[Tumor necrosis factor-related apoptosis-inducing ligand gene polymorphism and its plasma phenotype in relation to Crohn's disease].

Objectives: To investigate the associations of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) gene polymorphism and plasma soluble TRAIL level (sTRAIL) with Crohn's disease (CD) and to retrospectively analyze the effects of TRAIL gene variants and plasma sTRAIL levels on clinical response to infliximab (IFX). Methods: From January 2012 to January 2021, 312 CD patients [205 males, 107 females, average age (33.9±9.8) years] and 514 age-and gender-matched healthy controls [304 males, 210 females, average age (34.9±9.4) years] were recruited from the Department of Gastroenterology, the Second Affiliated Hospital of Wenzhou Medical University. Among them, 72 patients with active CD who were ineffective or intolerant to traditional drug therapy regularly received IFX (5 mg/kg) treatment. According to the changes in the Harvey-Bradshaw index (HBI) and the Simplified Endoscopic Score for Crohn's Disease (SES-CD) in the 14th week, these patients were classified into response group (a decrease in HBI≥3 or a decrease in SES-CD≥50%) and non-response group. TRAIL (rs1131568) gene polymorphism was analyzed by matrix-assisted laser desorption/ionization time of flight mass spectrometry technique. The plasma sTRAIL level was examined by enzyme-linked immunosorbent assay (ELISA). Based on the Montreal CD classification criteria, all CD patients were divided into different subgroups. Finally, a comprehensive analysis was performed to investigate the relationship between TRAIL (rs1131568) gene polymorphism, the plasma sTRAIL level and the risk of CD, the clinicopathological characteristics of CD patients, and the clinical response to IFX. Results: The recessive model analysis showed that the homozygous variant genotype (CC) was more prevalent in patients with moderately to severely active CD than in those with mildly active CD (45.34% vs 29.23%, P=0.005). Both variant allele (C) and homozygous variant genotype (CC) in patients with stricturing and penetrating CD were more frequent than those in patients with non-stricturing and non-penetrating CD (65.48% vs 57.53%, P=0.046; 49.21% vs 31.18%, P=0.001). The dominant model analysis showed that variant allele (C) and variant genotype (TC+CC) was higher in CD patients with perianal lesions than in those without perianal lesions (66.83% vs 58.17%, P=0.037; 92.31% vs 78.37%, P=0.002). The average plasma sTRAIL level was higher in CD patients than in healthy controls [(243.04±42.74) ng/L vs (194.16±31.14) ng/L, P<0.001]. Compared with the patients with mildly active CD, the plasma sTRAIL level was increased in those with moderately to severely active CD [263.47(242.09, 281.91) ng/L vs 231.13(211.11, 247.11) ng/L, P<0.001]. The same conclusion was also drawn for the patients with stricturing and penetrating CD in contrast to those with non-stricturing and non-penetrating CD [266.18 (246.68, 289.91) ng/L vs 227.19 (204.57, 249.59) ng/L, P<0.001]. The plasma sTRAIL level was also higher in patients with perianal disease than in those without perianal disease [(261.40±41.51) ng/L vs (233.86±40.41) ng/L, P<0.001]. Multiple linear regression analysis further showed that disease activity (ß=22.640, P<0.001) and homozygous variant genotype (CC) (ß=16.814, P<0.001) may be positively related to the plasma sTRAIL level in CD patients independently. At the 14th week of IFX treatment, the plasma sTRAIL level in the response group was lower than that in the non-response group [205.98(190.72, 214.56) ng/L vs (238.33±29.38) ng/L, P<0.001]. Compared with week 0, the plasma sTRAIL level was decreased in the response group in the 14th week [(205.98 (190.72, 214.56) ng/L vs (239.89±42.43) ng/L, P<0.001]. Non-conditional logistic regression analysis showed that variant allele (C) and variant genotype (TC+CC) were less frequent in the response group than in the non-response group (53.33% vs 70.83%, P=0.037; 70.00% vs 91.67%, P=0.036). Conclusions: The increased plasma sTRAIL level may be a risk factor for CD. TRAIL (rs1131568) gene variation and the increase of plasma sTRAIL level may be associated with the increased disease activity of CD and may be the risk factors for stenosis, penetration, and perianal lesions in CD patients. In addition, TRAIL (rs1131568) gene variation or the increase of plasma sTRAIL level may be related to no response to IFX treatment in CD patients.

[Epidemiological investigation on allergic diseases related to animal dander of cats, dogs and horses].

