Objective:To screen the host interaction proteins of the severe fever with thrombocytopenia syndrome virus(SFTSV)nonstructural protein(NSs)by immunoprecipitation combined with mass spectrometry analysis,to discuss the functions,subcellular localization,and biological pathways of these interaction proteins,and to provide the basis for clarifying the replication and pathogenic mechanism of SFTSV.Methods:The eukaryotic expression vectors pSFTSV-NSs-Flag(experimental group)and Flag-CMV-3(negative group)were transfected into the human embryonic kidney 293T cells,and contorl group(no treatment)was set up.The lysates of the cells in various groups were collected,and the expression and localization of SFTSV NSs in the host cells were verified by indirect immunofluorescence and Western blotting methods.The protein lysates were treated with protein A/G and immunoprecipitation was used to enrich host proteins binding to NSs.The captured interaction proteins were initially analyzed by silver staining and Coomassie brilliant blue staining to observe the differential protein bands in various groups;liquid chromatography-tandem mass spectrometry was used to obtain the information of protein sequences;the reliable proteins were retained and searched by UniProt database;Gene Ontology(GO)functional enrichment analysis,IPR,eukaryotic orthologous groups(KOGs)functional annotation,Kyoto Encyclopedia of Genes and Genomes(KEGG)signaling pathway enrichment analysis,subcellular localization,and transcription factor(TF)functional annotation were used to determine the subcellular structure,gene functions,and biological processes of the interaction proteins.Results:The immunofluorescence results showed that the SFTSV NSs expressed a single specific band at relative molecular mass 33 000 and was localized in the cytoplasm in a granular inclusion body-like manner.The silver staining and Coomassie brilliant blue staining results showed there were significant differential protein bands between experimental group and negative group.The mass spectrometry results identified 46 potential interaction proteins.The GO functional enrichment analysis,KOGs functional annotation,and KEGG signaling pathway enrichment analysis results showed that the biological pathways related to viral translation,cellular metabolism,and protein transport were enriched with a considerable number of proteins.Eight annotated proteins had intermediate filament domains.The highest percentage of subcellular localization was cytoplasmic proteins,consistent with the NSs localization site.The TF functional annotation analysis results showed one protein from the NF-Y family.Conclusion:The interaction proteins play roles in assisting the proper protein folding,participating in the cribosome translation,and forming the cytoskeleton,which may be involved in antiviral replication.These proteins can be used as candidate proteins for further study on the replication mechanism of SFTSV.

ObjectiveTo evaluate the clinical efficacy of modified Houpo Dahuangtang in moderate and severe acute respiratory distress syndrome （ARDS） patients with phlegm-heat accumulation，and monitor the pulmonary ventilation changes of patients before and after treatment by electrical impedance tomography（EIT）. MethodThe 62 cases of moderate and severe ARDS patients with phlegm-heat accumulation who required mechanical ventilation in the department of intensive care unit （ICU） in Chongqing Hospital of Traditional Chinese Medicine from September 2021 to June 2022 were selected，and divided into an experimental group（31 cases）and a control group（31 cases）using a random number table. On the basis of regular Western medicine treatment，the experimental group received modified Houpo Dahuangtang and the control group received warm water by a nasogastric tube for seven days. The changes in the clinical efficacy of traditional Chinese medicine（TCM），the oxygenation index［arterial oxygen partial pressure （PaO₂）/fractional inspired oxygen（FiO₂），P/F］，lactic acid（Lac），acute physiology and chronic health evaluation Ⅱ（APACHE Ⅱ） score，compliance，plateau pressure，gas distribution parameters monitored by EIT（Z1，Z2，Z3 and Z4），inflammatory factors［interleukin-6 （IL-6），IL-10， tumor necrosis factor-α （TNF-α） and C-reactive protein （CRP）］ of both groups before and after treatment were recorded. Besides， the mechanical ventilation time， length of stay in ICU， 28-day mortality and incidence of adverse reactions（delirium，abdominal pain and diarrhea）in the two groups were also observed. ResultThere was no significant difference in the baseline indexes of patients in the two groups，and thus the two groups were comparable. After treatment for one week， the total effective rate for TCM syndromes in the experimental group was 90.30%（28/31）， higher than the 67.74%（21/31）in the control group（Z=-2.415，P<0.05）.Compared with the same group before treatment， the plateau pressure and Lac decreased （P<0.01）and the compliance and P/F increased （P<0.01） in experimental group， while the Lac decreased （P<0.05）and the P/F increased （P<0.05）， and the compliance and plateau pressure did not change significantly in the control group. After treatment，the plateau pressure and inflammatory factors in the experimental group were lower than those in the control group（P<0.05）， but the compliance and P/F in the experimental group were higher than those in the control group（P<0.05）， and the gas distribution parameters Z1，Z2，Z3，Z4，Z1+Z2，and Z3+Z4 monitored by EIT in the experimental group were all higher than those in the control group （P<0.05）. There was no significant difference in mechanical ventilation time， ICU hospitalization time， 28-day mortality， delirium， abdominal pain， diarrhea and other adverse reactions between the two groups. ConclusionModified Houpo Dahuangtang can significantly improve the P/F，pulmonary ventilation in gravity-dependent regions and pulmonary compliance，reduce the release of inflammatory factors in moderate and severe ARDS patients. Compared with conventional methods，EIT can timely monitor the pulmonary ventilation changes in ARDS patients，which suggests its clinical feasibility.

