OBJECTIVE To investigate the mechanism of Shenling baizhu san （SLBZS） in ameliorating ulcerative colitis （UC）. METHODS Mice were randomly divided into normal group， model group， and low-， medium-， high-dose SLBZS groups （3， 6， 12 g/kg）， with 10 mice in each group. Except for the normal group， mice in other groups were given free access to 3% dextran sulfate sodium solution for 7 days to establish UC models. Simultaneously， mice in each group were intragastrically administered corresponding medicinal liquid or double-distilled water once a day for consecutive 7 days. After the last administration， the histopathological morphology of colon tissues was observed； the serum levels of inflammatory factors including tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β） and IL-18 were detected； the total bile acid contents in liver， gallbladder and colon tissues were determined； the expression level of NOD-like receptor family pyrin domain-containing protein 3 （NLRP3） in colon tissues was tested. The expression levels of flora metabolism-related proteins [G protein-coupled receptor 41（GPR41），GPR43，farnesoid X receptor （FXR）， G protein-coupled bile acid receptor 1 （TGR5）， flavin-containing monooxygenase 3 （FMO3）， cytochrome P450 family 27 subfamily A member 1 （CYP27A1）， CYP7A1] in liver or colon tissues， as well as the proteins related to Toll-like receptor 4 （TLR4）/nuclear factor-κB （NF-κB）/NLRP3 inflammatory axis [TLR4， NF-κB， phosphorylated NF-κB （p-NF-κB）， myeloid differentiation factor 88 （MyD88）， NLRP3， apoptosis-associated speck-like protein containing a CARD （ASC）， caspase-1] in colon tissues were measured. RESULTS Compared with the model group， the colon tissue injury and inflammatory cell infiltration were alleviated in medium- and high-dose SLBZS groups. The serum levels of TNF-α， IL-1β and IL-18， total bile acid contents in liver and colon tissues， as well as the protein expression levels of FMO3 and CYP7A1 in liver tissues， TLR4， p-NF-κB， NF-κB， MyD88， NLRP3， ASC and caspase-1 in colon tissues were significantly decreased （P＜0.05）. Meanwhile， the total bile acid content in gallbladder， the protein expression levels of GPR41 and GPR43 in colon tissues， along with CYP27A1， FXR and TGR5 in liver tissues were significantly increased （P＜0.05）. CONCLUSIONS SLBZS may relieve colonic inflammatory injury and ameliorate UC by regulating the expression of flora metabolism-related proteins and inhibiting the abnormal activation of TLR4/NF-κB/NLRP3 inflammatory axis.

Objective To explore the correlation between serum circularRNA-HOMER1(circHOMER1),microRNA(miR)-23a-3p levels with clinical stages and oxidative stress in patients with diabetic retinopathy(DR).Methods From January 2023 to July 2024,75 DR patients treated in Handan Central Hospital were included as the DR group.According to the clinical staging of DR,they were divided into non proliferative DR(NPDR group,n=43)and proliferative DR(PDR group,n=32).In addition,75 patients with simple type 2 diabetes who came to Handan Central Hospital were included as non DR group.The levels of serum circHOMER1,miR-23a-3p,malondialdehyde(MDA),superoxide dismutase(SOD),and reduced glutathione(GSH)were detect-ed.Clinical data of the subjects were collected.The TargetScan website was used to predict the targeting relationship between circHOMER1 and miR-23a-3p.Pearson method was used to analyze the correlation between serum circHOMER1,miR-23a-3p and MDA,SOD,GSH.Univariate and multivariate Logistic regression were used to analyze the influencing factors of progression of DR in type 2 diabetes patients.Receiver operating characteristic(ROC)carve was used to analyze the predictive value of serum circHOMER1 and miR-23a-3p in the progression of DR in patients with type 2 