Objective: A multicenter Chinese mainland survey was conducted to investigate the sensitization distribution characteristics of cat, dog and horse dander in patients with allergic diseases, so as to provide clinicians with epidemiological data of common animal allergens and useful information for the prevention and treatment of allergies in cats, dogs and horses. Methods: The epidemiological investigation and design was adopted. This study is based on the national epidemiological survey of allergic diseases led by the first affiliated Hospital of Guangzhou Medical University. From January to December in 2021, a total of 2 122 patients diagnosed with allergic diseases were included in the outpatient department of respiratory department/pediatrics/allergy department of 14 units such as the First Affiliated Hospital of Guangzhou Medical University, and 222 healthy subjects were included as controls from the physical examination center of the above units in the same period. All the subjects filled out the allergic disease questionnaire under the guidance of doctors, and the allergen-specific immunoglobulin E (sIgE) of cats, dogs and horses of all subjects were detected by magnetic particle chemiluminescence system. The epidemiological characteristics of three animal allergens in different diseases, ages and regions were analyzed. Chi square test was used to analyze the frequency difference between groups, t test or Mann Whitney U test was used to test the distribution difference between two groups, and one-way ANOVA or Kruskal Wallis H test was used to compare the distribution difference between multiple groups. Bar chart, Venn-plot and radar chart were drawn to show the sensitization distribution characteristics. A small number of missing values caused by subjects' omission have been excluded during the analysis. Results: The 2 122 patients with allergic diseases were 57.35% male (1 217/2 122) and 40.95% female (869/2 122), and 1.70% (36/2 122) patients had loss of gender information. The age of patients with allergic diseases was 9.0 (6.0, 28.0) years, while that of healthy controls was 29.0 (13.0, 39.0) years old, and there were 1.7% (36/2 122) and 0.9% (2/222) subjects with missing age information, respectively. The proportion of caesarean section in allergic patients was significantly higher than that in healthy controls (31.4% vs. 17.6%,χ2=16.582,P<0.001) [2.5% (54/2 122) of the patient group and 5.4% (12/222) of the control group had missing birth mode information], and the proportion of patients with allergic diseases who reported that both parents had allergic diseases was significantly higher than that of the control group (35.7% vs. 9.5%, χ2=65.171,P<0.001). Patients with allergic diseases are mainly school-age (6-12 years old) and adolescents (12-18 years old). 16.4% of patients with allergic diseases were sensitized to cat dander, 10% and 6% to dog and horse dander. The sensitization rate of cat dander in patients with rhinitis, asthma, conjunctivitis, food allergy and atopic dermatitis was the highest (16.4%-21.6%), followed by dog dander (10.2%-15.2%). The prevalence of allergic rhinitis was the highest among different animal sensitized populations. The proportion of cat, dog and horse allergens sensitized at the same time is between 10%-15%, and the proportion of any two or more animal dander sensitized at the same time is about 45%. Animal allergens are associated with respiratory allergic diseases, especially allergic rhinitis with allergic conjunctivitis. There were significant differences in the distribution of positive rates of three animal allergens in different regions, and the highest positive rate of cat dander was found in all provinces of the country. Conclusion: The sensitization rate of animal dander allergens increased significantly, and the highest was in children and adolescents. Cat dander is the most common animal allergen, followed by dog. Different animals show obvious cross or common sensitization due to their high homology.

[A preliminary study on the improved efficacy of mesalazine combined with vitamin D3 in ulcerative colitis].

Objective: To analyze the influence of vitamin D3 supplementation on the clinical efficacy of mesalazine in patients with ulcerative colitis (UC). Methods: From January 2015 to December 2020, patients with mild-to-moderate active UC were retrospectively and continuously enrolled, who accepted mesalazine treatment for at least 12 months at the Second Affiliated Hospital of Wenzhou Medical University. According to simultaneous supplement of vitamin D3 (125 IU/d), the patients were divided into study group and control group. Demographic and disease characteristics, serum 25-hydroxyvitamin D[25(OH)D] levels and other information were collected through retrieving hospital database. Student's t-test, Mann-Whitney U test and Chi-square test were applied for comparison of disease characteristics. The changes of modified Mayo scores[ΔMayo] and 25(OH)D[Δ25(OH)D] were compared before and after treatment by paired t-test, Wilcoxon signed rank test and Chi-square test. Multiple linear regression model was used to analyze the independent factors affecting ΔMayo and Δ25(OH)D, and variables with P-values less than 0.20 in the univariate analysis were allowed for further multivariate analysis. Results: A total of 74 UC patients (44 males, 30 females), with median age (range) 39.5 (20-76) years old, were analyzed and respectively assigned into study group (n=36) and control group (n=38). In study group, the average level of serum 25(OH)D was significantly increased at month 12 compared with that at baseline [(22.87±7.30) µg/L vs. (18.15±7.48) µg/L,P<0.001]. However, no significant elevation of serum 25(OH)D was found in control group [(19.17±8.49) µg/L vs. (19.82±9.47) µg/L,P=0.466]. Furthermore, there was a significant decrease of modified Mayo score [-3(-4.75, -1.25) vs.-2(-3.25, 0), P=0.034] and a higher clinical remission rate (55.6% vs. 28.9%, P=0.020) at month 12 in study group than those in control group. In addition, according to the baseline level of serum 25(OH)D before mesalazine treatment, 74 UC patients were divided into vitamin D deficiency group (n=38, serum 25(OH)D<20 µg/L) and non-deficiency group (n=36, serum 25(OH)D≥20 µg/L). At month 12 in vitamin D deficiency group, patients with vitamin D3 supplementation had a greater decline in modified Mayo score [-4(-5.75, -2) vs.-2(-4, 0), P=0.048] and a higher clinical remission rate (60.0% vs. 22.2%, P=0.019) compared with those without. Conclusions: In patients with mild-to-moderate active UC receiving mesalazine treatment, vitamin D3 supplementation may improve the clinical efficacy, especially in patients with vitamin D deficiency.

