Protective effect and mechanism of nanoantimicrobial peptide ND-C14 against Streptococcus pneumoniae infection / 世界急诊医学杂志（英文）

@#BACKGROUND: Streptococcus pneumoniae (S. pneumoniae) is a common pathogen that causes bacterial pneumonia. However, with increasing bacterial resistance, there is an urgent need to develop new drugs to treat S. pneumoniae infections. Nanodefensin with a 14-carbon saturated fatty acid (ND-C14) is a novel nanoantimicrobial peptide designed by modifying myristic acid at the C-terminus of human α-defensin 5 (HD5) via an amide bond. However, it is unclear whether ND-C14 is effective against lung infections caused by S. pneumoniae. METHODS: In vitro, three groups were established, including the control group, and the HD5 and ND-C14 treatment groups. A virtual colony-count assay was used to evaluate the antibacterial activity of HD5 and ND-C14 against S. pneumoniae. The morphological changes of S. pneumoniae treated with HD5 or ND-C14 were observed by scanning electron microscopy. In vivo, mice were divided into sham, vehicle, and ND-C14 treatment groups. Mice in the sham group were treated with 25 µL of phosphate-buffered saline (PBS). Mice in the vehicle and ND-C14 treatment groups were treated with intratracheal instillation of 25 µL of bacterial suspension with 2×108 CFU/mL (total bacterial count: 5×106 CFU), and then the mice were given 25 μL PBS or intratracheally injected with 25 μL of ND-C14 (including 20 µg or 50 µg), respectively. Survival rates were evaluated in the vehicle and ND-C14 treatment groups. Bacterial burden in the blood and bronchoalveolar lavage fluid were counted. The lung histology of the mice was assessed. A propidium iodide uptake assay was used to clarify the destructive effect of ND-C14 against S. pneumoniae. RESULTS: Compared with HD5, ND-C14 had a better bactericidal effect against S. pneumoniae because of its stronger ability to destroy the membrane structure of S. pneumoniae in vitro. In vivo, ND-C14 significantly delayed the death time and improved the survival rate of mice infected with S. pneumoniae. ND-C14 reduced bacterial burden and lung tissue injury. Moreover, ND-C14 had a membrane permeation effect on S. pneumoniae, and its destructive ability increased with increasing ND-C14 concentration.

Clinical study of serum human-βeta-defensin-1 level for evaluating short-term prognosis in patients with acute-on-chronic liver failure / 中华肝脏病杂志

Objective: To evaluate the diagnostic value of serum human-βeta-defensin-1 level (HBD-1) for short-term (28-day) prognosis in patients with acute-on-chronic liver failure (ACLF). Methods: Fifty cases diagnosed with ACLF were selected. 20 cases with decompensated cirrhosis and 20 cases with compensated cirrhosis who were admitted at the same time were included. Age, gender, serum HBD-1 level, C-reactive protein (CRP), procalcitonin (PCT), neutrophil count/lymphocyte ratio (NLR), blood routine, coagulation function, liver function, kidney function, and other indicators from the three groups of patients were collected. Patients with ACLF were screened for indicators related to the short-term (28-day) prognosis. Patients were divided into an improvement group and a worsening group according to the 28-day disease outcome. The serum HBD-1 level and other above-mentioned indicators were compared between the two patient groups. The receiver operating characteristic (ROC) curve was used to analyze the diagnostic efficacy of serum HBD-1 levels for short-term prognosis in patients with ACLF. PCT, NLR, and prothrombin activity (PTA) application as a mono indicator and HBD-1 in combination with NLR, PCT, and PTA were compared to evaluate diagnostic efficacy for short-term prognosis in patients with ACLF. The intergroup mean of measurement data was determined using a t-test or analysis of variance. χ (2) test was used for comparison of count data. Spearman's rank correlation analysis was used for correlation analysis. Results: There was no statistically significant difference in age and gender among the three groups: ACLF, decompensated cirrhosis, and compensated cirrhosis (P > 0.05). The expression levels of serum HBD-1 in the ACLF group, decompensated cirrhosis group, and compensated cirrhosis group were (319.1 ± 44.4) ng/ml, (264.5 ± 46.5) ng/ml and (240.1 ± 35.4) ng/ml, respectively, while the ACLF group expression levels were significantly increased, with statistical significance (P < 0.01).The serum HBD-1 level was significantly higher in the ACLF worsening group (346.2 ± 43.6) ng/ml than that in the improvement group (308.5 ± 40.6) ng/ml, and the difference was statistically significant (P < 0.05). Correlation analysis showed that HBD-1, NLR, PCT, prothrombin time (PT), and international standardized ratio (INR) were negatively correlated with the 28-day disease outcome (improvement) of patients (P < 0.05). PTA was positively correlated with 28-day disease outcome (improvement) (P < 0.05). The area under the receiver operating characteristic curve (AUC) for evaluating HBD-1's diagnostic efficacy for short-term prognosis in patients with ACLF was 0.774, with a sensitivity of 0.750, a specificity of 0.786, and a cut-off point of 337.96 ng/ml. PCT, NLR, and PTA had greater diagnostic efficacy. HBD-1 combined with PTA had the highest diagnostic efficacy, with an AUC of 0.802, a sensitivity of 0.778, and a specificity of 0.786. The diagnostic efficacy of HBD-1+PCT, HBD-1+NLR and HBD-1, PCT, and NCR was superior to PTA mono. Conclusion: The serum HBD-1 level gradually increases with the aggravation of liver function injury and is negatively correlated with the short-term prognosis in patients with ACLF. Serum HBD-1 level has high sensitivity and specificity in predicting short-term prognosis in patients with ACLF, and its diagnostic efficacy is superior to that of PCT, NLR, and PTA. The combined application of HBD-1 and PTA has higher diagnostic efficacy; however, when the serum HBD-1 level is greater than 337.96ng/ml, it indicates poor prognosis in patients.

