Constructing an efficient and easy-to-operate multigene expression system is currently a crucial part of plant genetic engineering. In this study, a fragment carrying three independent gene expression cassettes and the expression unit of the gene-silencing suppressor protein(RNA silencing suppressor 19 kDa protein, P19) simultaneously was designed and constructed. This fragment was cloned into the commonly used plant expression vector pCAMBIA300, and the plasmid pC1300-TP2-P19 was obtained. Each gene expression cassette consists of different promoters, fusion tags, and terminators. The target gene can be flexibly inserted into the corresponding site through enzymatic digestion and ligation or recombination and fused with different protein tags, which provides great convenience for subsequent detection. The enhanced green fluorescent protein(eGFP) reporter gene was individually constructed into each expression cassette to verify the feasibility of this vector system. The results of tobacco transient expression and laser-confocal microscopy showed that each expression cassette presented independent and normal expression. Meanwhile, the three key enzyme genes in the betanin synthesis pathway, BvCYP76AD, BvDODA1, and DbDOPA5GT, were constructed into the three expression cassettes. The results of tobacco transient expression phenotype, protein immunoblotting(Western blot), and chemical detection of product demonstrated that the three exogenous genes were highly expressed, and the target compound betanin was successfully produced. The above results indicated that the constructed multigene expression system for plants in this study was efficient and reliable and can achieve the co-transformation of multiple plant genes. It can provide a reliable vector platform for the analysis of plant natural product synthesis pathways, functional verification, and plant metabolic engineering.
Nicotiana/metabolism* ; Genetic Vectors/metabolism* ; Gene Expression Regulation, Plant ; Plant Proteins/metabolism* ; Plants, Genetically Modified/metabolism* ; Genetic Engineering/methods* ; Green Fluorescent Proteins/metabolism* ; Gene Expression

INTRODUCTION: Albeit comprising a small portion of the hospital population, persistent high utilisers (PHUs) contribute disproportionately to healthcare expenditures. Amid rising healthcare costs and an ageing population, this study examines factors associated with PHUs among residents in eastern Singapore. METHOD: This is a retrospective study of eligible patients at Changi General Hospital in Singapore between 1 January 2020 and 31 December 2022. The study included Singapore citizens who utilised any services offered by CGH. Patients were classified as PHUs if their annual healthcare expenditure exceeded SGD3700 for 3 consecutive years. Demographics, healthcare utilisation patterns and clinical profiles were compared, and multivariable analyses were conducted to identify factors associated with PHUs. RESULTS: There were 267,838 eligible patients identified, with 5316 (2%) classified as PHUs. PHUs accounted for 18.4% of the total healthcare expenditure, with the highest costs attributed to inpatient services, followed by outpatient services. PHUs were more likely to be older, male, non-Chinese and of lower socioeconomic status. Conditions with the strongest association with PHUs were mental health disorders, chronic obstructive pulmonary disease, osteoporosis, asthma and renal diseases. Inpatient discharges from renal medicine, psychological medicine and general/geriatric medicine wards had the strongest association with PHUs. Utilisation of allied health services had the highest odds of being a PHU in outpatient care. CONCLUSION This study identified key factors associated with PHUs, providing invaluable insights into the planning of population health services within the hospital's geographical region. Targeted service development and process improvements of medical care can help mitigate persistent high utilisation.
Humans ; Singapore/epidemiology* ; Male ; Female ; Retrospective Studies ; Middle Aged ; Aged ; Adult ; Health Expenditures/statistics & numerical data* ; Patient Acceptance of Health Care/statistics & numerical data* ; Population Health ; Aged, 80 and over ; Young Adult ; Adolescent

