Splenic adherent cells, stimulated in vitro, induce the reactive formation of lymphoid follicles and germinal centres in draining lymph nodes after subcutaneous transfusion into syngeneic mice

The reactive formation of lymphoid follicles and germinal centres in lymph nodes, induced by subcutaneous transfer of in vitro activated splenic adherent cells into syngeneic mice, were studied. Adherent cells were obtained by incubating spleen cell suspensions for 24 h and activated by incubating for 1 h in the medium containing keyhole limpet haemocyanin (KLH) absorbed onto alumina. Some of the treated adherent cells were irradiated with 10 Gy x-rays, while others were either not stimulated or were stimulated with alumina-KLH but killed by repeated freezing and thawing. Examination of adherent cell smears immunostained with antibodies against, F4/80, Mac-1, Mac-2 and NLDC-145 indicated that many adherent cells displayed macrophage markers but few displayed the interdigitating cell marker. Animals transfused with KLH-treated adherent cells with or without irradiation showed a marked increase in the number of lymphoid follicles and germinal centres in draining lymph nodes, whereas those transfused with adherent cells which had not been KLH-treated or which had been killed after KLH treatment displayed no significant change in the number of follicles. These results were interpreted as indicating that following transfusion, antigen-activated adherent macrophages migrated into the draining lymph nodes and induced the reactive formation of lymphoid follicles and germinal centres outside preexisting follicles.     

Structural and functional changes to lymph nodes in ageing mice

Lymph nodes (LN) are secondary lymphoid organs spread throughout the lymphatic system. They function to filter pathogenic material from the lymphatic fluid to maintain the health of the organism. Subcapsular sinus macrophages (SCSM) are among the first‐responders within the LN due to their strategic location within the subcapsular sinus region. These macrophages aid the delivery of immune complexes to B cells and follicular dendritic cells (FDC) within the LN. Here we show an increase in SCSM and other macrophage populations within aged LN. However, immune complex uptake by macrophages within LN was not altered with age, nor was immune complex uptake by B cells. LN stromal cell populations, important in immune responses and the localization and survival of leucocytes, were altered in their representation and distribution in aged LN. In particular, FDC regions were decreased in size and had decreased chemokine CXCL13 expression. Furthermore, the retention of immune complexes by FDC was decreased in aged LN at 24 hr post‐injection. As FDC are important in the maintenance of germinal centre responses, the decreased retention of immune complex in aged LN may contribute to the reduced germinal centre responses observed in aged mice.     

Formation of lymph follicles and germinal centers in the somatic and mesenteric lymph nodes of growing mice during ontogenesis

We investigated the age-dependent changes that occur in the numbers of lymph follicles and germinal centers in various lymph nodes in BALB/C and ICR mice aged between four days and 16 to 18 weeks. Young adult BALB/C mice have a relatively small body size, compared to ICR mice at the same stage, where there is a relatively large body size. In BALB/C mice somatic (popliteal, brachial, axillary, inguinal, submandibular and deep cervical) and mesenteric lymph nodes were examined. In ICR mice only the somatic (popliteal, brachial and axillary) lymph nodes were examined. In both BALB/C and ICR mice, the primary follicles were apparent in most somatic nodes by the 6th postnatal day. Up to 28 days of age, the number of follicles per node increased, reaching different levels in nodes from different locations. Thereafter, in most of the somatic nodes in BALB/C mice the number of follicles increased only slightly, although there was a substantial increase in ICR mice, reaching a peak or a plateau at 8 or 12 weeks of age. In the mesenteric (ileocecal) nodes in BALB/C mice, the primary follicles first appeared at 10 to 12 days, then there was a linear increase until a plateau level was reached at 8 weeks of age. Germinal centers appeared in the mesenteric nodes at 28 days and increased rapidly in number thereafter. In most somatic nodes germinal centers were scarcely observable until 8 weeks of age. Based on our observations we have three suggestions. Firstly, in BALB/C mice there were two different patterns of age-dependent changes in the numbers of lymph follicles in the somatic and the mesenteric nodes during ontogenesis. These different patterns are probably due to variations in the magnitude of the exogenous antigen stimulatory effect. Secondly, it seems likely that the variations in the numbers of lymph follicles that are produced in somatic nodes at different locations during the first 28 days after birth relate to the dimensions of the body regions that are drained by that particular somatic node at that stage of development. Thirdly, in the relatively small BALB/C mice, the ontogenetic production of lymph follicles in a somatic node is mostly completed during the first four weeks of life, whereas in the relatively larger ICR mice, this process may continue until the young adult stage of 8 weeks.     

