. The main factors associated with the six nutrient elements status were analyzed.Results:A total of 416 subjects were included, including 208 males and 208 females, whose average age was (72.43 ± 5.25) years, and body mass index (BMI) was (22.67 ± 3.49) kg/m 2. There were significant differences of blood Se, Zn, Cu, Mg, Ca and Fe levels between the areas with different Se levels ( Z/F = 288.30, 3.24, 14.81, 29.14, 131.28, 3.37, P < 0.05). Compared with Se-deficient and Se-sufficient areas, blood Se level was higher in Se-rich and Se-excessive areas and blood Zn level was lower in Se-excessive area ( P < 0.05); compared with Se-sufficient area, blood Cu level was lower in Se-deficient, Se-rich and Se-excessive areas, but blood Mg and Ca levels were higher ( P < 0.05), and the blood Fe level was lower in Se-excessive area ( P < 0.05). There were significant differences in the deficiency rates of Se, Zn, Cu, Mg, Ca and Fe among the elderly in different Se level areas (χ 2 = 140.83, 15.39, 31.90, 17.49, 157.60, 30.33, P < 0.01). There were significant differences in blood Zn, Cu, Ca and Fe levels between two gender groups ( P < 0.05); the blood Zn and Fe levels of the smokers were higher than those of the non-smokers, and the blood Cu level was lower than that of the non-smokers ( P < 0.05); the blood Zn and Fe levels of the drinkers were higher than those of the non-drinkers ( P < 0.05). Conclusions:The levels of six nutrient elements in the whole blood of the elderly in areas with different soil Se levels are different. To assess the health risks of the population in areas with different soil Se levels, it is necessary to consider the levels of multiple nutrient elements at the same time.

Objective:To evaluate the effect of patient-controlled intravenous analgesia (PCIA) with dexmedetomidine mixed with subanesthetic dose of ketamine on anxiety and depression in the patients with advanced cancer pain.Methods:Sixty patients of either gender with advanced cancer pain, aged 24-82 yr, with poor analgesic effect or obvious adverse reactions after three-step analgesic treatment, were selected and randomly divided into 2 groups ( n=30 each) using a random number table method: routine treatment group (group R) and dexmedetomidine mixed with ketamine group (group DK). The initial dose of morphine for PCIA was 1/3 of the oral dose in group R. In group DK, ketamine 5.4 mg/kg (90 μg·kg -1·h -1) and dexmedetomidine 6 μg/kg (0.1 μg·kg -1·h -1) were added on the basis of group R. Tropisetron 8 mg was added to analgesics and diluted to 200 ml with normal saline in both groups.The analgesic pump was programmed to deliver 4 ml with an initial dose of 4 ml, lockout interval of 15 min and background infusion at 4 ml/h.The numerical rating scale score, Ramsay sedation score, Chinese version of State-Trait Anxiety Inventory score and Beck Depression Inventory-Ⅱ score were recorded before PCIA and at 4, 12, 24 and 48 h of PCIA.The development of effective analgesia and satisfactory sedation, occurrence and degree of depression, score for patient's quality of life and satisfaction score, consumption of morphine and adverse reactions such as constipation, nausea and vomiting, agitation and respiratory depression were recorded within 48 h of PCIA. Results:Compared with group R, the NRS score was significantly decreased, the rate of effective analgesia was increased, Beck Depression Inventory-Ⅱscore and Chinese version of State-Trait Anxiety Inventory score were decreased, the incidence and degree of depression were decreased, incidence of nausea and vomiting and constipation, consumption of morphine and pressing times of PCIA pump were decreased, and the score for patient's quality of life and satisfaction score were increased in group DK ( P<0.05). Conclusion:PCIA with dexmedetomidine mixed with subanesthetic dose of ketamine can significantly enhance the analgesic effect, improve anxiety and depression, and raise the quality of life when used for the patients with advanced cancer pain.