diabetes.Results There was a targeted relationship between circHOMER1 and miR-23a-3p.The serum MDA(28.66±4.52ng/ml)and circHOMER1(1.24±0.16)levels in the DR group were higher than those in the non DR group(16.95±3.27ng/ml,1.02±0.11),while SOD(45.39±7.84U/L),GSH(135.82±21.23μg/mL)and miR-23a-3p(0.88±0.07)levels were lower than those in the non DR group(81.65±11.47U/L,207.44±25.95μg/mL,1.01±0.09),and differences were statistically significant(t=9.813～22.602,all P<0.001).The serum MDA(33.28±4.96ng/ml)and circHOMER1(1.36±0.20)levels in the PDR group were higher than those in the NPDR group(25.23±3.58ng/ml,1.15±0.17),while SOD(34.39±7.15U/L),GSH(113.50±20.17μg/ml)and miR-23a-3p(0.79±0.07)levels were lower than those in the NPDR group(53.27±8.44U/L,152.43±23.99μg/ml,0.94±0.08),and the differences were statistically significant(t=4.906～10.376,all P<0.001).Spearman analysis showed that serum MDA and circHOMER1 were positively correlated with the severity of DR(r=0.533,0.473,all P<0.001),while SOD,GSH,miR-23a-3p were negatively correlated with the severity of DR(r=-0.552,-0.515,-0.529,all P<0.001).Pearson analysis showed that serum circHOMER1 was negatively correlated with miR-23a-3p,SOD,GSH,and positively correlated with MDA(r=-0.475,-0.460,-0.455,0.462,all P<0.001).Serum miR-23a-3p was positively correlated with SOD and GSH,and negatively correlated with MDA(r=0.428,0.437,-0.439,all P<0.001).Logistic regression analysis showed that high MDA,low SOD,low GSH,high circHOMER1,low miR-23a-3p,high FPG and high HbA1c were the risk factors of progression of DR in type 2 diabetes patients(OR=0.214～3.556,all P<0.05).The area under curve(AUC)of serum circHOMER1 and miR-23a-3p alone and jointhy predicting the progression of DR in type 2 diabetes patients were 0.751,0.797 and 0.903 respectively.The combined prediction was higher than that of serum circHOMER1 and miR-23a-3p alone(Z=3.179,2.335,P=0.002,0.020).Conclusion Serum MDA and circHOMER1 levels are higher in DR patients,while serum SOD,GSH and miR-23a-3p levels are lower.Abnormal expression of circHOMER1 and miR-23a-3p in serum is associated with progression of DR and oxidative stress.Combined detection of circHOMER1 and miR-23a-3p in serum can predict the progression of DR in patients with type 2 diabetes.

Objective To explore the correlation between serum circularRNA-HOMER1(circHOMER1),microRNA(miR)-23a-3p levels with clinical stages and oxidative stress in patients with diabetic retinopathy(DR).Methods From January 2023 to July 2024,75 DR patients treated in Handan Central Hospital were included as the DR group.According to the clinical staging of DR,they were divided into non proliferative DR(NPDR group,n=43)and proliferative DR(PDR group,n=32).In addition,75 patients with simple type 2 diabetes who came to Handan Central Hospital were included as non DR group.The levels of serum circHOMER1,miR-23a-3p,malondialdehyde(MDA),superoxide dismutase(SOD),and reduced glutathione(GSH)were detect-ed.Clinical data of the subjects were collected.The TargetScan website was used to predict the targeting relationship between circHOMER1 and miR-23a-3p.Pearson method was used to analyze the correlation between serum circHOMER1,miR-23a-3p and MDA,SOD,GSH.Univariate and multivariate Logistic regression were used to analyze the influencing factors of progression of DR in type 2 diabetes patients.Receiver operating characteristic(ROC)carve was used to analyze the predictive value of serum circHOMER1 and miR-23a-3p in the progression of DR in patients with type 2 diabetes.Results There was a targeted relationship between circHOMER1 and miR-23a-3p.The serum MDA(28.66±4.52ng/ml)and circHOMER1(1.24±0.16)levels in the DR group were higher than those in the non DR group(16.95±3.27ng/ml,1.02±0.11),while