Neurotransmitters regulate the migration and cytotoxicity in natural killer cells

Natural killer (NK) cells and cytotoxic T lymphocytes (CTL), the functional coordination of which are governed by various signal substances, are crucial in the body’s defense of tumor and virus-infected cells. We investigated the role of various neurotransmitters and hormones on the regulation of functional parameters, including NK cell cytotoxicity, and the migration of NK cells and CTL within a three-dimensional collagen lattice. Using peripheral blood CTL and NK cells, we show that the neurotransmitters endorphin, histamine and substance P increase NK cell cytotoxicity, while norepinephrine inhibits cytotoxicity. Moreover, substance P reduces migratory activity, while norepinephrine increases NK cell and CTL migration. Furthermore, all three steroid hormones which were investigated, namely cortisone, testosterone, and estradiol, had regulatory influence on both cytotoxicity and migration of NK cells. These results further specify the functional basis of the complex interconnection between the immune and neuro-endocrine systems.     

Activated status of tumour-infiltrating lymphocytes and apoptosis in testicular seminoma.

Testicular seminoma is characterized by a prominent lymphoid infiltrate and an excellent prognosis. Cytotoxic T-lymphocytes (CTLs) infiltrating seminoma tumour nests constitute a major subset of the lymphoid infiltrate. The objective of this study was to determine whether CTLs express markers of cytotoxic potential and activity and whether the number of activated CTLs correlates with the extent of apoptosis in testicular seminomas, as opposed to non-seminomatous testicular germ cell tumours (NSTGCTs). Twenty cases of pure seminoma as well as 20 cases of NSTGCTs including 16 mixed germ cell tumours (MGCTs) were studied. Immunohistochemistry for the cytotoxic markers TIA-1 (cytotoxic potential) and granzyme B (cytotoxic activity) and the T-cell markers CD3 and CD8 was performed on formalin-fixed, paraffin-embedded sections. The apoptotic index (AI) was determined by the TUNEL method. The number of CD3(+), CD8(+), TIA-1(+), and granzyme B(+) cells in tumour cell nests was markedly increased in testicular seminomas, compared with NSTGCTs (p<0.01). Activated granzyme B(+) cells numbered 25.6+/-5.2 per high power field in seminomas and 8.9+/-3.2, 8.1+/-3.9, and 0.4+/-0.2 for embryonal carcinomas, yolk sac tumours, and immature teratomas, respectively. Double immunohistochemical staining for granzyme B and CD8 revealed that 82.6+/-8.5% of granzyme B-expressing cells were CD8(+). The tumour cell AI was significantly increased in embryonal carcinoma, compared with the seminoma, yolk sac tumour, and immature teratoma subgroups (6.7+/-1.3, 2.3+/-0.3, 3.0+/-1.1, and 2.3+/-1.1, respectively, p<0.001). TUNEL/CD3 double immunostaining revealed that a significant proportion of the apoptotic seminomatous tumour cells were in direct contact with one or more CD3(+) lymphocytes (47.2+/-6.2%). The number of activated granzyme B(+) CTLs showed a strong linear correlation with the AI in the seminoma group (r=0.71, p<0.0001) but not in other subgroups. TUNEL/granzyme B double immunolabelling revealed that a proportion of activated granzyme B(+) lymphocytes (20%) were often seen in close contact with apoptotic tumour cells. The presence of increased numbers of activated cytotoxic lymphocytes in testicular seminomas suggests that apoptotic tumour cell death in this neoplasm may be triggered by cytotoxic granule effectors. This phenomenon may be one of the key host immune mechanisms leading to the excellent prognosis in this tumour.     

Modeling the presentation of C3d-coated antigen by B lymphocytes: enhancement by CR1/2-BCR co-ligation is selective for the co-ligating antigen

We have used a set of single-chain variable fragment antibodies (sc) genetically fused with an influenza hemagglutinin-derived peptide as a means to investigate the role of CR1 and CR2 in antigen presentation by B cells. When incubated with the B cell lymphoma 2PK3, peptide-containing sc specific for either CR1 or CR1/2 mediated activation of the hemagglutinin peptide-specific T cell line IP-12-7, as assessed by IL-2 production. Efficient presentation was dependent on the binding of the constructs to CR1/2, implying that receptor-mediated endocytosis is responsible for the effect. Cross-linkage of CR1/2 or CD19 by mAb did not increase the extent of T cell activation. However, when CR1/2 was co-ligated with the BCR--using either polyclonal goat anti-mouse IgG or recombinant protein LA--the antigen concentration required to activate T cells decreased by two orders of magnitude. Moreover, this enhancement was selective for the antigen included in these complexes and did not affect the presentation of a free peptide or of antigen bound to CR1/2 excluded from the complexes. These results suggest that B cells may bind various C3d-coated antigens at a time, but only the one which reacts with the BCR will be processed with high efficiency. This mechanism may ensure the specificity of cognate T cell help.     