Objective:To analyze the efficacy and safety of nalbuphine for analgesia in patients with non-mechanical ventilation in intensive care unit (ICU).Methods:From December 2018 to August 2021, a multicenter randomized controlled clinical study was conducted to select non-mechanical ventilation patients with analgesic needs admitted to ICU of four hospitals in Henan Province and Guizhou Province. Patients were randomly assigned to nalbuphine group and fentanyl group. The nalbuphine group was given continuous infusion of nalbuphine [0.05~0.20 mg/(kg·h)], and the fentanyl group was given continuous infusion of fentanyl [0.5~2.0 μg/(kg·h)]. The analgesic target was critical-care pain observation tool (CPOT) score<2. The observation time was 48 hours. The primary endpoint was CPOT score, the secondary endpoints were Richmond agitation-sedation score (RASS), ICU length of stay, adverse events, and proportion of mechanical ventilation. The quantitative data of the two groups were compared by t test or Mann-Whitney U test. The enumeration data were compared by chi square test or Fisher exact probability method. The data at different time points between groups were compared by repeated measures analysis of variance. Results:A total of 210 patients were enrolled, including 105 patients in the nalbuphine group and 105 patients in the fentanyl group. There was no significant difference in baseline data between the two groups (all P>0.05). There was no significant difference in CPOT score between nalbuphine group and fentanyl group at each time point after medication ( P>0.05), the CPOT score of both groups at each time point after medication was significantly lower than that before medication, and the analgesic target could be achieved and maintained 2 hours after medication. There was no significant difference in RASS between the two groups at each time point after medication ( P>0.05), which was significantly lower than that before medication, and the target sedative effect was achieved 2 hours after medication. There was no significant difference in ICU length of stay between nalbuphine group and fentanyl group [5.0(4.0,7.5) d vs. 5.0(4.0,8.0) d, P=0.504]. The incidence of delirium, nausea and vomiting, abdominal distension, pruritus, vertigo and other adverse events in the nalbuphine group was lower than that in the fentanyl group (all P<0.05). There was no significant difference in the incidence of other adverse events such as deep sedation, hypotension and bradycardia between the two groups (all P>0.05). The incidence of respiratory depression in nalbuphine group was not significantly different from that in fentanyl group ( P>0.05), but the proportion of mechanical ventilation was significantly lower than that in the fentanyl group [1.9% (2/105) vs. 8.6%(9/105), P=0.030]. Conclusions:Nalbuphine could be used for analgesia in ICU patients with non-mechanical ventilation. The target analgesic effect could be achieved within 2 hours, and it had a certain sedative effect with a low incidence of adverse reactions.