小鼠PP、MLN和ILN中CD3~+、CD19~+细胞组成差异及四君子汤总多糖对其影响

对比研究口服四君子汤总多糖(SJTPS)对免疫抑制小鼠派氏集合淋巴结(PP)、肠系膜淋巴结(MLN)和腹股沟淋巴结(ILN)中淋巴细胞作用的差异。观察连续7 d口服1 g/kg.d SJTPS对100 mg/kg的环磷酰胺处理24 h后的小鼠PP、MLN、ILN中CD3+、CD19+细胞比例的变化。正常小鼠体内,PP中的CD19+淋巴细胞比例明显高于ILN和MLN中的CD19+淋巴细胞比例,而CD3+细胞的比例明显低于后两者,腹腔注射环磷酰胺后,小鼠PP、MLN、ILN中的CD3+细胞比例上升,CD19+细胞比例下降,口服SJTPS可以明显的对抗PP,MLN中的CD3+,CD19+细胞比例的变化,但对ILN中的CD3+、CD19+细胞比例变化的作用不显著。口服SJTPS对肠道黏膜免疫系统的作用强于对其他外周免疫系统的作用。     

FCA和FICA可增强淋巴滤泡捕捉抗原的能力

目的　探讨向体内注入非抗原类物质能否引起反应性淋巴滤泡形成 ,观察体内注入FCA(完全弗式佐剂 )、FICA(不完全弗式佐剂 )对滤泡树突状细胞 (FDC)捕捉抗原的能力有无影响。方法　将FCA、FI CA分别注入小鼠足底 ,数日后取出腘淋巴结 ,应用PNA B法及PAP免疫组化法染色 ,采取三维重塑方法 ,统计淋巴滤泡、FDC细胞群数量。结果　注入FCA、FICA后第 5日出现初级淋巴滤泡 ,第 3 5日数量达到高峰 ,之后逐渐减退。同时 ,滤泡树突状细胞 (FDC)与B细胞聚集同步出现。结论　动物体内注入有丝分裂物质FCA、FICA引起引流淋巴结内次级淋巴滤泡形成 ,初级滤泡形成过程中B细胞聚集与FDC形成是同步发生的     

PrP protein is associated with follicular dendritic cells of spleens and lymph nodes in uninfected and scrapie-infected mice.

Abnormal forms of a host protein, PrP, accumulate in the central nervous system in scrapie-affected animals. Here, PrP protein was detected immunocytochemically in tissue sections of spleen, lymph node, Peyer's patches, thymus, and pancreas from uninfected mice and from mice infected with a range of mouse-passaged scrapie strains and bovine spongiform encephalopathy (BSE). In the spleen, lymph node and Peyer's patches, PrP-positive cells were identified as follicular dendritic cells (FDC) by their location, appearance, and immune complex trapping function, whereas in the thymus they appeared to be two types of stromal cells: interdigitating cells (IDC) and cortical epithelial cells. In pancreas, PrP-containing cells were confined to the islets of Langerhans. Although the distribution of PrP immunolabelling was the same in tissues from scrapie-affected and uninfected mice, there was evidence that PrP accumulated in abnormal forms in FDC of infected mice. If, as is likely, PrP is essential for agent replication, our results suggest that FDC are the site of scrapie and BSE replication in the spleen and lymph node.     

Age- and gender-specific changes of hypocretin immunopositive neurons in C57Bl/6 mice

Loss of neurons or neuronal functions over time has been hypothesized to contribute to the dysregulation of autonomic functions observed in aging. In this study, we evaluated the total number of the hypothalamic hypocretin (orexin) immunopositive neurons in 100, 400, 800 and 1000-day-old male and female C57Bl/6 mice that are commonly used in aging studies in vertebrates. Males had 15–20% more hypocretin immunopositive neurons (HIN) than females at all ages examined. Neuronal number for both sexes was stable in the first 400 days of life, but started declining between 400 and 800 days with rates of approximately 1 neuron/day. The rate of loss doubled in males between 800 and 1000 days of age. The total average number of HIN for males was 2251 ± 139 at 100 days, 2235 ± 112 at 400 days, 1914 ± 81 at 800 days, and 1596 ± 301 at 1000 days. The total average number of HIN for females was 1805 ± 76 at 100 days, 1887 ± 118 at 400 days, and 1521 ± 181 at 800 days. Evaluation of the time-dependent decline in the number of hypocretin immunopositive neurons may help to explain the physiological changes in sleep or energy homeostasis regulation during aging.     