Objective:To study the effect of the injected bone marrow mesenchymal stem cells (BMSC) on rats with pulmonary fibrosis induced by paraquat (PQ) during different poisoning periods and explore the potential mechanism.Methods:From October to December 2018, BMSCs of SPF SD rats were isolated and purified by whole-bone marrow adherent culture method and cultured to the Third Generation (P3) . The surface antigens CD29, CD90, CD45 and CD34 of P3 BMSC were detected by Flow cytometry, the formation of alkaline phosphatase (ALP) , calcium nodules and fat droplets were observed by ALP, Alizarin Red staining and oil red O staining. At the same time, 36 SPF male rats were randomly divided into 6 groups: NC Group (Blank Control Group, injected with the same amount of saline) and PQ group (PQ model group, injected with 20% PQ solution 18 mg/kg intraperitoneally) , bMSC-A group, BMSC-B group, BMSC-C group and BMSC-D group were injected with BMSC suspension 1×10 6 cells/mice at 3 h、3 d、7 d and 14 d after PQ poisoning. After 28 days, the rats were killed, the lung organ coefficients were calculated, the hydroxyproline (HYP) content in lung tissue was calculated by alkaline hydrolysis, and the lung injury and fibrosis were observed by HE and Masson staining, serum TGF-1、TNF-α、MMP-9 and TIMP-1 were detected by Elisa. Results:High Purity BMSCs were successfully isolated and obtained. The P3 BMSC generation was positive expression of CD29、CD90、and negative expression of CD34、CD45, and had the potential of osteogenic and adipogenic differentiation. The results of HE staining and Masson staining showed that the alveolar structure in NC group was intact and homogeneous, in PQ group, the alveolar structure was severely damaged and a lot of collagen fibers and fibroblasts were deposited, and the degrees of lung injury in each BMSC intervention group were obviously less than in PQ group, in BMSC-A group and BMSC-B group, the degrees of reduction were obvious. Compared with NC group, the Lung organ coefficient, HYP content in lung tissue and TGF-β1, TIMP-1 levels in serum were significantly higher in PQ group ( P<0.05) , while TNF-α and MMP-9 had no significant difference ( P>0.05) . Compared with PQ group, the lung organ Coefficients, HYP, TGF-1 and TIMP-β1 in BMSC-A and BMSC-B groups were lower than those in PQ group ( P<0.05) . The Lung organ coefficients, TGF-β1 and TIMP-1 in BMSC-C and BMSC-D groups were lower than those in PQ group, there was no significant difference ( P>0.05) . Conclusion:Early BMSC injecting can alleviate pulmonary fibrosis induced by PQ. The mechanism may be that BMSC can reduce pulmonary fibrosis through reducing the level of TGF-β1 and regulating the balance of TIMP-1/MMP-9, threrby reducing inflammatory damage and increasing the degradation of extracellular matrix (ECM) .

Objective:To study the effect of the injected bone marrow mesenchymal stem cells (BMSC) on rats with pulmonary fibrosis induced by paraquat (PQ) during different poisoning periods and explore the potential mechanism.Methods:From October to December 2018, BMSCs of SPF SD rats were isolated and purified by whole-bone marrow adherent culture method and cultured to the Third Generation (P3) . The surface antigens CD29, CD90, CD45 and CD34 of P3 BMSC were detected by Flow cytometry, the formation of alkaline phosphatase (ALP) , calcium nodules and fat droplets were observed by ALP, Alizarin Red staining and oil red O staining. At the same time, 36 SPF male rats were randomly divided into 6 groups: NC Group (Blank Control Group, injected with the same amount of saline) and PQ group (PQ model group, injected with 20% PQ solution 18 mg/kg intraperitoneally) , bMSC-A group, BMSC-B group, BMSC-C group and BMSC-D group were injected with BMSC suspension 1×10 6 cells/mice at 3 h、3 d、7 d and 14 d after PQ poisoning. After 28 days, the rats were killed, the lung organ coefficients were calculated, the hydroxyproline (HYP) content in lung tissue was calculated by alkaline hydrolysis, and the lung injury and fibrosis were observed by HE and Masson staining, serum TGF-1、TNF-α、MMP-9 and TIMP-1 were detected by Elisa. Results:High Purity BMSCs were successfully isolated and obtained. The P3 BMSC generation was positive expression of CD29、CD90、and negative expression of CD34、CD45, and had the potential of osteogenic and adipogenic differentiation. The results of HE staining and Masson staining showed that the alveolar structure in NC group was intact and homogeneous, in PQ group, the alveolar structure was severely damaged and a lot of collagen fibers and fibroblasts were deposited, and the degrees of lung injury in each BMSC intervention group were obviously less than in PQ group, in BMSC-A group and BMSC-B group, the degrees of reduction were obvious. Compared with NC group, the Lung organ coefficient, HYP content in lung tissue and TGF-β1, TIMP-1 levels in serum were significantly higher in PQ group ( P<0.05) , while TNF-α and MMP-9 had no significant difference ( P>0.05) . Compared with PQ group, the lung organ Coefficients, HYP, TGF-1 and TIMP-β1 in BMSC-A and BMSC-B groups were lower than those in PQ group ( P<0.05) . The Lung organ coefficients, TGF-β1 and TIMP-1 in BMSC-C and BMSC-D groups were lower than those in PQ group, there was no significant difference ( P>0.05) . Conclusion:Early BMSC injecting can alleviate pulmonary fibrosis induced by PQ. The mechanism may be that BMSC can reduce pulmonary fibrosis through reducing the level of TGF-β1 and regulating the balance of TIMP-1/MMP-9, threrby reducing inflammatory damage and increasing the degradation of extracellular matrix (ECM) .