Extraction and bioactivity analysis of total protein of K326 tobacco leaves with high expression of Nicotiala alata defensin 1 gene / 中国生物制品学杂志

@#Objective To extract the total protein of K326 tobacco leaves with high expression of Nicotiana alata defensin 1(NaD1) gene and analyze its bioactivity.Methods Total proteins were extracted from Nicotiana alata flowers,wild type(WT) and K326 tobacco leaves(transgenic) with high expression of NaD1 gene,and determined for the concentrations by Bardford method,while for the antibacterial activity against fungi by filter paper method,and for the inhibition activity on cancer cells(HeLa cells) by CCK-8.Results The total protein concentrations of Nicotiana alata flowers,WT and transgenic K326 tobacco leaves were 11.25,10.33 and 10.14 mg/mL,respectively.The antibacterial activity of total protein from transgenic K326 tobacco leaves against Candida albicans was(85.68±3.08)%,which was 1.33 and 1.14 times that of total protein from WT K326 tobacco leaves and Nicotiana alata flowers,respectively(F=15 339,P <0.05);The antibacterial activity against Fusarium oxysporum was(148.48±2.47)%,which was 1.09 and 1.08 times that of total protein from WT K326tobacco leaves and Nicotiana alata flowers,respectively(F=4.927,P <0.05).The IC_(50) value of transgenic K326 tobacco leaf protein on HeLa cells was the smallest(6.11 mg/mL),and the inhibitory activity was 1.56 and 1.21 times that of total protein of WT K326 tobacco leaves and Nicotiana alata flowers,respectively(F=89.748,P <0.05).Conclusion The total protein of K326 tobacco with high expression of NaD1 gene has good antibacterial and anticancer bioactivities,which provides an experimental basis for producing antibacterial and anticancer biological agents with tobacco as bioreactor.

Clinical Efficacy of Maxingshigantang Enema in Treating Infants with Pneumonia and Asthma and Intervention Mechanism of Lung-intestine Microenvironment / 中国实验方剂学杂志

ObjectiveTo observe the clinical efficacy of Maxingshigantang enema in the treatment of infant viral pneumonia by comparing related indicators， and comprehensively evaluate the effect of traditional Chinese medicine （TCM） enema on the intestinal microenvironment. MethodSixty infants with viral pneumonia were selected and randomly divided into 3 groups. The dosage of enema drugs in high- （0.117 g·mL-1） and low-concentration （0.07 g·mL-1） TCM enema groups was same （3.5 g per time）， and the control group received normal saline enema， once a day for 7 days. Finally， the curative effect， total symptom score， salivary secretory immunoglobulin A （sIgA）， human beta defensin 2 （hBD2） and fecal calprotectin （CALP） of each group were statistically analyzed by SPSS 21.0， and the clinical efficacy of TCM enema in treating children with pneumonia and asthma was comprehensively evaluated. ResultThe curative effect of high-concentration TCM enema group （total effective rate 100%， χ2=7.059） was equivalent to that of low-concentration TCM enema group （total effective rate 95%， χ2=4.329）， higher than that of control group （total effective rate 70%） （P<0.017）. After treatment， compared with control group and low-concentration TCM enema group， high-concentration TCM enema group had higher total symptom score of children （P<0.05， P<0.01）. The proportion of coccobacillus was reduced in three groups， with high- and low-concentration TCM enema groups lower than control group （P<0.05）. The salivary sIgA concentration was increased in three groups （P<0.05）， with high-concentration TCM enema group higher than the other groups （P<0.01）. The hBD2 concentration was decreased in three groups， with high- and low-concentration TCM enema groups lower than control group （P<0.05）. The three groups reduced the fecal CALP concentration， and high-concentration TCM enema group had the highest reduction， followed by low-concentration TCM enema group （P<0.01）. ConclusionTCM enema outweighs western medicine in improving clinical symptoms， intestinal flora， and mucosal immune function， and reducing inflammation in children， and the high-concentration TCM enema group has better curative effect. Therefore， with easiness to operate， high compliance， and significant therapeutic effect， TCM enema is worthy of clinical promotion.

