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OBJECTIVE To explore the improvement effect and potential mechanism of total flavonoids from Alpinia zerumbet on gastric mucosa injury induced by absolute ethanol through microRNA-146a-5p （miR-146a-5p）. METHODS Using human gastric mucosa GES-1 cells as objects， the acute gastric ulcer model was established by absolute ethanol； based on the investigation of the effects of different concentrations of total flavonoids from A. zerumbet on cell activity and the selection of action concentration， the relative expression level of miR-146a-5p in GES-1 cells was detected， the protein expressions of tumor necrosis factor （TNF） receptor-associated factor 6（TRAF6）， nuclear factor-κB p65 （NF-κB p65） and TNF-α were detected， and the levels of interleukin- 1β （IL-1β）， IL-6 and prostaglandin E2 （PGE2） in cell supernatant were determined. The targeting relationship between miR-146a- 5p and TRAF6 was verified； the effects of overexpressed miR-146a-5p and TRAF6 knockdown on the levels of IL-1β， IL-6 and PEG2 in supernatant of model cells as well as the effects of miR-146a-5p knockdown on anti-gastric ulcer effect of total flavonoids from A. zerumbet were observed. RESULTS Compared with the blank group， the relative expression of miR-146a-5p in cells and the level of PGE2 in cell supernatant were decreased significantly in the model group （P＜0.01）， while the protein expressions of TRAF6， NF-κB p65 and TNF-α in cells and the levels of IL-1β and IL-6 in cell supernatant were increased significantly （P＜ 0.01）. Compared with the model group， the relative expression of miR-146a-5p in cells and the level of PGE2 in cell supernatant were increased significantly in model+A. zerumbet total flavonoids （60 mg/L） group （P＜0.01）， while the protein expressions of TRAF6， NF-κB p65 and TNF-α in cells and 82260767） the levels of IL-1β and IL-6 in cell supernatant were decreased significantly （P＜0.05 or P＜0.01）. There was a targeted relationship and a negative correlation between miR-146a-5p E-mail：3113836821@qq.com and TRAF6. After overexpression of miR-146a-5p or TRAF6 knockdown， the levels of IL-1β and IL-6 were decreased significantly in cell supernatant， while the level of PGE2 was increased significantly （P＜0.05）. After miR-146a-5p knockdown， the levels of IL-1β and IL-6 in cell supernatant and the protein expression of TRAF6 in cells administered with total flavonoids of A. zerumbet were increased significantly， while the level of PGE2 was decreased significantly （P＜0.05）. CONCLUSIONS Total flavonoids of A. zerumbet can improve the gastric mucosa injury induced by absolute ethanol. The mechanism may be related to up-regulating the expression of miR-146a-5p， inhibiting the expression of TRAF6， and further inhibiting the secretion of related inflammatory factors.
