Objective To compare the clinical efficacy of different doses of ultrafine extracted granule preparation (EGP) and traditional herbal decoction (THD) of Li Zhong Tang in treatment of epigastric pain. Methods Sixty cases of epigastric pain patients in accordance with the diagnostic criteria of deficiency and cold pattern of spleen and stomach in TCM were randomly divided into THD group, 1/3 dose group and 1/5 dose group, and were given THD, 1/3 dose of ultrafine EGP and 1/5 dose of ultrafine EGP, respectively. The clinical efficacy of the three groups after one course of medication was comparatively analyzed. Results There were no significant differences in age, course of disease, symptom score before treatment, epigastric pain efficacy and syndrome curative effect among the three groups, the differences had no statistical significance (P>0.05). The severity, frequency and duration of epigastric pain were all reduced in the three groups, with significant differences between before and after treatment (P<0.05). Conclusion There are no significant differences in the clinical efficacy on epigastric pain among THD, 1/3 dose of ultrafine EGP and 1/5 dose of ultrafine EGP. In addition, the effect of 1/3 dose group is very close to the THD group.

OBJECTIVE: To prepare clomipramine hydrochloride sustained release tablet and study its in vitro release rate.METHODS: Orthogonal experiment was carried out to optimize the formulation and the preparation was prepared taking by the formula dosage of HPMC,lactose and amylum pregelatinisatum as factors and the in vitro release rate as index.The in vitro drug release rate was investigated as well.RESULTS: The optimal formulation was as follows: HPMC 45 mg,lactose 35 mg,and amylum pregelatinisatum 40 mg.The preparation prepared in the optimal formulation had a sustained release of 24 h and the release behavior of the tablets followed the zero order equation.CONCLUSION: The formula of the sustained release tablets is reasonable and which had satisfactory sustained release efficacy.

Objective:To investigate the monocyte to high-density lipoprotein cholesterol (HDL-C) ratio (MHR) for the predictive value of early neurological deterioration (END) and poor outcome in patients with acute anterior circulation ischemic stroke (AACIS).Methods:Patients with AACIS admitted to Henan Provincial People's Hospital from January 2021 to January 2022 were included retrospectively. END was defined as the National Institutes of Health Stroke Scale (NIHSS) score within 7 d of onset increase ≥2 compred with baseline or the increase of motor function score ≥1. The patients were divided into END group and non-END group according to the presence or absence of END. The patients were also divided into good outcome group (0-2 points) and poor outcome group (3-6 points) according to the modified Rankin Scale score 3 months after onset. Multivariate logistic regression analysis was used to determine the independent risk factors for END and poor outcome, and the predictive value of MHR for END and poor outcome was evaluated by receiver operating characteristic (ROC) curve. Results:A total of 522 patients were enrolled, including 338 male (64.8%), aged 61.99±11.39 years old. One hundred and five patients (20.1%) had END, 123 (23.6%) had poor outcome. Multivariate logistic regression analysis showed that baseline NIHSS score (odds ratio [ OR] 1.075, 95% confidence interval [ CI] 1.017-1.137; P=0.010) and MHR (with the lowest quartile as the reference, the third quartile: OR 2.778, 95% CI 1.255-6.151, P=0.012; the fourth quartile: OR 12.645, 95% CI 5.942-26.912; P<0.001) were the independent risk factors for END; the baseline NIHSS score ( OR 1.075, 95% CI 1.021-1.132; P=0.006), END ( OR 2.306, 95% CI 1.010-6.261; P=0.047) and MHR (with the first quartile as reference, the fourth quartile: OR 2.769, 95% CI 1.167-6.569; P=0.021) were the independent risk factors for poor outcomes. ROC curve analysis showed that area under the curve of MHR for predicting END and poor outcome in patients with AACIS were 0.805 (95% CI 0.750-0.860; P<0.001) and 0.747 (95% CI 0.690-0.803; P<0.001) respectively. The best cutoff value was 0.435, the sensitivity was 73.3% and 64.2%, and the specificity was 79.6% and 78.7% respectively. The area under the curve of MHR for predicting END and poor outcome was higher than that of monocyte and HDL-C alone. Conclusion:MHR can be used as a predictor of END and poor outcome in patients with AACIS, and its predictive value is higher than that of monocytes or HDL-C.

