ObjectiveTo investigate the survival and adverse reactions of patients with unresectable cholangiocarcinoma after stereotactic body radiation therapy （SBRT）. MethodsA total of 27 patients with unresectable solitary cholangiocarcinoma without metastasis who underwent SBRT in The Fifth Medical Center of Chinese PLA General Hospital from February 2012 to July 2020 were enrolled. The prescribed dose to planning target volume was 42-60 Gy in 5-8 fractions， with 5-11 Gy/fraction. Among these patients， five patients were also treated with chemotherapy and transcatheter arterial chemoembolization. The 6-， 12-， 18-， and 24-month overall survival （OS） rates， progression-free survival （PFS） rates， and local control （LC） rates were used as the assessment indices for treatment outcome； Common Terminology Criteria for Adverse Events v.4.03 was used to evaluate adverse reactions； the Kaplan-Meier method was used to calculate OS， PFS， and LC rates. ResultsThe median follow-up time was 17 months. For all 27 patients， the 6-， 12-， 18-， and 24-month OS rates were 100%， 88%， 57.5%， and 47.9%， respectively； the 6-， 12-， 18-， and 24-month PFS rates were 74.1%， 58.6%， 47.9%， and 35.9%， respectively； the 6-， 12-， 18-， and 24-month LC rates were 96.3%， 91.9%， 84.8%， and 76.4%， respectively. No grade 3 or above toxic reactions were observed. Five patients were diagnosed with radiation-induced liver injury， but there was no death due to radiation-induced liver injury. ConclusionSBRT is safe and effective in the treatment of unresectable cholangiocarcinoma， with relatively high survival rate， PFS rate， and LC rate and low toxicity， and therefore， SBRT can be used as an alternative treatment method for patients with cholangiocarcinoma who are not candidates for surgery.

Objective:To investigate the detection rate and related factors of thyroid nodules in people with abnormal lipid metabolism.Methods:From September 4, 2016 to February 1, 2017, community residents living in Lanzhou City, Longnan City, Dingxi City and Linxia City of Gansu Province for more than 5 years were selected as the respondents. General data were recorded, venous blood was collected, blood lipid related biochemical indexes were detected, and thyroid ultrasound was performed. By comparing the general data and biochemical indexes, the detection of abnormal lipid metabolism and thyroid nodules were analyzed, and the risk factors of thyroid nodules in people with abnormal lipid metabolism were analyzed by logistic regression.Results:Two thousand and fifty-nine residents were included in this study (1 049 males and 1 010 females). The total detection rate of thyroid nodules was 23.17% (477/2 059). The detection rate of thyroid nodules in people with abnormal lipid metabolism [34.16%(151/442)] was significantly higher than that in people with normal lipid metabolism [20.16% (326/1 617) , P < 0.01], and the detection rate of thyroid nodules of women [43.37% (85/196) ] was higher than that of men [26.83% (66/246) , P < 0.01]. Among the people with abnormal lipid metabolism, the highest detection rate of thyroid nodules was in mixed hyperlipidemia [57.14% (16/28)], followed by hypertriglyceridemia [34.59% (92/266)]. The detection rates of thyroid nodules in the groups with elevated total cholesterol (TC) and low density lipoprotein cholesterol (LDL-C) levels [35.16% (32/91), 34.85% (23/66)] were higher than those in the marginal elevated group [27.04%(86/318), 30.42% (73/240)] and the normal groups [21.76% (359/1 650), 21.73% (381/1 753), P < 0.05]. The results of logistic regression analysis showed that the risk factors of thyroid nodules in people with abnormal lipid metabolism were increased age, elevated fasting blood glucose (FPG), elevated blood glucose 2 hours (2 h PG) after oral glucose tolerance test (OGTT) load and elevated glycosylated hemoglobin [HbA1c, odds ratio ( OR)=1.065, 1.387, 1.866, 1.384, P < 0.05]. Conclusions:The prevalence of TN is higher in populations with abnormal lipid metabolism. The control of blood sugar and blood lipid levels may play a role in the prevention of thyroid nodules.

