THE MIGRATION OF LYMPHOCYTES ACROSS SPECIALIZED VASCULAR ENDOTHELIUM

Lymphocytes were exposed in vitro to either trypsin or neuraminidase. The ability of the treated cells to migrate into tissues was measured (a) by i.v. injection into intact recipients and (b) by vascular perfusion through an isolated lymph-node preparation. The localization of trypsinized cells in the lymph-nodes of recipients was deficient when compared to untreated lymphocytes and there was a surplus of trypsinized cells in the blood. Trypsinized cells migrated into the isolated nodes in reduced numbers. By contrast, neuraminidase treated lymphocytes were markedly deficient in the blood of recipients early after injection; their localization in the spleen and lymph-nodes was also deficient but they were in surplus in the liver. Moreover they migrated into the isolated nodes in slightly increased numbers. By 24 hr after injection the perturbed localization pattern produced by either enzyme was partly restored to normal. In conclusion, trypsin interfered with the capacity of lymphocytes to migrate into lymph-nodes but neuraminidase did not; the latter promoted the hepatic sequestration of cells and the reduced localization in the blood and tissues was a consequence of this. The hypothesis that lymphocytes adhere to specialized endothelia in lymph-nodes because of specific glycoside sequences on their surface lacks experimental support.     

The migration of lymphocytes across specialized vascular endothelium. II. The contrasting consequences of treating lymphocytes with tryspin or neuraminidase.

Lymphocytes were exposed in vitro to either trypsin or neuraminidase. The ability of the treated cells to migrate into tissues were measured (a) by i.v. injection into intact recipients and (b) by vascular perfusion through an isolated lymph-node preparation. The localization of trypsinized cells in the lymph-nodes of recipients was deficient when compared to untreated lymphocytes and there was a surplus of trypsinized cells in blood. Trypsinized cells migrated into the isolated nodes in reduced numbers. By contrast, neuraminidase treated lymphocytes were markedly deficient in the blood of recipients early after injection; their localization in the spleen and lymph-nodes was also deficient but they were in surplus in the liver. Moreover they migrated into the isolated nodes in slightly increased numbers. By 24 hr after injection the perturbed localization pattern produced by either enzyme was partly restored to normal. In conclusion, tryspin interfered with the capacity of lymphocytes to migrate into lymph-nodes but neuraminidase did not; the latter promoted the hepatic sequestration of cells and the reduced localization in the blood and tissues was a consequence of this. The hypothesis that lymphocytes adhere to specialized endothelia in lymph-nodes because of specific glycoside sequences on their surface lacks experimental support.     

The use of trypsin to improve the localization of immunoglobulins in semi-thin frozen sections of the mammary gland

Summary Frozen sections, 0.5 m thick, of the lactating mouse mammary gland have been used to localize immunoglobulins A and G and serum albumin throughout the connective tissue stroma, in the lumina of blood vessels, in milk stored in the alveoli and in the lateral spaces between adjacent epithelial cells. In addition, the immunoglobulins were localized to their specific plasma cells in the connective tissue stroma. Serum albumin was further identified within the mammary epithelial cells as small spots of fluorescence scattered throughout the cytoplasm. The immunoglobulins were not localized within these cells in untreated sections, but in sections treated with trypsin and Soybean trypsin inhibitor, it was possible to identify a similar distribution to that for serum albumin. The spots of fluorescence representing the intracellular localization of the immunoglobulins and serum albumin were frequently found in association with the periphery of intracellular lipid droplets.     

