T cell response of I-Aq mice to self type II collagen: meshing of the binding motif of the I-Aq molecule with repetitive sequences results in autoreactivity to multiple epitopes

Type II collagen (CII) is of immunological interest because of its repetitive structure and properties as an autoantigen. The mouse gene has recently been cloned, thus enabling T cell-defined epitopes to be identified. Multiple novel epitopes on mouse CII are here detected in the autoreactive T cell response. The major response is directed to an epitope with residues 707-721 located on the CB10 fragment. Some 25 other epitopes are also recognized, including the autologous homologue of the 256-270 epitope which dominates in the response to foreign collagen. The cells reactive with mouse collagen peptides were of Th1 type, as judged by release of IFN-gamma. No significant reactivity was detected to mouse CII peptides during ongoing disease. Alignment of the mouse epitopes revealed a sequence motif with characteristic side chains at residues P1, P4 and P7, and to a lesser extent at P5, within a nonamer core sequence. Binding of these epitopes was simulated in a computer model of the I-Aq molecule, where peptides with anchor residues at P1, P4 and P7 were indeed found to fit the binding groove best. The spacing of pockets and the fine structure of the binding surface of the I-Aq molecule meshes with the repetitive structure of the collagen (X-Y-Gly), thus providing a likely explanation for the occurrence of multiple epitopes. Comparison with human DR binding motifs showed that the I-Aq motif resembles most closely that of the DR4 subtypes which predispose for rheumatoid arthritis.      

Cyclosporine and chloroquine synergistically inhibit the interferon-gamma production by CD4 positive and CD8 positive synovial T cell clones derived from a patient with rheumatoid arthritis.

OBJECTIVE: To investigate synergistic interaction between cyclosporine (Cy) and chloroquine (Chl) in an in vitro system, with regard to interferon-gamma (IFN) production by OKT3 activated T cell clones. METHODS. CD4+ and CD8+ T cell clones, derived from synovial tissue of a patient with rheumatoid arthritis (RA) were activated with plastic coated OKT3 monoclonal antibody in the presence or absence of various concentrations of Cy, Chl and their combinations. After 24 h of incubation the supernatants were assayed for IFN by ELISA. RESULTS. Cy as well as Chl were able to completely inhibit in a concentration dependent fashion the IFN production by CD4+ and CD8+ T cell clones. Combinations of Cy and Chl, which in themselves give minor inhibition of IFN production, were able to inhibit in a synergistically enhanced fashion the production of IFN by these clones. The synergy was formally proven by the construction of isoboles. This synergy was most pronounced when drug concentrations were used which individually gave minor inhibition of IFN production. CONCLUSION. We conclude that the results of our in vitro experiments may give rise to further investigation of the promising combination of Cy and Chl in the treatment of RA.     

Endothelial cell lysis induced by lymphokine-activated human peripheral blood mononuclear cells

In vitro exposure of human peripheral blood mononuclear cells (PBMC) to interleukin 2 results in the generation of lymphokine-activated killer (LAK) cells. Such LAK cells exhibit cytotoxicity against a spectrum of tumor target cell lines whereas they apparently do not affect normal tissues. In this report we show that PBMC that have been activated with T cell growth factor lyse trypsinized human umbilical cord venous endothelial cells as well as endothelial cell monolayers in a dose-dependent manner. Microscopic analysis showed that during the 4-h incubation period cell clumps containing detached endothelial cells and LAK cells were formed. When these clumps were evaluated with trypan blue the endothelial cells stained positive whereas LAK cells excluded the dye. No lysis occurred when fresh PBMC were added to target endothelial cells. The endothelial cell kill could not be blocked with an anti-LFA-1 antibody nor with intact OKT3 or F(ab')2 fragments of WT32. We conclude that lymphokine-activated PBMC exhibit cell-mediated endothelial cell detachment and lysis.     

