摘要:

We investigated intraepithelial T cells from the small intestine, SI (jejunum, ileum) and the large intestine, LI (colon) of euthymic (BALB/c, H–2d; C. B–17 +/+, H–2d; C57BL/6, H–2b) and athymic (C57BL/6nu/nu; BNXbg/bg nu/nu xid/xid) mice. From individual euthymic and athymic mice, 7 × 106intraepithelial lymphocytes (IEL) per mouse were isolated from the SI. Ten–fold lower numbers of IEL were obtained from the LI epithelium (4 × 105IEL per mouse). Thymus–dependent and ‐independent T cells represented>80% of SI–IEL but the fraction of T cells was reduced from 20% to 40% in LI–IEL. In euthymic mice, αβ T cells predominated in SI–IEL and in particular in LI–IEL populations, while SI–IEL and LI‐IEL populations of athymic mice contained predominantly αβ T cells. The intraepithelial T cell subset distribution was different in SI versus LI: mainly CD8+T cells were present in the SI, but a large CD4+T cell subset was present in the LI.‘Double positive’ CD4+CD8α+T cells were present mainly in the SI epithelium but were rare in the LI epithelium. In euthymic as well as athymic mice, T cells expressing the homodimeric CD8αβ isoform predominated in the SI epithelium, while T cells expressing the heterodimeric CD8αβ isoform predominated in the LI epithelium. LI–derived TCRα+IEL displayed the CD2+CD28+LPAM–1/2−M290+phenotype, and a fraction of them expressed the L–selectin LECAM–1. In contrast, a large fraction of TCRα+SI‐IEL was CD2−CD28−LPAM–1/2−M290+and LECAM–1−. RAG–1/2 expression was detectable by RT–PCR in IEL from the SI but not the LI. Striking differences in phenotype were thus apparent between thymus–dependent and thymus–independent T cel

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03541.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Flow cytometric measurement of circulating CD4+lymphocytes is important in the evaluation of disease progression in HIV–infected patients. Development of dyes that can be exited at 488 nm and have emission maximum in the far red area has made three–colour protocols, together with fluorescein isothiocyanate (FITC) and R–phycoerythrin (PE), possible in most clinical flow cytometers. We report here the comparison of a two–tube, three–colour protocol (including CD45/CD4/CD3 and CD8/CD4/CD3) with our conventional dual–colour protocol. No significant differences were found between percentage of CD3+lymphocytic cells determined with three different antibody combinations. When the CD8/CD4/CD3 combination was used a systematic overestimation of CD3+CD4+% cells was found. This turned out to be caused by the formation of ‘CD8–escapees’. These are clumps of CD8+cells that fall outside the lymphocyte gating region, principally because of high side scatter. The problem can be overcome by rigorous vortexing to loosen aggregates. The lymphocyte gating principle used in this protocol (gating on a side scatter/CD45 dot plot) is readily applicable to other antibody combinations. This was demonstrated by measuring CD5+B lymphocytes, a subset receiving increasing attention in the study of HIV–induced immune deviations. We conclude that our three–colour protocol for CD4+T–lymphocyte determinations offers significant advantages to the conventional dual–colour method, and we suggest that when possible anti–CD45 be added to dual–colour combinations in order

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03542.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Intradermal administration of glutaraldehyde–fixed Aujeszky's disease virus (ADV) infected autologous or allogeneic cells resulted in the induction of an interferon(IFN)–α/β response in pigs. Using a sensitive dissociation–enhanced lanthanide fluoroimmunoassay (DELFIA), IFN–α/β was detected in blood at 8 and 24 h after injection of ADV–infected cells. In parallel, by means ofin situhybridization, IFN–α/β mRNA containing cells were demonstrated in regional lymph nodes. Occasional IFN–α/β mRNA positive cells were also seen in injected dermal areas, but not in contralateral lymph nodes, spleen, bone marrow, blood or liver. The ability of leucocytes in whole blood cultures to produce IFN–α/β upon stimulation by ADV was markedly diminished 3–7 days after intradermal injection of ADV–infected cells. In contrast, cultures of purified peripheral blood mononuclear cells (PBMC) had intact IFN–α/β responses. Further, serum from ADV–injected pigs inhibited thein vitroADV–induced IFN‐α/β responses in PBMC from control pigs, most likely due to the demonstrated presence of anti–ADV antibodies. We suggest that the IFN‐α/β producing cells in lymph nodes may participate in the development of antiviral immunity and could be equi