[Research on allergen detection technology and allergic disease diagnosis strategy in China].

In China, the prevalence of allergic diseases is increasing, but allergy research and allergy diagnosis technologies are still in their infancy, posing certain hurdles to clinical diagnosis and treatment. The detection of allergen is essential for the prevention and treatment of allergic diseases. Clarifying the aetiology and risk factors of allergic diseases is critical in order to provide targeted environmental control and therapy for allergic diseases, as well as reduce patient pain and financial burden. In this study, allergen detection technologies and diagnosis strategies are described in order to provide direction for clinicians and laboratory technicians, improve allergic diseases diagnosis, and guide allergic disease therapy.

[Analysis of serum allergen-specific IgE in children with respiratory allergic disease in Guangdong].

Objective: To investigate the common allergens in children with respiratory allergic diseases in Guangdong and provide the basis for preventing and treating respiratory allergic diseases. Methods: A total of 254 serum samples were collected retrospectively from children with allergic rhinitis and (or) asthma in the pediatric clinic of the First Affiliated Hospital of Guangzhou Medical University or Boai Hospital of Zhongshan City from August 2019 to August 2020 and tested positive for at least one allergens by ImmunoCAP 1000 or skin prick test. There were 173 males and 81 females, with a median age (interquartile interval) of 7 (4.00, 9.75) years. Specific IgE against eighteen common allergen were tested by magnetic particle chemiluminescence immunoassay. Statistical analysis using χ2 test. Results: The positive rate of Dermatophagoides pteronyssinus (85.0%, 215/254) was the highest, followed by D. farinae (83.5%, 212/254), egg white (19.3%, 49/254), milk (14.6%, 37/254), German cockroach (14.2%, 36/254) and ox-eye daisy (12.6%, 32/254). Dust mites were the most common allergens in patients with allergic rhinitis / allergic asthma / allergic rhinitis and asthma. The positive rates of D. pteronyssinus (95.4% vs. 91.7% vs. 71.0%, χ2=23.257, P<0.001), D. farinae (95.4% vs. 90.6% vs. 67.7%, χ2=26.916, P<0.001), mugwort (10.8% vs. 9.4% vs. 1.1%, χ2=7.535, P=0.023), and ox-eye daisy (21.5% vs. 14.6% vs. 4.3%, χ2=10.876, P=0.004) in allergic rhinitis combined with asthma group and allergic rhinitis group were significantly higher than those in allergic rhinitis group. The positive rates of D. pteronyssinus (96.2% vs. 84.0% vs. 53.5%, χ2=46.343, P<0.001), D. farinae (96.2% vs. 81.5% vs. 48.8%, χ2=52.756, P<0.001) and German cockroach (17.7% vs. 14.8% vs. 2.3%, χ2=6.313, P=0.043) in > 6 years old group were significantly higher than those in > 3 years old group and > 3-6 years old group, while the sIgE positive rates of egg white (41.9% vs. 21.0% vs. 10.8%, χ2=20.281, P<0.001) and milk (41.9% vs. 16.0% vs. 4.6%, χ2=36.227, P<0.001) in ≤3 years old group were significantly higher than those in > 3-6 years old group and > 6 years old group. In addition, there were significant positive correlations between D. pteronyssinus and D. farinae, mugwort and ox-eye daisy, cat dander and horse dander, ox-eye daisy and dandelion, egg white and milk, timothy grass and sesame (rs≥0.511, P < 0.01). With the increase of the class of house dust mite allergen, the sIgE positive rates of mugwort (0.0% vs. 1.5% vs. 10.8%, χ2=9.714, P=0.008), ox-eye daisy (0.0% vs. 4.4% vs. 19.6%, χ2=16.195, P<0.001), cat dander (0.0% vs. 7.4% vs. 18.2%, χ2=11.459, P=0.003) and horse dander (0.0% vs. 1.5% vs. 15.5%, χ2=15.443, P < 0.001) increased significantly, while the sIgE positive rate of milk (29.0% vs. 16.2% vs. 10.1%, χ2=8.792, P=0.012) decreased significantly. Conclusions: Dust mite is the main allergen in children with respiratory allergic diseases in Guangdong, and ox-eye daisy is the main pollen allergen. The sensitization patterns of allergens varied in different ages and diseases, and children with respiratory allergic diseases should regularly detect allergens and pay attention to avoid them.

[Expert consensus on the strategy and measures to interrupt the transmission of schistosomiasis in China].