Objective:To explore the changes and clinical significance of inflammatory indices of urogenic sepsis with different severity.Methods:A retrospective case-control study was used to analyze the clinical data of 71 patients with urogenic sepsis admitted to 940th Hospital of PLA Joint Logistics Support Force from January 2010 to April 2018, including 34 males and 37 females, aged 39-96 years [(63.1±18.3)years]. The patients were divided into three groups according to the clinical diagnostic criteria for septic shock and sepsis according to the 2014 edition of the Chinese Urology Surgical Guidelines for Diagnosis and Treatment: 21 cases in sepsis group [sequential organ failure assessment (SOFA) score of 3.0 (2.0, 3.0)points], 21 cases in severe sepsis group [SOFA score of 9.0 (6.0, 11.0)points], and 29 cases in septic shock group [SOFA score of 15.0 (14.0, 16.0)points]. Spearman correlation analysis was used to analyze the correlation of inflammatory indicators with SOFA, including white blood cell count, percentage of neutrophils, C-reactive protein, interleukin-6, procalcitonin, fibrinogen, D-dimer, and platelet. Multiple linear regression analysis and stepwise regression weighted analysis were performed to analyze the relation between inflammatory indicators and sepsis severity. Levels of each inflammatory indicator was detected and compared among the groups.Results:① Spearman correlation analysis: percentage of neutrophils, D-dimer, interleukin-6, procalcitonin and SOFA scores were significantly positively correlated, with the r s value of 0.738, 0.712, 0.31, 0.795, respectively ( P<0.01); platelet and SOFA scores were significantly negatively correlated, with the r s value of -0.661 ( P<0.01). ② Multiple linear regression analysis: percentage of neutrophils, platelet, D-dimer, procalcitonin and SOFA score were significantly correlated ( P<0.01); Stepwise regression weighted analysis suggested that the model linear relationship and fit was good. ③ Inflammatory index comparison: percentage of neutrophils in sepsis group, severe sepsis group and septic shock group was 82.30 (76.25, 88.45), 90.50 (86.55, 93.85), 95.10 (92.05, 97.95), respectively; level of platelet was 183.01 (144.50, 246.50)×10 9/L, 149.11 (81.04, 207.00)×10 9/L, 81.26 (50.01, 93.50)×10 9/L, respectively; level of D-dimer was 0.98 (0.71, 1.74)mg/L, 3.45 (1.79, 5.56)mg/L, 7.19 (4.26, 11.63)mg/L, respectively; level of procalcitonin was 0.55 (0.21, 1.09)ng/ml, 5.45 (3.74, 11.80)ng/ml, 17.68 (13.97, 26.75)ng/ml, respectively. There were significant differences in above indicators among the groups ( P<0.05). Conclusions:The serum levels of procalcitonin, percentage of neutrophils, D-dimer and platelet are positively correlated with the severity of urogenic sepsis. While combined detection of those indicators can better predict the severity of the sepsis.