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OBJECTIVE@#To observe the therapeutic effect of acupuncture on cancer-related fatigue (CRF) and to explore its possible mechanism.@*METHODS@#A total of 80 patients with CRF were randomized into an observation group and a control group, and finally 67 patients completed the trial (36 patients in the observation group, 31 patients in the control group). Patients in the control group were treated with conventional chemoradiotherapy and symptomatic treatment, while no particular anti-fatigue intervention was adopted. On the basis of treatment in the control group, acupuncture was applied at Baihui (GV 20), Guanyuan (CV 4), Qihai (CV 6), Fengchi (GB 20), Zusanli (ST 36), Sanyinjiao (SP 6) in the observation group, once a day, 5 times as one course, with 2 days interval between each course, totally 4 courses were required. Before and after treatment, scores of functional assessment of cancer therapy-fatigue (FACT-F) in Chinese and McGill quality of life questionnaire (MQOL), serum levels of C-reactive protein (CRP), interleukin-6 (IL-6), tumor necrosis factor-α(TNF-α) and soluble TNF receptor-1 (sTNF-R1) were observed in the two groups.@*RESULTS@#①Compared before treatment, the FACT-F score was decreased after treatment in the observation group (<0.05), while there was no significant difference in the control group (<0.05). The change of the FACT-F score in the observation group was larger than that in the control group (<0.05). ②In the observation group, scores of physiological and psychological dimension were decreased (<0.05), score of social support dimension was increased after the treatment (<0.05). The score changes of physiological, psychological and social support dimension in the observation group were larger than those in the control group (all <0.05). ③After treatment, the serum levels of IL-6, TNF-α and sTNF-R1 were decreased in the observation group (<0.05), while the serum levels of CPR and IL-6 were increased in the control group (<0.05). The serum levels of CPR, IL-6 and TNF-α in the observation were lower than those in the control group (<0.05).@*CONCLUSION@#①Acupuncture can improve the related symptoms of depression, weakness and headache in patients with CRF, strengthen their cognition of the support from society and family, and boost the confidence in curing the disease. ②Acupuncture can effectively down-regulate serum levels of the relative inflammatory factors, which may be its possible mechanism on treating CRF.
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Humans , Acupuncture Points , Acupuncture Therapy , Biomarkers , Blood , C-Reactive Protein , Fatigue , Therapeutics , Interleukin-6 , Blood , Neoplasms , Therapeutics , Quality of Life , Receptors, Tumor Necrosis Factor, Type I , Blood , Tumor Necrosis Factor-alpha , BloodABSTRACT
Objective: To investigate the protective effect of human lipoxin A4 (LXA4) on N2a cell damage induced by β-amyloid protein 25-35 (Aβ25-35) and the underlying mechanism. Methods: Aβ25-35 was used to treat N2a cells to establish Alzheimer's disease (AD) cell injury model. Meanwhile, LXA4 was added to the experimental group at different concentrations (50, 100 and 200 nmol·L-1 ). MTT assay was used to detect the activity of N2a cells. The apoptosis was detected by Hoechst 33258-PI staining, the expression of P62 and TRAF6 mRNA was detected by RT-PCR, and the expression of P62 and TRAF6 protein was detected by Western blot. Results: Compared with that of the model group, the cell survival rate of LXA4 protective group (50,100 and 200 nmol·L-1 ) increased (P <0. 01) and the apoptosis of N2a cells induced by Aβ25-35 was reduced by LXA4 (100 and 200 nmol·L-1 ) . Compared with that of the model group, the expression of P62-mRNA and protein-P62 of N2a cells treated with Aβ25-35 increased (P <0. 05 or P <0. 01) and the expression of TRAF6-mRNA and protein-TRAF6 of N2a cells treated with Aβ25-35 were reduced (P <0. 05 or P <0. 01). Conclusion: LXA4 has protective effect on N2a cell damage induced by Aβ25-35 , and its mechanism may be related to the up-regulation of P62 gene and down-regulation of TRAF6 gene.
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AIM: To investigate the effects of TNF receptor-associated death domains (TRADD) of latent membrane protein 1 (LMP1) on the proliferation of nasopharyngeal cancer SP18 cells.METHODS: The SP18-LMP1 cells and SP18-LMP1TRADD cells, which expressed LMP1 and LMP1 TRADD proteins, respectively, were established.The proliferation of SP18 cells affected by LMP1TRADD was detected by cell counting to analyze the cell growth curve, and by colony formation assay, soft agar formation assay, and flow cytometry.Moreover, the expression profile of differential genes between SP18-LMP1 cells and SP18-LMP1TRADD cells was analyzed by gene chips.RESULTS: The cell growth curve, and the results of colony formation and soft agar formation displayed that the growth velocity and colony forming ability of SP18-LMP1 cells were stronger than those of SP18-LMP1TRADD cells (P<0.01).The results of flow cytometry analysis showed that the proliferation index of SP18-LMP1 cells was higher than that of SP18-LMP1TRADD cells (P<0.01).Sixty-three differentially expressed genes associated with cell proliferation were screened out, in which 33 genes were up-regulated and 30 were down-regulated in the SP18-LMP1TRADD cells.CONCLUSION: TRADD active region is an important functional site of LMP1 to promote the proliferation of SP18 cells.LMP1 may improve the cell proliferation index and induce the proliferation of SP18 cells through TRADD.
