Objective To explore the role of neogambogic acid in the characteristics of colorectal cancer stem cells (CRC-CSCs) through the Wnt/β-catenin signaling pathway. Methods The colorectal cells SW480 and HCT166 were divided into control group and neogambogic acid groups (1.5, 3, 6, and 12 μmol/L). The viability of CRC-CSCs was determined by MTT method, and spheroid and clone formation assays were used to assess the capacity of spheroid formation and self-renewal ability of the cells. The effects of neogambogic acid on the apoptosis and cell cycle of CRC-CSCs were evaluated by flow cytometry assays. Real-time quantitative PCR was used to detect the mRNA expression levels of relative markers (CD133, CD44, ALDH1, Oct4, and Nanog) of CRC-CSCs, and the protein expression levels of the self-renewal marker (PCNA), apoptosis markers (cleaved caspase-3 and cleaved caspase-9), and Wnt/β-catenin signaling pathway markers (p-GSK3β, GSK3β, β-catenin, and Wnt) were analyzed using Western blot. Results Compared with the control group, after neogambogic acid treatment, the viability of SW480 and HCT116 cells decreased (P<0.05), the spheroid forming ability and the clone numbers of CRC-CSCs decreased (P<0.001, P<0.01) but the cell apoptosis rate increased (P<0.01), and cell cycle was arrested in G₀/G₁ phase. Moreover, neogambogic acid downregulated the mRNA and protein expression of relative markers of CRC-CSCs (CD133, CD44, ALDH1, Oct4, and Nanog), PCNA, p-GSK3β, β-catenin, and Wnt (P<0.05) and upregulated the expression of cleaved caspase-3, cleaved caspase-9, and GSK3β (P<0.01). Conclusion Neogambogic can inhibit the stem cell properties of colorectal cells via inhibition of Wnt/β-catenin signaling pathway. As a result, neogambogic acid may be an attractive agent against colorectal cancer.

ObjectiveTo characterize the evidence distribution and methodological quality of randomized controlled trials （RCTs） on oral Chinese herbal medicine （CHM） for atopic dermatitis （AD） based on evidence mapping. MethodsSeven databases （CNKI， Wanfang Data， VIP， CBM， Cochrane Library， PubMed， and Embase） and the Chinese Clinical Trial Registry were searched for the RCTs in Chinese and English. Evidence distribution was presented graphically and textually， and methodological quality was assessed via the Cochrane Risk of Bias tool （ROB 1.0）. ResultsA total of 168 RCTs were included. The number of annual publications showing an increasing trend， and 72.6% RCTs had sample sizes of 51-100 participants. The studies evaluated 108 distinct CHM interventions categorized as decoctions， granules， Chinese patent medicines， and extracts. Compound Glycyrrhizin was the most frequently used， followed by Xiaofengsan and Chushi Weiling decoction. Among the RCTs， 57.1% had the treatment courses of 4-8 weeks. Outcome measures predominantly focused on clinical response rate， skin lesion severity scores， and adverse events， with less attention to TCM symptom scores， skin barrier function， and relapse rates. The overall risk of bias was generally high. ConclusionWhile CHM for AD is a research hotspot and demonstrates clinical advantages， the related studies have problems such as unclear clinical positioning， poor research standardization and methodological quality， and insufficient prominence of TCM clinical advantages. Large-sample， methodologically rigorous， and high-quality studies are needed to enhance the evidence base for CHM in treating AD.

ObjectiveTo characterize the evidence distribution and methodological quality of randomized controlled trials （RCTs） on oral Chinese herbal medicine （CHM） for atopic dermatitis （AD） based on evidence mapping. MethodsSeven databases （CNKI， Wanfang Data， VIP， CBM， Cochrane Library， PubMed， and Embase） and the Chinese Clinical Trial Registry were searched for the RCTs in Chinese and English. Evidence distribution was presented graphically and textually， and methodological quality was assessed via the Cochrane Risk of Bias tool （ROB 1.0）. ResultsA total of 168 RCTs were included. The number of annual publications showing an increasing trend， and 72.6% RCTs had sample sizes of 51-100 participants. The studies evaluated 108 distinct CHM interventions categorized as decoctions， granules， Chinese patent medicines， and extracts. Compound Glycyrrhizin was the most frequently used， followed by Xiaofengsan and Chushi Weiling decoction. Among the RCTs， 57.1% had the treatment courses of 4-8 weeks. Outcome measures predominantly focused on clinical response rate， skin lesion severity scores， and adverse events， with less attention to TCM symptom scores， skin barrier function， and relapse rates. The overall risk of bias was generally high. ConclusionWhile CHM for AD is a research hotspot and demonstrates clinical advantages， the related studies have problems such as unclear clinical positioning， poor research standardization and methodological quality， and insufficient prominence of TCM clinical advantages. Large-sample， methodologically rigorous， and high-quality studies are needed to enhance the evidence base for CHM in treating AD.

