ABSTRACTDendritic cells (DCs) are professional antigen-presenting cells that can prime T cells and polarize the cellular immune response. Because Th1-type immune responses have been connected to success in combating viral infection, a promising therapeutic application of DCs would be their differentiation in vitro and injection back into the host to boost an immune response in infected animals. This study was aimed both at developing a protocol to cultivate feline DCs in the absence of exogenous proteins for their use in vivo and at investigating what might be the most appropriate stimulus to induce their maturation in vitro and finding correlates of maturation. We generated DCs from peripheral blood monocytes in the presence of feline interleukin-4 and granulocyte-macrophage colony stimulating factor, and after 5 days their maturation was induced with either lipopolysaccharide, human recombinant tumor necrosis factor alpha, poly(I:C), or activated feline platelets. After 48 h, their CD14, CD1a, major histocompatibility complex class II, and B7.1 surface expression was analyzed in parallel with their ability to uptake antigen or prime a mixed leukocyte reaction. The results presented show that feline DCs cultured in autologous plasma differentiate and are able to mature in the presence of stimuli similar to the ones currently used for other species. The present work sets the grounds for future use of DCs obtained by the protocol described for in vivo vaccination and immunotherapy of feline immunodeficiency virus-infected cats.

2001-05-01·Veterinary Immunology and Immunopathology3区 · 农林科学

Regulation of bovine E-selectin expression by recombinant tumor necrosis factor alpha and lipopolysaccharide

3区 · 农林科学

Article

作者: B.A Mallard ; Corinne Van Kampen

Induction of adhesion molecules by cytokines and LPS is an important mechanism of regulating leukocyte migration into tissue. Expression and regulation of E-selectin may be differentially influenced by the stimuli involved with effects on mRNA or surface protein kinetics. Surface protein and mRNA expression kinetics of bovine E-selectin were measured and compared in primary cultures of bovine aortic endothelial cells (BAEC) stimulated for various periods of time with recombinant bovine tumor necrosis factor alpha (rbTNF-alpha) or Escherichia coli lipopolysaccharide (LPS). E-selectin mRNA expression was measured via quantitative reverse transcription polymerase chain reaction (Q-RT-PCR) using a construct that contained multiple synthetic oligonucleotides for several bovine adhesion molecules and cytokines. Surface expression of E-selectin was measured by flow cytometry. Unstimulated BAECs expressed minimum or no E-selectin on the surface. A low number of endothelial cells expressed surface E-selectin as early as 1h post-stimulation and surface expression was sustained after both stimuli for 24-72h. Mean fluorescence intensity (MFI) indicated peak surface concentration of E-selectin at 6 h post-stimulation after LPS followed by a gradual decrease to 72h without returning to baseline values. Mean fluorescence intensity following stimulation with TNF-alpha increased slightly between 0 and 72h. The pattern of mRNA expression differed between stimuli. LPS-stimulated BAECs expressed peak amounts of E-selectin mRNA at 6 h, followed by a decline to baseline by 24 h. Conversely, BAECs stimulated with rbTNF-alpha expressed significantly (p pound 0.05) higher amounts of mRNA at 1h than compared to unstimulated controls (0 h), but this decreased to below baseline levels by 6h; followed by a gradual increase and eventually a sharp increase between 18 and 72 h. To account for the lack of correlation between mRNA and protein expression, it was hypothesized that shedding of surface E-selectin accounted at least in part, for the large increase in mRNA expression seen at 18-72h. Culture supernatants from rbTNF-alpha-treated BAECs were harvested, and tested for the presence of shed E-selectin using ELISA. Unstimulated culture supernatants contained little or no E-selectin. Between 6 and 48 h, the concentration of E-selectin in culture supernatants from rbTNF-alpha-stimulated BAECs increased approximately two-fold, suggesting that the sharp increase in E-selectin mRNA expression around 18 h may be related to significant loss of surface E-selectin during this period.

2000-10-20·The European Journal of Surgery

Antisense Oligodeoxynucleotides to Human Inducible Nitric Oxide Synthase Selectively Inhibit Induced Nitric Oxide Production by Human Vascular Endothelial Cells: An Experimental Study

Article

作者: Nariyuki Hayashi ; Katsuhisa Tanjoh ; Ryouichi Tomita

OBJECTIVETo find out whether the antisense oligodeoxynucleotide designed against the complementary DNA to human inducible NO synthase (iNOS) would block the translation from iNOS mRNA to NO in human vascular endothelial cells.DESIGNProspective controlled study.SETTINGResearch laboratory, Japan.MATERIALCultured human vascular endothelial cells.INTERVENTIONSHuman vascular endothelial cells were cultured for 24 hours, and divided into two groups, one of which was exposed to antisense oligodeoxynucleotides and the other to sense oligodeoxynucleotides at 100, 200, 400 and 800 micromol/L, respectively, for 18 hours. They were then exposed to interferon-gamma (1000 units/ml) and recombinant human tumour necrosis factor alpha (500 units/ml) to stimulate NO production. Each experiment was repeated ten times.RESULTSClear bands expressing cDNA of iNOS mRNA were identified in agarose gels in all cultured cells in both groups. The mean nitrite concentration in the supernatants of cultured cells in the group with the addition of antisense oligodeoxynucleotides was significantly lower than that in the group with sense oligonucleotides added at 200, 400, and 800 micromol/L.CONCLUSIONThese findings indicate that the antisense oligodeoxynucleotides inhibited the NO production dependently, by preventing only their translation but not the processing, before their transcription from DNA into iNOS mRNA.

100 项与 Recombinant mutant human tumor necrosis factor alpha(State Key Laboratory Of Cancer Biology) 相关的药物交易

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适应症	最高研发状态	国家/地区	公司	日期
结直肠癌	临床2期	-	State Key Laboratory Of Cancer Biology	-
头颈部肿瘤	临床2期	-	State Key Laboratory Of Cancer Biology	-
肺癌	临床2期	-	State Key Laboratory Of Cancer Biology	-
肾细胞癌	临床2期	-	State Key Laboratory Of Cancer Biology	-
胃癌	临床2期	-	State Key Laboratory Of Cancer Biology	-