Vitamin D and COVID-19

The ongoing COVID -19 pandemic is caused by severe acute respiratory syndrome corona virus -2 (SARS-CoV-2). Since its emergence in Wuhan in Hubei province of China in December 2019, the virus has spread to every continent except Antartica. Currently, there is no registered treatment or vaccine for the disease. In the current scenario of the deadly virus spreading across continents and the absence of a specific treatment of novel corona virus, there is an urgent need to search for alternative strategies to prevent and control the rapid replication of virus. Vitamin D supplementation may reduce the incidence, severity and risk of death from pneumonia (consequent to the cytokine storm) in the current COVID pandemic. Through its effect on innate and adaptive immunity, vitamin D can reduce the risk of viral respiratory tract infections. 1, 25(OH) vitamin D directly stimulates the production of anti-microbial peptides like defensin and Cathelicidin that can reduce the rate of viral replication. In addition, it can also reduce the concentration of pro-inflammatory cytokines that are responsible for causing cytokine storm and resultant fatal pneumonia. In order to reduce the risk of infection especially in developing country like India, it is recommended that people at risk of COVDI19 may be considered for vitamin D supplementation.

Serum defensin levels in patients with systemic sclerosis

Abstract Background: Systemic sclerosis (SSc) is an autoimmune disease characterized by fibrosis of skin and lung as well as involvement of kidney, gastrointestinal system and heart. Aetiology and exact mechanism of disease is poorly understood. The association between antimicrobial peptides (AMPs) and other diseases such as idiopathic pulmonary fibrosis, diffuse panbronchiolitis, pulmoner alveolar proteinosis and psoriasis have been reported. A small number of studies have examined the role of AMPs on autoimmune diseases which has not been studied in scleroderma yet. We aimed to investigate AMP serum levels and their association with disease characteristics of SSc. Methods: Forty-two patients (40 female, mean age 42 years) and 38 healthy subjects (32 female, mean age 38 years) were enrolled. For SSc patients, the following data were recorded: disease subset (limited/diffuse), autoantibodies (antinuclear, anti-centromere (ACA), and anti-SCL-70), blood tests, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP), modified Rodnan skin score, presence and history of digital ulcers, kidney, gastrointestinal disease and lung involvement assessed by computed tomography and pulmonary function tests. Association between serum AMPs and disease characteristics were analysed. Results: Twenty-nine of the patients had diffuse (69%) and 13 of the patients had limited (31%) systemic sclerosis. Average disease duration was 5.5 years. Pulmonary involvement was detected in 20 patients (47.6%). Serum concentration of alpha defensin was higher than healthy subjects (563 ± 415 vs 377 ± 269 ng/mL, p = 0.02). However, no difference was observed for beta-1 and beta-2 defensins in SSc patients and healthy controls. In sub-group analysis patients with interstitial lung disease had higher levels of alpha defensin than those without lung involvement (684 ± 473 vs 430 ± 299 ng/ml, p = 0.04). There was also correlation between alfa defensin serum concentrations and CRP (r = 0.34). Conclusions: Alpha defensin levels are increased in scleroderma patients and correlated with lung involvement indicating a role in the pathogenesis of disease. Trial registration: This study is not a clinical trial study.(AU)

Antineoplastic mechanism of antimicrobial peptides: Selective membrane destruction and non-membrane dissolution / 中国组织工程研究