Protection of germinal centres from complement attack: Decay-accelerating factor (DAF) is a constitutive protein on follicular dendritic cells. A study in reactive and neoplastic follicles

The development of B-cell memory is linked to the presence of germinal centres. This process is dependent on the presence of antigen, usually in the form of immune complexes with antibody, on the surface of the follicular dendritic cells (FDCs) that form a network in the germinal centre. The presence of immune complexes poses a constant danger of activating complement. Decay accelerating factor (DAF, CD55) and the membrane attack complex (MAC) inhibitor (CD59) are two cell proteins whose sole function is to protect cells from the action of complement, the former affecting the earlier components of the complement cascade, and the latter the terminal ones; both are bound to the cell surface via a glycosylphosphatidylinositol link. DAF but not CD59 could be demonstrated on FDCs. DAF is also present on the FDCs in follicular lymphomas despite the absence of complement (C3) in neoplastic follicles. This indicates that DAF is constitutive to FDCs but does not preclude the possibility that its expression is increased when immune complexes are deposited.     

The superantigen-induced polarization of T cells in rat peripheral lymph nodes is influenced by genetic polymorphisms in the IL-4 and IL-6 gene clusters

In recent years, it has become clear that the polarization of T cells depends on the genetic background. However, due to the complexity of the genetic background of each animal, a direct comparison of the phenotype is difficult. In this study, a new rat strain LEW.BN-4-10 carrying the chromosomal regions on chromosomes 4 and 10, which harbor IL-6 and IL-4 gene clusters of BN, has been bred on the genetic background of LEW. It was asked whether these two gene clusters influence the polarization of T cell responses. As a model, the Mycoplasma arthritidis mitogen (MAM)-induced inflammation was used focusing on the microenvironment of the draining lymph node (LN). The effect of differences in these regions was tested by comparing LEW.BN-4-10 and LEW rats under steady-state conditions and upon injection of MAM into the forepaw. Under steady-state conditions, the two strains showed differences in the dendritic cell (DC) subset composition. When MAM was injected, the number of T cells in LEW.BN-4-10 rats producing T(h)2 cytokines such as IL-4 and IL-13 was significantly increased compared with LEW. The data suggest that these differences in the microenvironments in LN of LEW and LEW.BN-4-10 rats resulted in different susceptibility to the disease (increase of cells in LN and paw swelling). In addition, deviations in the distribution and function of injected effector T cells were found in the LN of LEW and LEW.BN-4-10 rats after MAM treatment. The data indicate that the IL-6 and IL-4 gene clusters are involved in polarizing T cell responses in vivo.     

HSP70 is selectively over-expressed in the blast cells of the germinal centres and paracortex in reactive lymph nodes

AIMS: We evaluated by immunohistochemistry HSP70 expression in reactive lymph nodes since its morphological expression and location have not been previously described and correlated with lymphocyte kinetics. METHODS AND RESULTS: Ninety-six cases of non-specific lymphadenitis were immunostained for HSP70, CD20, CD3, Ki67, Bcl-2, CD21. The type and the location of HSP70-positive cells were determined. Their number out of 2000 cells in each germinal centre and in each paracortical area was counted at 60x magnification with the help of a quantitative grid. Seventeen percent of germinal centre cells and 7.6% of the paracortex cells were positive. This difference was highly significant. The positively reacting cells were B-cells and had a blast (centroblast or immunoblast) morphology, with negative mantle and marginal lymphocytes and T-cells. Lymphoplasmacytoid cells and plasma cells reacted only weakly or were negative. Germinal centre antigen-presenting cells and interdigitating dendritic cells reacted from lightly to moderately. CONCLUSIONS: HSP70 was selectively over-expressed by B-blasts mainly located within germinal centres with a lower number in the paracortex. The difference in the mean number between the two sites was statistically highly significant. No correlation was found with bcl-2 and Ki67 expression. Mantle, marginal and T-lymphocytes were always negative. The biological meaning and role of this over-expression in centroblasts and immunoblasts remain to be elucidated.     