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Objective To observe the effects of Ginkgo biloba extract 50 (GBE50) on hemodynamics and myocardial tissue TNF-α pathway in rats with acute blood stasis syndrome; To discuss its mechanism of action. Methods A model of acute blood stasis syndrome was established by subcutaneous injection of epinephrine hydrochloride and ice bath. Male SD rats were divided randomly into normal group, model group and GBE50 group. After medicine was administrated by gavage for 11 days, the acute blood stasis model was established. The hemodynamic changes were observed in the next day. The expressions of TNF-α, TNFR1 and TNFR2 mRNA were detected by RT-PCR. The expressions of TNF-α, TNFR1 and TNFR2 protein were tested by radioimmunoassay and immunohistochemistry. Results The hemodynamics of the model group was lower than that of the normal group. Compared with model group, GBE50 could increase mLVSP, DP and decrease t-dp/dtmax(P<0.05). The protein expressions of TNF-α, TNFR1, and TNFR2 in GBE50 group were higher than those in model group, and TNFR2 mRNA expression increased significantly in GBE50 group (P<0.05). Conclusion GBE50 can ameliorate the decline of hemodynamics of rats with acute blood stasis syndrome, and regulate the myocardial TNF-α pathway. Increasing the expression of protective receptor TNFR2 may be one of the mechanisms of protecting cardiac function.
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Objective To observe the effects of Ginkgo biloba extract 50 (GBE50) on hemodynamics and myocardial tissue TNF-α pathway in rats with acute blood stasis syndrome; To discuss its mechanism of action. Methods A model of acute blood stasis syndrome was established by subcutaneous injection of epinephrine hydrochloride and ice bath. Male SD rats were divided randomly into normal group, model group and GBE50 group. After medicine was administrated by gavage for 11 days, the acute blood stasis model was established. The hemodynamic changes were observed in the next day. The expressions of TNF-α, TNFR1 and TNFR2 mRNA were detected by RT-PCR. The expressions of TNF-α, TNFR1 and TNFR2 protein were tested by radioimmunoassay and immunohistochemistry. Results The hemodynamics of the model group was lower than that of the normal group. Compared with model group, GBE50 could increase mLVSP, DP and decrease t-dp/dtmax(P<0.05). The protein expressions of TNF-α, TNFR1, and TNFR2 in GBE50 group were higher than those in model group, and TNFR2 mRNA expression increased significantly in GBE50 group (P<0.05). Conclusion GBE50 can ameliorate the decline of hemodynamics of rats with acute blood stasis syndrome, and regulate the myocardial TNF-α pathway. Increasing the expression of protective receptor TNFR2 may be one of the mechanisms of protecting cardiac function.
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Objective: To study the expression of tumor necrosis factor receptor-associated protein 1 (TRAP1) in gastric cancer tissue and its correlation with malignant biology. Methods: Gastric cancer tissue and adjacent normal tissue were collected, and mRNA content and protein content of TRAP1 were detected; gastric cancer cell lines SGC7901, BGC823, AGS and MGC803 were cultured, and mRNA contents and protein contents of TRAP1, CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF were detected. Results: mRNA and protein expression levels of TRAP1 in gastric cancer tissue were significantly higher than those in adjacent normal tissue, and mRNA and protein expression levels of TRAP1 in gastric cancer tissue with muscularis and serosa infiltration, lymph node metastasis, distant organ metastasis and TNM III/[U+2163] stage were significantly higher than those in gastric cancer tissue with mucosa and submucosa infiltration, non-lymph node metastasis, non-distant organ metastasis and TNM [U+2160]/III stage. mRNA and protein expression levels of TRAP1, CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF in MGC803 were the highest, and mRNA and protein expression levels of TRAP1, CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF in SGC7901 were the lowest. mRNA and protein expression levels of TRAP1 in gastric cancer cell lines were positively correlated with mRNA and protein expression of CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF. Conclusions: The expression of TRAP1 significantly increases in gastric cancer tissue; TRAP1 may regulate the malignant biology of cells by increasing the expression of CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF, thereby resulting in the occurrence and development of gastric cancer.
