Objective To establish a bioiflm (BF) models of Haemophilus inlfuenza in vitro, and to observe the changes of antibiotic susceptibility after the BF fromation. Methods Thirty strains Haemophilus inlfuenzae isolated from adenoids of children with adenoidal hypertrophy and cultured in a 96-well plate. The BF was identiifed by crystal violet staining and scanning electron microscopy (SEM). The minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC) and the minimum bioiflm bacteria bioiflm clear concentration (MBEC) of ampicillin (AMP), ceftriaxone (CRO), levolfoxacin (LVFX) and azithromycin (AZM) were individually detected. Result All of 30 strains of Haemophilus inlfuenzae formed various BF. After BF is formed, the increase of MBEC for different antibiotics was inconsistent with the increase of MIC and MBC. The difference was statistically signiifcant (MBEC/MBC, H=91.54;MBEC/MIC, H=87.91;all P<0.001). The MBEC of AMP was the highest, up to 100 times than the MBC and MIC. The MBEC of CRO was dozens of times than the MBC and MIC. The MBEC of LVFX and AZM were most close to those of MBC and MIC. Conclusion After the formation of BF, resistance to antibiotics of Haemophilus inlfuenzae is enhaced. LVFX and AZM showed more favorable effect on Haemophilus infuenzae BF.

Objective To study the expression and significance of tumor metastasis suppressor gene-1(TMSG1) in esophageal squamous cell carcinoma (ESCC) and EC109 cells.Methods Immunohistochemistry S-P method was used to examine the expression of TMSG-1 protein in 136 cases of ESCC and 37 cases of normal esophageal mucosa.We analyzed the relationship between TMSG-1 and clinicopathological data of ESCC patients.EC109 cells were treated with 3 μg/mL of cisplatin (CDDP) in vitro for 24 h (the intervention group) and the control group was set up at the same time.The proliferation-inhibitory capability was analyzed with MTT assay.RT-PCR was used to examine the expression of TMSG-1 in the intervention group and the control group.Results The positive rate of TMSG-1 in ESCC and normal esophageal mucosa was 52.2％ (71/136) and 94.6％ (35/37),respectively.The expression of TMSG-1 in ESCC was significantly lower than that in normal esophageal mucosa (P＜0.05).The expression of TMSG-1 was related to TNM stage,differentiation degree and lymph node metastasis (P＜0.05).After EC 109 cells were treated with CDDP for 24 h,the proliferation inhibition rate was increased significantly compared with the control group (P＜0.01).RT-PCR results showed that the expression of TMSG-1 in the cells of the intervention group was significantly higher than that in the control group (P＜ 0.01).Conclusion The abnormal expression of TMSG-1 may play a role in the development and metastasis of ESCC.Examination of TMSG-1 may be useful for making diagnosis and guiding clinical therapy of ESCC.

Objective To research the molecular biology characteristics and transient expression in BHK-21 cells of E geue segment from Japanese encephalitis virus(JEV) and construct an eukaryotic expression vector pEGFP-C1-JEV.Methods E gene segment of JEV was amplified by RT-PCR,construct the recombinant vector pEGFP-C1-JEV,which could express EGFP label proteins.Transfect pEGFP-C1-JEV vector into BHK-21 via LipofectAMINETM 2000,to observe expressing of EGFP label protein and transcription of aim gene,and to check up localization and antigenicity of expressed E protein by Immunohistochemistry and Western blot.Results It showed that the recombinant plasmid pEGFP-C1-JEV was successfully constructed and transfected to BHK-21 cells,the normal expression of green fluorescent protein expression rate was higher.RT-PCR showed that gene transcription in BHK-21 and normal expression,expression protein was mainly distributed in the cytoplasm of BHK-21 cells and the envelope in,and can with guinea pig anti-JEV antibody binding.Conclusion pEGFP-C1-JEV vector in BHK-21 cells was normal expression and there were no effect on cell growth and morphology.Meanwhile,on eukaryotic antigens was good antigenicity.This research as a base foundation for E protein gene of JEV eukaryotic expression and function in vitro and applied research.

