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El complejo BAFF (factor activador de células B) compuesto por la citocina BAFF, APRIL y sus receptores -BAFF-R (BR3), TACI y BCMA- influyen en la sobrevida periférica, en la maduración de los linfocitos B y en el cambio de clase de las inmunoglobulinas, con múltiples implicaciones clínicas potenciales. Las funciones biológicas de BAFF y su relevancia en varios desórdenes clínicos -autoinmunes, neoplásicos, infecciosos, incluyendo las terapias BAFF dirigidas- son revisadas y discutidas en el presente artículo. Los niveles séricos de BAFF/APRIL se encuentran incrementados en las enfermedades autoinmunes en las que sus concentraciones se relacionan con los títulos de anticuerpos, actividad, progresión de la enfermedad e incluso compromiso orgánico, haciendo de su inhibición un blanco terapéutico atractivo
The BAFF complex (B cell activator factor) composed by the BAFF cytokine, APRIL and their receptors -BAFF-R (BR3), TACI, BCMA- influences B-lymphocyte maturation, peripheral survival and immunoglobulins class isotype switching, with multiple potential clinical implications. In this review we discuss BAFF biologic functions and it relevance in several clinical disorders -autoimmune, neoplastic, infectious and BAFF therapies-. BAFF/APRIL
Subject(s)
Humans , Autoimmune Diseases , B-Cell Activating FactorABSTRACT
Objective To investigate the expression of a proliferation-inducing ligand (APRIL) in urothelial carcinoma tissue and its clinical significance. Methods We investigated the expression of APRIL mR-NA in urothelial carcinoma tissue of 33 patients in urothelial carcinoma in Cangzhou Central Hospital from July 2013 to March 2015, Among of them, 23 cases of adjacent pericancerous tissues were detected by qRT-PCR. Immunohistochemistry and western blot were used to detect the APRIL protein expression level in urothelial carci-noma tissue and adjacent pericancerous tissues. We analyzed the relationship between APRIL expression and clinical pathology in patients with urothelial carcinoma by statistical methods. Results The qRT-PCR revealed that the expression of APRIL in urothelial carcinoma tissue was significantly higher than that in the adjacent peri-cancerous tissues (P 0.05). Conclusion The APRIL is a high abnormal expression in urothelial carcinoma tissues , and maybe related to the occurrence and development of urothelial carcinoma.
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Objective To investigate the role of APRIL in SW480 cell line.Methods The CRC model was established in the nude mice,all the mice were divided into 3 groups,the mice were separately treated with APRIL siRNA,pGC-vector and PBS solution.The APRIL mRNA was detected by RT-PCR and the APRIL protein was surveyed by the way of immunohis-tochemistry (IHC).The proteins of TIMP-3,Syndecan-1and MMP-9 also were assessed by IHC.Results ①Tumor mass in the group of nude mice injected with PBS (2.15±0.30 g)was significantly higher than the injection APRIL siRNA group (0.95±0.15 g,P0.05)compared with the injection of empty vector group (2.20±0.25 g).②APRIL mRNA/18S rRNA ratio (2.48±0.25)in the group of mice injected with PBS was signifi-cantly higher than the injection APRIL siRNA group (0.39±0.15,P0.05)compared with the injection of empty vector group (2.51±0.30).③SW480 cells injected with APRIL siRNA signifi-cantly inhibited invasion and metastasis.TIMP-3 Allred scores in three groups were 7.70±0.35,1.10±0.16 and 1.15± 0.12,Syndecan-1 protein was 7.80±0.30,1.05±0.20 and 1.10±0.22 MMP-9 protein was 1.20 ±0.10,8.00±0.25 and 8.20±0.20,respectively.Conlusion APRIL was closely connected with the growth and metabasis of CRC.