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OBJECTIVE@#To study the expression of tumor necrosis factor receptor-associated protein 1 (TRAP1) in gastric cancer tissue and its correlation with malignant biology.@*METHODS@#Gastric cancer tissue and adjacent normal tissue were collected, and mRNA content and protein content of TRAP1 were detected; gastric cancer cell lines SGC7901, BGC823, AGS and MGC803 were cultured, and mRNA contents and protein contents of TRAP1, CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF were detected.@*RESULTS@#mRNA and protein expression levels of TRAP1 in gastric cancer tissue were significantly higher than those in adjacent normal tissue, and mRNA and protein expression levels of TRAP1 in gastric cancer tissue with muscularis and serosa infiltration, lymph node metastasis, distant organ metastasis and TNM Ⅲ/Ⅳ stage were significantly higher than those in gastric cancer tissue with mucosa and submucosa infiltration, non-lymph node metastasis, non-distant organ metastasis and TNM Ⅰ/Ⅱ stage. mRNA and protein expression levels of TRAP1, CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF in MGC803 were the highest, and mRNA and protein expression levels of TRAP1, CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF in SGC7901 were the lowest. mRNA and protein expression levels of TRAP1 in gastric cancer cell lines were positively correlated with mRNA and protein expression of CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF.@*CONCLUSIONS@#The expression of TRAP1 significantly increases in gastric cancer tissue; TRAP1 may regulate the malignant biology of cells by increasing the expression of CyclinB1, CyclinD1, CyclinE, MMP-2 and VEGF, thereby resulting in the occurrence and development of gastric cancer.
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Osteoclasts are multinucleated cells formed mainly on bone surfaces in response to cytokines by fusion of bone marrow-derived myeloid lineage precursors that circulate in the blood. Major advances in understanding of the molecular mechanisms regulating osteoclast formation and functions have been made in the past 20 years since the discovery that their formation requires nuclear factor-kappa B (NF-kappaB) signaling and that this is activated in response to the essential osteoclastogenic cytokine, receptor activator of NF-kappaB ligand (RANKL), which also controls osteoclast activation to resorb (degrade) bone. These studies have revealed that RANKL and some pro-inflammatory cytokines, including tumor necrosis factor, activate NF-kappaB and downstream signaling, including c-Fos and nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), and inhibition of repressors of NFATc1 signaling, to positively regulate osteoclast formation and functions. However, these cytokines also activate NF-kappaB signaling that can limit osteoclast formation through the NF-kappaB signaling proteins, TRAF3 and p100, and the suppressors of c-Fos/NFATc1 signaling, IRF8, and RBP-J. This paper reviews current understanding of how NF-kappaB signaling is involved in the positive and negative regulation of cytokine-mediated osteoclast formation and activation.