INTRODUCTION: Obstructive sleep apnoea (OSA) is common in Singapore, with moderate to severe OSA affecting around 30% of residents. These consensus statements aim to provide scientifically grounded recommendations for the management of OSA, standar-dise the management of OSA in Singapore and promote multidisciplinary collaboration. METHOD: An expert panel, which was convened in 2024, identified several areas of OSA management that require guidance. The expert panel reviewed the current literature and developed consensus statements, which were later independently voted on using a 3-point Likert scale (agree, neutral or disagree). Consensus (total ratings of agree and neutral) was set a priori at ≥80% agreement. Any statement not reaching consensus was excluded. RESULTS: The final consensus included 49 statements that provide guidance on the screening, diagnosis and management of adults with OSA. Additionally, 23 statements on the screening, diagnosis and management of paediatric OSA achieved consensus. These 72 consensus statements considered not only the latest clinical evidence but also the benefits and harms, resource implications, feasibility, acceptability and equity impact of the recommendations. CONCLUSION The statements presented in this paper aim to guide clinicians based on the most updated evidence and collective expert opinion from sleep specialists in Singapore. These recommendations should augment clinical judgement rather than replace it. Management decisions should be individualised, taking into account the patient's clinical characteristics, as well as patient and caregiver concerns and preferences.
Humans ; Sleep Apnea, Obstructive/diagnosis* ; Singapore ; Consensus ; Adult

Objective To explore the effectiveness of"near peer learning"(NPL)in the electromyo-graphy(EMG)teaching module for neurology residents.Methods The Department of Neurology,Peking Union Medical College Hospital implemented an NPL instructional design for a course on EMG for residents from November 2020 to March 2024.This teaching session was held annually,in which senior residents in-structed juniors who were 1 or 2 years earlier in their training.The residents participated in the pre-course/post-course tests and completed a feedback survey at the end of the session.This evaluation method was used to un-derstand the effectiveness of the NPL intervention in EMG teaching.Results Over four years,a total of 83 residents participated.Among them,there were 24 postdoctoral students,52 postgraduates and 7 junior resi-dents.The results showed that the post-course test scores were significantly improved compared with pre-course test scores(74.33±2.43 vs.70.11±2.49,P=0.005),with the most remarkable improvements seen for"tu-tees"(73.84±20.53 vs.70.29±21.46,P=0.020),postgraduates(74.04±22.51 vs.68.97±21.40,P=0.009),first-year residents(70.19±4.02 vs.63.59±3.59,P=0.040)and first-time participating resi-dents(65.23±3.24 vs.60.97±3.21,P=0.030).The post-program feedback showed that both tutors and tu-tees thought highly of NPL,believing that it enabled them to gain knowledge and helped them to improve teaching skills.Conclusions The NPL intervention is suitable for the teaching of EMG,because of its contri-bution to knowledge acquisition and basic clinical skills improvement.The NPL is worth replicating in other teaching and learning programs.

Objective To investigate the neuroprotective effects and mechanisms of abscisic acid(ABA)in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)-induced Parkinson's disease(PD)mouse models.Methods Eight-week-old C57BL/6J mice were randomly divided into three groups,control group(Ctrl),MPTP group,and MPTP+ABA group,12 mice in each group.Except for the control group,mice in the other groups were intraperitoneally injected with MPTP 25 mg/kg daily for 8 consecutive days to establish a subacute PD model.The MPTP+ABA group received intraperitoneal injections of ABA 25 mg/kg daily for 11 consecutive days,starting 3 days prior to MPTP administration.Behavioral tests were performed 24 hours after the last administration.On day 3,the expression of tyrosine hydroxylase(TH)and glial fibrillary acidic protein(GFAP)in the substantia nigra pars compacta(SNc)and striatum(STR)was analyzed by Western blotting,and mRNA levels of inflammatory factors were measured by Real-time PCR.Immunofluorescent staining was used to detect the expression of TH,GFAP,and ionized calcium-binding adapter molecule 1(Iba1).Results Compared with the control group,MPTP-treated mice exhibited impaired motor function,a reduced number of TH-positive dopaminergic neurons in the SNc,down-regulated TH protein expression in both the SNc and striatum,up-regulated GFAP protein expression,increased numbers of GFAP-and Iba1-positive cells,and elevated levels of pro-inflammatory factors.In contrast,the MPTP+ABA group showed improved motor function,increased TH-positive neurons in the SNc,up-regulated TH protein expression,down-regulated GFAP protein expression,reduced numbers of GFAP-and Iba1-positive cells,and decreased pro-inflammatory factor levels compared to the MPTP group.Conclusion ABA ameliorates motor dysfunction in MPTP-induced PD model mice,reduces degeneration and death of dopaminergic neurons in the SNc,suppresses the proliferation and activation of astrocytes and microglia in the SNc and striatum,and alleviates neuroinflammation.These results suggest that ABA exerts neuroprotective effects in MPTP-induced PD model mice.