Activation of a procollagenase by low-molecular-weight angiogenesis factor

Avascular tumours have the ability to establish a blood supply for themselves by secreting a humoral factor which stimulates their host's endothelial cells to proliferate and to migrate towards the tumour source. The mechanism of action of such a humoral anglo-genesis factor is more than that of an endothelial-cell growth factor since it requires an oriented migration of cells towards the tumour. We report here the activation of pure skin-fibroblast procollagenase by a low-molecular-weight angiogenesis factor capable of stimulating endothelial-cell growth in vitro. The activation was observed when either Type I or III collagen was used as substrate. It is suggested that at least one function of angiogenesis factor is to promote limited degradation of the connective tissue through which it passes causing channeling in the matrix along which stimulated endothelial cells may     

Immunofluorescent study of the origin of connective tissue cells in xenogenic radiation chimeras under normal conditions and in aseptic inflammation]

The origin of elements of the focus of aseptic inflammation and the normal subcutaneous connective tissue in the xenogenic (mouse-rat) radiation chimaeras was investigated by means of indirect Coons method with antiserum to the rat bone marrow cells. The cells of the imflammation focus (leucocytes, macrophages, fibroblasts or fibroblast-like cells, polynuclear giant cells of foreign bodies), as well as leucocytes, macrophages and some fibroblasts of the normal subcutaneous connective tissue, were shown to take their origin from the transplanted bone marrow cells of the donor.     

The leukocyte podosome

Myeloid leukocytes are the first line of host defence. When they sense perturbations in tissue homeostasis such as infection, inflammation and ischemia, they respond by trafficking. Whilst neutrophils and macrophages migrate to sites of infection, dendritic cells (DC) migrate from tissue-resident sites back into lymph nodes where they activate T and B lymphocytes. The directed migration of these leukocytes through peripheral tissues is thus crucial for their function. This article considers recent advances in our understanding of the adhesive and motile behaviour of macrophages and DC, with particular emphasis on the podosomes that appear to be required for normal migration through extracellular matrices.     

Immunolocalization of 67 kDa elastin-binding protein in perinatal rat lungs

Summary 67 kDa elastin-binding protein (RL-67EBP) has been isolated from neonatal rat lungs by the use of an elastin-coupled affinity column, followed by elution with either lactose or synthetic elastin hexapeptide (VGVAPG), and immunohistochemistry has been used on perinatal rat lungs to determine the tissue localization of this protein. No immunoreactive structures occur in fetal lungs, or in the lungs of day-1 and-4 neonates. On day-7 after birth, immunoreactive cells appear in the subepithelial connective tissue of the intrapulmonary airways, from day-10 on, these cells become evenly distributed in the alveolar parenchyma. Occasionally, some cells occur in the alveolar air space, being free from the surface of the alveolar septum. Unpermeabilized cells obtained by bronchoalveolar lavage, show cell surface immunoreactivity, indicating that RL-67EBP is expressed on the surface membrane of the cells. From these findings, it is suggested that the immunoreactive cells are blood-borne monocytes, and that RL-67EBP may function as an elastin peptide receptor by which monocytes mobilize through interstitial connective tissue during their migration from blood to alveolar air space, where they eventually differentiate into alveolar macrophages.     

Drosophila primordial germ cell migration requires epithelial remodeling of the endoderm

Trans-epithelial migration describes the ability of migrating cells to cross epithelial tissues and occurs during development, infection, inflammation, immune surveillance, wound healing and cancer metastasis. Here we investigate Drosophila primordial germ cells (PGCs), which migrate through the endodermal epithelium. Through live imaging and genetic experimentation we demonstrate that PGCs take advantage of endodermal tissue remodeling to gain access to the gonadal mesoderm and are unable to migrate through intact epithelial tissues. These results are in contrast to the behavior of leukocytes, which actively loosen epithelial junctions to migrate, and raise the possibility that in other contexts in which migrating cells appear to breach tissue barriers, they are actually exploiting existing tissue permeability. Therefore, the use of active invasive programs is not the sole mechanism to infiltrate tissues.     