Subject(s)
Cytokines , NF-kappa B , NFATC Transcription Factors , Osteoclasts , RANK Ligand , Receptor Activator of Nuclear Factor-kappa B , TNF Receptor-Associated Factor 3 , Tumor Necrosis Factor-alphaABSTRACT
Aims:As differences in promoter activity and CD30 surface expression between CD30+ lymphoid cell-lines and peripheral blood leukocytes have been shown previously to be independent of the size of the CD30 promoter microsatellite, in this study we investigate the instability within the region in a range of neoplasms including malignant lymphoma, breast and colon carcinoma and lung adenocarcinoma. Study Design:A representative sample of CD30+ and CD30- lymphomas and cell lines as well as non-haemopoietic malignancies and normal tissues were typed for CD30 microsatellite length and compared. Place and Duration of Study: Department of Anatomical Pathology, Pathwest Laboratory Medicine WA, Perth, Australia and School of Chemistry & Biochemistry, University of Western Australia, 2000-2012. Methodology: DNA was prepared from archived biopsy specimens and used to PCR ampify the CD30 microsatellite region prior to size determination using an Genescan instrument (Applied Biosystems). Results: This study has identified instability within the CD30 promoter microsatellite region in DNA from the tumour tissue of cases of malignant lymphoma, colon carcinoma and lung adenocarcinoma but not in DNA from benign lymphoid cells or normal tissues. Conclusion: These findings indicate that variability in the length of the CD30 microsatellite may be a general characteristic of the neoplastic phenotype and may reflect defects in the mismatch repair system in these malignancies rather than a specific feature of CD30- positive neoplasms. Our results suggest that the CD30-MS may be a useful marker to distinguish between normal lymphoid tissue and lymphoid malignancy.
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The herpes virus entry mediator (HVEM) is a member of the tumor necrosis factor receptor superfamily (TNFRSF), and therefore it is also known as TNFRSF14 or CD270 (1,2). In recent years, we have focused on understanding HVEM function in the mucosa of the intestine, particularly on the role of HVEM in colitis pathogenesis, host defense and regulation of the microbiota (2,3,4). HVEM is an unusual TNF receptor because of its high expression levels in the gut epithelium, its capacity to bind ligands that are not members of the TNF super family, including immunoglobulin (Ig) superfamily members BTLA and CD160, and its bi-directional functionality, acting as a signaling receptor or as a ligand for the receptor BTLA. Clinically, Hvem recently was reported as an inflammatory bowel disease (IBD) risk gene as a result of genome wide association studies (5,6). This suggests HVEM could have a regulatory role influencing the regulation of epithelial barrier, host defense and the microbiota. Consistent with this, using mouse models, we have revealed how HVEM is involved in colitis pathogenesis, mucosal host defense and epithelial immunity (3,7). Although further studies are needed, our results provide the fundamental basis for understanding why Hvem is an IBD risk gene, and they confirm that HVEM is a mucosal gatekeeper with multiple regulatory functions in the mucosa.
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Animals , Humans , Mice , Colitis , Epithelium , Genome-Wide Association Study , Immune System , Immunity, Innate , Immunity, Mucosal , Immunoglobulins , Inflammation , Inflammatory Bowel Diseases , Intestines , Ligands , Microbiota , Mucous Membrane , Receptors, Tumor Necrosis Factor , Virus InternalizationABSTRACT
Objective To study the correlation between single nucleotide polymorphism (SNP) of downstream mononucleotide in signal transduction of Toll-like receptors and predisposing genes of rheumatoid arthritis (RA).Methods One hundred and sixty-two RA patients were selected from northern part of Han people in China,and 188 healthy subjects were enrolled as normal controls.Rs7744 of Myd88,rs5030445 of TRAF6,rs11265618 and rs4845626 of IL-6R were analyzed.The data of genotypic frequency and allele genotypic frequency were statistical analyzed by x2 test between the two groups.Results The difference of allele frequency of TRAF6 rs5030445 between the two groups were remarkable (x2=5.716,P＜0.05),and so did the IL-6R rs11265618 (x2=5.704,P＜0.05).But the difference of genotypic frequency was not statistically significant for locus of Myd88 rs7744,TRAF6 rs5030445 and IL-6R rs11265618 (P＞0.05).And the differenceof allele frequency and genotypic frequency between the two groups had no statistical significance in locus of Myd88 rs7744 and IL-6R rs4845626 (P＞0.05).Conclusion There is significant correlation between single nucleotide polymorphism and predisposing genes of RA.The G allele of TRAF6 rs5030445 gene locus is a possible predisposing gene for RA.The T allele of IL-6R rs11265618 is a possible predisposing gene of RA.