Objective:To investigate the influences of age-adjusted Charlson comorbidity index (ACCI) on prognosis of patients undergoing laparoscopic radical gastrectomy.Methods:The retrospective cohort study was conducted. The clinicopathological data of 242 gastric cancer patients who underwent laparoscopic radical gastrectomy in 19 hospitals of the Chinese Laparoscopic Gastrointestinal Surgery Study Group-04 study, including 54 patients in Fujian Medical University Union Hospital, 32 patients in the First Hospital of Putian City, 32 patients in Renji Hospital Affiliated to Shanghai Jiaotong University School of Medicine, 31 patients in Zhangzhou Affiliated Hospital of Fujian Medical University, 17 patients in Nanfang Hospital of Southern Medical University, 11 patients in the First Affiliated Hospital with Nanjing Medical University, 8 patients in Qinghai University Affiliated Hospital, 8 patients in Meizhou People′s Hospital, 7 patients in Fujian Provincial Hospital, 6 patients in Zhongshan Hospital of Fudan University, 6 patients in Longyan First Hospital, 5 patients in the First Affiliated Hospital of Xinjiang Medical University, 5 patients in the First Hospital Affiliated to Army Medical University, 4 patients in the Second Affiliated Hospital of Guangzhou University of Traditional Chinese Medicine, 4 patients in West China Hospital of Sichuan University, 4 patients in Beijing University Cancer Hospital, 3 patients in the First Affiliated Hospital of Xiamen University, 3 patients in Guangdong Provincial People′s Hospital, 2 patients in the First Affiliated Hospital of Xi′an Jiaotong University, from September 2016 to October 2017 were collected. There were 193 males and 49 females, aged 62(range, 23?74)years. Observation indicators: (1) age distribution, comorbidities and ACCI status of patients; (2) the grouping of ACCI and comparison of clinicopathological characteristics of patients in each group; (3) incidence of postoperative early complications and analysis of factors affecting postoperative early complications; (4) follow-up; (5) analysis of factors affecting the 3-year recurrence-free survival rate of patients. Follow-up was conducted using outpatient examination or telephone interview to detect postoperative survival of patients up to December 2020. Measurement data with normal distribution were represented as Mean± SD, and comparison between groups was conducted using the t test. Measurement data with skewed distribution were represented as M( Q1, Q3) or M(range), and comparison between groups was conducted using the Mann-Whitney U test. Count data were described as absolute numbers or percentages, and comparison between groups was conducted using the chi-square test or Fisher exact probability. Comparison of ordinal data was conducted using the nonparametric rank sum test. The X-Tile software (version 3.6.1) was used to analyze the best ACCI grouping threshold. The Kaplan-Meier method was used to calculate survival rates and draw survival curves. The Log-Rank test was used for survival analysis. The Logistic regression model was used to analyze the factors affecting postoperative early complications. The COX proportional hazard model was used for univariate and multivariate analyses of factors affecting the 3-year recurrence-free survival rate of patients. Multivariate analysis used stepwise regression to include variables with P<0.05 in univariate analysis and variables clinically closely related to prognosis. Results:(1) Age distribution, comor-bidities and ACCI status of patients. Of the 242 patients, there were 28 cases with age <50 years, 68 cases with age of 50 to 59 years, 113 cases with age of 60 to 69 years, 33 cases with age of 70 to 79 years. There was 1 patient combined with mild liver disease, 1 patient combined with diabetes of end-organ damage, 2 patients combined with peripheral vascular diseases, 2 patients combined with peptic ulcer, 6 patients combined with congestive heart failure, 8 patients combined with chronic pulmonary diseases, 9 patients with diabetes without end-organ damage. The ACCI of 242 patients was 2 (range, 0-4). (2) The grouping of ACCI and comparison of clinicopathological characteristics of patients in each group. Results of X-Tile software analysis showed that ACCI=3 was the best grouping threshold. Of the 242 patients, 194 cases with ACCI <3 were set as the low ACCI group and 48 cases with ACCI ≥3 were set as the high ACCI group, respectively. Age, body mass index, cases with preoperative comorbidities, cases of American Society of Anesthesiologists classification as stage Ⅰ, stage Ⅱ, stage