BACKGROUND: Antimicrobial peptides, an extract from nature, are a basic component of host immunity that make toxic effect on highly proliferative cells. They have attracted extensive attention of scientists. The understanding of the antineoplastic mechanism of antimicrobial peptides can contribute to its application in clinical practice. OBJECTIVE: To summarize the research advances in antineoplastic mechanism of antimicrobial peptides. METHODS: The first author conducted a computer-based retrieval of PubMed, Springerlink, Web of Science, and ScienceDirect databases for relevant articles published from January 2015 to May 2019. The keywords were “antimicrobial, peptide, antitumor mechanisms, antitumor activity and anti-neoplastic”. The articles concerning antineoplastic mechanism of antimicrobial peptides and research progress were selected. RESULTS AND CONCLUSION: Cationic antimicrobial peptides synthesized by ribosomes and the host defense peptides can interact with the membrane of bacteria, which showed a broad-spectrum antimicrobial activity. Compared with normal cells, the proportion of phosphatidylserine on the surface of cancer cells, which is negatively charged, is increased dramatically. As a result, the cationic amphiphilic peptides are good candidate for the antineoplastic drugs, and possess a high selectivity. There are two major antitumor mechanism of antimicrobial peptides, which are selective membrane destruction and non-membrane dissolution (α-defensin-1 and lactoferrin B). The clinical application of antimicrobial peptides against tumors is mainly restricted by their stability and the ways to administration. By optimizing its structure and drug delivery systems, these antimicrobial peptides will play a critical role in the cancer treatment.

Expression of Ietalurus punetaus β-defensin based on recombinant Pichia pastoris / 生物工程学报

β-defensin is a primary protein immune factor in channel catfish's (Ietalurus punetaus) resistance to pathogenic microorganisms. Its primary structure contains a signal peptide composed of 24 amino acid residues at the N-terminal and a mature peptide composed of 43 amino acid residues at the C-terminal. The mature peptide region is responsible for the biological activity of β-defensin. In the present study, a recombinant strain of Pichia pastoris that produces channel catfish β-defensin, was constructed to realize the biosynthesis of channel catfish β-defensin based on eukaryotic expression. First, the β-defensin gene "IPBD" was isolated from the skin of channel catfish by RT-PCR. After linking it with the expression vector pPICZA, pPICZA-IPBD was transferred into competent P. pastoris X-33 cells to obtain recombinant P. pastoris strains. The yeast transformants with multi-copy gene inserts were obtained by using the culture medium containing 1 000 μg/mL zeocin. Using BMM culture medium (without amino nitrogen culture medium) instead of BMMY culture medium (with amino nitrogen culture medium), the fermentation and culture conditions of the recombinant strain were optimized, and the optimal conditions for producing channel catfish β-defensin were determined as follows: the expression was induced for 96 h with 1.0% methanol at 28 °C , 250 r/min. Purified protein with molecular weight of 5.98 kDa was obtained by nickel affinity chromatography, and MALDI-TOF/TOF mass spectrometry proved that it was the expected recombinant IPBD. The antibacterial test results showed that the inhibitory rates of recombinant IPBD on Gram-positive Staphylococcus aureus and Listeria monocytogenes and Gram-negative Pseudomonas aeruginosa were 69.6%, 71.6% and 65.8%, respectively. This study provides a recombinant DNA technique for the development of small molecule natural antibacterial peptide from fish.

Research progress in prognostic biomarkers of liver failure / 中国综合临床

Liver failure is a group of serious clinical syndrome, which develops rapidly and has a dangerous prognosis.In recent years, there are many biomarkers to evaluate the prognosis of patients with liver failure at home and abroad, such as neutrophil/lymphocyte ratio, alpha fetoprotein, galactose lectin-3, osteopontin, Golgi protein 73, human β - defensin-1, etc.these biomarkers are of great significance for early identification of patients with liver failure, accurate evaluation of their prognosis, and formulation of effective treatment plan.

The Relation Between Decay Missing Filled-Teeth (DMF-t), Body Mass Index (BMI) with Salivary Human Beta-3 (HBD-3) Secretion in Children with Caries and Free Caries