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Objective To investigate the effects of curcumin on mRNA expression of cytokines related to Toll-like receptor (TLR) 4 signaling in THP-1 cells.Methods After pretreatment with different concentrations (50,25,12.5 mg/L) of curcumin or dexamethasone for 12 hours,THP-1 cells were stimulated by lipopolysaccharide (LPS.1 mg/L) for 4 hours followed by the collection of cells.Then total RNA was isolated from these cells and subjected to reverse transcription-polymerase chain reaction (RT-PCR) for the detection of mRNA expression of tumor necrosis factor receptor-associated factor 6 (TRAF6),interleukin-1 receptorassociated kinase (IRAK1) and nuclear factor (NF)-κB.THP-1 cells without pretreatment or stimulation served as negative control,and those only stimulated with LPS served as LPS group.Results After stimulation with LPS (1 mg/L) for 4 hours,the mRNA expressions of TRAF6,IRAK1 and NF-κB were significantly upregulated in THP-1 cells compared with negative control cells (f=38.69,39.13,23.99,all P<0.01).Curcumin of 50 mg/L and 25 mg/L significantly inhibited the mRNA expressions of TRAF6.IRAK 1 and NF-κB upregulated by LPS with an inhibition rate of more than 50% (all P<0.0 1).Conclusions Certain concentrations of curcumin can inhibit the mRNA expressions of TRAF6.IRAK1 and NF-κB.which demonstrates the regulatory effect of curcumin on the mRNA expressions of TLR4 signaling pathway-associated cytokines.
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Herpesvirus saimiri (HVS), a member of the gamma-herpesvirus family, encodes an oncoprotein called Saimiri Transforming Protein (STP) which is required for lymphoma induction in non-human primates. However, a detailed mechanism of STP-A11-induced oncogenesis has not been revealed yet. We first report that STP-A11 oncoprotein interacts with TNF-alpha receptor-associated factor (TRAF) 6 in vivo and in vitro. Mutagenesis analysis of the TRAF6-binding motif 10PQENDE15 in STP-A11 reveals that Glu (E)12 residue is critical for binding to TRAF6 and NF-kappaB activation. Interestingly, co-expression of E12A mutant, lack of TRAF6 binding, with cellular Src (Src) results in decreased transcriptional activity of Stat3 and AP-1, a novel target of STP-A11 compared to that of wild type. Furthermore, the presence of STP-A11 enhances the association of TRAF6 with Src and induces the translocation of both TRAF6 and Src to a nonionic detergent-insoluble fraction. Taken together, these studies suggest that STP-A11 oncoprotein up-regulates both NF-kappaB and AP-1 transcription activity through TRAF6, which would ultimately contribute cellular transformation.