SOD(45.39±7.84U/L),GSH(135.82±21.23μg/mL)and miR-23a-3p(0.88±0.07)levels were lower than those in the non DR group(81.65±11.47U/L,207.44±25.95μg/mL,1.01±0.09),and differences were statistically significant(t=9.813～22.602,all P<0.001).The serum MDA(33.28±4.96ng/ml)and circHOMER1(1.36±0.20)levels in the PDR group were higher than those in the NPDR group(25.23±3.58ng/ml,1.15±0.17),while SOD(34.39±7.15U/L),GSH(113.50±20.17μg/ml)and miR-23a-3p(0.79±0.07)levels were lower than those in the NPDR group(53.27±8.44U/L,152.43±23.99μg/ml,0.94±0.08),and the differences were statistically significant(t=4.906～10.376,all P<0.001).Spearman analysis showed that serum MDA and circHOMER1 were positively correlated with the severity of DR(r=0.533,0.473,all P<0.001),while SOD,GSH,miR-23a-3p were negatively correlated with the severity of DR(r=-0.552,-0.515,-0.529,all P<0.001).Pearson analysis showed that serum circHOMER1 was negatively correlated with miR-23a-3p,SOD,GSH,and positively correlated with MDA(r=-0.475,-0.460,-0.455,0.462,all P<0.001).Serum miR-23a-3p was positively correlated with SOD and GSH,and negatively correlated with MDA(r=0.428,0.437,-0.439,all P<0.001).Logistic regression analysis showed that high MDA,low SOD,low GSH,high circHOMER1,low miR-23a-3p,high FPG and high HbA1c were the risk factors of progression of DR in type 2 diabetes patients(OR=0.214～3.556,all P<0.05).The area under curve(AUC)of serum circHOMER1 and miR-23a-3p alone and jointhy predicting the progression of DR in type 2 diabetes patients were 0.751,0.797 and 0.903 respectively.The combined prediction was higher than that of serum circHOMER1 and miR-23a-3p alone(Z=3.179,2.335,P=0.002,0.020).Conclusion Serum MDA and circHOMER1 levels are higher in DR patients,while serum SOD,GSH and miR-23a-3p levels are lower.Abnormal expression of circHOMER1 and miR-23a-3p in serum is associated with progression of DR and oxidative stress.Combined detection of circHOMER1 and miR-23a-3p in serum can predict the progression of DR in patients with type 2 diabetes.

L-valine is an important branched-chain amino acid widely used in the food, pharmaceutical, and feed industries. Microbial fermentation has become the primary production method for L-valine. However, current industrial production still faces issues such as inefficient carbon flux utilization, imbalance in cofactor supply and demand, and suboptimal fermentation processes, which limit the efficient synthesis of L-valine. To further enhance the production performance of L-valine, In this study, metabolic engineering was conducted for a previously constructed Escherichia coli strain with a high yield of L-valine to optimize carbon flux distribution and balance cofactor consumption. Dual-phase oxygen-controlled fermentation was carried out to enhance L-valine production. Firstly, to address the pyruvate loss, we knocked out multiple competing pathway genes (ldhA, poxB, pflB, frdA, and pta), which resulted in a 48% increase in flask yield of the constructed strain VL-04. Next, we optimized the cofactor supply and demand balance by replacing ilvE with bcd (NADH-preferential) from Bacillus subtilis to construct the strain VL-06, which achieved a flask yield of 22.80 g/L, a further improvement of 25.8%. Subsequently, the fermentation conditions of VL-06 were optimized in a 5 L bioreactor with dual-phase oxygen-controlled fermentation. After optimization, the L-valine production reached 86.44 g/L in 26 h, with a glucose-to-acid conversion rate of 44.08% and a production intensity of 3.32 g/(L·h). This study not only shortens the time for L-valine production but also improves the economic efficiency, providing insights for similar fermentation processes employing dual-phase oxygen control.