@#Introduction: Dental caries in children is a major problem of mouth disease throughout the world, so too is there currently an increase in health problems in children due to obesity. Human Beta defensing(HBD) has been found in saliva and from several studies stated that HBD aside from being a broad-spectrum antimicrobial can act as an immunomodulator. The purpose of this study is to reveal whether there is a relationship between obesity and HBD-3 salivary concentration in caries patients and caries-free patients. Methods: This cross-sectional observational study was involved 62 children with caries and caries-free, aged 9-11 years, students at Qommarudin Islamic Boarding School, Gresik, East Java Indonesia. dental caries examination, carried out in accordance with World Health Organization (WHO) diagnostic criteria. Body mass index (BMI) was measured from the height and weight of individuals, HBD3 concentrations were tested with an ELISA kit from Bioassay Technology Laboratory (China) from saliva samples. Evaluate the results with the Kruskal Wallis test, followed by the Mann-Whitney test. The level of significance used in this statistical test was 0.05. Results: there was a relationship between BMI level and HBD-3 concentration in the caries group (p <0.05, p = 0.009) with a moderate level of association. but there was no significant relationship in the caries-free group (p> 0.05, p = 0.189). Conclusion: There was an association between BMI and HBD-3 salivary concentration in caries patients but there was no relationship in the caries-free group.

Effect of icariin on early βdefensin2 and T cell subsets in rats after tracheotomy / 中南大学学报(医学版)

OBJECTIVES@#To investigate the effect of icariin (ICA) on early β-defensin-2 and T cell subsets in rats after tracheotomy.@*METHODS@#A total of 54 SPF male Sprague-Dawley rats were randomly divided into a normal control group (group A), a model group (group B), and a model+ICA treatment group (group C), with 18 rats in each group. A tracheotomy intubation model of the B and C group was prepared. After 6 h of surgery, ICA intervention was given to group C. Groups A and B were given the same amount of normal saline. Lung tissue, alveolar lavage fluid and peripheral blood were taken at 24 h, 72 h and 168 h, respectively. The expression of rat β-defensin-2 mRNA in lung tissue was detected by RT-PCR. The content of β-defensin-2 in alveolar lavage fluid and peripheral blood serum was detected by ELISA. The content of peripheral blood T cell subsets (CD3, CD4, CD8) was detected by flow cytometry, and the ratio of CD4/CD8 was calculated.@*RESULTS@#After tracheotomy, the levels of β-defensin-2 mRNA and β-defensin-2 in lung tissue from the group B were increased significantly at 24 h, then they were decreased gradually, and decreased most significantly at 168 h (0.05). The level of CD3 T cells in peripheral blood was significantly lower than that in the group A (0.05). After ICA intervention in group C: lung tissue, alveolar lavage fluid, peripheral blood serum β-defensin-2 content, and peripheral blood CD3 and CD4 T cell levels were gradually increased, significantly higher than those in the group B (<0.05). CD8 T cell level was significantly lower than that in the group A at 24 h (<0.05), the CD4/CD8 ratio was significantly higher at 168 h than those in the group A or B (both <0.01).@*CONCLUSIONS@#ICA can improve the early lung immune function in rats with tracheotomy, which might be related to up-regulation of β-defensin-2 in lung tissue and alveolar lavage fluid, concomitant with increases in CD3 and CD4 T cells and CD4/CD8 ratio in peripheral blood while reduction in CD8 cells.

Recombinant expression of Crassostrea gigas defensin in Pichia pastoris and its antibacterial activity / 生物工程学报

Defensins are endogenous cationic antimicrobial peptides rich in arginine and cysteine residues. They are important immune factors resisting pathogenic bacteria infection for mollusks. The 43 amino acid residues near the carboxyl terminal for Crassostrea gigas defensin (CgD) form its mature peptide region, responsible for the biological activity of CgD. First, two target genes, CgDH⁺ (with 6×His-tag at 3' end) and CgDH- (without 6×His-tag at 3' end) were separated and amplified by RT-PCR with specific primers from Crassostrea gigas mantle. These two target genes were ligated to the expression vector pPICZαA to construct recombinant expression vectors, pPICZαA-CgDH⁺ and pPICZαA-CgDH-, which were transformed into competent Pichia pastoris X-33 cells by electroporation respectively. The recombinant target proteins, CgDH⁺ and CgDH-, were induced for 72 h with 1% methanol at 29 °C and 250 r/min. The recombinant CgDH⁺ (5.78 kDa) was purified by immobilized metal affinity chromatography (IMAC), and identified by MALDI-TOF-TOF analysis, demonstrating that it was the expected target protein. Based on the concentration of the purified product, the estimated yield of recombinant CgDH⁺ was 2.32 mg/L. Antimicrobial assay showed that the culture medium supernatant containing recombinant CgDH⁺ and recombinant CgDH-, respectively, had activities against Staphylococcus aureus and Pseudomonas aeruginosa, indicating that the existence of 6×His tag in the recombinant proteins do not affect their biological activities.