香菇多糖口服和注射给药对DTH小鼠耳后淋巴结和小肠派氏结中T淋巴细胞的影响

目的:通过比较口服和注射香菇多糖对DTH小鼠耳后淋巴结和小肠派氏结中T淋巴细胞的影响,探讨肠道粘膜免疫系统在口服药物发挥免疫调节作用中的重要性。方法:28只6～8周龄NIH小鼠随机分为对照组、环磷酰胺免疫抑制模型组、香菇多糖灌胃组、香菇多糖注射组,各组小鼠用2,4-二硝基氟苯背部致敏,4天后耳廓攻击。攻击30小时后,对比各组小鼠耳肿胀差异,并应用流式细胞术检测被攻击耳廓的耳后淋巴结和小肠派氏结中T淋巴细胞的比例变化和活化水平。结果:与环磷酰胺免疫抑制模型组比较,香菇多糖能有效增强小鼠DTH反应,降低耳后淋巴结和小肠派氏结的T细胞比例,提高T细胞活化水平,且口服组的作用明显优于注射组。结论:对比注射给药,口服香菇多糖可以更有效地调节肠道粘膜免疫系统和整体免疫系统功能,提示肠道粘膜免疫系统在口服药物发挥免疫调节作用中扮演着重要的角色。     

Sensitivity of immunocompetent cells of DBA/2 and C57Bl/6 mice to cyclophosphamide

T cells were most sensitive to cyclophosphamide in DBA/2 mice, while in C57Bl/6 mice both T and B cells were sensitive. The formation of antibody-producing cells and the production of specific antibodies were delayed in DBA/2 mice immunized after pretreatment with antitumor drug. Accumulation of antibody-producing cells in the spleen was more active in immunized C57Bl/6 mice treated with cyclophosphamide compared to animals not treated with cyclophosphamide.     

Determination of oxidation phenotype in inbred C57Bl/6 and BALB/C mice

Research Laboratory of Pharmacogenetics, aand Research Laboratory of Pharmacokinetics, Institute of Pharmacology, Academy of Medical Sciences of the USSR, Moscow. (Presented by Academician of the Academy of Medical Sciences of the USSR A. V. Val'dman.) Translated from Byulleten' Éksperimental'noi Biologii i Meditsiny, Vol. 110, No. 11, pp. 491–493, November, 1990.     

Morphofunctional characteristic of mast cells in BALB/c and C57Bl/6 mice during cold exposure

Mast cells of the mesentery and subcutaneous tissue in BALB/c and C57Bl/6 mice were studied after single and repeated cold exposure (−20°C, 3 min). Immediate adaptive reactions of mast cells in BALB/c and C57Bl/6 mice did not differ after single cold exposure and were manifested in increased degranulation. Repeated cold exposure of BALB/c mice was followed by an adaptive reaction, which included an increase in the count of mast cells in subcutaneous tissue and normalization of the degranulation index. In C57Bl/6 mice the count of mast cells in subcutaneous tissue decreased, while the degranulation index remained high. These changes reflect the disadaptive response of mast cells to repeated cold exposure. Translated fromByulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 138, No. 8, pp. 207–209, August, 2004     

Variants of secondary immune response in CBA and C57Bl/6 mice

The manifestation of a secondary immune response to intraperitoneal and subcutaneous injection of sheep red cells in various doses was studied in CBA and C57Bl/6 mice. The parameters under study were the delayed-type hypersensitivity reaction and antibody production assessed from the levels of antibody-producing cells of classes M and G in the lymph node and spleen. The manifestation and correlations between delayed-type hypersensitivity and antibody production were found to depend on the route of antigen administration and its first immunizing dose, interval between the two immunizations, and genetic control of the immune response. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 118, N ^o 11, pp. 486–488, November, 1994 Presented by K. P. Kashkin, Member of the Russian Academy of Medical Sciences     

Antioxidant enzymes and lipid peroxidation in C57Bl/6 and BALB/c mice

Superoxide dismutase and catalase activities and levels of thiobarbituric acid-reactive lipid peroxidation (LPO) products were estimated in the liver of C57B1/6 and BALB/c mice. The results indicate that although antioxidant enzymes are more active in BALB/c mice, compensation of oxidation processes in this strain is possible only if LPO-inducing agents are absent or present at low levels, and that these agents, including exogenous ones, may be expected to activate lipid oxidation in this strain to a greater extent than in C57Bl/6 mice. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 120, N ^o 12, pp. 580–583, December, 1995     