Since 2015 when the transmission of schistosomiasis was controlled in China, the country has been moving towards elimination of schistosomiasis, with the surveillance-response as the main interventions for schistosomiasis control. During the period of the 13th Five-Year Plan, the transmission of schistosomiasis had been interrupted in four provinces of Sichuan, Jiangsu, Yunnan and Hubei and the prevalence of schistosomiasis has been at the historically lowest level in China. As a consequence, the goal set in The 13th Five-Year National Schistosomiasis Control Program in China is almost achieved. However, there are multiple challenges during the stage moving towards elimination of schistosomiasis in China, including the widespread distribution of intermediate host snails and complicated snail habitats, many types of sources of Schistosoma japonicum infections and difficulty in management of bovines and sheep, unmet requirements for the current schistosomiasis control program with the currently available tools, and vulnerable control achievements. During the 14th Five-Year period, it is crucial to consolidate the schistosomiasis control achievements and gradually solve the above difficulties, and critical to provide the basis for achieving the ultimate goal of elimination of schistosomiasis in China. Based on the past experiences from the national schistosomiasis control program and the challenges for schistosomiasis elimination in China, an expert consensus has been reached pertaining to the objectives, control strategy and measures for The 14th Five-Year National Schistosomiasis Control Program in China, so as to provide insights in to the development of The 14th Five-Year National Schistosomiasis Control Program in China.

[Assessment of schistosomiasis transmission risk along the Yangtze River basin after the flood disaster in 2020].

OBJECTIVE: To evaluate the impact of the flood disaster on schistosomiasis transmission along the Yangtze River basin in 2020, so as to provide insights into schistosomiasis prevention and control in flood-affected areas. METHODS: The data pertaining to the endemic situation of schistosomiasis were collected from 5 provinces of Hunan, Hubei, Jiangxi, Anhui and Jiangsu from 2013 to 2019, including Schistosoma japonicum infections in humans and livestock and snail distribution, and the warning water levels and actual water status were collected in water regions locating in these 5 provinces. The cumulative numbers of S. japonicum egg-positive individuals and bovines during the period from 2013 to 2019, the area of snail habitats in 2019 and the water level on July 12, 2020 were estimated at a county level and employed as parameters for classification of schistosomiasis transmission risk. Then, the cumulative value of each risk index was calculated to assess the risk of schistosomiasis transmission risk. RESULTS: After the flood disaster along the Yangtze River basin in 2020, there were 10, 5 and 9 counties (districts) at high risk of schistosomiasis transmission in 5 provinces of Hunan, Hubei, Jiangxi, Anhui and Jiangsu based on number of egg-positive individuals, number of egg-positive bovines and snail distribution, respectively. Based on comprehensive risk indices, there were 10 (8 in Dongting Lake regions of Hunan Province and 2 in Poyang Lake regions of Jiangxi Province) and 15 counties (districts) (4 in Hubei Province, 7 in Hunan Province and 4 in Jiangxi Province) identified at grades 5 and 4 risk of schistosomiasis transmission. CONCLUSIONS: Dongting Lake regions and Poyang Lake regions are the most severely flood-affected schistosomiasis-endemic foci of China in 2020, and the flood disaster may facilitate the transmission of schistosomiasis in affected areas. Therefore, schistosomiasis control requires to be intensified after the flood disaster to prevent the rebound of the disease.

LncRNA CASC2 regulates high glucose-induced proliferation, extracellular matrix accumulation and oxidative stress of human mesangial cells via miR-133b/FOXP1 axis.

OBJECTIVE: Diabetic nephropathy (DN) is one of the primary complications of diabetes. Long non-coding RNA cancer susceptibility candidate 2 (CASC2) has been established to function in DN, while its role in high glucose (HG)-induced human mesangial cells (HMCs) remains limited. MATERIALS AND METHODS: The expression level of CASC2 and miR-133b was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation was assessed using cell counting kit-8 (CCK-8) assay. Extracellular matrix (ECM) accumulation was monitored through the expression levels of collagen IV (Col IV) and fibronectin (FN) using qRT-PCR and western blot analyses. Oxidative stress was observed through the expression of NADPH oxidase2 (NOX2) and the activity of malondialdehyde (MDA) and superoxide dismutase (SOD) using western blot or corresponding detection kit. The expression of forkhead box P1 (FOXP1) at mRNA and protein levels was determined by qRT-PCR and Western blot, respectively. The relationship between miR-133b and CASC2 or FOXP1 was predicted by online bioinformatics tools and verified by dual-luciferase reporter assay or RNA pull-down. RESULTS: The expression of CASC2 was reduced in serum from DN patients and HG-induced HMCs. CASC2 upregulation inhibited HG-induced HMCs proliferation, ECM accumulation and oxidative stress. MiR-133b was a target of CASC2 with a high level in serum from DN patients and HG-induced HMCs, and its enrichment reversed the effects of CASC2 upregulation. Besides, FOXP1 was a target of miR-133b with a low level in HG-induced HMCs, and its knockdown abolished the impacts of CASC2 upregulation. CONCLUSIONS: CASC2 upregulation suppressed HG-induced proliferation, ECM accumulation and oxidative stress of HMCs through miR-133b /FOXP1 regulatory axis, suggesting that CASC2 was a novel biomarker for DN treatment.