Tandem expression, purification and biological activity of recombinant multimeric β-defensin130 in Escherichia coli / 生物工程学报

To improve and broaden the antimicrobial activity of β-defensin130, 3 copies of β-defensin130 encoding sequences were synthesized and cloned into pET28a (+) expression vector, and expressed in Escherichia coli BL21 (DE3) as a 25 kDa soluble protein. The affinity purified 3×β-defensin 130 displayed antimicrobial activity against not only Gram-positive strains including Staphylococcus aureus (ATCC 25923) (45 μg/mL) and Listeria monocytogenes (ATCC 221633) (80 μg/mL) but also Gram-negative strains. Furthermore, the antimicrobial activity of β-defensin130 was not affected by temperature, pH and proteinase digestion. In addition, E. coli-derived 3×β-defensin130 was not toxic to HEK 293 cells and showed a relatively low hemolytic activity against rabbit erythrocytes. Our study proves 3×β-defensin130 expressed in E. coli is stable, non-cytotoxic and low-hemolytic active with great potential as alternative antibiotics.

Expressions of intestinal alpha-defensin-5, beta-defensin-2 in acute liver failure / 中华实用儿科临床杂志

Objective To investigate the dynamic changes in intestinal alpha-defensin-5 (RD-5),beta-defensin-2 (BD-2) mRNA after acute liver failure(ALF),and to explore their role in ALF.Methods A total of 60 C57BL5 mice were divided into 4 groups by means of random number table method:normal control group,ALF group,E.coli via gavage group and ALF + E.coli via gavage group.Intraperitoneal injection of D-galactosamine (500 mg/kg) and lipopolysaccharide(10 μg/kg) to make the model,in addition,ALF mice were fed with E.coli,and the observation time was 6 hours,12 hours,and 24 hours after modeling,and each time point had 6 specimens.Real-time PCR was used to test the RD-5 mRNA and BD-2 mRNA levels in the ileum tissue.Results The levels of RD-5 and BD-2 showed dynamic change in the experiment of ALF.Compared with the levels of RD-5 and BD-2(11.25 ±0.74,23.86 ±0.39) of the normal control group,the levels of RD-5 and BD-2 in ALF group and E.coli via gavage group increased at 6 hours after modeling(14.19 ±0.39,26.79 ± 0.36 and 12.57 ± 0.68,26.45 ± 0.85),and the differences were significant(all P＜0.05);at 12 hours after modeling,the RD-5 and BD-2 reached to the maximum concentration(15.76 ±0.33,29.10 ± 0.61 and 12.90 ± 0.96,27.42 ± 0.71),and the differences were statistically signi-ficant (all P ＜ 0.05).The degree of elevation of BD-2 was higher than RD-5.Later,they gradually declined.Conclusions RD-5 and BD-2 may play an important role in the pathogenesis of intestinal endotoxemia in experimental ALF.

Role of human βdefensin 2 on preventing oxidized low-density lipoprotein induced monocyte foaming / 中华传染病杂志

Objective To clarify the role of human β-defensin2 ( hBD2) on preventing oxidized low-density lipoprotein (OX-LDL) induced human leukemic monocyte (THP-1) foaming.Methods The monocyte foaming model was established using THP-1 cell induced by OX-LDL and the model was identified by oil red staining.The hBD2 was overexpressed on THP-1 cells by using lentivirus system and the effect of hBD 2 overexpression on THP-1 cell foaming induced by OX-LDL was detected.The levels of inflammatory factors including tumor necrosis factor (TNF)-α, interleukin (IL)-1βand IL-6 in cell supernatant of each group were detected by enzyme-linked immunosorbent assay ( ELISA).Differences between the groups were compared by using the t test.Results The gene transfection efficiency of the cells was close to 100%at 72 h after infection. The hBD2 protein levels were 0.122 ±0.024 in the control group, 0.123 ±0.022 in Lv-control infection group and 0.981 ±0.183 in Lv-hBD2 infection group; and the level in control group was statistically higher than that in hBD-2 infection group (t＝-3.175, P＝0.007).The relative levels of hBD2 mRNA at 72 h after virus infection were 0.131 ±0.021 in control group, 0.128 ±0.022 in Lv-control group and 1.001 ±0.105 in Lv-hBD2 infection group; and the level in control group was statistically higher than that in hBD-2 infection group (t＝-7.213, P＝0.003).The results of oil red staining showed that OX-LDL inducing THP-1 cells for 72 h could significantly induce lipid accumulation in cells.Overexpression of hBD2 could effectively inhibit lipid accumulation in THP-1 cells induced by OX-LDL.The expression of hBD2 mRNA in THP-1 group was significantly higher than that in THP-1+OX-LDL group (t＝3.237, P＝0.004); and the difference was also significant when comparing THP-1+Lv-hBD2+OX-LDL group with THP-1+OX-LDL group (t＝-6.021, P＝0.003).The level of hBD2 protein in THP-1 group was significantly higher than that in THP-1+OX-LDL group (t＝0.314, P＝0.006); and the difference was also significant when comparing THP-1+Lv-hBD2+OX-LDL group with THP-1+OX-LDL group (t＝-4.061,P＝0.007).The levels of TNF-α, IL-1βand IL-6 in the supernatant of THP-1 cells induced by OX-LDL for 72 h were significantly increased compared with those in THP-1group (t＝-3.825,-2.017 and -3.551, respectively; P＝0.007, 0.004 and 0.005, respectively). The levels of TNF-α, IL-1βand IL-6 in THP-1+Lv-hBD2+OX-LDL group were significantly lower than those in THP-1+OX-LDL group ( t＝4.132, 3.681, and 2.991, respectively; P＝0.003, 0.002, and 0.007, respectively).Conclusions hBD2 can effectively inhibit THP-1 foaming induced by OX-LDL, which may be related to its inhibition of inflammatory response.

