1.Influences of transforming growth factor-?1 on scavenger receptor class A and class B (CD36) in THP-1 derived macrophages
Xuguang LIU ; Jun AN ; Jian SUO ; Fanglei HAN ; Qicheng CUI
Journal of Jilin University(Medicine Edition) 2006;0(02):-
Objective To investigate the influence of transforming growth factor-?1(TGF-?1) on macrophage scavenger receptor (ScR) class A and B(ScR-B,CD36) in order to provide theoretical foundation for expounding the formation and therapy of atherosclerosis(AS).Methods THP-1 derived macrophages were divided into control group and experimental group,the cells in experimental group were treated with 3.0 mg?L-1Anti TGF-?1 Ab,the cells in control group were treated with 3.0 mg?L-1 IgG. The 125I-acetylated low density lipoprotein (125I-Ac-LDL for ScR-A) and 125I-oxidized low density lipoprotein (125I-Ox-LDL for CD36) uptaking (including 4℃ binding,37℃ association and degradation) were measured respectively.Influence of anti TGF-?1 antibody (Anti TGF-?1 Ab) on the ScR-A and CD36 mRNA expressions in THP-1 derived macrophages was measured meanwhile,respectively.Results Compared with control group ( binding: 8.23 ?g?g-1 ?1.24 ?g?g-1 protein,association:45.69 ?g?g-1 ?6.92 ?g?g-1 protein,and degradation: 112.18 ?g?g-1 ?20.15 ?g?g-1 protein),Anti TGF-?1 Ab increased 125I-Ac-LDL binding(48.67 ?g?g-1 ?6.52 ?g?g-1 protein),association (412.30 ?g?g-1?12.21 ?g?g-1 protein),and degradation (896.48 ?g?g-1 ?32.74 ?g?g-1 protein) significantly (P
2.Study on the antigenicity of Streptococcus pneumonia polysaccharide, its derivatives and conjugates by three immunological assays
Fanglei LIU ; Yali HOU ; Bing WU ; Jia LIU ; Xinru WANG ; Xubo YU ; Yanhong MU ; Aiping LIU ; Zhiqiang ZHAO ; Guilin XIE
Chinese Journal of Microbiology and Immunology 2014;(6):471-475
Objective To monitor and analyze the antigenicity of Streptococcus pneumonia polysac-charide, its derivatives and conjugates by three immunological assays .Methods Inhibition ELISA and rate nephelometry(RN) were established for this study.Antigenicity of serotype 23F pneumococcal conjugates and their intermediates were analyzed by double immunodiffusion assay , inhibition ELISA and RN .The re-sults derived from three assays were comparatively analyzed to evaluate the changes of antigenicity during the preparing process of serotype 23F conjugate.Results Double immunodiffusion assay, inhibition ELISA and RN were all applicable to antigenicity analysis during the process of conjugate preparation .Inhibition ELISA could quantitatively detect a slight difference of polysaccharide antigenicity during the preparing process . Conclusion The antigenicity of polysaccharide during the preparing process of pneumococcal conjugates could be analytically monitored by using three immunological assays .This study provided evidence for suc-cessfully using immunological assays as the quality control means during the preparing process of pneumococ -cal conjugates .
3.Study on the molecular size distribution and the structural characteristics of group B meningococcal cap-sular polysaccharides
Zhiqiang ZHAO ; Yingying YANG ; Xubo YU ; Yiyang FENG ; Ani LI ; Hongchun FANG ; Ruijie QIAO ; Bing WU ; Fanglei LIU ; Guilin XIE
Chinese Journal of Microbiology and Immunology 2014;(5):381-387
Objective To investigate the molecular size distribution and the structure of group B me-ningococcal capsular polysaccharides for the development of vaccines .Methods The molecular size distribution of group B meningococcal capsular polysaccharides was analyzed by chromatography on a Sepharose CL -4B col-umn.The molecular weight of repeat units were measured by matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF-MS).The structural characteristics of group B meningococcal capsular polysaccharides were analyzed by nuclear magnetic resonance ( NMR) based on the chemical shift of all charac-teristic protons by using group C meningococcal capsular polysaccharides and sialic acid as the controls .Results The KD value of group B meningococcal capsular polysaccharides extracted from 15 strains were ranged from 0.60 to 0.76.The molecular weight of repeat units was 284, which was identical to the theoretical value .The group B meningococcal capsular polysaccharides were 2→8 linked homopolymers of sialic acid lacking O-acetyl groups.Conclusion The group B meningococcal capsular polysaccharides had lower molecular weights , which might result in their poor immunogenicity .The structure of group B meningococcal capsular polysaccharides could be quickly and accurately analyzed by NMR technology .