Dynamic infection of Verticillium dahliae in upland cotton.

Verticillium wilt, an infection caused by the soilborne fungus Verticillium dahliae, is one of the most serious diseases in cotton. No effective control method against V. dahliae has been established, and the infection mechanism of V. dahliae in upland cotton remains unknown. GFP-tagged V. dahliae isolates with different pathogenic abilities were used to analyse the colonisation and infection of V. dahliae in the roots and leaves of different upland cotton cultivars, the relationships among infection processes, the immune responses and the resistance ability of different cultivars against V. dahliae. Here, we report a new infection model for V. dahliae in upland cotton plants. V. dahliae can colonise and infect any organ of upland cotton plants and then spread to the entire plant from the infected organ through the surface and interior of the organ. Vascular tissue was found to not be the sole transmission route of V. dahliae in cotton plants. In addition, the rate of infection of a V. dahliae isolate with strong pathogenicity was notably faster than that of an isolate with weak pathogenicity. The resistance of upland cotton to Verticillium wilt was related to the degree of the immune response induced in plants infected with V. dahliae. These results provide a theoretical basis for studying the mechanism underlying the interaction between V. dahliae and upland cotton. These results provide a theoretical basis for studying the mechanism underlying the interaction between V. dahliae and upland cotton.

Regulation of Lnc-NTF3-5 on islet ß-cell dysfunction in high glucose environment and related mechanisms.

OBJECTIVE: The pathogenesis of diabetes is closely related to islet ß-cell dysfunction. Lnc-NTF3-5 participates in the occurrence and development of various diseases. However, Lnc-NTF3-54's effect on islet ß-cell dysfunction in high glucose environment remains unclear. MATERIALS AND METHODS: The islet ß cell MIN6 cells were cultured in vitro and randomly divided into control group, high glucose group, NTF3-5 siRNA group, and NTF3-5 group, which was respectively transfected with Lnc-NTF3-5 siRNA and Lnc-NTF3-5 plasmid under high glucose condition. Lnc-NTF3-5 expression was measured by real time PCR and cell proliferation was assessed by MTT assay. In addition, Caspase 3 activity, SOD activity, and ROS content were also detected along with the secretion of IL-10 and IL-1 by enzyme-linked immunosorbent assay (ELISA). RESULTS: Compared with control group, Lnc-NTF3-5 expression in MIN6 cells was significantly increased in high glucose environment (p<0.05). In high glucose environment, Lnc-NTF3-5 plasmid transfection up-regulated Lnc-NTF3-5 expression, inhibited cell proliferation, increased Caspase 3 activity, and decreased SOD activity. Meanwhile, Lnc-NTF3-5 plasmid also increased ROS content and IL-1 level, and decreased IL-10 level and insulin secretion. Compared with high glucose group, the differences were statistically significant (p<0.05). However, transfection of Lnc-NTF3-5 siRNA down-regulated Lnc-NTF3-5 expression under high glucose environment and reversed the above changes. Compared with high glucose group, the differences were statistically significant (p<0.05). CONCLUSIONS: Lnc-NTF3-5 expression is increased in high glucose environment. Targeting Lnc-NTF3-5 can inhibit islet cell apoptosis, oxidative stress, and promote islet cell proliferation and insulin secretion.

[Comparison of gut microbiotal compositional analysis of patients with irritable bowel syndrome through different bioinformatics pipelines].

OBJECTIVE: To assess whether the same biological conclusion, diagnostic or curative effects regarding microbial composition of irritable bowel syndrome (IBS) patients could be reached through different bioinformatics pipelines, we used two common bioinformatics pipelines (Uparse V2.0 and Mothur V1.39.5)to analyze the same fecal microbial 16S rRNA high-throughput sequencing data. METHODS: The two pipelines were used to analyze the diversity and richness of fecal microbial 16S rRNA high-throughput sequencing data of 27 samples, including 9 healthy controls (HC group), 9 diarrhea IBS patients before (IBS group) and after Rifaximin treatment (IBS-treatment, IBSt group). Analyses such as microbial diversity, principal co-ordinates analysis (PCoA), nonmetric multidimensional scaling (NMDS) and linear discriminant analysis effect size (LEfSe) were used to find out the microbial differences among HC group vs. IBS group and IBS group vs. IBSt group. RESULTS: (1) Microbial composition comparison of the 27 samples in the two pipelines showed significant variations at both family and genera levels while no significant variations at phylum level; (2) There was no significant difference in the comparison of HC vs. IBS or IBS vs. IBSt (Uparse: HC vs. IBS, F=0.98, P=0.445; IBS vs. IBSt, F=0.47,P=0.926; Mothur: HC vs.IBS, F=0.82, P=0.646; IBS vs. IBSt, F=0.37, P=0.961). The Shannon index was significantly decreased in IBSt; (3) Both workshops distinguished the significantly enriched genera between HC and IBS groups. For example, Nitrosomonas and Paraprevotella increased while Pseudoalteromonadaceae and Anaerotruncus decreased in HC group through Uparse pipeline, nevertheless Roseburia 62 increased while Butyricicoccus and Moraxellaceae decreased in HC group through Mothur pipeline.Only Uparse pipeline could pick out significant genera between IBS and IBSt, such as Pseudobutyricibrio, Clostridiaceae 1 and Clostridiumsensustricto 1. CONCLUSION: There were taxonomic and phylogenetic diversity differences between the two pipelines, Mothur can get more taxonomic details because the count number of each taxonomic level is higher. Both pipelines could distinguish the significantly enriched genera between HC and IBS groups, but Uparse was more capable to identity the difference between IBS and IBSt groups. To increase the reproducibility and reliability and to retain the consistency among similar studies, it is very important to consider the impact on different pipelines.