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Objective To investigate the effects of a proliferation-inducing ligand(APRIL) on migration and invasion of colorectal cancer (CRC) and matrix metalloproteinases (MMPs) expression in order to observe the role of APRIL in CRC metastasis.Methods The siRNA plasmid vector targeting APRIL gene (siRNA-APRIL) was transfected into SW480 cells and recombinant human APRIL(rhAPRIL) was used to stimulate HCT-116 cells.Tumor cell migration and invasion were measured by Transwell chambers.RT-PCR and ELISA were applied to examine the expression level of MMPs.Results Metastatic and invasive capacities of siRNA-APRIL transfected SW480 were significantly inhibited,and these capacities of APRIL stimulated HCT-116 cells were significantly enhanced compared with their respective controls( all P＜0.05 ),accompanied with the alterations of MMPs mRNA and secreted protein expression( P＜0.05).The number of invading cells of SW480 control and rhAPRIL stimulated HCT-116 was significantly decreased by a MMP inhibitor GM6001 ( P＜0.05 ).Conclusion APRIL facilitates migration and invasion of CRC via regulation of MMPs,which suggests that APRIL might be used as a new target for the intervention and treatment of CRC metastasis.
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BACKGROUND: BAFF (B cell-activating factor) and APRIL (a proliferation-inducing ligand) are members of the tumor necrosis factor family and promote B cell survival and proliferation. We evaluated the correlation between serum concentration of BAFF or APRIL and severity of acute graft-versus-host disease (GVHD). METHODS: Fifteen patients who received allogeneic hematopoietic stem transplantation for leukemia and developed acute GVHD were enrolled. We determined serum concentrations of BAFF and APRIL at the onset of the first clinical manifestation of GVHD by enzyme-linked immunosorbent assay. RESULTS: Nine patients had grade 2 acute GVHD, and 6 had grade 3-4 acute GVHD. The BAFF serum concentration was higher in patients with grade 3-4 acute GVHD (1,093.42 in grade 2 vs. 2,171.99 pg/mL in grade 3-4), although the difference was not significant (P=0.077). However, the ratio of BAFF serum concentration to absolute lymphocyte count (ALC) (BAFF/ALC) was significantly higher in patients with grade 3-4 acute GVHD (P=0.045). The APRIL serum concentration and APRIL/ALC ratio showed similar results (P=0.077 and P=0.013, respectively). CONCLUSION: Patients with grade 3-4 acute GVHD had higher BAFF/ALC and APRIL/ALC ratios than patients with grade 2 acute GVHD. These findings suggest that B cells might play an important role in the development of acute GVHD, and that the BAFF and APRIL concentrations in serum might be significant predictive factors for estimating the severity of acute GVHD. Their clinical significance should be further evaluated in a larger patient population.
Subject(s)
Humans , B-Lymphocytes , Cell Survival , Graft vs Host Disease , Hematopoietic Stem Cell Transplantation , Hematopoietic Stem Cells , Leukemia , Lymphocyte Count , Transplants , Tumor Necrosis Factor-alphaABSTRACT
Objective To investigate mRNA and protein expression of aproliferation-inducing ligand (APRIL) in peripheral blood mononuclear cell and plasma of patients with B cells non-Hodgkin lymphoma (B-NHL) pre- or post- chemotherapy and to explore the role of APRIL in the B-NHL. Methods The mRNA and protein expression of APRIL were detected by real-time fluorescence quantitative polymerase chain reaction (RFQ-PCR) and enzyme-linked immunosorbent assay(ELISA), respectively. According to the standard curves,the quantitative levels of target mRNA and protein were determined. Results The detection linear range of targeted mRNA by RFQ-PCR was 101-109 pg/ml, and the coefficient of variation values for both intra- and inter-experimental reproducibility ranged 1.69 %-5.99 % and 6.35 %-10.12 %, respectively. To detect expression of APRIL protein by ELISA, the correlation coefficient of standard curves reached 0.9922. Before or after chemical treatment, the expression levels of APRIL mRNA and protein in patients with B-NHL were significantly higher than those in normal control (P ＜0.01), while the expression levels in post-treatment patients with Ⅲ and Ⅳ stage were lower than those of pre-treatment patients with corresponding stage (P ＜0.05, respectively), but those of pre- and post-treatment patients with Ⅰ / Ⅱ stage were not different (P ＞0.05).Conclusion The expression level of APRIL mRNA is similar to that of APRIL protein. APRIL may be involved in pathogenesis and development of B-NHL. Moreover, APRIL may be related to the burden of B-NHL and it may be considered as a targeted molecule for B-NHL.