Hepatocellular carcinoma(HCC)is one of common cancer that seriously endangers human health.Designing methods for early,rapid,and accurate diagnosis of HCC has become the key point.Golgi protein 73(GP73),a novel potential biomarker for HCC,is crucial for diagnosis and treatment of HCC.In this study,a colorimetric sensor with rapidity,smplicity and high specificity was established for detection of GP73 based on peroxidase-like activity of hemin-reduced graphene oxide-manganese dioxide(H-rGO-MnO2).The H-rGO-MnO2-GP73Apt1 signal probe was synthesized by carboxyl of H-rGO-MnO2 nanozyme and amination of GP73 aptamer(GP73Apt1)though amide reaction.In the presence of GP73,the sulfhydryl-modifed GP73 aptamer(GP73Apt2),as the capture probe,and the signal probe both specifically recognized GP73,forming a sandwich structure(GP73Apt2-GP73-H-rGO-MnO2-GP73Apt1).This structure could catalyze the oxidation of H2O2 to produce hydroxyl radical(·OH),thereby oxidizing the colorless phthalenediamine(OPD)into the yellow 2,3-diaminophenazine(DPA).The quantitative detection of GP73 was achieved by measuring the characteristic absorbance of DPA at 450 nm.In the GP73 concentration range of 10-150 ng/mL,there was a good linear relationship between the DPA absorbance at 450 nm(A450 nm)and the GP73 concentration under optimal conditions.The linear equation was A450 nm=0.00321CGP73+0.8988,with the correlation coefficient(R2)of 0.9960 and the detection limit(LOD)of 5.38 ng/mL.The colorimetric sensor was applied to detection of GP73 in human serum samples,with recoveries of 88.4%?98.8%.This sensor showed high specificity,sensitivity,and stability,and had potential for clinical detection of GP73,providing a new approach for the early diagnosis of HCC.

This study analyzed the burden and trends in enteric infections in China from 1990 to 2021 from a One Health perspec-tive.Data on mortality associated with enteric infections were extracted from the 2021 Global Burden of Disease(GBD)database.The analysis focused on assessing the mortality rates of enteric infectious diseases attributed to various etiologies and risk factors,along with the age and sex distribution,from 1990 to 2021.Average annual percentage change(AAPC)was used to assess the total changes in disease burden.The age-standardized mortality rate of intestinal infections in China decreased from 9.642/100 000 in 1990 to 0.439/100 000 in 2021,with an AAPC of-57.103%(95%CI:-57.118%to-57.088%).In 2021,Rotavirus,Norovirus,and Crypto-sporidium were the top three etiologies contributing to disease burden,with mortality rates of 1.020/100 000,0.040/100 000 and 0.079/100 000,respectively.A significant variation in etiology distribution was observed across age groups:Rotavirus,Shigella,and Crypto-sporidium dominated among children under 5 years of age,whereas Cryptosporidium,Norovirus,and Clostridioides difficile were more prevalent in older populations.Risk factor analysis indicated that unsafe water sources and poor sanitation accounted for 73.394%of all enteric disease-related deaths.In conclusion,the burden of enteric infections in China markedly declined from 1990 to 2021,and sig-nificant variations in the etiological spectrum and disease burden were observed across age groups.The persistent effects of unsafe wa-ter sources and poor sanitation underscore the need for targeted interventions to further decrease the burden of these diseases.Our find-ings highlight the success of public health interventions in decreasing the burden of enteric infections in China,while emphasizing the need for targeted measures to address disparities in high-risk populations and improve environmental sanitation.