Prenatal and early postnatal development of the glial cells in the median eminence of the rat

Summary The development of the glial cells of the rat median eminence (ME), including the supraependymal cells, was investigated from embryonic day (ED) 14 through postnatal day (PD) 7, and pituicyte development from ED 12 through ED 17. The anlage of the ME and neurohypophysis shows a neuroepithelial-like structure at ED 12. From ED 13 to 15, the cells of both regions start to differentiate. At the ultrastructural level, only one cell type appears. At the beginning of ED 16, glioblasts of the oligodendrocyte and astrocyte series migrate laterally (from the region of the arcuate nucleus) into the ME. Also at this time the first distinctive structural features appear in the neurohypophysial anlage, the cells of which later develop into pituicytes. Starting at ED 18, tanycytes and astrocytic tanycytes arise in the ME from local glial cells, and somewhat later oligodendroblasts and astroblasts are formed from immigrant glioblasts. Due to their common features, the pituicytes, tanycytes and astrocytic tanycytes apparently represent different forms of the same parent cell type. Microglial and supraependymal cells are first seen at ED 12. Initially, they resemble the prenatal phagocytic connective tissue cells and mature in the fetus into typical electron-dense microglia and macrophage-like supraependymal cells. Both cell types are apparently of mesodermal origin. The microglial elements of the ME probably migrate from the mesenchyma through the basement into the nervous tissue. The intraventricular macrophages of the infundibular region may originate from microglia, epiplexal cells and subarachnoid macrophages.Dedicated to Prof. I. Törö, Budapest, on the occasion of his 80th birthday     

Biophysical studies on the differentiation of human CD14+ monocytes into dendritic cells

Dendritic cells (DCs), which are the most efficient antigen-presenting cells (APCs) currently known, can be derived from CD14⁺ monocytes (DC predecessor cells) in vitro. Immature DCs actively take up antigens and pathogens, generate major histocompatability complex-peptide complexes, and migrate from the sites of antigen acquisition to secondary lymphoid organs to become mature dendritic cells that interact with and stimulate T-lymphocytes. During this process, the cells must undergo deformation to translocate through several barriers, including the basement membrane and interstitial connective tissue in the blood vessel wall. To further understand the mechanisms of the activation of immunological responses and the migration from peripheral tissue to secondary lymphoid organs, we have applied biophysical and microrheological methods to study the development processes of DCs in vitro. The results showed that membrane fluidity, osmotic fragility, membrane viscoelastic properties, infrared spectroscopy, and cytoskeleton organization of DCs exhibit significant differences in different developmental stages. These authors contributed equally to this work.     

The Biomechanical Properties of 3d Extracellular Matrices and Embedded Cells Regulate the Invasiveness of Cancer Cells

The malignancy of tumors depends on the biomechanical properties of cancer cells and their microenvironment, which enable cancer cells to migrate through the connective tissue, transmigrate through basement membranes and endothelial monolayers and form metastases in targeted organs. The current focus of cancer research is still based on biological capabilities such as molecular genetics and gene signaling, but these approaches ignore the mechanical nature of the invasion process of cancer cells. This review will focus on how structural, biochemical and mechanical properties of extracellular matrices (ECMs), and adjacent cells regulate the invasiveness of cancer cells. In addition, it presents how cancer cells create their own microenvironment by restructuring of the ECM and by interaction with stromal cells, which then further contribute to the progression of cancer disease. Finally, this review will point out that mechanical properties are a critical determinant for the efficiency of cancer cell invasion and the progression of cancer which might affect the future development of new cancer treatments.     

Dolichos biflorus agglutinin (DBA) reacts selectively with mast cells in human connective tissues