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03543.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

We report a novel mechanism ofIgG2deficiency. Several investigators have reported patients withIgGsubclass deficiencies due to homozygous deletion of immunoglobulin heavy chain constant region genes. However, it is unclear what mechanism is responsible forIgGsubclass deficiency in cases where no gene deletions have been detected and which are accompanied by recurrent infections due to aberrant immunoregulation. In the present study, we have focused our attention on production by peripheral blood mononuclear cells (PBMCs) of interferon‐gamma (IFN‐γ), which is known to induceIgG2expression. PBMCs from four patients with IgG2 deficiency and their families were studied. Mitogen‐induced IFN‐γ production byPBMCswas decreased in all of the patients, although the proliferative responses of PBMCs and the percentages ofCD3, CD4, andCD8T cell subsets were not decreased.IgG2production byPBMCswas restored upon addition of IFN‐γ and mitogen to the PBMCs of the patients withIgG2deficiency though it was not restored in the patients with common variable immunodeficiency. We conclude that defects in production of IFN‐γ play an important role i

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03544.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

The epitope repertoire of B cells, due to their selective ability to process their specific antigen and the potential bias imposed on the resulting peptides by the surface immunoglobulins bound to the antigen, may influence the T‐helper repertoire.Immunization of C57B1/6 mice withTorpedoacetylcholine receptor (TAChR) causes experimental autoimmune myasthenia gravis (EAMG). Anti‐TAChRCD4+cells recognize epitopes within three sequence regions of theTAChRa subunit (“dominant epitopes’). Immunization of mice with denatured or syntheticTAChRantigens sensitizes CD4+cells to otherTAChRsequence regions (‘cryptic epitopes’).We investigated here whether clustering of B and T epitopes within the same short sequence segments occurs during the anti‐TAChRresponse, as previously described for the response to hexogenous antigens unrelated to homologous self proteins.Twelve 19–20 residue synthetic sequences of theTAChRα, β and δ subunits, containing dominant or cryptic CD4+epitopes for C57B1/6 mice, were tested for ability to induce anti‐peptide antibody production. C57B1/6 mice were immunized with the individual peptides. Ten peptides stimulated antibody production. Therefore>80% of these shortTAChRsequences also contain B epitopes.Therefore also in the anti‐TAChRresponse leading toEAMGT and B cell epitopes frequently reside within the same

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03545.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

We have examined for the presence or absence of T cell receptor V‐alpha (VA) and V‐beta (VB) gene expression in infiltrating T lymphocytes (ITL) isolated from Graves' thyroid glands in comparison to paired peripheral blood lymphocyte (PBL) samples using a qualitative based polymerase chain reaction (PCR) assay. Sequence specific oligonucleotides for VA and VB T cell receptor gene (TCR) families that had previously been validated in other studies, were used for the PCR analysis, followed by Southern blot hybridization with a labelled, internal C‐region primer. A total of seven Graves' disease patients who had been treated with carbimazole were studied. T ceil receptor VA and VB gene usage was examined in freshly isolated, unstimulated ITLs from five patients. A widespread usage of VA and VB gene families with 12 to 18 families being used was apparent. Use of oiigo‐dT or C‐region priming of the mRNA prior to reverse transcription of the mRNA did not have any significant affect on the results nor did the use of whole Graves' thyroidmRNA as the starting material (n = 2) or perfusion of one gland with saline to remove as much of the contaminating blood from the gland. Our results contrast with those of Davies and colleagues who have previously shown a restricted repertoire of VA gene families m ITLs in comparison to autologous PBLs, and are much more in line with other recent reports indicating a diverse VA repertoire of the infiltrating T cells in Graves' thyroid glands derived from patients treated with anti‐th

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03546.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