Towards the Application of Human Defensins as Antivirals

Defensins are antimicrobial peptides that participate in the innate immunity of hosts. Humans constitutively and/or inducibly express α- and β-defensins, which are known for their antiviral and antibacterial activities. This review describes the application of human defensins. We discuss the extant experimental results, limited though they are, to consider the potential applicability of human defensins as antiviral agents. Given their antiviral effects, we propose that basic research be conducted on human defensins that focuses on RNA viruses, such as human immunodeficiency virus (HIV), influenza A virus (IAV), respiratory syncytial virus (RSV), and dengue virus (DENV), which are considered serious human pathogens but have posed huge challenges for vaccine development for different reasons. Concerning the prophylactic and therapeutic applications of defensins, we then discuss the applicability of human defensins as antivirals that has been demonstrated in reports using animal models. Finally, we discuss the potential adjuvant-like activity of human defensins and propose an exploration of the ‘defensin vaccine’ concept to prime the body with a controlled supply of human defensins. In sum, we suggest a conceptual framework to achieve the practical application of human defensins to combat viral infections.

Expression of human β-defensin 1 in peripheral blood of children with community-acquired pneumonia in terms of clinical features / 中国感染与化疗杂志

Objective To investigate the expression of human beta defensin 1 (HBD-1) in peripheral blood of children with community acquired pneumonia (CAP) and its clinical implication.Methods A total of 122 CAP children were included in this study from February 2015 to October 2015.The patients were stratified in terms of age,etiology,and disease severity.Additional 52 patients were included as control in the same period.All patients were classified according to the clinical feature after admission.Serum HBD-1 level was determined by ELISA method.Results Serum HBD-1 level was significantly higher in CAP group than in control group (P＜0.001).HBD-1 level did not vary significantly with pathogen or sex in CAP group.HBD-1 level did not show significant difference between severe pneumonia and mild pneumonia.The HBD-1 level in 6-12 months age group was significantly higher than that in other age groups.HBD-1 level was not correlated to CRP or neutrophils in CAP children.Conclusions HBD-1 plays a role in anti-infective process as part of body innate immunity.It boasts non-specific and broad-spectrum anti-infective immunity against various pathogens.

Clinical Study on Pidotimod in the Treatment of Bronchial Asthma Complicated with Recurrent Respiratory Tract Infection in Children / 中国药房

OBJECTIVE:To investigate the clinical effects of pidotimod in the bronchial asthma complicated with recurrent re-spiratory tract infection,and its effects on immunoglobulin and related indexes. METHODS:A total of 120 bronchial asthma pa-tients with recurrent respiratory tract infection selected from our hospital during Mar. 2011-May 2013 were divided into trial group and control group according to random number table,with 60 cases in each group. Control group received routine corticosteroid therapy,and trial group was additionally given Pidotimod oral solution 0.4 g,po,bid,for 14 d,on the basis of control group. Clinical indexes(the times of respiratory infection,the duration of fever,cough,wheezing attack and antibiotics use),serum in-dexes [β-defensin-1(hBD-1),immunoglobulin A(IgA),IgG,IgM,UREA,ALT],the results of pharynx test before and after treatment,and the occurrence of ADR were observed in 2 groups. RESULTS:Before treatment,there was no statistical signifi-cance in clinical indexes,serum indexes,the results of pharynx test between 2 groups(P>0.05). After treatment,clinical indexes of trial group were significantly lower or shorter then before treatment or control group,while serum levels of hBD-1,IgA and IgG were significantly higher than before treatment or control group,with statistical significance(P0.05). The types and number of pathogenic bacteria of respiratory tract infection were decreased significantly in 2 groups,and the trial group was significantly less then the control group,with statistical significance(P<0.05). No obvious ADR was found in 2 groups. CONCLUSIONS:Pidotimod shows good clinical effects on bronchial asthma complicated with recurrent re-spiratory tract infection,can improve immunity and reduce the types and number of pathogenic bacteria with good safety.