To evaluate the impact of admission diagnosis on seizure outcome in patients with autoimmune epilepsy (AE).
 Methods: We conducted a retrospective study on 40 AE patients at Department of Neurology, Xiangya Hospital, Central South University from Jan. 1st, 2017 to Nov. 21st, 2018. According to their admission diagnosis, these patients were further assigned into 2 groups: An AE diagnosed group and an AE undiagnosed group. Demographic data, clinical characteristics, cerebrospinal fluid (CSF), electroencephalogram, and brain imaging were compared between the 2 groups. Favorable seizure outcome was defined as >50% reduction of seizure frequency at the first follow-up. The impact of admission diagnosis on seizure outcome of AE patients was analyzed.
 Results: The ages of 40 AE patients were (33.23±16.41) years, comprising 19 males and 21 females. No significant difference was found between the AE diagnosed group and the AE undiagnosed group in gender, age, central nervous system-specific Ab profile, CSF, and brain imaging. Duration of symptom onset to Ab detection was significantly longer in the AE undiagnosed group than that in the AE diagnosed group (P<0.01). Duration of symptom onset to immunotherapy was also significantly longer in the AE undiagnosed group than that in the AE diagnosed group (P<0.001). There were more patients with favorable seizure outcome in the AE diagnosed group than the AE undiagnosed group (P<0.05).
 Conclusion: Admission diagnosis for patients with AE is associated with seizure outcome after immunotherapy. For adult-onset epilepsy or epilepsy with unknown etiology, the diagnosis of AE should be considered. Early diagnosis of AE and prompt initiation of immunosuppressive treatment are crucial to increase the likelihood of achieving favorable seizure outcome.
Adolescent ; Adult ; Electroencephalography ; Epilepsy ; Female ; Humans ; Immunotherapy ; Male ; Middle Aged ; Retrospective Studies ; Seizures ; Young Adult

The gut microbiota plays an important role in the development and progression of colorectal cancer (CRC). To learn more about the dysbiosis of carcinogenesis, we assessed alterations in gut microbiota in patients with CRC. A total of 23 subjects were enrolled in this study: 9 had CRC (CRC group) and 14 had normal colons (normal group). The microbiome of the mucosal-luminal interface of each subject was sampled and analyzed using 16S rRNA gene amplicon sequencing. We also used Phylogenetic Investigation of Communities by Reconstruction of Unobserved States (PICRUSt) to predict microbial functional profiles. The microbial composition of the mucosal lumen differed between the groups, and the presence of specific bacteria may serve as a potential biomarker for colorectal carcinogenesis. We identified a significant reduction in Eubacterium, which is a butyrate-producing genera of bacteria, and a significant increase in Devosia in the gut microbiota of CRC patients. Different levels of gut microflora in healthy and CRC samples were identified. The observed abundance of bacterial species belonging to Eubacterium and Devosia may serve as a promising biomarker for the early detection of CRC.

Objective To study the role of autophagy in proliferation,migration and tube formation of retinal vascular endothelial cells (RVECs) under the condition of hypoxia in vitro.Methods Mouse RVECs were isolated from 50 C57BL/6J mice primarily cultured using explant culture method,and the cells were identified by immunofluorescence of CD34.Well cultured RVECs were divided into normal control group,hypoxia control group and hypoxia+ 3-methyladenine (3-MA) group.The cells in the normal control group were cultured in the normal environment for 24 hours,and the cells in the hypoxia control group were cultured 1％ O2 environment for 24 hours,and the cells in the hypoxia+3-MA group were pretreated with 5 mmol/L 3-MA for 4 hours and then exposed to 1％ O2 environment for 24 hours.Microtubule-related protein 1 light chain 3 (LC3-Ⅱ/LC3-Ⅰ) and Beclin-1 protein contents in the cells were detected by Western blot analysis;the ultrastructure of autophagosome was examined under the transmission electron microscope.The proliferation,migration and tube formation of the cells were detected by Click-iTEdU kit,scratch assay and matrigel,respectively.Results Primarily culture cells grew well with the cobblestone-like appearance 5-7 days after culture and showed positive response for CD34.The autophagosome number was increased in the hypoxia control group compared with the hypoxia+3-MA group and normal control group.The LC3-Ⅱ/LC3-Ⅰ ratio was 0.243 ± 0.030,0.658 ± 0.032 and 0.405 ± 0.095;the relative expression of Beclin-1 protein in the cells was 0.260±0.040,0.650±0.071 and 0.461±0.089;the proliferation rate of the cells was (45.93± 6.39) ％,(22.74± 2.35) ％ and (24.12 ± 3.59) ％;the scratch healing rate of the cells was (36.02 ± 5.84) ％,(57.26±11.98) ％ and (18.16±9.73) ％;the number of tube formation was 29.20±6.10,41.40±4.04 and 22.00± 2.92 in the normal control group,hypoxia control group and hypoxia + 3-MA group,respectively,with significant differences among the groups (F =35.86,23.53,34.28,21.12,23.27;all at P＜0.01).Compared with the normal control group,the ratio of LC3-Ⅱ/LC3-Ⅰ and the expression of Beclin-1 protein,scratch healing rate and the number of tube formation were significantly increased,and the proliferation rate was significantly reduced in the hypoxia control group (all at P ＜ 0.05).Compared with the hypoxia control group,the ratio of LC3-Ⅱ/LC3-Ⅰ and the expression of Beclin-1 protein,scratch healing rate and the number of tube formation were significantly decreased in the hypoxia + 3-MA group.Conclusions Hypoxia environment activates autophagy of mouse RVECs,which enhances the migration and tube formation abilities of the cells.3-MA inhibits the angiogenesis abilities of mouse RVECs.