Activation of rat complement by soluble and insoluble rat IgA immune complexes

The ability of rat monoclonal IgA, specific for 2,4-dinitrophenyl (DNA), to activate the complement (C) system of the rat was investigated using aggregated IgA or IgA immune complexes (IC). IgA was coated onto a solid phase, and tested for its capacity to bind C3 upon incubation at 37 degrees C in normal rat serum (NRS) in the presence of Mg-EGTA. Binding of C3 was observed dependent on the dose of dimeric (d-), polymeric (p-) and secretory IgA tested. In contrast, little C3 fixation was observed in this system with monomeric (m-) rat IgA or with mouse m- and d-IgA (MOPC315). Soluble and insoluble rat IgA IC were prepared using dinitrophenylated rat serum albumin (DNP8RSA) as antigen (Ag), and assessed for C activation. It was shown that insoluble IC (immune precipitates; IP) containing m-, d- or pIgA of rat origin activate the alternative pathway of rat C, as demonstrated by their capacity to induce C consumption in NRS in the presence of Mg-EGTA. When p- and m-IgA IP were compared for their capacity to activate C, it was found that p-IgA activated C four times as efficiently as m-IgA IP (at 2 mg/ml). Soluble rat IgA IC were prepared in an excess of DNP8RSA, fractionated by gel filtration on Sepharose 6B, and analyzed for C activation and antibody (Ab)/Ag ratio. In contrast to m-IgA IP, soluble m-IgA did not activate C. On the other hand soluble d-IgA IC activated C dependent on their concentration and size: at a concentration of 0.1 mg/ml high-molecular weight d-IgA IC with a high Ab/Ag ratio were four times as efficient as low-molecular weight IC with a low Ab/Ag ratio, and twice as efficient as IP prepared at equivalence. To demonstrate the induction by IgA of the assembly of the terminal membrane attack complex, trinitrophenyl (TNP)-conjugated rat red blood cells (TNP-RRBC) coated with d- or p-IgA were shown to be lysed in NRS in the presence of Mg-EGTA. No lysis of m-IgA-coated TNP-RRBC was observed. The results in this study demonstrate that both soluble and insoluble rat IgA IC activate the alternative pathway of homologous rat C. Alternative pathway activation by soluble rate IgA IC is dependent on the size of the IC. The degree of polymerization of the IgA Ab itself also influences C activation.     

The mouse Mid1 gene: implications for the pathogenesis of Opitz syndrome and the evolution of the mammalian pseudoautosomal region

We have recently reported isolation of the gene responsible for X- linked Opitz G/BBB syndrome, a defect of midline development. MID1 is located on the distal short arm of the human X chromosome (Xp22. 3) and encodes a novel member of the B box family of zinc finger proteins. We have now cloned the murine homolog of MID1 and performed preliminary expression studies during development. Mid1 expression in undifferentiated cells in the central nervous, gastrointestinal and urogenital systems suggests that abnormal cell proliferation may underlie the defect in midline development characteristic of Opitz syndrome. We have also found that Mid1 is located within the mouse pseudoautosomal region (PAR) in Mus musculus , while it seems to be X- specific in Mus spretus. Therefore, Mid1 is likely to be a recent acquisition of the M. musculus PAR. Genetic and FISH analyses also demonstrated a high frequency of unequal crossovers in the murine PAR, creating spontaneous deletion/duplication events involving Mid1. These data provide evidence for the first time that genetic instability of the PAR may affect functionally important genes. In addition, we show that MID1 is the first example of a gene subject to X-inactivation in man while escaping it in mouse. These data contribute to a better understanding of the molecular content and evolution of the rodent PAR.      

p53 immunoreactivity in hepatocellular adenoma, focal nodular hyperplasia, cirrhosis and hepatocellular carcinoma

The prolonged half-life of mutant p53 makes feasible its immunocytochemical detection. In order to assess the pathogenetic role of mutant p53 in regenerative and neoplastc liver disease we studied its immunohistochemical expression in cases of hepatic cirrhosis, hepatocellular carcinoma (HCC), cirrhosis with areas of HCC, hepatocellular adenoma and focal nodular hyperplasia. The study included needle and wedge biopsies of 50 cirrhotic livers, 59 HCCs (36 of them with associated cirrhosis), six adenomas and two focal nodular hyperplasias. Sixty-five HCC fineneedle cytology specimens were also included in the study. There was no immunohistochemical evidence of mutant p53 expression in any of the cases of cirrhotic liver (except for one instance associated with HCC) adenoma or focal nodular hyperplasia. In contrast p53 was detected in 8.5% of HCC cases in the biopsy series and 24% of HCC cases in the fine needle aspiration series. In addition, mutant p53 expression in HCC was positively correlated with tumour grade. According to grade, the distribution of p53 positive immunoreactivity among HCCs was as follows: Grade I-II, 0% of cases in the biopsy series and 9% in the fine needle aspirates; Grade III, 18% in the biopsy series and 55% in the fine needle aspirates; and Grade IV, 40% in the biopsy series. Therefore, mutant p53 expression does not seem to be associated with benign liver lesions but seems to correlate with the progression of HCC through various grades of increasing malignancy.     

Formation in the presence of C3 nephritic factor (C3NeF) of an alternative pathway C3 convertase containing uncleaved B.