Dolichos biflorus agglutinin (DBA) binds to N-acetyl-D-galactosamine (GalNAc) residues in glycoconjugates and agglutinates erythrocytes carrying blood group antigen A. In cryostat sections of various tissues from blood group-specified humans, fluorochrome-coupled DBA bound preferentially to fusiform connective tissue cells and to certain epithelial cells. The connective tissue cells were identified as mast cells by their typical metachromasia in consecutive staining with toluidine blue. Double labeling with DBA and conjugated avidin revealed two distinct populations of mast cells. In several tissues the DBA-reactive cells likewise displayed uniform avidin reactivity. In intestinal mucosa, however, morphologically distinct DBA-binding mast cells were found, which were labeled with the avidin conjugates only in specially fixed paraffin sections. DBA did not bind to vascular endothelial cells, which could be identified by double staining with antibodies to factor VIII-related antigen. Labeling with Helix pomatia agglutinin (HPA), another blood group A-reactive lectin, resulted in distinct blood group-dependent fluorescence of the endothelia. Sophora japonica agglutinin (SJA), a blood group B-reactive lectin, labeled vascular endothelial cells in tissues from blood group A, AB, and B donors. HPA and SJA reacted with small mast cells in the gastrointestinal mucosa but failed to label large mast cells in any of the tissues. These results indicate that the blood group reactivity of lectins, as determined by erythroagglutination, is not necessarily consistent with their reactivity with blood group determinants in tissue sections. Moreover, DBA conjugates appear to be a reliable probe for detection of mast cells in various human connective tissues.     

Extraepithelial cells expressing distinct olfactory receptors are associated with axons of sensory cells with the same receptor type

During critical phases of mouse development, axons from olfactory sensory neurons grow out of the nasal neuroepithelium and navigate through the connective mesenchyme tissue towards their targets in the developing telencephalic vesicle. Between embryonic days E11 and E16, populations of cells are located in the mesenchyme which express distinct olfactory receptor genes along with the olfactory marker protein (OMP); thus they express markers characteristic for mature olfactory sensory neurons. These extraepithelial cells are positioned along the axon tracts, and each population expressing a given receptor gene is specifically associated with the axons of those olfactory sensory neurons with the same receptor type. The data suggest that they either might be guide posts for the outgrowing axons or migrate along the axons into the brain.     

Localization of intermediate filament proteins in various structures of the human placenta as revealed by immunoperoxidase technique.

Formalin-fixed, paraffin embedded fragments of normal human placenta were subjected to immunoperoxidase technique with the use of Dako Kits appropriate for detection of vimentin, desmin, and cytokeratins (K 660, 530, and 518 respectively). Distribution of the intermediate filament proteins in syncytiotrophoblast, connective tissue of villous stroma, walls of blood vessels as well as in cells of basal plate and placental islets was compared. General pattern of staining and its localization was characteristic for each of the 3 types of antibodies. In spite of that, most structures observed revealed the presence of more than one type of intermediate filament proteins, with the exception of syncytiotrophoblast containing cytokeratins only. Staining of blood serum and of connective tissue matrix with anti-desmin and anti-cytokeratin antibodies was also observed, whereas fibrinoid remained always negative.     

Lymphocyte-fibroblast adhesion induced by interferon-gamma

Adhesion of lymphocytes to vascular endothelium is thought to be of importance in regulating the passage of lymphocytes from the circulation to areas of inflammation. Evidence suggests the presence of site-specific lymphocyte receptor molecules on the endothelial cell surface which can be modulated by soluble immune factors. The factors responsible for maintaining lymphocyte infiltration at tissue sites are unknown. We have examined the adherence of human peripheral blood T lymphocytes to human fibroblast monolayers in vitro and the role of interferon-gamma in enhancing adherence. Treatment of fibroblasts with interferon-gamma resulted in an increase in the number of adherent T cells in a dose- and time-dependent manner. Enhanced adhesion was noted as early as 4 hr after interferon stimulation (291 +/- 7 T cells/field vs 51 +/- 10 without IFN stimulation) and binding was further increased by lengthening the exposure time of fibroblasts to interferon up to 72 hr (475 +/- 86 T cells/field). Kinetic and inhibition experiments using monoclonal antibody to HLA-DR demonstrated that adhesion of T lymphocytes to interferon-stimulated fibroblasts proceeds by a mechanism independent of DR induction. In addition, adherence was not histocompatibility antigen-restricted, as adherence to autologous and allogeneic fibroblast monolayers was not significantly different. Nonadherent T cells, collected at the end of adhesion assays, were deficient in their capacity to bind to a second interferon-treated monolayer, suggesting the depletion of a subpopulation of T cells responsible for adhesion. Alterations of fibroblasts in vivo by immune cell-derived cytokines may be an important mechanism for the localization of lymphocytes at sites of connective tissue inflammation.     