To study the biological function of DNAH2 (Homo sapiens dynein, axonemal, heavy chain 2) gene, we constructed human stable U2OS cell line of DNAH2 gene knockout through CRISPR/Cas9n double nick system. The A, B sgRNAs (Single guide RNA) and complementary strands were designed and synthesized. The double-stranded structures were formed during annealing, and connected with BbsⅠ cohesive ends-containing pX462 linear vector to construct the recombinant eukaryotic expression plasmids, including pX462-DNAH2-A and pX462-DNAH2-B. After the co-transfection of the two plasmids into U2OS cells, the addition of puromycin and limiting dilution method were used to obtain positive monoclonal cell line. Western blotting assay was then performed to detect the expression of DNAH2 protein, and PCR-sequencing technology was finally utilized to analyze the mutation feature. The results showed that A, B sgRNAs duplex was successfully inserted into pX462 vector, and DNAH2 protein was not expressed and DNAH2 gene suffered from the frame-shift mutation in U2OS-DNAH2-KO monoclonal cell line. These demonstrated that DNAH2 knockout U2OS stable cell line was successfully constructed through CRISPR/Cas9n double nick system, which providing a useful tool for the study of DNAH2 gene.

Objective To establish a homothermal and fast detecting method on pathogenic bacteria by combining recombinase-aid amplification (RAA) with molecular beacon.Methods The establishment of the methodology.Staphylococcus aureus specific primers were designed from the relative region of the staphylococcal protein A (SPA).Asymmetry amplification was optimized by adjusting the primer concentration ratios.The results of amplification and hybridization were visualized and analyzed by agarose gel electrophoresis and fluorescence detection.The sensitivity was identified by detecting dilute positive plasmids.And the specificity was determined using RAA method by detecting 72 pathogenic bacteria,including Staphylococcus aureus and other Staphylococcus spp.from the Department of Clinical Laboratory of Daping Hospital in December 2016.Besides,the Kappa analysis and the clinical diagnosis efficiency were investigated by analyzing 39 extra strains in the laboratory in December 2016.Results When the concentration ratio of restrictive and non-restrictive primer was 1:20,the yield efficiency of single-stranded DNA (ssDNA) reached the peak.And as for the hybridization efficiency,the asymmetry amplification was higher than symmetry amplification.Twenty copies/μl was proposed as the limits of detection by testing dilute plasmids.And the RAA hybridization method could distinguish Staphylococcus aureus with other Staphylococcus spp.Comparing with traditional detection methods with a Kappa index of 0.860,this method shows a good consistency.By analyzing the 111 bacteria,the sensitivity of the method is 92.5％ (37/40),the specificity is 97.2％ (69/71),the positive predictive value is 94.9％ (37/39),the negative predictive value is 95.8％ (69/72),the positive likelihood radio is 33.04,the negative likelihood radio is 0.077,the Youden index is 0.897 and the Kappa index is 0.902.Conclusion Through the combination of asymmetry recombinase-aid amplification optimization and molecular beacon probe,a new method of detecting bacteria DNA with RAA hybridization technique is established,providing the foundation for its clinical application.