C3 nephritic factor (C3NeF) interacts with native C3 and B in the absence of D to generate a C3 convertase containing an uncleaved form of B. Dose response studies with C3NeF and B, respectively, revealed incremental C3 inactivation without loss of B. These findings are in agreement with the previous isolation from such reaction mixtures of a 10S complex containing haemolytically inactive C3 and active B and manifesting C3 convertase activity. Functional contamination of C3 with C3b was negated by demonstrating that pretreatment of C3 with C3bINA had no effect on its subsequent interaction with B and C3NeF to generate C3 convertase activity, while pretreatment of C3b eliminated its effective interaction with B and C3NeF. Relatively higher concentrations of C3bINA present during interaction of C3, B and C3NeF suppressed C3 inactivation, indicating its dependence on amplification by utilization of the initial C3b generated. Trace quantities of D were not found by functional analyses of C3, B and C3NeF and pretreatment of these proteins with a concentration of DFP sufficient to suppress D activity had no effect on their effective interaction. The introduction of D to mixtures of C3NeF, B, and C3 resulted in B clevage and more efficient expression of C3 convertase function as defined by a reduced requirement for C3NeF.     

Clearance kinetics and tissue distribution of aggregated human serum IgA in rats.

In the present study the clearance kinetics and tissue distribution of human polyclonal heat-aggregated serum IgA (AIgA) of different sizes in rats was studied after intravenous administration of 125I-AIgA. The 125I-AIgA of different sizes disappeared from the circulation in a biphasic manner with an initial rapid half-life (T1/2) and a second slower T1/2. The first T1/2 was related to the size of the 125I-AIgA: high molecular weight (MW) 125I-AIgA was cleared much faster than 125I-AIgA with a low MW. Relatively more degradation products were observed in blood when high MW 125I-AIgA were injected as compared to low MW 125I-AIgA. The AIgA were mainly taken up by the liver. Eight minutes after injection of high MW 125I-AIgA, 90% of the injected dose was found in the liver, whereas less than 2% was detected in the spleen. Very little activity was detectable in other organs, such as lungs, heart and kidneys. In the present study 1-3% of the injected 125I-AIgA were found in the bile. Analysis of this material revealed that low MW 125I-AIgA were transported more efficiently to the bile than high MW 125I-AIgA. To obtain more insight into the receptors involved in the clearance of 125I-AIgA, rats were pretreated with ovalbumin or asialofetuin. The clearance of 125I-AIgA of different sizes was inhibited when rats were pretreated with asialofetuin. Pretreatment with ovalbumin had no effect on the clearance rates of 125I-AIgA. These results suggest a role for carbohydrate receptors, which recognize glycoprotein-containing galactose terminal residues on Kupffer cells, in the clearance of 125I-AIgA.     

电离辐射对放射工作者职业健康的影响

目的 分析放射工作者外周血象、淋巴细胞微核及染色体畸变情况,为放射工作者职业防护和健康监测提供依据。方法 对2015年、2017年和2019年连续3次接受健康检查的127名放射工作者进行淋巴细胞微核、染色体及血象分析,将其设为放射组。另外选取133名无射线接触史的医务人员设为对照组;结果 放射组中淋巴细胞微核率和染色体畸变率高于对照组,白细胞和血小板计数低于对照组,均具有统计学意义（P < 0.05）。127名放射工作者外周血白细胞总数随着接触电离辐射时间的增长逐渐降低,染色体畸变率逐渐增加,均具有统计学意义（P < 0.05）。损害工龄大于20年的放射工作者染色体畸变率高于低工龄组,不同损害工龄之间比较无统计学意义（P > 0.05）。核医学与介入治疗工种染色体畸变率高于其他工种,具有统计学意义（P < 0.05）。结论 长时间接触低剂量电离辐射可使放射工作者白细胞总数降低和淋巴细胞染色体畸变率增加,应加强放射工作者防护措施以备降低电离辐射损伤程度,特别要加强核医学和介入治疗放射工作人员的职业防护。     

Tubular epithelial cells: A critical cell type in the regulation of renal inflammatory processes

Most chronic human kidney diseases are characterized by a final common pathway consisting of interstitial inflammation and ultimately leading to interstitial fibrosis. Within this process, tubular epithelial cells (TECs) play a critical role. Both in vitro and in vivo it has been demonstrated that TECs are an important source of various cytokines, chemokines, growth factors, adhesion molecules and extracellular matrix components. In the present review we will outline the capacity of TECs to produce inflammatory mediators and discuss the different mechanisms involved in the regulation of production of these mediators.