[A cross-sectional study on application of glucocorticoid in systemic lupus erythematosus patients].

Objective: To explore the status of glucocorticoid application in patients with systemic lupus erythematosus (SLE) in China. Methods: Epidemiological survey was used. The SLE patients who met the 1997 classification criteria of American College of Rheumatology were enrolled. The usage of glucocorticoid and related adverse reactions were recorded and analyzed. Results: A total of 400 SLE patients were enrolled, including 35 men and 365 women. The average age was (37.4±14.0) years old, and the average duration of disease was (6.7±5.8) years. There were 310 patients using glucocorticoid as maintenance. Sixty-one percent (n=244) patients started using medium dose (prednisone 30-<60 mg/d) as the initial treatment of glucocorticoid, which lasted for(37±11)days.The time of drug duration in patients with low dose prednisone (7.5-<30 mg/d)and high dose (60-100 mg/d) was(92±20)and(17±3)days respectively (P<0.05 between 3 groups). However, patients receiving different initial dosage were of no discrepancy in the maintenance therapy. During maintenance, even though 51.0% (n=158) patients were on prednisone 2.5-5 mg/d, the duration of drug use in >5-10 mg/d groupwas longer[(29.9±3.3) months]. Patients with involvement of internal organs had a higher tendency to use 60-100 mg/d prednisone or pulse-dose therapy in the initial treatment, nevertheless these two groups had no difference of maintenance dosage. Among all 400 patients, 62 patients withdrew glucocorticoid, including 17 patients with disease remission (4.3%), 44 by self-withdrawal and one with adverse reaction. Conclusion: In China, the medium dosage of glucocorticoid is the most common initial treatment in patients with SLE.Prednisone 2.5-5 mg/d was the most common choicefor maintenance therapy. Currently, the proportion of glucocorticoid withdrawal remains low in SLE patients achieving remission.

Molecular cloning, overexpression, purification, and sequence analysis of the giant panda (Ailuropoda melanoleuca) ferritin light polypeptide.

The complementary DNA (cDNA) of the giant panda (Ailuropoda melanoleuca) ferritin light polypeptide (FTL) gene was successfully cloned using reverse transcription-polymerase chain reaction technology. We constructed a recombinant expression vector containing FTL cDNA and overexpressed it in Escherichia coli using pET28a plasmids. The expressed protein was then purified by nickel chelate affinity chromatography. The cloned cDNA fragment was 580 bp long and contained an open reading frame of 525 bp. The deduced protein sequence was composed of 175 amino acids and had an estimated molecular weight of 19.90 kDa, with an isoelectric point of 5.53. Topology prediction revealed one N-glycosylation site, two casein kinase II phosphorylation sites, one N-myristoylation site, two protein kinase C phosphorylation sites, and one cell attachment sequence. Alignment indicated that the nucleotide and deduced amino acid sequences are highly conserved across several mammals, including Homo sapiens, Cavia porcellus, Equus caballus, and Felis catus, among others. The FTL gene was readily expressed in E. coli, which gave rise to the accumulation of a polypeptide of the expected size (25.50 kDa, including an N-terminal polyhistidine tag).

Development and Application of Diagnostics in the National Schistosomiasis Control Programme in The People's Republic of China.

Schistosomiasis, caused by Schistosoma japonicum infection to human, has a documented history of more than 2100years in The People's Republic of China. In spite of great progress in controlling the disease, it is still one of the most serious parasitic diseases in the country. The study and use of diagnostic techniques play an important role in the targeting of chemotherapy that has been continuously applied in the national schistosomiasis control programme for several decades. This paper reviews the development and application of parasitological, immunodiagnostic and molecular diagnostic technology for S. japonicum in The People's Republic of China with a brief mention of diagnostic imagery, such as ultrasound and radiology. When analysing the efficacy and performance characteristics of the main diagnostic techniques in current use, it becomes apparent that approaches that worked well in the past are less suitable now as successful control has shifted the endemic situation towards control and interruption of transmission. The conclusion is that a mutable approach must be adopted choosing the most appropriate diagnostic technique for each control stage (and area), thus modifying the methodology according to the prevailing diagnostic needs in terms of sensitivity and specificity.