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Objective To investigate the effects of a proliferation-inducing ligand(APRIL)gene silencing by small interfering RNA(siRNA)on cell cycle and proliferation of colon carcinoma SW480 cells.Methods The siRNA plasmid vector targeting APRIL gene,named as siRNA-APRIL,was transfected into SW480 cells,transfected with scrambled vector as a nontargeting control and nontransfected group as another control.APRIL mRNA and protein expression were examined by real-time PCR and Western blot,respectively.Cell proliferation activity was analyzed by cell counting kit-8(CK-8),cell cycle was detected by flow cytometry,and p21 together with p27,two important regulatory genes in cell cycle,were measured by RTPCR.Results Compared with nontargeting control and nontransfected control,APRIL expression was inhibited significantly at both mRNA and protein level by siRNA-APRIL being transfected in SW480 cells(P ＜0.05).Cell proliferation ability was drastically repressed after siRNA-APRIL being transfected at 48 h,72 h and 96 h(P ＜ 0.05).After transfected 48 h,the percent of Go/G1 phase cell was significantly increased,S and G2/M phase cell were significantly decreased,the number of cell in apoptosis was increased and the expression of p21 and p27 mRNA were up-regulated(P ＜ 0.05).There was no significant difference when compared the two control groups each other(P ＞ 0.05).Conclusion siRNA-APRIL can effectively knockdown the expression of APRIL gene in SW480 cells,moreover,it can inhibit the cell proliferation and induce G0/G1 phase cell cycle arrest,which occurrence may involve in upregulation the mRNA expression of p21 and p27.
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BACKGROUND: APRIL, originally known as a cytokine involved in B cell survival, is now known to regulate the inflammatory activation of macrophages. Although the signal initiated from APRIL has been demonstrated, its role in cellular activation is still not clear due to the presence of BAFF, a closely related member of TNF superfamily, which share same receptors (TACI and BCMA) with APRIL. METHODS: Through transfection of siRNA, BAFF-deficient THP-1 cells (human macrophage-like cells) were generated and APRIL-mediated inflammatory activities were tested. The expression patterns of APRIL were also tested in vivo. RESULTS: BAFF-deficient THP-1 cells responded to APRIL-stimulating agents such as monoclonal antibody against APRIL and soluble form of TACI or BCMA. Furthermore, co-incubation of the siBAFF-deficient THP-1 cells with a human B cell line (Ramos) resulted in an activation of THP-1 cells which was dependent on interactions between APRIL and TACI/BCMA. Immunohistochemical analysis of human pathologic samples detected the expression of both APRIL and TACI in macrophage-rich areas. Additionally, human macrophage primary culture expressed APRIL on the cell surface. CONCLUSION: These observations indicate that APRIL, which is expressed on macrophages in pathologic tissues with chronic inflammation, may mediate activation signals through its interaction with its counterparts via cell-to-cell interaction.