Rapid tyrosine phosphorylation of Grb2 and Shc in T cells exposed to anti-CD3, anti-CD4, and anti-CD45 stimuli: differential effects of aging

Two adapter proteins, Grb2 and Shc, have recently been implicated in the transmission of activation signals from the stimulated T cell receptor to Ras. We show here that in vitro stimulation of mouse splenic T cells with crosslinked anti-CD3 antibody leads within 30 s to phosphorylation of both Grb2 and Shc. Treatment with crosslinked anti-CD45 antibody leads to phosphorylation of Grb2 and also to a slight retardation in the mobility of this protein in an SDS polyacrylamide gel; both changes are seen within 30 s of crosslinking. Crosslinked anti-CD4 antibody leads to phosphorylation of Shc and to the phosphorylation of a 30-kDa protein that cross-reacts with anti-Grb2 antibodies. Aging leads to a decline in CD3-stimulated phosphorylation of Shc (but not Grb2), and to an increase in CD4-stimulated phosphorylation of Grb2, Shc, and the 30-kDa Grb2-like protein. Increased tyrosinephosphorylation of Grb2 after exposure to either anti-CD3 or anti-CD45 suggests that Grb2 may be a common substrate for both CD3-linked kinases and the CD45 phosphatase. The differences between T cells from young and old mice suggest that aging may lead to a set of alterations in kinase/substrate coupling that contribute to immune dysfunction in the elderly, and that activation of the Ras pathway might be impaired by aging in T lymphocytes.     

Aggressive large granular lymphocyte lymphomas in five dogs: a clinical cytohistological and immunological study

Among 276 canine lymphomas referred to the Haematology–Cytology–Immunology laboratory of the Lyon Veterinary School between 1997 and 2003, there were five aggressive large granular lymphocyte (LGL) malignancies. The five dogs were clinically examined and followed up. Cytological and histological analyses of the liver, spleen, lymph nodes, intestine and bone marrow were performed. The immunophenotype and proliferation index were established. The most significant clinical finding was that of an aggressive clinical course, the presence of hepatomegaly and/or splenomegaly, an abdominal lymphadenopathy, anaemia in four cases and blood and bone-marrow involvement in two cases. The cytological presentation was a diffuse infiltration of atypical, large granular lymphoid cells. Two cases were of the null type (CD3–, CD79a–, CD4–, CD8–), and three were of the T-cell type (CD3+, CD79a–, CD4–, CD8+). The proliferation index was high in all cases, with a median of 54.4%. The histological presentation of the null-type cases was an infiltration of the livers portal triads and the spleens red pulp. The T CD8+ cases showed two different patterns, characterised by infiltration: in the first, of all the intestines layers, the livers portal triads, the spleens red pulp and the lymph nodes and, in the second, infiltration of the livers sinusoids, the spleens red pulp. Although these aggressive LGL lymphomas are still poorly known, they may be compared to three types of human lymphoma: aggressive NK cell lymphoma/leukemia, hepatosplenic T-cell lymphoma and enteropathy-type T-cell lymphoma.     

Antibodies to acetylcholine receptors in myasthenia gravis. In vitro synthesis by peripheral blood lymphocytes before and after thymectomy.

Pokeweed mitogen (PWM)-driven in vitro synthesis of antibodies to the acetylcholine receptor (PSA) was studied in non-thymoma patients with myasthenia gravis. In a group of 46 patients, the occurrence of PSA was related to the presence of the thymus or, in operated patients, the absence of a clinical effect of thymectomy. Sixteen patients were followed before and soon after thymectomy. PSA disappeared in all patients, at least temporarily, between 6 weeks and 1 year afterwards, independent of the clinical course and eventual clinical effect of the operation. A recurrence was found only in one of the five patients who derived no benefit from the operation. These findings support the hypothesis that the therapeutic effect of thymectomy can be explained by removal of a source of autoreactive lymphocytes. There was no correlation between the changes in serum levels of a-AChR and clinical improvement, suggesting a minor role of circulating peripheral blood lymphocytes (PBL) and the thymus in the total production of a-AChR.     