Ⅲ, tumor diameter, cases with tumor histological type as signet ring cell or poorly differentiated adenocarcinoma and cases with tumor type as moderately or well differentiated adenocarcinoma, cases with tumor pathological T staging as stage T1, stage T2, stage T3, stage T4, chemotherapy cycles were (58±9)years, (22.6±2.9)kg/m 2, 31, 106, 85, 3, (4.0±1.9)cm, 104, 90, 16, 29, 72, 77, 6(4,6) in the low ACCI group, versus (70±4) years, (21.7±2.7)kg/m 2, 23, 14, 33, 1, (5.4±3.1)cm, 36, 12, 3, 4, 13, 28, 4(2,5) in the high ACCI group, showing significant differences in the above indicators between the two groups ( t=-14.37, 1.98, χ2=22.64, Z=-3.11, t=-2.91, χ2=7.22, Z=-2.21, -3.61, P<0.05). (3) Incidence of postoperative early complications and analysis of factors affecting postoperative early complications. Of the 242 patients, 33 cases had postoperative early complications, including 20 cases with local complications and 16 cases with systemic complica-tions. Some patients had multiple complications at the same time. Of the 20 patients with local complications, 12 cases had abdominal infection, 7 cases had anastomotic leakage, 2 cases had incision infection, 2 cases had abdominal hemorrhage, 2 cases had anastomotic hemorrhage and 1 case had lymphatic leakage. Of the 16 patients with systemic complications, 11 cases had pulmonary infection, 2 cases had arrhythmias, 2 cases had sepsis, 1 case had liver failure, 1 case had renal failure, 1 case had pulmonary embolism, 1 case had deep vein thrombosis, 1 case had urinary infection and 1 case had urine retention. Of the 33 cases with postoperative early complications, there were 3 cases with grade Ⅰ complications, 22 cases with grade Ⅱ complications, 5 cases with grade Ⅲa complications, 2 cases with grade Ⅲb complications and 1 case with grade Ⅳ complica-tions of Clavien-Dindo classification. Cases with postoperative early complications, cases with local complications, cases with systemic complications were 22, 13, 9 in the low ACCI group, versus 11, 7, 7 in the high ACCI group, respectively. There were significant differences in cases with postoperative early complications and cases with systemic complications between the two groups ( χ2=4.38, 4.66, P<0.05), and there was no significant difference in cases with local complications between the two groups ( χ2=2.20, P>0.05). Results of Logistic regression analysis showed that ACCI was a related factor for postoperative early complications of gastric cancer patients undergoing laparoscopic radical gastrectomy [ odds ratio=2.32, 95% confidence interval ( CI) as 1.04-5.21, P<0.05]. (4) Follow-up. All the 242 patients were followed up for 36(range,1?46)months. During the follow-up, 53 patients died and 13 patients survived with tumor. The 3-year recurrence-free survival rate of the 242 patients was 73.5%. The follow-up time, cases died and cases survived with tumor during follow-up, the 3-year recurrence-free survival rate were 36(range, 2-46)months, 29, 10, 80.0% for the low ACCI group, versus 35(range, 1-42)months, 24, 3, 47.4% for the high ACCI group. There was a significant difference in the 3-year recurrence-free survival rate between the two groups ( χ2=30.49, P<0.05). (5) Analysis of factors affecting the 3-year recurrence-free survival rate of patients. Results of univariate analysis showed that preoperative comorbidities, ACCI, tumor diameter, histological type, vascular invasion, lymphatic invasion, neural invasion, tumor pathological TNM staging, postoperative early complications were related factors for postoperative 3-year recurrence-free survival rate of gastric cancer patients undergoing laparoscopic radical gastrectomy [ hazard ratio ( HR)=2.52, 3.64, 2.62, 0.47, 2.87, 1.90, 1.86, 21.77, 1.97, 95% CI as 1.52-4.17, 2.22-5.95, 1.54-4.46, 0.27-0.80, 1.76-4.70, 1.15-3.12, 1.10-3.14, 3.01-157.52, 1.11-3.50, P<0.05]. Results of multivariate analysis showed that ACCI, tumor pathological TNM staging, adjuvant chemotherapy were indepen-dent influencing factors for postoperative 3-year recurrence-free survival rate of gastric cancer patients undergoing laparoscopic radical gastrectomy ( HR=3.65, 11.00, 40.66, 0.39, 95% CI as 2.21-6.02, 1.40-86.73, 5.41-305.69, 0.22-0.68, P<0.05). Conclusions:ACCI is a related factor for post-operative early complications of gastric cancer patients undergoing laparos-copic radical gastrectomy. ACCI, tumor pathological TNM staging, adjuvant chemotherapy are indepen-dent influencing factors for postoperative 3-year recurrence-free survival rate of gastric cancer patients undergoing laparoscopic radical gastrectomy.