Objective To investigate the expression of Situin 1 ( SIRT1) in 5 strains of human lung adenocarcinoma cell lines, inclu-ding HCC827, H1650, H1975, A549 and H1299, and its relation to the susceptibility of nedaplatin ( NDP ) . Methods The SIRT1 mRNA and protein levels in 5 strains of human lung adenocarcinoma cells were detected by real-time quantitative PCR and Western blot, respectively. The viability of cells treated with NDP was detected by the CCK-8 method and the half growth inhibition concentra-tion ( IC50 ) was calculated. After the expressions of SIRT1 in A549, H1299, H1650 and H1975 cells were down-regulated by the siR-NA interference, the effects of NDP on the viability and apoptosis of these cells were determined by the CCK-8 method and flow cytom-etry, respectively.Results The expression levels of SIRT1 mRNA (4.53 ± 0.74, 3.11 ± 0.64, 15.76 ± 2.28 and 18.09 ± 1.17) and protein (0.23 ± 0.03, 0.21 ± 0.02, 0.52 ± 0.11 and 0.56 ± 0.08) in H1650, H1975, A549 and H1299 cells were significantly higher than that in HCC827 cells (1.00 for SIRT1 mRNA and 0.11 ± 0.02 for SIRT1 protein, F=122.10 and 26.50, respectively, P<0.01). The susceptibility of A549 and H1299 cells to NDP [IC50=(7.38 ± 1.59) and (8.14 ± 1.43) μmol/L, respectively] was significantly higher than that of HCC827, H1650 and H1975 cells [IC50=(26.16±4.35),(22.29±3.26) and (24.41 ± 2.58), respectively, F=30.86, P<0.01].The survivals of A549 and H1299 cells transfected by siSIRT1 and treated with NDP were significantly higher than that in the NC group ( F=235.10 and 39.20, respectively,P<0.01) , and the apoptotic rates were the reverse ( t=7.29 and 6.68, re-spectively, P<0.05) . However, the survivals of H1650 and H1975 cells transfected by siSIRT1 and treated with NDP were significantly lower than that in the NC group ( F=185.40 and 60.09, respectively,P<0.01) , and the apoptotic rates were the reverse ( t=6.15 and 31.36, respectively,P<0.01).Conclusion The expression of SIRT1 in A549 and H1299 cells with high expression of SIRT1 increases their susceptibility to NDP , while that in H1650 and H1975 cells with moderate expression of SIRT1 decreases their susceptibility to NDP, indicating that SIRT1 may play dual roles in the resistance of human lung adenocarcinoma cells to platinum.

Objective: To evaluate the clinical value of suprapubic Trocar puncture lithotripsy in the treatment of benign prostatic hyperplasia and bladder stones. Methods: From February 2016 to August 2018, 60 patients with benign prostatic hyperplasia and bladder stones in the Second Hospital of Pujiang County were randomly selected.The patients were divided into transurethral prostatic bipolarplasma electrotomy group(control group, 30 cases) and transurethral prostatic bipolar excision procedure combined with suprapubic Trocar puncture cystinolithotomy group(study group, 30 cases) according to the operation methods.The operation time, stone extraction time, removal of bladder fistula time, catheter extraction time, success of one stone extraction, postoperative complications of the two groups were statistically analyzed. Results: The operation time, stone removal time, bladder fistula removal time and catheter removal time of the study group were significantly shorter than those of the control group(t=6.965, 4.541, 3.365, 3.306, all P<0.05). The success rate of the first stone extraction in the study group was 100.0%(30/30), which was significantly higher than 86.7%(26/30) in the control group (χ²=12.83, P<0.05). The incidence rate of postoperative complications in the study group was 10.0%(3/30), which was significantly lower than 23.3%(7/30) in the control group(χ²=13.34, P<0.05). Conclusion Suprapubic Trocar puncture lithotripsy has high value in the treatment of benign prostatic hyperplasia and bladder stones.

Blastocystis sp. is a kind of protozoa living in the intestinal tract of human and animals, which will cause intestinal diseases such as diarrhea, abdominal distension and vomiting. This paper was aimed to understand the infection of Blastocystis sp. In golden monkeys and the transmission path in North China. Thirty-seven feces samples from golden monkeys and 116 cockroach samples from Shijiazhuang Zoo were collected from July to October 2019 for PCR analysis of Blastocystis sp. Genetic diversity analysis was further conducted on the samples with positive PCR results. The results showed that the infection rate was 48.7% (18/37) in golden monkeys and 82.8% (96/116) in cockroaches, respectively. The genetic evolution analysis based on small subunit ribosomal RNA demonstrated that three subtypes (ST) of Blastocystis sp. including ST1, ST2, and ST3 existed in the intestinal tract of golden monkeys, while only ST2 was detected in the intestinal tract of cockroaches. This paper may provide supports for the quarantine and control of Blastocystis sp. for the zoo in Northern China.