Induction of Ly-6A/E expression by murine lymphocytes after in vivo immunization is strictly dependent upon the action of IFN-alpha/beta and/or IFN-gamma

Ly-6A/E is a phosphatidylinositol-linked membrane protein which mediates murine T and B cell signalling. IFN-gamma, IFB-alpha/beta, LPS, and IL-4 have all been reported to induce or upregulate Ly-6A/E by normal lymphocytes. Since no systematic study has addressed the stimulant selectivity of Ly-6A/E expression by murine lymphocytes nor investigated its induction and regulation during primary in vivo immune responses we analyzed in vitro Ly-6A/E expression after murine stimuli and during a number of distinct in vivo immunizations. We show that LPS induces B cell Ly-6A/E in vitro by stimulating the release of IFN-alpha/beta by 'contaminating' adherent cells. In the presence of anti-IFN-gamma + anti-IFN-alpha/beta antibodies, no Ly-6A/E was induced upon addition of multiple cytokines, including IL-4, or mitogenic doses of anti-Ig antibody. Furthermore, IFN-gamma-containing, CD4+ T cell (Th1) supernatants potently induced Ly-6A/E by murine B cells whereas IL-4-containing (Th2) supernatants were either weak or ineffective; anti-IFN-gamma + anti-IFN-alpha/beta inhibited Ly-6A/E induction by both Th1 and Th2 supernatants. Immunization of mice with Brucella abortus or poly (I).poly (C) resulted in induction of Ly-6A/E expression by virtually all B and T cells, whereas injection of G alpha M delta led to peak induction of Ly-6A/E by approximately 50% of both B and T cells. Lymphocytes from mice infected with the nematode parasites Nippostrongylus brasiliensis or Heligmosomoides polygyrus expressed no Ly-6A/E.(ABSTRACT TRUNCATED AT 250 WORDS)     

实弹射击考核对不同特质焦虑新兵T淋巴细胞亚群及ACTH的影响

目的：研究急性应激对不同特质焦虑新兵的T淋巴细胞亚群及ACTH的影响。方法：状态一特质焦虑问卷（STAI）将新兵分成高、低特质焦虑组各16人。用流式细胞仪及ELISA测定被试基础期（打靶前4d）、应激期（打靶时）、恢复期（打靶后5d）的CD4^＋、CCD8^＋及ACTH浓度。结果：①高特质组3个时期的CD4^＋、CD4^＋／CCD8^＋比值均低于低特质组。CD8＋高于低特质组；高特质组应激期CD4^＋显著下降（P〈0．05）、CD4^＋／CCD8^＋比值与恢复期有显著差异（P〈0．01）．而低特质组仅应激期CD4^＋定量与恢复期有显著差异（P〈0．01）。②实弹射击前、中、后的ACTH浓度，高特质组变化呈极显著差异（P〈0．001），而低特质组应激期与基础期呈显著差异（P〈0．05）。结论：①急性应激影响新兵的CD4^＋、CCD8^＋及其比值，其程度与特质焦虑有关。②应激时高特质组ACTH浓度下降显著大于低特质组，且随应激源消除不易恢复。     

Cutaneous lymphocyte antigen expression on human effector B cells depends on the site and on the nature of antigen encounter

In contrast to T cells, information on skin-homing B cells expressing the cutaneous lymphocyte antigen (CLA) is sparse. CLA expression on human B cells was investigated among circulating immunoglobulin-secreting cells (ISC) and among antigen-specific antibody-secreting cells (ASC) elicited by parenteral, oral or rectal primary immunization, or by parenteral or oral secondary immunization with Salmonella typhi Ty21a. CLA expression was examined by combining cell sorting with an enzyme-linked immunospot assay. Among all ISC, the proportion of CLA(+) cells was 13-21%. Parenteral immunization induced antigen-specific ASC of which 13% were CLA(+), while oral and rectal immunizations were followed by only 1% of CLA(+) ASC (p<0.001). Oral re-immunization was followed by an up-regulation of CLA (34-48%) regardless of the route of priming. Parenteral re-immunization elicited ASC of which 9-14% were CLA(+). In conclusion, the expression of CLA on human effector B cells depends on the site of antigen encounter: intestinal stimulation elicits cells with no CLA, while parenteral encounter elicits significant numbers of CLA(+) cells. Even though primary antigen encounter in the intestine failed to stimulate CLA expression, up-regulation of CLA was found upon intestinal antigen re-encounter. These findings may be of relevance in the pathogenesis of some cutaneous disorders.     

从人外周血B淋巴细胞中应用PCR扩增人抗体基因

本文用常规ＰＣＲ法和半套式ＰＣＲ法，以一组人抗体重链和轻链引物，直接从人外周血淋巴细胞中扩增出抗体重链Ｆｄ基因和轻链基因。一些常规ＰＣＲ法不能直接扩增的人抗体基因，用半套式ＰＣＲ法扩增却得到了阳性结果。扩增的抗体基因的分子量与国内外同类报道一致。本文结果提示，在建立抗体基因文库时，半套式ＰＣＲ法能进一步丰富扩增的抗体基因的多样性