OBJECTIVE: To analyze the association of the clinical inflammatory indices with the severity of urinary sepsis. METHODS: We reviewed the clinical data of 70 patients with urinary sepsis treated in our hospital between January, 2013 and April, 2018. All the patients were diagnosed in line with the Guidelines for Diagnosis and Treatment of Urological Diseases in China (2014 edition), including 22 patients with sepsis, 12 with hypotension and severe sepsis, 17 with septic shock, and 19 with critical septic shock. White blood cell count (WBC), neutrophil percentage (N%), platelets (PLT), fibrinogen (FIB), Ddimer, interleukin-6 (IL-6), procalcitonin (PCT) and C-reactive protein (CRP) were examined in all the cases and compared among the 4 groups. The correlations of these inflammatory markers with the severity of sepsis were analyzed using logistic regression analysis. RESULTS: The 4 groups of patients showed significant differences in N%, PLT, D-dimer, and PCT ( < 0.05) but not in CRP (>0.05). Kruskal-Wallis Pairwise comparisons showed that the N% and PCT in patients with sepsis differed significantly from those in the other 3 groups; platelets in patients with sepsis differed significantly from those in patients with septic shock and critical septic shock; D-dimer differed significantly between patients with sepsis and those with septic shock. Among the 4 groups, the median levels of PLT decreased and PCT and N% increased with the worsening of sepsis. Logistic regression analysis indicated that PCT (=0.186, =0.000), N% (=0.047, =0.035) and PLT (=-0.012, =0.003) were significantly correlated with the severity of sepsis in these patients. CONCLUSIONS PCT, PLT and N% are all significantly correlated with the severity of sepsis, and their combined detection can be informative for assessing the severity of sepsis to facilitate clinical decisions on treatment.
Biomarkers ; blood ; C-Reactive Protein ; analysis ; China ; Fibrin Fibrinogen Degradation Products ; analysis ; Fibrinogen ; analysis ; Humans ; Interleukin-6 ; blood ; Leukocyte Count ; Platelet Count ; Procalcitonin ; blood ; Sepsis ; blood ; diagnosis ; Severity of Illness Index ; Shock, Septic ; blood ; diagnosis ; Statistics, Nonparametric ; Urinary Tract Infections ; diagnosis

L-asparaginase (L-ASN) is widely applied in the treatment of malignant tumor and low-acrylamide food production, however, the low expression level hampers its application. Heterologous expression is an effective strategy to increase the expression level of target enzymes, and Bacillus is generally used as the host for efficient production of enzymes. In this study, the expression level of L-asparaginase in Bacillus was enhanced through optimization of expression element and host. Firstly, five signal peptides (SP_SacC, SP_AmyL, SP_AprE, SP_YwbN and SP_WapA) were screened, among which SP_SacC showed the best performance, reaching an activity of 157.61 U/mL. Subsequently, four strong promoters (P43, P_ykzA-P43, P_Ubay and P_bacA) from Bacillus were screened, and tandem promoter P_ykzA-P43 showed the highest yield of L-asparaginase, which was 52.94% higher than that of control strain. Finally, three Bacillus expression hosts (B. licheniformis Δ0F3 and BL10, B. subtilis WB800) were investigated, and the maximum L-asparaginase activity, 438.3 U/mL, was reached by B. licheniformis BL10, which was an 81.83% increase compared with that of the control. This is also the highest level of L-asparaginase in shake flask reported to date. Taken together, this study constructed a B. licheniformis strain BL10/P_ykzA-P43-SP_SacC-ansZ capable of efficiently producing L-asparaginase, which laid the foundation for industrial production of L-asparaginase.