Objective To understand the epidemiological characteristics of norovirus detection based on the diarrhea syndromic surveillance in Pudong ,Shanghai .Methods Diarrhea syndromic surveillance program was conducted in outpatient and emergency departments of 12 sentinel hospitals during 2012 -2016 .The clinical and epidemiological data of diarrheal patients were collected .The fecal specimens were also sampled . The detections for norovirus by polymerase chain reaction and gene sequencing were performed .Chi-square test or Fisher exact probability test was used to compare the detection rate .Binary logistic regression was used to explore the impact factors of norovirus infection among diarrheal patients . Results The detective rate of norovirus among diarrheal patients was 21 .59％ ,peaking from October to next March .Among all the age groups ,the detection rate was highest among patients with 25 - 64 years old .The patients with more severe diarrhea symptoms (> 5 times a day) were more likely to be infected with norovirus than those with diarrhea 3 - 5 times a day (χ2 = 21 .167 ,P< 0 .01) .Vomiting was also an indicator of norovirus infection .Patients presented with vomiting had a higher norovirus detection rate (χ2 = 198 .543 , P < 0 .01) . Norovirus G Ⅱ was the predominant genotype .Conclusions The recent epidemic of norovirus infection in diarrheal patients in Pudong new district has an apparent seasonality peaked from October to next March .Adult ,patients with vomiting and more severe diarrhea symptoms are at risk of norovirus infection .The long-term surveillance is critical for the norovirus infection control .

Objective To explore the effects of luteolin on cognition function in pentylenetetrazol (PTZ)-induced epileptic rats and related mechanism.Methods Fifty male SD rats were randomly divided into a normal control group(n=8), a model group(n=12), and groups of 25, 50 mg/kg luteolin(both ofn=11), as well as 100 mg/kg luteolin group(n=8). Those rats were given different doses of luteolin (25, 50 and 100 mg/kg, daily, intragastric administration) for 36 consecutive days. Similarly, rats of the normal control group and the model group were given 0.5% sodium carboxymethyl cellulose suspension liquid via intragastric administration. Thirty minutes later, a model of epilepsy was induced using PTZ (40 mg/kg, daily) via intraperitoneal injection except the control group. Learning and memory of rats were evaluated by Morris water maze and novel objective recognition trials(including escape latency and recognition index). The levels of CaM and CaMPK were determined by ELISA methods, and expression of Ras proteins in the hippocampus were detected by Western Blot.Results Compared with the model group, luteolin treatment groups significantly shorten the escape latency(28.51 ± 3.84 s, 19.77 ± 5.41 s, 14.86 ± 2.76 svs. 37.08 ± 5.18 s) in the Morris water maze, and increased recognition index(18.77% ± 2.02%, 25.06% ± 4.32%, 31.92% ± 2.65%vs. 13.87% ± 2.14%) in the novel objection trial(P<0.05 orP<0.01). Meanwhile, CaM(140.33 ± 13.52 ng/L, 124.26 ± 9.97 ng/L, 113.52 ± 11.57 ng/Lvs. 158.36 ± 10.68 ng/L) and CaMPK(8.25 ± 1.37 ng/ml, 7.69 ± 0.84 ng/ml, 6.74 ± 0.93 ng/mlvs. 9.87 ± 1.02 ng/ml) were significantly decreased(P<0.05 orP<0.01). What’s more, the expression of Ras proteins(0.99 ± 0.08, 0.76 ± 0.07, 0.52 ± 0.07vs. 1.58 ± 0.12) was obviously decreased compared with the model group(P<0.05 orP<0.01).Conclusion Luteolin could effectively improve the cognition dysfunction of epileptic rats, and the mechanism might be relevant to regulate the CaM-CaMPK signaling pathway via down-regulation of CaM, CaMPK, as well as Ras protein.