Objective:There is no standard method for esophageal remnant gastric reconstruction for proximal gastrectomy. Reflux esophagitis caused by esophagogastrostomy remains a difficult surgical problem. To report the preliminary surgical results of novel esophagus-conical remnant gastric side overlap anastomosis (CGEO) , with particular emphasis on postoperative esophageal reflux.Methods:In June 2022, we developed a novel CGEO for laparoscopic proximal gastrectomy on two patients with Siewert type II esophagogastric junction adenocarcinoma. Surgical procedures for CGEO: (1) Laparoscopic proximal gastrectomy and preparation of conically shaped gastric remnant; (2) Determining anastomotic site of residual stomach and esophagus; (3) Side-to-side anastomosis of right esophageal wall to anterior of conical gastric remnant; (4) Valvuloplasty of esophageal stump.Results:Case 1 was a 71-year-old man with an operation time of 305 minutes and was successfully discharged from the hospital on the 9th day after surgery, and the postoperative pathology was T3N0M0. Case 2 was an 82-year-old man with an operation time of 325 minutes. He was discharged on the 10th day after surgery. In both cases, only mild esophageal mucosal changes were seen in gastroscopy, there were no obvious symptoms of esophageal reflux. There was also no significant weight change at half a year after operation.Conclusion:CGEO is moderately safe in radical surgery for proximal gastric cancer, and may have a preventive effect on the occurrence of postoperative esophageal reflux, but long-term results need to be confirmed by further studies with follow-up.

Objective:There is no standard method for esophageal remnant gastric reconstruction for proximal gastrectomy. Reflux esophagitis caused by esophagogastrostomy remains a difficult surgical problem. To report the preliminary surgical results of novel esophagus-conical remnant gastric side overlap anastomosis (CGEO) , with particular emphasis on postoperative esophageal reflux.Methods:In June 2022, we developed a novel CGEO for laparoscopic proximal gastrectomy on two patients with Siewert type II esophagogastric junction adenocarcinoma. Surgical procedures for CGEO: (1) Laparoscopic proximal gastrectomy and preparation of conically shaped gastric remnant; (2) Determining anastomotic site of residual stomach and esophagus; (3) Side-to-side anastomosis of right esophageal wall to anterior of conical gastric remnant; (4) Valvuloplasty of esophageal stump.Results:Case 1 was a 71-year-old man with an operation time of 305 minutes and was successfully discharged from the hospital on the 9th day after surgery, and the postoperative pathology was T3N0M0. Case 2 was an 82-year-old man with an operation time of 325 minutes. He was discharged on the 10th day after surgery. In both cases, only mild esophageal mucosal changes were seen in gastroscopy, there were no obvious symptoms of esophageal reflux. There was also no significant weight change at half a year after operation.Conclusion:CGEO is moderately safe in radical surgery for proximal gastric cancer, and may have a preventive effect on the occurrence of postoperative esophageal reflux, but long-term results need to be confirmed by further studies with follow-up.

Objective : To screen the RNA binding proteins interacting with NBV nonstructural protein σNS in host cells and to analyze their bioinformatics functions. Methods : In this study , the eukaryotic expression vector pEF⁃HA⁃MB⁃S3 of NBV σ NS was constructed and transfected into HEK293T cells after verification. After RNase A treatment , the obtained protein lysate was enriched by immunoprecipitation to enrich σNS binding proteins , identified and analyzed by LC⁃MS/MS mass spectrometry , and the properties and specific functions of proteins were discovered with the help of related Bioinformatics tools. Results : In this study , 32 candidate RNA binding proteins interacting with NBV σNS proteins were successfully screened , and the results of bioanalysis showed that these proteins were mainly located in cytoplasm and nucleus , and were mainly involved in biological processes such as cell metabolism , biological regulation , virus translation and transcription. Conclusion This study preliminarily analyzes the function of RNA binding proteins interacting with NBV σNS , which lays a foundation for further study on the mechanism of σNS protein in NBV life cycle.