Objective To observe the effect of transcranial direct current stimulation (tDCS) with mirror neuronal rehabilitation training system (MNST-V1.0) in post-traumatic unconscious patients after severe craniocerebral injury. Methods A prospective, self controlled and open-label method was used. Thirty-six post-traumatic unconscious patients with severe craniocerebral injury from January 2016 to July 2017 were selected. Four cases of the patients did not complete the treatment and the last 32 cases completed the study. All patients were given routine wake-up therapy, and tDCS combined with MNST-V1.0 (20 min/time, 1 time/d, 6 times/week, a total of 8 weeks) was given at the same time. The Glasgow coma scale (GCS), JFK coma recovery scale and Four coma rating scale before treatment and 2, 4, 8 weeks after treatment were recorded. Results The scores of open reaction, language and motor response score of GCS 2, 4, 8 weeks after treatment were significantly higher than those before treatment:(1.56 ± 0.82), (2.06 ± 1.01) and (3.11 ± 1.45) scores vs. (1.00 ± 0.45) scores, (2.23 ± 1.06), (2.56 ± 1.08) and (3.02 ± 1.04) scores vs. (1.00 ± 0.61) scores, (2.79 ± 1.12), (3.22 ± 1.33) and (4.44 ± 1.07) scores vs. (1.00 ± 0.54) scores, and there were statistical differences (P ＜ 0.01 or ＜0.05). The scores of hearing, vision, movement, speech response, communication and arousal of JFK coma recovery scale 2, 4, 8 weeks after treatment were significantly higher than those before treatment, and there were statistical differences (P＜0.01). The scores of open reaction, sport reaction, brainstem response of Four coma rating scale 2, 4, 8 weeks after treatment were significantly higher than those before treatment, and there were statistical differences (P＜0.05); there was no statistical difference in respiratory score of Four coma rating scale before and after treatment (P＞0.05). Conclusions The tDCS combined with MNST-V1.0 can improve the consciousness level in post-traumatic unconscious patients with severe craniocerebral injury, and have the effect of promoting awakening.

Objective To investigate the effects of icariin on high glucose-induced autophagy and apoptosis of podocytes,and the regulating effects on mammalian target of rapamycin(mTOR)/serine-threonine kinase(Akt)/cyclic adenosine monophosphate response element binding protein(CREB)pathway.Methods The mouse podocytes MPC5 were taken and divided into five groups:normal control group(5.5 mmol·L-1 glucose),high glucose group(30 mmol·L-1 glucose),icariin group(30 mmol·L-1glucose+5 μmol·L-1icariin),GDC-0349 group(30 mmol·L-1glucose+50 μmol·L-1 GDC-0349),icariin+GDC-0349 group(30 mmol·L-1 glucose+5 μmol·L-1 icariin+50 μmol·L-1 GDC-0349).Cultured for 48 hours,the tetramethylazozolium salt method was used to detect the viability of MPC5 cells;acridine orange staining was used to observe the autophagy of MPC5 cells;apoptosis of MPC5 cells was detected by flow cytometry;Western blotting was used to detect the expression of autophagy[microtubule associated protein one light chain 3(LC3)II,LC3Ⅰ,autophagy-related protein(Beclin-1)],apoptosis[Bcl-2 related X protein(Bax),B cell lymphoma-2(Bcl-2)]and mTOR/Akt/CREB pathway-related proteins of MPC5 cells.Results Compared with the normal control group,the cell viability,expression levels of Bcl-2,phosphorylated mTOR(p-mTOR)/mTOR,phosphorylated Akt(p-Akt)/Akt,phosphorylated CREB(p-CREB)/CREB protein of MPC5 cells in the high glucose group were significantly decreased(P＜0.05),the autophagy ability was enhanced,the autophagosome showed orange fluorescence,and the apoptosis rate,LC3Ⅱ/LC3Ⅰ,Beclin-1,Bax protein expression levels were significantly increased(P＜0.05).Compared with the high glucose group,the cell viability,LC3Ⅱ/LC3Ⅰ,Beclin-1,Bcl-2,p-mTOR/mTOR,p-Akt/Akt,p-CREB/CREB protein expression levels of MPC5 cells in icariin group were significantly increased,the autophagy ability was further enhanced,the number of autophagosomes was increased,the autophagosomes showed brick red fluorescence(P＜0.05),the apoptosis rate and Bax protein expression level were significantly decreased(P＜0.05),and the cell viability,LC3Ⅱ/LC3Ⅰ,Beclin-1,Bcl-2,p-mTOR/mTOR,p-Akt/Akt and p-CREB/CREB proteins expression levels of MPC5 cells in GDC-0349 group were significantly decreased,the autophagy ability was weakened,the number of autophagosomes was reduced,the autophagosomes showed orange fluorescence(P＜0.05),and the apoptosis rate and Bax protein expression level were significantly increased(P＜0.05);icariin+GDC-0349 could reverse the effect of icariin on high glucose induced MPC5 cells(P＜0.05).Conclusion Icariin promotes elevated glucose-induced podocyte autophagy and inhibits apoptosis by activating the mTOR/Akt/CREB pathway.