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Humans , Transcription, Genetic , Transcription Factor AP-1/agonists , TNF Receptor-Associated Factor 6/metabolism , Solubility , STAT3 Transcription Factor/metabolism , Proto-Oncogene Proteins pp60(c-src)/metabolism , Protein Binding , Oncogene Proteins, Viral/metabolism , NF-kappa B/agonists , Ions , Herpesvirus 2, Saimiriine/metabolism , Detergents , Cell LineABSTRACT
OBJECTIVE: Small interfering RNA (siRNA) triggers RNA interference in mammalian somatic cells. TNF-alpha is a proinflammatory cytokine implicated in the pathogenesis of inflammatory arthritis including rheumatoid arthritis (RA). This study was to use TNF receptor 1 (TNFRI)- specific siRNA to inhibit the TNF-alpha mediated signaling in RA fibroblast like synoviocytes (FLS) and chondrocytes. METHODS: TNFRI specific siRNA was produced by targeting 3 nucleotide sequences at 474~494, 562~582 and 668~688. Reverse transcriptase polymerase chain reaction (RT-PCR) and Western blot were performed to optimize the silencing effects of TNFRI siRNA in cultured FLS and chondrocytes. The inhibition of TNF-alpha mediated signaling was determined by ELISA assay of metalloproteinase 1 secretion induced by TNF-alpha. RESULTS: The TNFRI siRNA inhibited the expression of TNFRI mRNA and protein in both RA FLS and chondrocytes. MMP-1 secretion induced by TNF-alpha was significantly downregulated by TNFRI siRNA. CONCLUSION: TNFRI siRNA can inhibit the expression and signaling downstream of TNFRI in both RA FLS and chondrocytes efficiently. This suggests that RNA interference technique by siRNA could be considered as a potential therapeutic target for RA.
Subject(s)
Arthritis , Arthritis, Rheumatoid , Base Sequence , Blotting, Western , Chondrocytes , Enzyme-Linked Immunosorbent Assay , Fibroblasts , Receptors, Tumor Necrosis Factor , Reverse Transcriptase Polymerase Chain Reaction , RNA Interference , RNA , RNA, Messenger , RNA, Small Interfering , Tumor Necrosis Factor-alphaABSTRACT
Objective To detect the concentrations of (soluble TNF receptor type,sTNFR-I, sTNFR-II) and (soluble vascular cell adhesion molecule-1,sVCAM-I) in deep burn wound fluids and the number of macrophages in different stage of human deep burn wound, and to analyse the correlation between them. Methods The wound fluid was collected with sponge during the dressing change, and then the concentrations of sTNFR-I, sTNFR-II and sVCAM-I were detected by ELISA. The tissues of burn wound were collected at operation, and the tissue slices were dyed with HE.The macrophages were marked by immunohistochemistry with antibody of CD68, and then the number of macrophages was counted under the microscope. Results The concentrations of sTNFR-I and sTNFR-II in burn wound fluid and the number of macrophages in burn wound were significantly increased two days postburn, which maintained at high levels until the wound healed. The concentration of sVCAM-I was decreased immediately after the burn until the wound healed. The correlation coefficient between the concentrations of sTNFR-I, sTNFR-II, sVCAM-I and the number of macrophages were 0.95, 0.97 and -0.37, respectively. Conclusion The number of macrophages has a strong correlation with sTNFR-I and sTNFR-II in human deep burn wound, and has a weak correlation with sVCAM-I,which suggests that the macrophages may play a cooperative role with sTNFR in burn wound healing.
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OBJECTIVE To investigate the effects of N,N'-Dinitrosopiperazine (DNP) on the cell proliferation of nasal and nasopharyngeal epithelia and the expression levels of TRAF2 and p16 genes in TgN (p53mt-LMP1)/HT transgenic mice and the relationships between them.METHODS The epithelial proliferating characteristics of nasal cavity and nasopharynx in TgN (p53mt-LMP1)/HT cancerous lesion inducing group treated by DNP (TI),TgN (p53mt-LMP1)/HT controlling group (TC), C57BL/6J cancerous lesion inducing group (CI) treated by DNP and C57BL/6J controlling group (CC) were observed for pathological evaluation by HE staining,and the expression levels of TRAF2 and p16 genes in these tissue samples were determined by immunohistochemical methods.RESULTS The occurring rates of precancerous lesions in nasal and/or nasopharyngeal epithelia in TI,TC,CI and CC groups were 90%,10%,0 and 0,respectively (P
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By searching an EST database, we identified two TNF receptor superfamily members (named mTNFRH1 and mTNFRH2). Amino acid sequences are highly conserved between the two receptors (78% identity). The chromosomal loci of mTnfrh1 and mTnfrh2 genes are found in distal chromosome 7 in the mouse. mTNFRH1 and mTNFRH2 do not contain the cytoplasmic domain, indicating that they might function as decoy receptors. Furthermore, an alternatively spliced form of mTNFRH1 was found which contains neither the transmembrane domain nor the cytoplasmic domain, thus presumably existing as a soluble form. Northern blot analysis showed that mTnfrh1 mRNA was negligibly expressed in tissues, while mTnfrh2 mRNA was strongly expressed in spleen, lung, liver, kidney, and testis. When the extracellular domains of mTNFRH1 and mTNFRH2 were expressed in bacteria, their molecular weight of extracellular region was approximately 15 kDa. Both of the soluble forms were effective in inhibiting T-cell proliferation stimulated by anti-CD3 monoclonal antibody. Our data suggest that mTNFRH1 and mTNFRH2 may be implicated in exerting a modulatory role in the immune response.