[The influence of depression degree on regulatory T cells in patients with non-M3 acute myeloid leukemia].

Objective: To investigate the influence of depression levels on regulatory T cells (Tregs) in patients with non-M3 acute myeloid leukemia (AML). Methods: A total of 63 patients with primarily diagnostic non-M3 AML and 25 healthy controls were enrolled, and the levels of depression by using HADM score and the percentages of Tregs by flow cytometry were evaluated in pre-treatment and post-treatment, respectively. Results: After every course of chemotherapy, the percentages of Tregs of PBMNC in AML showed the higher level of (6.48±1.81)% than those of (4.99±1.29)% in control (P= 0.001). There was no difference among different levels of depression groups after the first cycle. However, the percentages of Tregs increased with the worse of depression after the second and third cycles. Partial correlation analysis after adjusting age indicated that the percentages of Tregs showed no correlation with the levels of depression after the first cycle (correlation coefficient, 0.120, P=0.345), and showed the positive correlation with depression levels after the second and third cycles (correlation coefficient, 0.619 and 0.614, all P values <0.05). Conclusion: The depression levels showed an association with the percentages of Tregs in patients with non-M3 AML, which could be observed only after the second cycles of chemotherapy.

Impaired function of CD19(+) CD24(hi) CD38(hi) regulatory B cells in patients with pemphigus.

BACKGROUND: Pemphigus is an organ-specific autoimmune bullous disease. OBJECTIVES: To determine the role of regulatory B cells (Bregs) in patients with pemphigus. METHODS: The frequency of the occurrence of CD19(+) CD24(hi) CD38(hi) Bregs was detected from 34 patients with pemphigus and 20 healthy controls. Interleukin (IL)-10 secretion was processed after stimulating B cells. Specific antidesmoglein antibody (Ab) titres and their subclasses were also measured. Ab response and cytokine production from peripheral blood mononuclear cells (PBMCs) with or without Bregs were analysed. RESULTS: The number of Bregs was significantly increased in patients with pemphigus compared with healthy controls (15 ± 7% vs. 9 ± 3%; P < 0·01) and the proportion of Bregs in the active groups (newly diagnosed and chronic active patients) was significantly higher than in remittent individuals (16 ± 7% vs. 13 ± 8%; P = 0·04). The IL-10-producing B cells were significantly increased upon stimulation both in patients and in healthy controls. However, the increase ratio of IL-10-producing B cells between short- and long-term stimulation was significantly lower in patients with pemphigus (1·0-fold vs. 2·6-fold increase in control group; P < 0·01). Strikingly, Bregs from the controls were able to suppress interferon (IFN)-γ expression and T helper cell 1 (Th1) immune response (26% inhibition rate), while the suppressive function of Bregs from patients with pemphigus was significantly decreased (9% inhibition rate). There was no difference in Ab levels from PBMCs with or without Bregs after stimulation. CONCLUSIONS: Bregs in patients with pemphigus are elevated but with defective regulatory function on Th1 cells.

Overexpression, purification, and pharmacologic evaluation of anticancer activity of ribosomal protein L24 from the giant panda (Ailuropoda melanoleuca).

The ribosomal protein L24 (RPL24) belongs to the L24E family of ribosomal proteins and is located in the cytoplasm. The purpose of this study was to investigate the structure and anti-cancer function of RPL24 of the giant panda (Ailuropoda melanoleuca). The complementary DNA of RPL24 was cloned successfully using reverse transcription-polymerase chain reaction technology. We constructed a recombinant expression vector containing RPL24 complementary DNA and overexpressed it in Escherichia coli using pET28a plasmids. The expression product obtained was purified using Ni-chelating affinity chromatography. The results indicated that the length of the fragment cloned is 509 bp, and it contains an open-reading frame of 474 bp encoding 157 amino acids. Primary structure analysis revealed that the molecular weight of the putative RPL24 protein is 17.78 kDa with a theoretical isoelectric point of 11.86. The RPL24 gene is readily expressed in E. coli, and the RPL24 fused with the N-terminal histidine-tagged protein to give rise to the accumulation of an expected 23.51-kDa polypeptide. The inhibitory rate in mice treated with 0.1 mg/mL RPL24, the highest of 3 doses administered, can reach 67.662%, which may be comparable to the response to mannatide. The histology of organs with tumors showed that the tissues in the RPL24 group displayed a looser arrangement compared with that in the control group. Furthermore, no obvious damage was apparent in other organs, such as heart, lung, and kidney. The data showed that the recombinant RPL24 had time and dose dependency on the cell growth inhibition rate. Human laryngeal carcinoma Hep-2 cells treated with 0.3125-10 µg/mL RPL24 for 24 h displayed significant cell growth inhibition (P < 0.05; N = 6) in assays using 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyltetrazolium bromide compared with that in control (untreated) cells. By contrast, human hepatoma Hep G-2 cells displayed no significant change (P > 0.05; N = 6) from control (untreated) cells. RPL24 has time and dose dependency on Hep-2 cell growth inhibition. The data indicate that the effect at low concentrations is better than that at high concentrations, and the concentration of 0.625 µg/mL provides the best rate of growth inhibition. Further research is ongoing to determine the bioactive principles of recombinant RPL24 protein that are responsible for its anticancer activity.