OBJECTIVES: In this study, we explored how adiponectin mediated urotensin II (UII)‍-induced tumor necrosis factor-‍α (TNF-‍α) and α‍-smooth muscle actin (α‍-SMA) expression and ensuing intracellular signaling pathways in adventitial fibroblasts (AFs). METHODS: Growth-arrested AFs and rat tunica adventitia of vessels were incubated with UII and inhibitors of signal transduction pathways for 1‍‒‍24 h. The cells were then harvested for TNF-α receptor (TNF-‍α-R) messenger RNA (mRNA) and TNF-‍α protein expression determination by reverse transcription-polymerase chain reaction (RT-PCR) and enzyme-linked immunosorbent assay (ELISA), respectively. Adiponectin and adiponectin receptor (adipoR) expression was measured by RT-PCR, quantitative real-time PCR (qPCR), immunohistochemical analysis, and cell counting kit-8 (CCK-8) cell proliferation experiments. We then quantified TNF-α and α-SMA mRNA and protein expression levels by qPCR and immunofluorescence (IF) staining. RNA interference (RNAi) was used to explore the function of the adipoR genes. To investigate the signaling pathway, we applied western blotting (WB) to examine phosphorylation of adenosine 5'-monophosphate (AMP)‍-activated protein kinase (AMPK). In vivo, an adiponectin (APN)‍-knockout (APN-KO) mouse model mimicking adventitial inflammation was generated to measure TNF-α and α‍-SMA expression by application of qPCR and IF, with the goal of gaining a comprehensive atlas of adiponectin in vascular remodeling. RESULTS: In both cells and tissues, UII promoted TNF-α protein and TNF-α-R secretion in a dose- and time-dependent manner via Rho/protein kinase C (PKC) pathway. We detected marked expression of adipoR1, T-cadherin, and calreticulin as well as a moderate presence of adipoR2 in AFs, while no adiponectin was observed. Globular adiponectin (gAd) fostered the growth of AFs, and acted in concert with UII to induce α-SMA and TNF-α through the adipoR1/T-cadherin/calreticulin/AMPK pathway. In AFs, gAd and UII synergistically induced AMPK phosphorylation. In the adventitial inflammation model, APN deficiency up-regulated the expression of α-SMA, UII receptor (UT), and UII while inhibiting TNF-‍α expression. CONCLUSIONS From the results of our study, we can speculate that UII induces TNF‍-‍α protein and TNF-‍α‍-R secretion in AFs and rat tunica adventitia of vessels via the Rho and PKC signal transduction pathways. Thus, it is plausible that adiponectin is a major player in adventitial progression and could serve as a novel therapeutic target for cardiovascular disease administration.
Mice ; Rats ; Animals ; Adventitia/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Calreticulin/metabolism* ; Vascular Remodeling ; AMP-Activated Protein Kinases/metabolism* ; Cells, Cultured ; RNA, Messenger/genetics* ; Inflammation

Background::Sepsis is a serious disease caused by infection. Aminophylline has anti-asthma and anti-inflammatory effects. We aimed to explore the safety and effect of aminophylline in sepsis.Methods::We conducted a clinical randomized controlled trial involving 100 patients diagnosed with sepsis within 48 h after intensive care unit (ICU) admission in two sites. All patients were randomized in a 1:1 ratio to receive standard therapy with or without aminophylline. The primary clinical outcome was all-cause mortality at 28 days.Results::From September 27, 2018 to February 12, 2020, we screened 277 septic patients and eventually enrolled 100 patients, with 50 assigned to the aminophylline group and 50 to the usual-care group. At 28 days, 7 of 50 patients (14.0%) in the aminophylline group had died, compared with 16 of 50 (32.0%) in the usual-care group ( P = 0.032). Cox regression showed that the aminophylline group had a lower hazard of death (hazard ratio = 0.312, 95% confidence interval: 0.129-0.753). Compared with the usual-care group, patients in the aminophylline group had a longer survival time ( P = 0.039 by the log-rank test). The effects of aminophylline on vasopressor dose, oxygenation index, and sequential organ failure assessment score were time-dependent with treatment. There were no significant differences in total hospitalization days, ICU hospitalization days, and rates of serious adverse events (all P > 0.05). No adverse events were observed in the trial. Conclusions::Aminophylline as an adjunct therapy could significantly reduce the risk of death and prolong the survival time of patients with sepsis.Trial registration::ChiCTR.org.cn, ChiCTR1800019173.