Effect of ultraviolet B irradiation on immobilization stress-induced changes in the protective systems of C57Bl/6 mice

We studied the effect of ultraviolet B irradiation on superoxide dismutase activity, ceruloplasmin level in the plasma, and steroid hormone concentration in the adrenal glands of C57Bl/6 mice subjected to immobilization stress. Ultraviolet B irradiation did not abolish the increase in superoxide dismutase activity, but decreased ceruloplasmin level in the plasma and corticosteroid concentration in the adrenal glands of mice exposed to immobilization stress. __________ Translated from Byulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 141, No. 2, pp. 180–183, February, 2006     

Airways infection with virulent Mycobacterium tuberculosis delays the influx of dendritic cells and the expression of costimulatory molecules in mediastinal lymph nodes

Despite tuberculosis resurgence and extensive dendritic cell (DC) research, there are no in vivo studies evaluating DC within regional lymphoid tissue during airways infection with virulent Mycobacterium tuberculosis (Mtb) H37Rv. Using DC-specific antibodies, immunocytochemistry, flow cytometry and Ziehl-Neelsen (ZN) for bacilli staining, we searched for Mtb and DC changes within mediastinal lymph nodes, after intratracheal (ITT) inoculation of virulent Mtb. ZN and immunocytochemistry in frozen and paraffin sections of mediastinal lymph nodes identified Mtb until day 14 after ITT inoculation, associated with CD11c(+) and Dec205(+) DC. Analysing CD11c, MHC-CII, and Dec205 combinations by flow cytometry in MLN suspensions revealed that CD11c(+)/MHC-CII(+) and CD11c(+)/Dec205(+) DC did not increase until day 14, peaked on day 21, and sharply declined by day 28. No changes were seen in control, saline-inoculated animals. The costimulatory molecules evaluated in CD11c(+) DCs followed a similar trend; the CD80 increase was negligible, slightly surpassed by CD40. CD86 increased earlier and the three markers peaked at day 21, declining by day 28. While antigen-specific proliferation was not evident for MLN CD4(+) T cells at 2 weeks postinfection, delayed-type hypersensitivity responses upon ITT inoculation revealed that, as early as day 3 and 7, both the priming and peripheral systemic immune responses were clearly established, persisting until days 14-21. While airways infection with virulent Mtb triggers an early, systemic peripheral response maintained for three weeks, this seems dissociated from regional events within mediastinal lymph nodes, such as antigen-specific T-cell reactivity and a delay in the influx and local activation of DC.     

Dendritic cell subsets in lymph nodes are characterized by the specific draining area and influence the phenotype and fate of primed T cells

Dendritic cells (DC) are important in differential T-cell priming. Little is known about the local priming by DC in the microenvironment of different lymph nodes and about the fate of the imprinted T cells. Therefore, freshly isolated rat DC from mesenteric lymph nodes (mLN) and axillary lymph nodes (axLN) were phenotyped and cultured with blood T cells in the presence of the superantigen Mycoplasma arthritidis mitogen (MAM). The phenotype, proliferation and apoptosis of the primed T cells were analysed. Our data show that a common DC population exists in both mLN and axLN. In addition, region-specific DC with an organotypical marker expression imprinted by the drained area were found. Coculture of T cells with DC from mLN or axLN resulted in a distinct shift in the CD4 and CD8 expression of T cells and their phenotype. Furthermore, when these differentially primed mLN and axLN T cells were injected into recipients, mLN-primed T cells survived longer in other lymphoid organs. The results show that the region-specific DC have a unique phenotype and an impact on the ratio of CD4 : CD8 T cells during an immune response in vivo.     

Multidimensional assessment of differences in monoamine metabolism in C57Bl/6 and BALB/c mice

Monoamine metabolism in the hypothalamus and striatum of BALB/c and C57Bl/6 mice (intact and stressed in the open field test) was studied using single-and multidimensional statistical methods. It is suggested that the revealed difference in neurotransmitter metabolism is associated with genetically controlled behavior of these animals under conditions of emotional stress. The results of discriminant analysis suggest that the regulation of monoamine metabolism during emotional stress is genetically determined. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 129, No. 5, pp. 574–577, May, 2000