Metabolic Engineering/methods* ; Escherichia coli/genetics* ; Valine/biosynthesis* ; Fermentation ; Bacillus subtilis/genetics*

Phenylpyruvic acid (PPA) is used as a food and feed additive and has a wide range of applications in the pharmaceutical, chemical and other fields. At present, PPA is mainly produced by chemical synthesis. With the green transformation of the manufacturing industry, biotransformation will be a good alternative for PPA production. The L-amino acid deaminase (PmiLAAD) from Proteus mirabilis has been widely studied for the production of PPA. However, the low yield limits its industrial production. To further enhance the production of PPA and better meet industrial demands, a more efficient synthesis method for PPA was established. In this study, PmiLAAD was heterologously expressed in Escherichia coli. Subsequently, a colorimetric reaction method was established to screen the strains with high PPA production. The semi-rational design of PmiLAAD was carried out, and the obtained triple-site mutant V18 (V437I/S93C/E417A) showed a 35% increase in catalytic activity compared with the wild type. Meanwhile, the effect of N-terminal truncation on the catalytic activity of the V18 mutant was investigated. After the optimization of the whole-cell conditions for the obtained mutant V18-N7, fed-batch conversion was carried out in a 5-L fermenter, and 44.13 g/L of PPA was synthesized with a conversion rate of 88%, which showed certain potential for industrial application. This study lays foundation for the industrial production of phenylpyruvic acid and also offers insights into the biosynthesis of other chemicals.
Escherichia coli/metabolism* ; Proteus mirabilis/genetics* ; Phenylpyruvic Acids/metabolism* ; Protein Engineering/methods* ; Recombinant Proteins/biosynthesis* ; Bacterial Proteins/metabolism*

L-tyrosine is one of the 20 amino acids that make up proteins and is an essential amino acid for mammals, often used as a nutritional supplement. The conventional methods for synthesizing L-tyrosine have some problems such as the production of many by-products, high requirements for production conditions, and environmental pollution. In this study, we designed and constructed a multi-enzyme cascade for the synthesis of L-tyrosine with alanine, glutamate, ammonium chloride, and phenol as substrates. Initially, the sources of glutamate oxidase, alanine aminotransferase, and tyrosine phenol lyase were screened and analyzed, which was followed by the identification of the rate-limiting enzyme in the reaction process. A colorimetric screening method was established, and the rate-limiting enzyme DbAlaA was engineered to enhance its activity by 40.0%. Subsequently, the reaction conditions, including temperature, pH, cell concentration, and surfactant and coenzyme dosages, were optimized. After optimization, the yield of L-tyrosine reached 9.93 g/L, with a alanine conversion rate of 54.90%. Finally, a feed-batch fermentation strategy was adopted, and the yield of L-tyrosine reached 56.07 g/L after 24 h, with a alanine conversion rate of 65.22%. This study provides a reference for the whole-cell catalytic synthesis of L-tyrosine and its industrialization.
Tyrosine/biosynthesis* ; Escherichia coli/metabolism* ; Tyrosine Phenol-Lyase/genetics* ; Multienzyme Complexes/metabolism* ; Fermentation

Objective: To investigate the factors affecting the development of non-proliferative diabetic retinopathy (NPDR) among patients with type 2 diabetic mellitus (T2DM), so as to provide insights into manangement of NPDR. Methods: T2DM patients without obvious eye discomfort at ages of 18 years and older admitted to Department of Endocrinology, Jining No. 1 People's Hospital during the period from December 2019 to May 2021 were enrolled. Participants' demographics, smoking, alcohol consumption, medical history of diabetes and use of medicines were collected, and the height, weight and blood pressure were measured. The levels of glycosylated hemoglobin (HbA1c), fasting blood glucose, blood C-peptide, lipid and creatinine were tested, and retinopathy was examined with a non-mydriatic fundus camera. The factors affecting the development of NPDR were identified among T2DM patients using a multivariable logistic regression model. Results: A total of 486 T2DM patients were enrolled, including 354 men (72.84%), with a median age of 48.00 (15.25) years, and median diabetes duration of 35.00 (104.25) months. The prevalence of NPDR was 19.34% among the participants. multivariable logistic regression analysis identified an educational level of senior high school and above (OR=0.546, 95%CI: 0.325-0.918), duration of diabetes (OR=1.008, 95%CI: 1.005-1.012), HbA1c (OR=1.183, 95%CI: 1.034-1.354) and use of non-sulfonylurea insulin secretagogues (OR=1.859, 95%CI: 1.082-3.196) as factors affecting the risk of NPDR among T2DM patients. Conclusion A high risk of NPDR is found among T2DM patients with a low educational level, long duration of diabetes, poor HbA1c control and use of non-sulfonylurea insulin secretagogues.