This study analyzed the burden and trends in enteric infections in China from 1990 to 2021 from a One Health perspec-tive.Data on mortality associated with enteric infections were extracted from the 2021 Global Burden of Disease(GBD)database.The analysis focused on assessing the mortality rates of enteric infectious diseases attributed to various etiologies and risk factors,along with the age and sex distribution,from 1990 to 2021.Average annual percentage change(AAPC)was used to assess the total changes in disease burden.The age-standardized mortality rate of intestinal infections in China decreased from 9.642/100 000 in 1990 to 0.439/100 000 in 2021,with an AAPC of-57.103%(95%CI:-57.118%to-57.088%).In 2021,Rotavirus,Norovirus,and Crypto-sporidium were the top three etiologies contributing to disease burden,with mortality rates of 1.020/100 000,0.040/100 000 and 0.079/100 000,respectively.A significant variation in etiology distribution was observed across age groups:Rotavirus,Shigella,and Crypto-sporidium dominated among children under 5 years of age,whereas Cryptosporidium,Norovirus,and Clostridioides difficile were more prevalent in older populations.Risk factor analysis indicated that unsafe water sources and poor sanitation accounted for 73.394%of all enteric disease-related deaths.In conclusion,the burden of enteric infections in China markedly declined from 1990 to 2021,and sig-nificant variations in the etiological spectrum and disease burden were observed across age groups.The persistent effects of unsafe wa-ter sources and poor sanitation underscore the need for targeted interventions to further decrease the burden of these diseases.Our find-ings highlight the success of public health interventions in decreasing the burden of enteric infections in China,while emphasizing the need for targeted measures to address disparities in high-risk populations and improve environmental sanitation.

INTRODUCTION: Since 2016, several therapies have been approved for treating atopic dermatitis (AD) in Singapore, including biologics, oral Janus kinase (JAK) inhibitors and topical crisaborole. This study supplements the 2016 Singapore treatment guidelines for AD, focusing on newer therapies for moderate-to-severe disease, while revisiting older treatment regimens to accommodate changes in knowledge and practice. METHOD: A modified Delphi panel was held, led by 2 co-chairs. The voting expert panel consisted of 12 dermatologists experienced in managing AD in Singapore. Delphi survey rounds were conducted between 24 July and 27 October 2023. Panellists indicated their agreement with drafted statements using a 5-point Likert scale. Consensus was defined as ≥80% agreement. An expert meeting was held to facilitate the consensus process between rounds 1 and 2 of voting. RESULTS: All expert panellists participated in both survey rounds, with a 100% response rate. Thirty-nine statements, classified into general principles, conventional treatments, biologics and JAK inhibitors, were proposed. Of these, 27 statements reached consensus at the end of round 1. After the expert meeting, 17 statements were included in round 2, of which 16 statements reached consensus. One statement did not reach consensus. Key updates are the inclusion of dupilumab and JAK inhibitors as potential first-line treatments for moderate-to-severe AD, in certain populations. CONCLUSION This modified Delphi study generated consensus among Singapore dermatology experts, to update treatment guidelines in moderate-to-severe atopic dermatitis. The consensus statements developed are intended to supplement the 2016 Singapore treatment guidelines for AD. Further revisions may be required when new evidence and/or treatments become available.
Dermatitis, Atopic/drug therapy* ; Humans ; Singapore ; Janus Kinase Inhibitors/therapeutic use* ; Biological Products/therapeutic use* ; Delphi Technique ; Consensus ; Antibodies, Monoclonal, Humanized/therapeutic use* ; Severity of Illness Index ; Bridged Bicyclo Compounds, Heterocyclic/therapeutic use* ; Dermatologic Agents/therapeutic use* ; Practice Guidelines as Topic ; Pyrimidines/therapeutic use* ; Boron Compounds

To explore the effectiveness of "near peer learning" (NPL) in the electromyography(EMG)teaching module for neurology residents.