Subject(s)
Animals , Mice , Alternative Splicing/genetics , Amino Acid Sequence , Base Sequence , Cell Division/physiology , Databases, Nucleic Acid , Gene Expression/genetics , Molecular Sequence Data , Receptors, Tumor Necrosis Factor/biosynthesis , Recombinant Proteins/biosynthesis , T-Lymphocytes/cytologyABSTRACT
Objective:To construct and express the recombinant of human soluble TNF receptor I by E coli Methods:The cDNA coded for extracellular region of human TNFRI was amplified by RT PCR and inserted into a expression vector, pET 28a Then, the recombinant sTNFRI/pET 28a was transfected and expressed in E coli Results:After the stimulation with IPTG, sTNFRI fusion protein was effectively produced in E coli BL 21 transfected with the exogenous gene A unique band was found at 27 kD by SDS PAGE and its expression was about 31% of total protein in E coli The purified sTNFRI fusion protein was shown to be able to suppress the cytotoxicity of TNF on L929 cells and found by indirect immune fluorescence to inhibit specifically the binding of TNF with TNFR on target cells Conclusion:The recombinant human soluble TNFR I have been obtained by using the genetic engineering technology
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BACKGROUND AND OBJECTIVES: It has been reported that various inflammatory and immune reactions are involved in the development and progression of atherosclerosis. We tried to investigate whether the TNF receptor superfamilies are involved in the development and progression of atherosclerosis. MATERIALS AND METHOD: Thirteen carotid atheroma specimens(frozen sections : 10 cases, paraffin section : 5 cases) were obtained from the patients who underwent carotid endarterectomy at Samsung Medical Center and one normal aortic tissue was obtained from a transplantation donor in brain death. In the carotid endarterectomy specimens and a normal aortic tissue , the expressions of R110(TR1), 139(TR2) and DR3(TR3), members of the TNF receptor superfamilies were evaluated by immunohistochemical staining with monoclonal antibodies. Simultaneously, we evaluated the expressions of foam cells, smooth muscle cells, T-lymphocytes and B-lymphocytes. RESULTS: Immunohistochemical analysis identified a strong expressions of foam cells and smooth muscle cells in all atheroma. But, the expression of T-lymphocytes was minimal and that of B-lymphocytes was rare. The expression of DR3(TR3) was seen in all atheroma as strongly positive. The expression of 139(TR2) was observed well in frozen sections, but not in paraffin sections. Whereas, that of R110(TR1) was observed in paraffin sections as weakly positive, but not in frozen section. The areas where the TNF receptor superfamilies were expressed correlated to the area of foam cell presence. The expression of DR3 also correlated with expression of smooth muscle cells. In normal aortic tissue, the expression of inflammatory cells or TNF receptor superfamilies was not observed except smooth muscle cells which were observed in normal artery. CONCLUSION: Foam cells and smooth muscle cells were abundantly present in atheroma. The TNF receptor superfamilies are expressed in the atheroma and the region of expression was coincident with the presence of foam cells.