Verticillium Wilt of Redbud in China Caused by Verticillium dahliae.

Chinese redbud (Cercis chinensis Bunge), a member of the Fabaceae, is an important ornamental plant native to China with reported desirable medicinal effects, including stimulating blood circulation, detumescence, and detoxification (1). In October 2011, wilt symptoms of gradual leaf yellowing, wilting, scorching (marginal browning), and twig dieback were observed on plants in Yangling, Shaanxi, China. The incidence of diseased plants was about 20% in two main areas (about 20 ha in Zijingshan Park in Zhengzhou, Henan Province; and about 3,000 ha in Taiping National Forest Park in Xi'an, Shaanxi Province). Wilted leaves of diseased plant senesced and died, but defoliation was not observed. Brown discoloration was observed in vascular tissues of petioles, twigs, and stems of diseased plants, sometimes in a ring pattern. The symptoms were often restricted to the lower part of the tree or a few branches. To identify the causal agent, six twigs (each approximately 50 mm in diameter and 10 cm long) sampled from an infected tree in Yangling were rinsed in running water, surface-sterilized with 75% ethanol for 2 min, rinsed in sterilized water three times, dried, cut into 1 cm long segments, and the segments put onto potato dextrose agar (PDA) medium. A fungal isolate was recovered from diseased vascular tissues of each sample when cultured on PDA in the dark at 25°C. After 5 days, colonies changed from white to black as a result of production of microsclerotia. Microscopic observation revealed that conidiophores were hyaline and verticillate, with three to four phialides at each node. Conidia were ellipsoidal, hyaline, single-celled, and 2.5 to 7.5 × 1.25 to 4.5 µm. On the basis of these morphological characteristics, the fungus was identified as Verticillium dahliae (3). To prove Koch's postulates, the roots of 10 healthy, 1-year-old C. chinensis plants were each irrigated in a greenhouse with 50 ml of a conidial suspension (1.0 × 107 spores/ml) of an isolate recovered from an infected plant (2); five control plants were inoculated similarly with sterilized water. Fifteen days after inoculation, the same wilt symptoms observed on the original plants had developed on 9 of the 10 inoculated plants, whereas the control plants remained healthy. The pathogen was recovered 15 days after inoculation by isolating from petiole and stem tissues of symptomatic plants, but was not isolated from the control plants. The internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2) of the nuclear ribosomal DNA was PCR-amplified with primers ITS1 and ITS4 (4), and sequenced. BLAST analysis of the ITS sequence (GenBank Accession No. AB735536) showed 100% homology with that of an isolate of V. dahliae (FJ572050). To our knowledge, this is the first report of Verticillium wilt on C. chinensis in China. References: (1) Y. Li et al. J. Integr. Plant Biol. 47:1021, 2005. (2) H. A. Melouk and C. E. Horner. Phytopathology 65:767, 1975. (3) G. F. Pegg and B. L. Brady. Verticillium Wilts, CABI Publishing, Oxford, UK, 2002. (4) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. M. A. Innis et al., eds. Academic Press, San Diego, CA, 1990.

Pretreatment with bilirubin protects islet against oxidative injury during isolation and purification.

BACKGROUND: A high yield of pure, viable islets is one of the most important prerequisites for successful islet transplantation. However, during isolation and purification, many factors may cause oxidative stress, impacting islet viability. Accumulating evidence indicates that bilirubin (BR) not only has antioxidative but also has cytoprotective activities. In this study, we investigated whether pretreatment with bilirubin would protect islets against oxidative damage during isolation and purification. METHODS: Wistar rats were randomly divided into control and BR groups. The latter rats received an injection of BR 2 hours before islet isolation, whereas the controls received vehicle. Islet purity was determined using a dithizone stain. Survival rate and viability were determined using acridine orange and propidium iodide staining and the Cell Counting Kit-8 Kit. Islet function was quantified by testing glucose-stimulated insulin secretion. Islet damage caused by oxidative stress was quantified by measuring the malondialdehyde (MDA) in freshly isolated islets. RESULTS: Pretreatment with bilirubin did not enhance the purity, but significantly enhanced the survival rate and viability of the islets. Islet function in the BR group was significantly better than that in the control cohort. The MDA level was 0.62 ± 0.23 nmol/L/µg protein in the BR group, which was significantly lower (P < .05) than that in controls (1.31 ± 0.34 nmol/L/µg protein). CONCLUSIONS: We concluded that oxidative stress during islet isolation and purification can be mitigated by BR pretreatment. BR exerts antioxidant and cytoprotective properties by reducing lipid peroxidation (MDA) and enhancing islet viability and function. Pretreatment with BR may become a simple, clinical applicable means to improve human islet isolation and transplantation outcomes.