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Humans , Cell Communication , Cell Line , Cell Survival , Diphenylamine , Inflammation , Macrophages , RNA, Small Interfering , TransfectionABSTRACT
Objective To construct of shRNA lentiviral expression vector targeting APRIL (aproliferation-inducing ligand) gene in CFPAC-1 cell of human pancreatic cancer. Methods We used gene engineering to screen RNA interference targeting sequence of APRIL gene. The complementary DNA containing both sense and antisense Oligo DNA of the targeting sequence was designed, synthesized and cloned into the pGCL-GFP vector. The resulting lentiviral vector containing shAPRIL were named LV-shAPRIL. Then it was conformed by PCR and DNA sequencing identification. 293T cells were eotransfected with LV-shAPRIL,pHelper 1.0 and pHelper 2.0 to product ientivirus. The titer of virus was tested according to the expression level of GFP in the 293T cells. After recombinant lentivirus infection into CFPAC-1 cells, we used real-time RT-PCR and Western blotting to examine APRIL mRNA and protein expression at different cell culture period.Results PCR analysis and DNA sequencing conformed that shAPRIL DNA was successfully inserted into the lentiviral vector. The titer of concentrated virus were 5 × 107 TU/ml. APRIL expression in CFPAC-1 cells were inhibited significantly at both mRNA and protein level. APRIL mRNA expression were decreased 73%, 70%and 71% , respectively, after the infection of 4 days, 4 weeks and 8 weeks by LV-shAPRIL. APRIL protein expression were decreased 66%, 63% and 62%, respectively , after the infection of 4 days , 4 weeks and 8weeks by LV-shAPRIL. Conclusions ShRNA lentiviral expression vector targeting APRIL gene has been successully constructed, and it can effectively inhibit the expression of APRIL gene in CFPAC-1 ceils. This study lays a foundatin for in vivo research APRIL gene scilence in pancreatic cancer cell using the model of nude mice.
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Objective To investigate the inhibitive effect of shRNA (short hairpin RNA) targeting APRIL gene on the pancreatic cancer cells in vitro and in vivo, in order to explore the feasibility of gene therapy for pancreatic cancer. Methods The LV-shAPRIL targeting APRIL gene had been constructed before, and was used to infect the CFPAC-1 cells. Cell proliferation and apoptosis were examined by MTT and flow cytometry. Then CFPAC-1 cells were used to construct the model of transplantation tumor into the nude mice, the tumor growth was assessed after LV-shAPRIL treatment. Results 96 hours after the LV-shAPRIL infection into CFPAC-1 cells, the cell proliferation was significantly inhibited when compared with control group and lentivirus infection group (P＜0.05 ). Flow cytometry showed the apoptosis ratio of the CFPAC-1 cells was (17.35±0.96)% in LV-shAPRIL group, which was higher than that in control group and lentivirus infection group (P＜0.05 ). After LV-shAPRIL injection into the model of nude mice, the tumor growth was slower than that in the two control groups. The tumor's volume of the LV-shAPRIL group was(821.8±123.3) mm3 and the mass was (2.16±0.18)g at 27 day, and were obviously depressed, when compared with two control groups (P±0.05). Conclusions LV-shAPRIL targeting APRIL gene can inhibit the growth of the CFPAC-1 cells in vitro and vivo. This may provide a new gene therapy approach for pancreatic cancer.
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Objective To study the expression and significance of B-cell activating factor(BAFF) and a proliferation-inducing ligand(APRIL) in patients with idiopathic thrombocytopenic purpur(ITP). Methods The serum levels of BAFF and APRIL in 27 cases with ITP were tested by ELISA.Results ①The serum levels of BAFF in ITP were higher than those of control group (3.92?1.88?g/ml vs 2.90?0.52?g/ml,P
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Objective:To construct a eukaryotic vector containing human TACI-linker-BR3-IgGFc double receptor fusion gene and to investigate its expression in COS-7 cells.Methods: The standard cloning technology was employed to construct the(title) vector,which was then used to transfect COS-7 cells by lipofectamine 2000.The expression of target protein and their interaction with BAFF and APRIL protein were examined by Western blot,ELISA,and immunohistochemistry methods.Results: The expression of the fusion protein was observed in transiently transfected COS-7 cells, with the transfection efficiency being 48%.The fusion protein was detected in the supernatant.It suggested that the fusion protein interacted well with BAFF and(APRIL) proteins;the IgGFc tag did not influence the natural assembly and the biological activity of fusion proteins.Conclusion: A vector containing TACI-linker-BR3 double receptor and IgGFc fusion gene has been successfully constructed and the(subsequent) fusion protein is bioactive,which paves a way for further research on the pathogenesis of autoimmune diseases and biotherapy strategy.