不同分化阶段的树突状细胞的细胞膜生物物理特性变化

DCs是迄今所知最有效的抗原呈递细胞,在体外可以用CD14 的单核细胞诱导分化而得到.imDCs能够主动地摄取抗原和病原体,产生MHC-抗原肽复合物,并且从抗原获取位点向二级淋巴组织迁移,逐渐分化成mDCs,mDCs与幼稚的T细胞相互作用,从而导致免疫应答或耐受.在这些过程中,DCs必须经历数次变形和转位以通过基底膜和血管壁等屏障,并且在二级淋巴组织内与幼稚的T细胞发生直接的物理性接触.为了更好地理解DCs从外周组织向二级淋巴组织迁移的过程和启动免疫应答的机制,通过研究体外DCs分化过程中细胞膜的生物物理特性,包括细胞膜的粘弹性、表面电荷及其分布和流动性,结果发现DCs细胞膜粘弹性逐渐增加,mDCs的电泳率最大,表面电荷分布出现明显的不对称现象,并且膜流动性也逐渐增加,说明DCs的细胞膜生物物理特性在其行使生理功能的过程中发挥着重要的作用,这对更深入地理解DCs的迁移和与幼稚T细胞相互作用以及免疫应答的启动过程具有非常重要的意义.     

New insights into CD4+ T cell abnormalities in systemic sclerosis

Systemic sclerosis (SSc) is an autoimmune connective tissue disease that is characterized by vasculopathy and excessive deposition of extracellular matrix, which causes fibrosis of the skin and internal organs and eventually leads to multiorgan dysfunction. Studies have shown that CD4⁺ T cell activation is a key factor in the pathogenesis of scleroderma because activated T cells can release various cytokines, resulting in inflammation, microvascular damage and fibrosis. T helper cell 17 (Th17) and regulatory T (Treg) cell activities are a hallmark SSc, as Th17-type cytokines can induce both inflammation and fibrosis. More recently, several studies have reported new T cell subsets, including Th9 and Th22 cells, along with their respective cytokines in the peripheral blood, serum and skin lesions of individuals with SSc. Herein, we review recent data on various CD4⁺ T helper cell subsets in SSc, and discuss potential roles of these cells in promoting inflammation and fibrosis.     

Migrating mast cells in the gallbladder epithelium of cattle and sheep

Summary This paper reports the existence of mast cells in an epithelial location in the gallbladders of both cattle and sheep. The histochemical studies performed on these cells showed that their cytoplasmic granules contain heparin and biogenic amines in both species. Optical- and electron microscopic observations demonstrated that, in both species, mast cells from the connective tissue of the gallbladder diapedese across the basal lamina and migrate through the epithelium all the way to the luminal surface, and that a degranulation process takes place during this migration. The biochemical results showed a correlation between the number of mast cells present in the epithelium and the amount of heparin detected in the different regions of the gallbladders of the species studied. Unusually high contents of heparin were found in both cattle and sheep gallbladders, suggesting that they should be studied as possible commercial sources of this polimer.     

A possible contribution of undifferentiated cells to post-traumatic osteogenesis.

The contribution of undifferentiated cells to callus formation is difficult to observe during fracture healing and is therefore debated. In adult mongrel dogs these cells can easily be observed in a narrow space separating a freshly inserted screw from the surrounding compact bone. Undifferentiated connective tissue cells invade these spaces from the medullary and, less so, from the periosteal site. They seem to resemble histologically and more so ultrastructurally B-type lymphocytes. They migrate to the site of repair or are brought there by the blood stream. Locally these cells proliferate and differentiated into osteoblasts. The authors prefer to call these cells lymphocyte-like cells.