Objective:To investigate and analyze the clinical efficacy of salvage liver transplantation (SLT), rehepatectomy (RH), local ablation (LA), and prognostic risk factors in patients with postoperative recurrence of hepatocellular carcinoma.Methods:Clinical data of 145 patients with recurrent liver cancer in the 900th Hospital of the Joint Logistics Support Force of the People's Liberation Army from January 2005 to June 2018 were retrospectively collected. SLT group, RH group, and LA group included 25, 44, and 76 cases, respectively. Follow-up and statistics were recorded on the overall survival rate, relapse-free survival rate, and complications of the three groups of patients at 1, 2, and 3 years after surgery. Univariate and multivariate COX analyses were used to analyze the prognostic risk factors in patients with recurrent HCC.Results:The overall survival rates of 1, 2, and 3 years following surgery in the SLT, RH, and LA groups were 100.0%, 84.0%, 72.0%, 95.5%, 77.3%, 65.9%, 90.8%, 76.3%, and 63.2%, respectively, when the recurrence of liver cancer met the Milan criteria. The overall survival rate did not differ statistically between SLT and RH ( P = 0.303) or between RH and LA ( P = 0.152). There were statistically significant differences in recurrence-free survival between SLT and RH or RH and LA ( P = 0.046). There was no statistically significant difference in the incidence of complications between SLT and RH or RH and LA ( P > 0.017). Age > 65 years was an independent risk factor affecting the overall survival rate in patients with recurrent HCC. Age > 65 years and recurrence time < 24 months were independent risk factors affecting the recurrence-free survival rate in patients with recurrent HCC. Conclusion:SLT is the best treatment option when the recurrence of HCC meets Milan's criteria. RH and LA are the appropriate treatment plans for recurrent HCC when the liver source is limited.

Objective:To characterize the histopathological subtypes and their clinicopathological parameters of gender and onset age by common, rare and sparse primary esophageal malignant tumors (PEMT).Methods:A total of 272 437 patients with PEMT were enrolled in this study, and all of the patients were received radical surgery. The clinicopathological information of the patients was obtained from the database established by the State Key Laboratory of Esophageal Cancer Prevention & Treatment from September 1973 to December 2020, which included the clinical treatment, pathological diagnosis and follow-up information of esophagus and gastric cardia cancers. All patients were diagnosed and classified by the criteria of esophageal tumor histopathological diagnosis and classification (2019) of the World Health Organization (WHO). The esophageal tumors, which were not included in the WHO classification, were analyzed separately according to the postoperative pathological diagnosis. The χ 2 test was performed by the SPSS 25.0 software on count data, and the test standard α=0.05. Results:A total of 32 histopathological types were identified in the enrolled PEMT patients, of which 10 subtypes were not included in the WHO classification. According to the frequency, PEMT were divided into common (esophageal squamous cell carcinoma, ESCC, accounting for 97.1%), rare (esophageal adenocarcinoma, EAC, accounting for 2.3%) and sparse (mainly esophageal small cell carcinoma, malignant melanoma, etc., accounting for 0.6%). All the common, rare, and sparse types occurred predominantly in male patients, and the gender difference of rare type was most significant (EAC, male∶ female, 2.67∶1), followed with common type (ESCC, male∶ female, 1.78∶1) and sparse type (male∶ female, 1.71∶1). The common type (ESCC) mainly occurred in the middle thoracic segment (65.2%), while the rare type (EAC) mainly occurred in the lower thoracic segment (56.8%). Among the sparse type, malignant melanoma and malignant fibrous histiocytoma were both predominantly located in the lower thoracic segment (51.7%, 66.7%), and the others were mainly in the middle thoracic segment.Conclusion:ESCC is the most common type among the 32 histopathological types of PEMT, followed by EAC as the rare type, and esophageal small cell carcinoma and malignant melanoma as the major sparse type, and all of which are mainly occur in male patients. The common type of ESCC mainly occur in the middle thoracic segment, while the rare type of EAC mainly in the lower thoracic segment. The mainly sparse type of malignant melanoma and malignant fibrous histiocytoma predominately occur in the lower thoracic segment, and the remaining sparse types mainly occur in the middle thoracic segment.