Mechanism of immune interferon production in vitro: interaction between immune interferon-producing cells and antigenic cells.

Various processes of in vitro immune interferon production by sensitized spleen cells stimulated with allogeneic cells were investigated. When L cells, an interferon-inducing antigen, were fixed with methyl alcohol or paraformaldehyde, the ability to induce immune interferon disappeared. In this immune interferon production system, the majority of sensitized spleen cells adhered to target cells within 1 h of cocultivation. Adherence of immune interferon-producing cells to target cells was observed only when L cell-sensitized spleen cells were cocultured with L cells or with mouse embryo cells derived from C3H mice. Fixation of antigenic cells with methyl alcohol or paraformaldehyde significantly reduced cell adherence. When L cells alone or sensitized spleen cells alone were pretreated separately with cytochalasin D, neither cell type could bind to partner cells. Specific adherence did not take place at 4 degrees C, nor in the presence of dinitrophenol or sodium azide. Continuous protein synthesis in both cells was not required for immune cell adherence. Divalent cations, Ca2+ or Mg2+, were required for this immune specific adherence to take place. However, once stable adherence was established, treatment with cytochalasin D, ethylenediaminetetraacetic acid, or sodium azide, or simple reduction of temperature, did not disrupt the binding. Interaction between immune interferon-producing cells and antigenic cells can be subdivided into two phases according to the requirement for divalent cations: (i) lymphocytes and antigenic cells interact transiently, and divalent cations are required to maintain the binding; (ii) lymphocytes and antigenic cells form a stable interaction, and deprivation of divalent cations does not disrupt the binding. Colchicine showed an inhibitory effect in the period after cell-to-cell adherence. Colchicine did not inhibit the release of interferon. On the other hand, vinblastine, another antimicrotubule agent inhibited the secretion of immune interferon. Since interferon synthesis was not stopped immediately after addition of cycloheximide, continued protein synthesis of sensitized spleen cells was not required for interferon secretion. The present study showed that adherence of immune interferon-producing cells to antigenic cells was a complex phenomenon involving a series of successive events.     

Electron microscopic evidence of a viral nature for osteoclast inclusions in Paget's disease of bone

Circumstantial evidence from electron microscopic and immunological studies support the view that Paget's disease of bone represents a slow virus infection. However, there is only limited information available regarding its electron microscopic, enzyme and immunocytochemical characteristics. Two cases were studied using electron microscopy with particular emphasis on the inclusions in osteoclasts. Detailed ultrastructural and cytochemical studies including immuno-electron microscopy were performed. Some osteoclasts demonstrated specific virus-like structures composed of aggregations of microtubules in the nucleus and cytoplasm. The structures were easily digested by trypsin or protease, and were sensitive to RNase, which provided substantial evidence of a proteinaceous nature and inclusion of ribonucleic acid. Immunocytochemical examination identified binding of anti-respiratory syncytial virus and anti-measles virus antibodies in the tissue obtained from one of the two cases examined. The presence of viral antigens in structures in the cytoplasm of Pagetic osteoclasts supports the theory of paramyxovirus involvement in this disease.     

Immunohistochemical and electron microscopic observations of stromal cells in the human oviduct mucosa

Stromal cells in the lamina propria of the human oviduct mucosa are unique cells that can differentiate into decidual cells during ectopic pregnancy in the oviduct. The nature of stromal cells is still unknown. In the present study, we investigated human oviductal stromal cells with transmission electron microscopy and immunohistochemistry and revealed that they had ultrastructural features similar to myofibroblasts and expressed alpha-smooth muscle actin, a marker used to identify myofibroblasts. Primary cilia were also one of the characteristic profiles of the stromal cells. These findings showed that the connective tissue-stromal cells in the human oviduct mucosa are myofibroblasts. They are considered to play an important role in the transport of oocytes by bringing about contraction of the mucosal folds.     

Gene-inducing program of human dendritic cells in response to BCG cell-wall skeleton (CWS), which reflects adjuvancy required for tumor immunotherapy

Adjuvants induce the expression of a number of genes in dendritic cells (DCs), which facilitate effective antigen-presentation and cytokine/chemokine liberation. It has been accepted that the toll-like receptor (TLR) family governs the adjuvant activity in DCs. An adjuvant with a long history is mycobacteria in an oil-in-water emulsion, namely Freund's complete adjuvant. Since the active center for the adjuvancy in mycobacteria is the cell-wall skeleton (CWS), we used the bacillus Calmette-Guerin cell-wall skeleton (BCG-CWS) to test DC maturation by GeneChip analysis. We identified the genes supporting an efficient DC response and output. Approximately 2000 genes were up-regulated by BCG-CWS stimulation. BCG-CWS-, peptidoglycan (PGN)- and lipopolysaccharide (LPS)-stimulation generally up-regulated some gene clusters including genes for inflammatory cytokines (TNF, IL1alpha, IL1beta, IL6, IL12 p40, IL23 p19, etc.), chemokines (CCL20, IL8, etc.), cell adhesion molecules (ICAM-1, etc.), apoptosis-related proteins (GADD45B, BCL2A1, etc.), metabolic enzymes (PTGS2, SOD2, etc.) and miscellaneous proteins (EHD1, TNFAIP6, etc.). LPS-stimulation, but not BCG-CWS- or PGN-stimulation, up-regulated the interferon-inducible antiviral proteins, including IFIT1, IFIT2, IFIT4, CXCL10, ISG15, OASL, IFITM1 and MX1. We also found that the BCG-CWS- or PGN-stimulation up-regulated CXCL5, MMP1, etc. We discussed their properties in association with TLRs and recently discovered TLR adapters.     