Objective:To evaluate the utility of pulse oximetry-derived parameters—specifically, the pulse oximetry plethysmographic waveform area under the curve (POP AUC) and the peripheral perfusion index (PPI)—in assessing disease severity and predicting prognosis in patients with sepsis. Methods:In this prospective descriptive study, 68 patients with sepsis were categorized based on illness severity into septic shock and non-shock groups, and by 28-day outcome into survival and non-survival groups. POP AUC, PPI, and lactate (Lac) levels were recorded at 0, 24, 48, 72, and 96 hours after admission. APACHEⅡ and SOFA scores were calculated within the first 24 hours. The prognostic value of these parameters was evaluated. Results:Significant differences were observed between the septic shock and non-shock groups in POP AUC, PPI, Lac (all P < 0.05 except at 96 h), APACHEⅡ, and SOFA scores (all P < 0.05). These differences were most pronounced at admission: POP AUC0 (2475.1 ± 899.0) vs. (4260.3 ± 1028.5), PPI 0 (0.78 ± 0.74) vs. (3.13 ± 2.18), Lac 0 (4.95 ± 4.32) vs. (2.07 ± 1.55), APACHE Ⅱ (16.78 ± 5.59) vs. 11.82 ± 4.89), and SOFA (8.89 ± 3.25) vs. (5.06 ± 2.60). Optimal prognostic cut-off values were 2741.43 for POP AUC, 0.97 for PPI, 2.05 for Lac, 12.5 for APACHEⅡ, and 5.5 for SOFA. ROC curve analysis showed that at 24 hours, POP AUC and PPI had significantly larger AUC values than Lac ( P < 0.05), while no significant differences were found among other parameters. Significant differences between non-survivors and survivors were also found in POP AUC, PPI (at 0, 24, and 48 h), APACHE II, and SOFA (all P < 0.05). No significant differences were observed in PPI (72 h and 96 h) or Lac between the two outcome groups. Conclusions:POP AUC and PPI, as derived from pulse oximetry, are non-inferior to Lac, SOFA, and APACHEⅡ scores in evaluating disease severity and predicting 28-day mortality in sepsis patients. These parameters show promise as practical and non-invasive tools for clinical assessment in sepsis.

Since the National Health Commission launched the pilot work of " Internet+ nursing service" in 2019, medical institutions at all levels in China keep innovating their nursing service models to provide targeted high-quality care for discharged patients or special groups suffering from diseases and poor mobility. In April 2021, a tertiary hospital in Qingdao carried out the " Internet+ " whole course management practice with specialized nurses as the main service subjects, which center on discharged patients as the main service objects. By estabusing a management team and the construction of " Internet+ " information exchange platform, the hospital implemented the whole course of disease management process including patients′ pre-hospital management, in-hospital estimate, and post-hospital follow-up rehabilitation, to provide specialized nursing services such as PICC dressing change care, stoma care, and mother and infant care for patients at home. By February 2022, the " Internet+ " whole course management service had expanded to 50 kilometers away from hospital, with a total of 1 181 specialized nursing services. This management highlighted the characteristics of specialized nursing services in tertiary hospitals, accurately matched the needs of patients, reflected the concept of holistic care, and provided references for the rapid promotion of the development of " Internet+ nursing service" in China.

Tyrosine is an important aromatic amino acid. Besides its nutritional value, tyrosine is also an important precursor for the synthesis of coumarins and flavonoids. Previously, our laboratory constructed a Saccharomyces cerevisiae strain LTH0 (ARO4K229L, ARO7G141S, Δaro10, Δzwf1, Δura3) where tyrosine feedback inhibition was released. In the present study, heterologous expression of betaxanthins synthesis genes DOD (from Mirabilis jalapa) and CYP76AD1 (from sugar beet B. vulgaris) in strain LTH0 enabled production of yellow fluorescence. The engineered strain LTH0-DOD-CYP76AD1 was subjected to UV combined with ARTP mutagenesis, followed by flow cytometry screening. Among the mutants screened, the fluorescence intensity of the mutant strain LTH2-5-DOD-CYP76AD1 at the excitation wavelength of 485 nm and emission wavelength of 505 nm was (5 941±435) AU/OD, which was 8.37 times higher than that of strain LTH0-DOD-CYP76AD1. Fourteen mutant strains were subjected to fermentation to evaluate their tyrosine producing ability. The highest extracellular tyrosine titer reached 26.8 mg/L, which was 3.96 times higher than that of strain LTH0-DOD-CYP76AD1. Heterologous expression of the tyrosine ammonia lyase FjTAL derived from Flavobacterium johnsoniae further increased the titer of coumaric acid to 119.8 mg/L, which was 1.02 times higher than that of the original strain LTH0-FjTAL.
Flavobacterium ; High-Throughput Screening Assays ; Mirabilis ; Saccharomyces cerevisiae/genetics* ; Tyrosine