Gangliosides modulate the expression ofCD4molecules on the cell surface of T lymphocytes. We report here that treatment of human peripheral blood lymphocytes with exogenous monosialogangliosideGM3induces a rapid down‐modulation of theCD4molecules on the plasma membrane ofCD4+T lymphocytes, as assessed by cytofluorimetric analysis and quantitative immunoelectron microscopy. TheCD4down‐modulation was gang Hoside‐dose dependent and was already evident after 5 min of treatment, reaching the maximum after 20 min. The expression of other surface antigens was not affected byGM3treatment. The immunoelectron microscopic analysis showed that, followingGM3addition, gold labelledCD4molecules were rapidly redistributed on the cell surface, clustered and internalized via endocytic pits and vesicles. These results indicate thatCD4down‐modulation induced byGM3occurs through an endocytic mechanism. A persistent low level ofCD4expression on the cell surface up to 24 h afterGM3treatment, compared with a stable expression of eitherCD4in untreated cells andCD3inGM3‐treated cells, suggests intracellular degradation of the internalizedCD4

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03547.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

TheMRL‐lpr/lprandMRL‐++ mice were studied for the expression of cytokines in the spleen, lymph node., thymus, kidney and brain through the reverse transcription‐polymerase chain reaction (RT‐PCR). The frequencies of IL‐4 and TNF‐a expression in the thymus and spleen were significantly higher inMRL‐lpr/lprmice than inMRL‐++ mice from the age of 17 to 32 weeks. More importantly, IL‐4 transcript was demonstrated in the early rather than in the terminal stage of the lupus disease. At the 20th week,MRL‐lpr/lprmice with active disease exhibited higher concentrations of IL‐1α, IL‐6 andTNF‐a in serum thanMRL‐++ mice. Interestingly, inMRL‐lpr/lprbut notMRL‐++ mice, the IL‐6 concentration in culture supernatants of the thymic cells was significantly higher than that of the splenic or lymph node cells. On the other hand, IL‐6 and IL‐l/? were expressed in the brain and kidney ofMRL‐lpr/lprmice but not ofMRL‐++ mice. CulturedMRL‐lpr/lprmesangial cells could also express IL‐6 but to a lesser extent. These results suggest that the abnormal splenic and thymic IL‐4 andTNF‐α expression may predispose the development of autoimmune reactions. The expression of IL‐1ß and IL‐6 in the brain and kidney may be implic

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03548.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

The expression of lymphokine genes during infection of virulent (Tulahuén) or mild (CA‐I) strains of T. cruzi was studied in mice lackingCD4and/orCD8molecules. The increased susceptibility ofCD4−andCD4−CD8−mice to infection withCA‐Ior Tuiahuen was parallelled by diminished IFN‐γmRNAlevels. Nitric oxide release and inducible nitric oxide synthasemRNAaccumulation by cells from Tulahuen infectedCD4−mice was also diminished.CD8−(but notCD4−CD8−mice) showed an increasedIL‐4andIL‐10mRNAaccumulation upon infection with both strains ofT. cruzi.A Th2‐like’ response (higherIL‐4andIL‐10mRNAto IFN‐γmRNAratio), was also observed when cells from non‐infectedCD8‐ m

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03549.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY

摘要:

The inflammatory nature of multiple sclerosis (MS) implicates the participation of immunoregulatory cytokines, including the Th2 relatedIL‐10. We describe the use of in situ hybridization withcDNAoligonucieotide probes to detect and enumerate mononuclear cells (MNC) expressingmRNAforIL‐10which is known to down‐regulate Thl cell related cytokines such as interferon‐γ. Expression ofIL‐10was studied in bloodMNCofMSand blood and cerebrospinal fluid (CSF)MNCof optic neuritis (ON) patients without culture and after culture in the presence of myelin basic protein (MBP), the control antigen acetylcholine receptor (AChR), and without antigen. Numbers ofIL‐10mRNAexpressingMNCwere elevated in theMSpatients' blood both when enumerated without culture and after culture with MBP. Control patients with myasthenia gravis had elevated numbers ofAChR‐reactiveIL‐10mRNAexpressing cells, while numbers ofMBP‐reactiveIL‐10positive ceils did not difl'er from numbers registered in cells without antigen. Patients withON, in many instances representing early MS, hadIL‐10positive bloodMNCthat were elevated to the same extent as in clinically deflniteMS, and further increased in theCSF.ONpatients examined within 1 month after onset had lower numbers ofMBPinducedIL‐10mRNAexpressing bloodMNCcompared with patients examined later suggesting thatIL‐10is related to the degree of inf

ISSN:0300-9475    

DOI:10.1111/j.1365-3083.1995.tb03550.x

出版商:Blackwell Publishing Ltd    

年代:1995

数据来源: WILEY