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BACKGROUND: To clarify the clinicoradiological correlation and prognosis of acute ischemic stroke involving para-median territory of pons. METHODS: We studied 37 patients with first-ever ischemic stroke involving paramedian terri-tory of pons and divided them based on the shape and level of lesion shown in their MRI. The clinical features, MRI findings, and prognosis were assessed. RESULTS: The paramedian infarctions extending to the basal surface were found in 28 patients (76%), and small infarctions separated from the basal surface were found in 9 patients (24%). In patients with infarction extending to the basal surface, 23 patients (82%) had progressive or fluctuating onset, whereas all patients with small infarction separated from the basal surface had non-progressive onset. In the group with upper pon-tine lesion (14 patients), dysarthria-clumsy hand syndrome was found in 4 patients, ataxic hemiparesis (AH) in 3, pure motor hemiparesis (PMH) in 2, and pure sensory stroke in 1. In the group with middle and lower pontine lesion (22 patients), PMH was found in 9, AH in 3, and sensory motor stroke in 2. The mean Modified Rankin Disability Scale scores on admission and after follow-up (mean 29 months) of the group with upper pontine lesion were 2.36 +/-0.50 and 1 . 0 0 +/-0.55, those with mid-lower pontine lesions, 3.48 +/-0.51 and 1.17 +/-0.49 (P0.05 respectively). CONCLUSIONS: Paramedian pontine infarction extending to the basal surface usually presents with progressive onset. Paramedian pontine infarction most often produces classic lacune syndrome of which PMH is the most common. In our study, patients with mid-lower paramedian pontine infarction had more severe initial neurological deficits than those with upper paramedian pontine infarction. However, a late outcome was found to be favorable in both groups.
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Humans , Follow-Up Studies , Hand , Infarction , Magnetic Resonance Imaging , Paresis , Pons , Prognosis , StrokeABSTRACT
Objective To establish a SYBR Green Ⅰ quantitative real-time PCR method for detecting the expression of APRIL gene(a proliferation-inducing ligand,APRIL) in peripheral blood of patients with auto-immune diseases,e.g.,systemic lupus erythematosus(SLE) and rheumatoid arthritis(RA),and investigate the relationship of APRIL mRNA expression with pathogenesis and prognosis of auto-immune diseases.Methods Plasmid PGEM-T easy-APRIL was cloned as the standard template.SYBR Green Ⅰ quantitative real-time PCR was set up to examine the expression of APRIL mRNA in peripheral blood of 58 patients with auto-immune diseases and 20 healthy controls using Line Gene FQD-33A Detection System.Results The obtained data were normalized by dividing the copy number of target cDNA by those of GAPDH(glyceraldehycle-3-phosphate dehydrogenase,GAPDH).APRIL expression levels ranges from 3.95 to 192 and mean value was 29.68?4.5.APRIL expression of twenty healthy controls showed range from 3.1 to 18.7 and mean value of 10.56?2.0.APRIL expression levels in patients with auto-immune diseases were higher than those in healthy controls.In auto-immune diseases group APRIL expression levels of untreated patients were higher than those of the other patients,and a statistical significance was found.Conclusions APRIL mRNA was successfully detected by SYBR Green Ⅰ quantitative real-time PCR and the method was accurate and reliable.The expression of APRIL mRNA of auto-immune disease patients was higher than those of healthy controls,and the expression of untreated patients was the highest.This method may be used for further study on the high-level expression of APRIL mRNA in mechanism of auto-immune diseases as well as the development and prognosis of diseases.