Construction of rat beta defensin-2 eukaryotic expression vector and expression in the transfected rat corneal epithelial cell / 国际眼科杂志(Guoji Yanke Zazhi)

@#AIM: To construct a recombinant eukaryotic expression vector of rat beta defensin-2(rBD-2), transfect it into the rat corneal epithelial cells with lipofection, determine the expression of target gene in the transfected cells, and discuss the potentiality of recombinant plasmid expressed in corneal epithelial cells, hoping to provide an experimental foundation for further study on the antimicrobial activity of rBD-2 <i>in vitro </i>and <i>in vivo</i> and to assess the probability of defensins as a new application for infectious corneal diseases in the future. <p>METHODS: The synthetic rBD-2 DNA fragment was inserted between the <i>Xho</i>I and <i>BamHI</i> restriction enzyme cutting sites of eukaryotic expression vector pIRES2-ZsGreen1 to construct the recombinant plasmid pIRES2-ZsGreen1-rBD-2, then transformed it into <i>E.coli DH5α</i>, positive clones were screened by kanamycin and identified with restriction endonucleases and sequencing analysis. Transfection into the rat corneal epithelial cells was performed by lipofection. Then the experiment was divided into three groups: rat corneal epithelial cell was transfected with the recombinant plasmid pIRES2- ZsGreen1-rBD-2, rat corneal epithelial cell was transfected with the empty plasmid pIRES2-ZsGreen1 and the non-transfected group. The inverted fluorescence microscope was used to observe the transfection process. At last, the level of rBD-2 mRNA expressed in the transfected cells and the control groups are compared by the real-time fluoresence relative quantitative PCR. <p>RESULTS: The recombinant eukaryotic expression vector of pIRES2-ZsGreen1-rBD-2 was successfully constructed. The level of rBD-2 mRNA in transfected cells was significantly higher than that in control groups through the real-time fluorescence relative quantitative PCR. <p>CONCLUSION: The recombinant eukaryotic expression vector pIRES2-ZsGreen1-rBD-2 could be transfected into rat corneal epithelial cells, and exogenous rBD-2 gene could be transcripted into mRNA in the transfected cells.

Antifungal effects of synthetic human β-defensin 3-C15 peptide

OBJECTIVES: The purpose of this ex vivo study was to compare the antifungal activity of a synthetic peptide consisting of 15 amino acids at the C-terminus of human β-defensin 3 (HBD3-C15) with calcium hydroxide (CH) and Nystatin (Nys) against Candida albicans (C. albicans) biofilm. MATERIALS AND METHODS: C. albicans were grown on cover glass bottom dishes or human dentin disks for 48 hr, and then treated with HBD3-C15 (0, 12.5, 25, 50, 100, 150, 200, and 300 µg/mL), CH (100 µg/mL), and Nys (20 µg/mL) for 7 days at 37℃. On cover glass, live and dead cells in the biomass were measured by the FilmTracer Biofilm viability assay, and observed by confocal laser scanning microscopy (CLSM). On dentin, normal, diminished and ruptured cells were observed by field-emission scanning electron microscopy (FE-SEM). The results were subjected to a two-tailed t-test, a one way analysis variance and a post hoc test at a significance level of p = 0.05. RESULTS: C. albicans survival on dentin was inhibited by HBD3-C15 in a dose-dependent manner. There were fewer aggregations of C. albicans in the groups of Nys and HBD3-C15 (≥ 100 µg/mL). CLSM showed C. albicans survival was reduced by HBD3-C15 in a dose dependent manner. Nys and HBD3-C15 (≥ 100 µg/mL) showed significant fungicidal activity compared to CH group (p < 0.05). CONCLUSIONS: Synthetic HBD3-C15 peptide (≥ 100 µg/mL) and Nys exhibited significantly higher antifungal activity than CH against C. albicans by inhibiting cell survival and biofilm.