Lycopene, as a high value-added terpene compound, has been widely concerned by researchers at home and abroad. Firstly, the ability of lycopene synthesis of Saccharomyces cerevisiae model strains S288c and YPH499 was analyzed and compared. The results showed that YPH499 was more suitable for lycopene synthesis as yeast chassis. Subsequently, the effects of constitutive promoters GPDpr, TEF1pr and inducible promoters GAL1pr, GAL10pr on Lycopene synthesis were compared. The results showed that when GPDpr and TEF1pr were used as promoters of crtE, crtB and crtI in lycopene synthesis pathway, the production of lycopene was 15.31 mg/L after 60 h fermentation in shaking flask. When GAL1pr and GAL10pr were used as promoters, the production was 123.89 mg/L, which was 8.09 times higher. In addition, the methylvaleric acid (MVA) pathway was further modified to overexpress the key enzyme gene of N-terminal truncation, tHMG1 (3-hydroxy-3-methylglutaryl coenzyme A reductase). The lycopene production was 265.68 mg/L, and the yield per cell was 72.79 mg/g. The Saccharomyces cerevisiae strain designed and constructed in this study can express lycopene in high yield per cell, thus could be used in the industrial production of lycopene after further construction and optimization.
Biosynthetic Pathways ; genetics ; Fermentation ; Industrial Microbiology ; Lycopene ; metabolism ; Saccharomyces cerevisiae ; genetics ; metabolism ; Species Specificity

Clostridium difficile (CD),a gram-positive,spore-forming,obligate anaerobic bacillus,is a leading cause of antibiotic-associated diarrhea (AAD) worldwide.With the widespread use of broad-spectrum antibiotics,the incidence of clostridium difficile infection in children is rising,while recurrent clostridium difficile infection (RCDI) requires prolonged treatment and higher medical costs.Malignancy,recent surgery and antibiotic exposures have been identified as the risk factors in children.While the toxigenic strains culture and the cell cytotoxicity neutralization assay are gold standard for the diagnosis,new diagnostic approaches such as nucleic acid amplification method have become available.The use of antibiotics,fecal microbiota transplantation (FMT) or monoclonal antibodies are included in the current treatments for RCDI.This review will cover published studies to discuss the risk factors,diagnosis and treatment of RCDI in children.

Few tools of gene editing have been developed in Bacillus licheniformis at present. In order to enrich the tools, an FLP/FRT gene editing system that can repeatedly use a single selectable marker was constructed in Bacillus licheniformis, and the system was verified by knocking out an alpha amylase gene (amyL), an protease gene (aprE) and knocking in an exogenous Vitreoscilla hemoglobin gene (vgb). First, knock-out plasmids pNZTT-AFKF of amyL and pNZTT-EFKF of aprE were constructed using thermosensitive plasmid pNZT1 as a carrier. The two knock-out plasmids contained respective homology arms, resistance genes and FRT sites. Then the knock-out plasmids were transformed into Bacillus licheniformis and the target genes were replaced by respective deletion cassette via twice homologous exchange. Finally, an expression plasmid containing FLP recombinase reading frane was introduced and mediated the excision of resistance marker. In order to expand the practicability of the system, knock-in plasmid pNZTK-PFTF-vgb was constructed, with which knock-in of vgb at pflB site was carried out successfully. The results showed that amyL and aprE were successfully knocked out and the marker kanamycin cassette exactly excised. The activities of amylase and protease of deletion mutants were reduced by 95.3% and 80.4% respectively. vgb was successfully knocked in at pflB site and the marker tetracycline cassette excised. The expression of integrated vgb was verified via real-time PCR. It is the first time to construct an FLP/FRT system for gene editing in Bacillus licheniformis, which could provide an effective technical means for genetic modification.
Bacillus licheniformis ; Gene Editing ; Plasmids ; Sequence Deletion

Objective To analyze the time of removing setons by observing 30 cases of the infliximab (IFX)combined drainage setons therapy in the treatment of perianal fistulising Crohn's disease.Methods Thirty patients of perianal fistulising Crohn's disease of Department of Colorectal Surgery,Jiangsu Prorince Hospital of TCM in Aug.2015 to Jul.2017 were treated with drainage setons followed by intravenous IFX injections according to the standard,and setons were removed after the third injection of IFX.The method of repeated measurement were adopted to record.Clinical symptoms,perianal manifestations,laboratory indexes,healing rate and general linear model and nonparametric test of SPSS 23.0 software were used for statistical analysis.Results The differences of WBC,CRP,ESR,HB,PLT,BMI,CDAI and PDAI between post treatment and pretreatment were statistically significant (P ＜ 0.05).WBC,CRP and ESR will rerise at some time after removing the rubber band (sixth week).The closure rate of fistular at the 14th week was 53.3％,and after the follow-up in the ward at 22th week and 30th week,the clinical efficacy of fistular was not statistically significant (P ＞0.05) at the 14th week,22th week and 30th week.Conclusion IFX combined drainage setons which was removed when the induction therapy finished can induce Crohn's disease remission,improve laboratory index and promote fistula closure.the clinical efficacy in short time was stable.