4.Establishment of a quantitative ELISA assay recommended by WHO for the detection of human IgG antibodies specific for Streptococcus pneumoniae capsular polysaccharides (Pn PS ELISA)
Xinru WANG ; Jichun SHI ; Jisheng LIN ; Maoguang LI ; Chune WANG ; Fanglei LIU ; Qiang YE ; Zhiqiang ZHAO ; Guilin XIE
Chinese Journal of Microbiology and Immunology 2013;(10):783-788
Objective To establish a standardized quantitative enzyme-linked immunosorbent as-say ( ELISA) recommended by WHO for the detection of human IgG antibodies specific for Streptococcus pneumoniae capsular polysaccharides ( Pn PS ELISA ) .Methods According to the WHO recommended standard Pn PS ELISA protocol , capsular polysaccharide concentrations of 13 serotypes (1, 3, 4, 5, 6A, 6B, 7F, 9V, 14, 18C, 19A, 19F, 23F) of Streptococcus pneumonia for coating were optimized;the ELISA plates and AP conjugated goat anti-human IgG antibody for the detection were experimentally selected .Using the established assay parameters assured by testing quality control standards ( QCs) provided by WHO refer-ence laboratory , a panel of 16 LIBP ( Lanzhou Institute of Biological Products Co .Ltd.) QCs were measured for comparison analysis between WHO reference laboratory and LIBP laboratory .In the meantime , the range of IgG concentrations of an internal QC panel 907 for 13 serotypes of pneumococcal capsular polysaccharide were established for routine QC work in LIBP laboratory , and inter-assay precision within LIBP laboratory was assessed as well .Results The standardized Pn PS ELISA assay established in LIBP laboratory met the criteria required by WHO .There was a high correlation between the data collected by WHO reference labora -tory and LIBP laboratory (slope=0.94, coefficient of correlation r=0.97, P<0.05).Eighty one percent of IgG concentrations measured by LIBP laboratory were within the range of ±40%of those measured by WHO reference laboratory , which met the criteria of 75%data falling within the ambit of ±40%of assigned values commonly used for comparison .The ranges of IgG concentration in LIBP QC 907 for 13 serotypes had been established.The inter-assay precision in LIBP laboratory was high with coefficiency of variation ( CV) less than 30%.Conclusion LIBP laboratory has successfully established the standardized ELISA recommended by WHO for quantitative detection of human IgG antibodies against pneumococcal capsular polysaccharides (Pn PS ELISA).The range of IgG concentrations in LIBP QC 907 for 13 serotypes of pneumococcal capsular polysaccharide are also established for routine quality-control practice .