Experimental study on secondary amyloidosis associated with chronic arthritis

Various organs from rabbits immunized with heat-killed E. coli 0:14 to induce chronic polyarthritis resembling rheumatoid arthritis (RA) were examined for amyloid deposition since amyloidosis is a frequent feature of long-standing RA. Amyloids were confirmed mainly by polarizing as well as nonpolarizing light microscopy after Congo red staining and partly by electron microscopy and immunoperoxidase technique. Amyloid deposits after an additional 2 months from the first appearance of arthritis were most frequently found in the spleen and kidneys suggesting secondary amyloidosis. Amyloidosis was observed in 36% of 75 arthritic rabbits. The incidence of amyloidosis was significantly higher in the arthritic rabbits than in the non-arthritic rabbits (p less than 0.05). This suggests that prolonged sensitization with heat-killed E. coli 0:14 containing endotoxic substance participates in the formation of both arthritis and amyloidosis.     

Founder effect in spinal and bulbar muscular atrophy (SBMA)

We analyzed the polymorphic (CAG)n and (GGC)n repeats of the androgen receptor gene in 113 unrelated X-linked spinal and bulbar muscular atrophy (SBMA) X chromosomes and 173 control X chromosomes in Japanese males. The control chromosomes had an average CAG repeat number of 21 +/- 3 with a range from 14-32 repeat units, and SBMA chromosomes had a range from 40-55 with a median of 47 +/- 3 copies. The control chromosomes had seven different alleles of the (GGC)n repeat with the range of 11 to 17; the most frequent size of (GGC)n was 16 (79%), while (GGC)17 was very rare (1%). However, in SBMA chromosomes only two alleles were seen; the most frequent size of (GGC)n was 16 (61%) followed by 17 (39%). (GGC)n size distribution was significantly different between SBMA and control chromosomes (P < 0.0001), indicating the presence of linkage disequilibrium. There was no allelic association between the (CAG)n and (GGC)n microsatellites among control subjects as well as SBMA patients, which suggests that a founder effect makes a more significant contribution to generation of Japanese SBMA chromosomes than new mutations.      

Possible role of autoantibodies against nephrin in an experimental model of chronic graft-versus-host disease

Nephrin, a product of the NPHS1 gene, is a component of the slit diaphragms that are found between glomerular foot processes and is a crucial element for glomerular filtration barrier. Recently, nephrin has been focused in a number of studies of proteinuria development including various types of acquired glomerular diseases including minimal change nephrotic syndrome and membranous nephropathy. However, the precise role of nephrin in such acquired glomerular diseases is still unknown. To analyse the role of nephrin further, two kinds of anti-nephrin antibodies were raised in the rabbits and applied to an experimental mouse model of chronic graft-versus-host disease, in which (C57BL/10 x DBA/2) F1 mice developed clinically apparent severe proteinuria with significant glomerular lesions 7 weeks after parental DBA/2 cell transfer. Antibody-sandwich ELISA detected anti-nephrin antibodies during week 2 to week 6, with the peak at week 2 or week 4. Colocalization of nephrin and IgG on week 4, week 6, and week 8 was revealed by confocal microscopic analysis, suggesting that in situ immune complex formation with nephrin in glomerular lesion. Taken together, it seems to be suggested nephrin and its autoantibody have a certain role in the development of glomerular lesion in our model mice.     

Development of the rat meninx: experimental study using bromodeoxyuridine

The development of the rat meninx from the viewpoint of cell proliferation was studied microscopically and immunohistochemically using bromodeoxyuridine (BUdR). A compact cell layer around the neural tube, the meninx primitiva, was observed in 12- and 13-day fetuses. A reticular structure resembling the subarachnoid space appeared in the 14-day fetus. The ectomeninx, consisting of a collagen fiber layer, part of which became the dura mater, appeared in 15-day fetuses, allowing discrimination of the endomeninx, the arachnoid cell layer. The primordium of the choroid plexus also appeared in the lateral ventricle on the same day. Bone appeared in the primitive dura mater, and stratification of the meninx was almost complete in 21-day fetuses. BUdR-positive cells were confirmed in the meninx from day 12 of gestation to day 15 postpartum. The number of BUdR-positive cells was greatest in fetuses aged about 12 or 13 days, reaching nearly 50%, but decreased gradually toward the neonatal period. The findings of this study suggest that, after the migration of neural crest cells, marked cell proliferation in the meninx begins. Differentiation into various layers then follows and is almost complete before birth, whereas the proliferation of arachnoid cells continues even in the early neonatal period.