Objective:To compare and analyze the differences in the setup accuracy of different immobilization method in breast cancer radiotherapy after breast-conserving surgery.Methods:A retrospective study was conducted on 60 patients who received radiotherapy after breast-conserving surgery from January to August, 2021. These patients were divided into two groups. One group consisted of 30 cases who were immobilized using a modified body thermoplastic membrane combined with a multifunction body board during the breast cancer radiotherapy and was called the modified body thermoplastic membrane group. The other group comprised 30 cases immobilized using a vacuum cushion during breast cancer radiotherapy and was referred to as the vacuum cushion group. The setup errors, 3D vector errors, the proportion of errors of > 5 mm, and the dosimetric differences in the planning target volume (PTV) and the clinical target volume (CTV) before and after simulated treatment bed moving (including the PTV_ V100, PTV_ V95, and CTV_ V95 before simulated treatment bed moving and the PTV_ V100 S, PTV_ V95 S, and CTV_ V95 S after simulated treatment bed moving) were compared between two groups. Moreover, for the modified body thermoplastic membrane group, the changes in the average setup errors at different radiotherapy stages were also analyzed. Results:A total of 369 cone-beam CT scans were conducted for 60 patients, including 195 CT scans for the modified body thermoplastic membrane group and 174 CT scans for the vacuum cushion group. The setup errors in the x, y, and z directions (right-left, anterior-posterior, and superior-inferior, respectively) of the modified body thermoplastic membrane group were (2.59±1.98) mm, (2.38±2.04) mm, and (1.45±1.16) mm, respectively, while those of the other group were (2.24±1.63) mm, (2.78±2.17) mm, and (2.70±1.88) mm, respectively. The 3D vector errors of both groups were (4.32±2.28) mm and (5.13±2.14) mm, respectively. Therefore, the setup error in direction z and the 3D vector error of the modified body thermoplastic membrane group were less than those of the vacuum cushion group ( t = -7.77, -3.41, P<0.05). Moreover, the proportion of setup errors of > 5 mm in the x direction of the vacuum cushion group was lower than that of the modified body thermoplastic membrane group ( χ2 = 7.13, P<0.05), while such proportion in the z direction of the modified body thermoplastic membrane group was lower than that of the vacuum cushion group ( χ2= 5.90, P<0.05). After the simulated treatment bed moving, the PTV_ V100 S of the modified body thermoplastic membrane group was better than that of the vacuum cushion group ( t = 2.47, P < 0.05). Furthermore, for the modified body thermoplastic membrane group, the setup errors in the x direction in the first week were higher than those in the 2-3 weeks and 4-5 weeks ( P<0.05). Conclusions:The modified body thermoplastic membrane combined with a multifunction body board yield better immobilization effects than a vacuum cushion. However, it produces high setup errors in the x direction in the first week of the radiotherapy, to which special attention should be paid.

Objective To investigate the pollution level of 9,10-anthraquinone in teas sold in Wuhan, and to assess the dietary exposure of 9,10-anthraquinone ingested through tea. Methods The content of 9,10-anthraquinone in teas collected from local tea markets in Wuhan was analyzed by GC-MS/MS. Results 9,10-anthraquinone was detected in all 36 tea samples with a concentration ranging from 0.0080 mg/kg to 0.137 mg/kg. The standard limit for 9,10-anthraquinone has not yet been set in China. Referring to EU standards, the total over-standard rate was 86.11% (31/36). Generally, the concentration of residual 9,10-anthraquinone was higher in highly fermented teas. The highest average concentration of residual 9,10-anthraquinone was found in fully/post-fermented tea (0.0762 mg/kg), and the over-standard rate was 100%. The second highest was in semi-fermented tea (0.0452 mg/kg), and the over-standard rate was 86.7%. The concentration of 9,10-anthraquinone in non-fermented tea was 0.0262 mg/kg, and the over-standard rate was 42.9%. According to people^’s tea-drinking habits, tea samples were brewed with boiling water（1:50）for 5 minutes, and the concentration of 9,10-anthraquinone in tea soup was 0.0004 mg/kg～0.01 mg/kg, with an average leaching rate of 7.2%. The average daily intake exposure of tea for an adult was 0.0551μg/kg bw/day. Conclusion There was a certain degree of 9,10-anthraquinone pollution in the teas sold in Wuhan, and the rate exceeding the standard was high. Since the acceptable daily intake (ADI) for 9,10-anthraquinone has not been established, it is impossible to assess the health risks of 9,10-anthraquinone ingested through tea.