5.Preparation of a human meningococcal reference serum and standardization of IgG concentrations to capsular polysaccharides and bactericidal activities against serogroup A, C, Y and W135 strains
Zhiqiang ZHAO ; Xinru WANG ; Yanan LI ; Hao WANG ; Fanglei LIU ; Fei YUAN ; Ruijie QIAO ; Jingming JIANG ; Yanlin HE ; Jisheng LIN ; Qiang YE ; Guilin XIE
Chinese Journal of Microbiology and Immunology 2014;(6):459-464
Objective To prepare a human meningococcal reference serum and standardize IgG concentrations to capsular polysaccharides and in vitro bactericidal activities of the reference serum against serogroup A, C, Y and W135 strains.Methods Twenty healthy adults were recruited and given one dose of immunization with tetravalent (serogroups A, C, Y and W135) meningococcal polysaccharide vaccine . Plasma samples were collected and gone through a series of process treatments including defibrination , filtra-tion, and lyophilization to prepare the meningococcal reference serum Men 10.The IgG concentrations of Men10 to capsular polysaccharides of serogroups A , C, Y and W135 were calibrated by using an internation-al reference serum CDC1992 as the standard in enzyme-linked immunosorbent assay (ELISA).Provisional IgG concentrations of Men10 were intensively validated by testing a panel of 12 calibration serum samples from Centers for Disease Control and Prevention , USA ( US CDC) and a panel of 56 serum samples immu-nized with A, C, Y and W135 meningococcal polysaccharide vaccine from Lanzhou Institute of Biological Products Co., Ltd.(LIBP) with the assays using Men10 and CDC1992 as the standard and/or test sam-ples, respectively.The bactericidal titers against serogroup A , C, Y and W135 strains were measured by se-rum bactericidal assay (SBA).Results Four thousand vials (0.5 ml/vial)of lyophilized human meningo-coccal reference serum Men10 were successfully prepared with 2.5%of residual moisture .Reference serum Men10 was sterile and free from contamination by hepatitis B virus , hepatitis C virus , human immunodefi-ciency virus and syphilis .Provisional IgG concentration of Men 10 to capsular polysaccharide of serogroups A, C, Y and W135 was calibrated by using CDC1992 as the standard.Furthermore, IgG concentrations of both panels of 12 CDC calibration serum samples and 56 LIBP serum samples calibrated by using Men 10 as the standard correlated well with those by using CDC1992 as the standard (r=0.99,P<0.05).The IgG concentrations of CDC1992 as calibrated by using Men10 as the standard showed significant correlation with its previously determined values with variation <10%.SBA titers for serotype A , C, Y and W135 strains were established as well .Conclusion A panel of new human meningococcal reference serum Men 10 with accurately calibrated IgG concentration against capsular polysaccharide of serogroups A , C, Y and W135 as well as SBA titers was successfully established .
6.PHF5A Promotes Proliferation and Migration of Non-Small Cell Lung Cancer by Regulating of PI3K/AKT Pathway.
Houhui WANG ; Fanglei LIU ; Chunxue BAI ; Nuo XU
Chinese Journal of Lung Cancer 2023;26(1):10-16
BACKGROUND:
There have been many significant advances in the diagnosis and treatment of non-small cell lung cancer (NSCLC). However, the mechanism underlying the progression of NSCLC is still not clear. Plant homodomain finger-like domain-containing protein 5A (PHF5A) plays an important role in processes of chromatin remodeling, morphological development of tissues and organs and maintenance of stem cell pluripotency. This study aims to investigate the role of PHF5A in the proliferation and migration of NSCLC.
METHODS:
A549 and PC-9 PHF5A overexpression cell lines were constructed. PHF5A expression was decreased in H292 and H1299 cells by using siRNA. Flow cytometry was used to detect the cell cycle. MTT assay and clone formation assay were used to examine the proliferative ability of NSCLC, while migration assay and wound healing assay were performed to evaluate the ability of migration. Western blot analysis was used to measure the expressions of PI3K, p-AKT and the associated downstream factors.
RESULTS:
Up-regulation of PHF5A in A549 and PC-9 cells increased the proliferation rate, while down-regulation of PHF5A in H292 and H1299 cells inhibited the proliferation rate at 24 h, 48 h and 72 h (P<0.05). The metastatic ability was elevated in the PHF5A-overexpresion groups, while reduced in the PHF5A-down-regulation group (P<0.05). In addition, reduced expression of PHF5A induced cell cycle arrest at G1/S phase (P<0.05). Furthermore, decreased expression of PHF5A reduced the expression levels of PI3K, phosphorylation of AKT, c-Myc (P<0.05) and elevated the expression of p21 (P<0.05).
CONCLUSIONS
These results demonstrated that PHF5A may play an important role in progression of NSCLC by regulating the PI3K/AKT signaling pathway.
Humans
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Carcinoma, Non-Small-Cell Lung/pathology*
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Lung Neoplasms/pathology*
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Proto-Oncogene Proteins c-akt/metabolism*
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Phosphatidylinositol 3-Kinases/metabolism*
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Cell Line, Tumor
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Cell Proliferation/genetics*
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Cell Movement/genetics*
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Gene Expression Regulation, Neoplastic
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Trans-Activators/genetics*
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RNA-Binding Proteins/metabolism*