Objective: To analyz the current situation of the diagnosis, treatment and prevention of methylmalonic acidemia, the phenotypes, biochemical features and genotypes of the patients in the mainland of China, were investigated. Methods: Tottally 1 003 patients of methylmalonic acidemia from 26 provinces and municipalities of the mainland of China were enrolled. The clinical data, biochemical features and gene mutations were studied. Blood aminoacids and acylcarnitines, urine organic acids, and plasma total homocysteine were determined for the biochemical diagnosis. Gene analyses were performed for the genetic study of 661 patients. The patients were treated with individual intervention and long-term follow up. Prenatal diagnoses were carried out for 165 fetuses of the families. Results: Among 1 003 patients (580 boys and 423 girls), 296 cases (29.5%) had isolated methylmalonic acidemia; 707 cases (70.5%) had combined homocysteinemia; 59 patients (5.9%) were detected by newborn screening; 944 patients (94.1%) had the onset at the ages from several minutes after birth to 25 years and diagnosed at 3 days to 25 years of age. The main clinical presentations were psychomotor retardation and metabolic crisis. Multi-organ damage, including hematological abnormalities, pulmonary hypertension, kidney damage, were found. MMACHC, MUT, MMAA, MMAB, HCFC1, SUCLG1, SUCLA2 mutations were found in 631 patients (96.6%) out of 661 patients who accepted gene analysis. MMACHC mutations were detected in 460 patients (94.7%) out of 486 cases of methylmalonic acidemia combined with homocysteinemia. MUT mutations were found in 158 (90.3%) out of 169 cases of isolated methylmalonic acidemia. The development of 59 patients detected by newborn screening were normal; 918 cases (97.2%) were diagnosed after onset accepted the treatment. Forty-five of them completely recovered with normal development. Twenty-six patients (2.7%) died; 873 (92.5%) patients had mild to severe psychomotor retardation. Methylmalonic acidemia were found in 35 out of 165 fetuses by metabolites assay of amniotic fluid and amniocytes gene analysis. Conclusion Combined methylmalonic acidemia and homocysteinemia is the common type of methylmalonic acidemia in the mainland of China. CblC defect due to MMACHC mutations is the most common type of methylmalonic acidemia combined with homocysteinemia. MUT gene mutations are frequent in the patients with isolated methylmalonic acidemia. Newborn screening is key for the early diagnosis and the better outcome. Combined diagnosis of biochemical assays and gene analysis are reliable for the prenatal diagnosis of methylmalonic acidemia.

Most of the current research on depression focuses on neuronal regulation,while the astrocytic mechanism of depression is far from explored.Astrocytes are the most numerous and widely distributed glial cells in the central nervous system.With a complex structural morphology,astrocytes play an important role in a variety of neuropsychiatric disorders by interacting with neuronal synapses,vasculature and other glial cells.Recent studies have shown that astrocytes may be involved in depression by regulating monoamine transmitters,glutamate cycle,synaptic plasticity,energy metabo-lism,and neuroinflammation.This review is intended to inspire new ideas for the treatment of depres-sion and the development of novel drugs based on astrocyte regulation.