Bacillus licheniformis/metabolism* ; Asparaginase/genetics* ; Bacillus/genetics* ; Protein Sorting Signals ; Promoter Regions, Genetic/genetics* ; Bacillus subtilis/genetics* ; Bacterial Proteins

Bacitracin is a broad-spectrum polypeptide antibiotic, which is formed by 11 amino acids residues. Precursor amino acids supply might be the limit factor during bacitracin fermentation. First, our results demonstrated that increasing Ile and Leu supplies were regarded as the efficient strategies for the enhanced titer of bacitracin. Then, the amino acid permease YhdG, which was identified as the BCAA permease, was deleted and overexpressed in DW2, respectively. Our results showed that knocking out of permease YhdG could improve bacitracin production remarkablely. The bacitracin titer of the yhdG deficient strain DW2ΔyhdG reached 917.35 U/mL by flask fermentation, increased by 11% compared with that of DW2. In addition, the bacitracin titer was decreased by 25% in the YhdG overexpressed strain. Meanwhile, the intracellular concentrations of BCAA were higher than DW2 during the biosynthesis of bacitracin. The above results suggested that the permease YhdG might act as an exporter for branched chain amino acids in B. licheniformis DW2. Taken together, the increasing intracellular concentrations of branched chain amino acids by deleting amino acid permease YhdG could improve bacitracin titer. This study provided a new strategy for high-level production of bacitracin.

Bacitracin is a broad-spectrum antibiotics mainly produced by Bacillus, and is used as veterinary medicine in the fields of livestock and poultry breeding. Insufficient supply of precursor amino acids might be an important factor that hinders high-level microbial production of bacitracin. We investigated the effect of strengthening L-cysteine supply on bacitracin production by an industrial bacitracin producer, Bacillus licheniformis DW2. Overexpression of cysK encoding L-cysteine synthase led to a 9.17% increase of the bacitracin titer. Moreover, overexpression of cysE encoding L-serine acetyltransferase and cysP encoding thiosulfate/sulfate intracellular transporter increased the bacitracin titers by 7.23% and 8.52%, respectively. Moreover, overexpression of a putative cystine importer TcyP led to a 29.19% increase of intracellular L-cysteine, and bacitracin titer was increased by 7.79%. Subsequently, the strong promoter PbacA was used to replace the promoters of genes cysP, cysE and tcyP in strain DW2::ysK, respectively. The resulted strain CYS4 (DW2::cysK-PbacA-(cysP)-PbacA(cysE)- PbacA(tcyP) produced 910.02 U/mL bacitracin, which was 21.10% higher than that of the original strain DW2 (747.71 U/mL). Together with the experiments in 3 L fermenters, this research demonstrated that enhancing intracellular L-cysteine supply is an effective strategy to increase bacitracin production of B. licheniformis.
Amino Acids ; Bacillus licheniformis/genetics* ; Bacitracin ; Cysteine ; Metabolic Engineering

Bacitracin is a broad-spectrum cyclic peptide antibiotic, and mainly produced by Bacillus. Energy metabolism plays as a critical role in high-level production of target metabolites. In this study, Bacillus licheniformis DW2, an industrial strain for bacitracin production, was served as the original strain. First, our results confirmed that elimination of cytochrome bd oxidase branch via deleting gene cydB benefited bacitracin synthesis. Bacitracin titer and ATP content were increased by 10.97% and 22.96%, compared with those of original strain, respectively. Then, strengthening cytochrome aa3 oxidase branch via overexpressing gene qoxA was conducive to bacitracin production. Bacitracin titer and ATP content were increased by 18.97% and 34.00%, respectively. In addition, strengthening ADP synthesis supply is also proven as an effective strategy to promote intracellular ATP accumulation, overexpression of adenosine kinase DcK and adenylate kinase AdK could all improve bacitracin titers, among which, dck overexpression strain showed the better performance, and bacitracin titer was increased by 16.78%. Based on the above individual methods, a method of combining the deletion of gene cydB and overexpression of genes qoxA, dck were used to enhance ATP content of cells to 39.54 nmol/L, increased by 49.32% compared to original strain, and bacitracin titer produced by the final strain DW2-CQD (DW2ΔcydB::qoxA::dck) was 954.25 U/mL, increased by 21.66%. The bacitracin titer produced per cell was 2.11 U/CFU, increased by 11.05%. Collectively, this study demonstrates that improving ATP content was an efficient strategy to improve bacitracin production, and a promising strain B. licheniformis DW2-CQD was attained for industrial production of bacitracin.
Bacillus licheniformis ; metabolism ; Bacitracin ; biosynthesis ; Energy Metabolism ; genetics ; Industrial Microbiology ; methods