Optic neuritis in chronic relapsing experimental allergic encephalomyelitis in Biozzi ABH mice: demyelination and fast axonal transport changes in disease.

The encephalitogenicity of optic nerve tissue was demonstrated in Biozzi ABH (H-2(dq1)) mice. Acute experimental allergic encephalomyelitis (EAE) occurred in 11/14 animals and 4/5 exhibited relapse. The involvement of the optic nerve in spinal cord homogenate induced chronic relapsing EAE (CREAE) was demonstrated by mononuclear cell infiltration and myelin degradation in the optic nerve prior to and during clinical disease. During the relapse phase gross pathological assessment revealed swollen and translucent plaques on the optic nerves. Advanced lesions showed widespread demyelination, astrocytic gliosis and fibrotic changes of the blood vessels. Physiologically, the fast axonal transport of proteins from the retina to the optic nerve and superior colliculus was significantly decreased during relapse. The association of inflammation and demyelination with physiological deficit in the optic nerve highlights the usefulness of this model in the study of multiple sclerosis in which acute monosymptomatic unilateral optic neuritis is a common manifestation. Furthermore, the novel induction of CREAE with optic nerve homogenate suggests that optic neuritis is a common significant role in the pathophysiology and progression of neurological disease in CREAE which may be relevant to studies of optic neuritis in multiple sclerosis.

Serological responses to human papillomavirus type 6 and 16 virus-like particles in patients with cervical neoplastic lesions.

Serum samples from 36 cervical carcinoma patients, 33 patients with high-grade squamous intraepithelial lesions, and 31 cytologically normal women were tested by enzyme-linked immunosorbent assay (ELISA) using human papilloma virus type 6 (HPV 6) and HPV 16 virus-like particles as antigens. Forty serum specimens from 1-year-old children were used to assign cutoff points. When serum samples from the subjects infected with HPV 16 were tested in an HPV 16 ELISA detecting immunoglobulin A (IgA), IgG, and IgM binding, 61% showed IgA, 44% showed IgG, and 39% showed IgM reactivity. Of HPV 6- or 11- or HPV 18-infected subjects. fewer than 17% showed IgA or IgG responses and 33% showed IgM reactivity. In contrast, 13% showed IgA, 10% showed IgG, and 16% showed IgM reactivity in the HPV DNA-negative controls. The results suggest that the IgA and IgG responses are HPV 16 specific and the IgM response is cross-reactive to different HPV types. On the other hand, the serological responses to HPV 6 did not differ in the patient and control groups. The percentages of patients positive for both IgA and IgG antibodies were significantly higher in the groups with high-grade squamous intraepithelial lesions (12% [4 of 33]; P = 0.04) and cancer (17% [6 of 36]; P = 0.02) than in the healty women (0% [0 of 31]), and the percentages for either IgA or IgG were higher for the cancer group (47% [17 of 36]; P = 0.01) than in the normal group (19% [6 of 31]). Most sera positive for IgA and IgG in the patient groups showed higher titers than those in the normal group. All these results suggest that high IgA and IgG responses are good indicators for estimating HPV 16 infection.

PECAM-1: its expression and function as a cell adhesion molecule on hemopoietic and endothelial cells.

PECAM-1, the platelet-endothelial cell adhesion molecule, is expressed on a variety of mature hemopoietic cell types (including neutrophils, monocytes and T cell subsets) and is also present on endothelia. In such cases, this glycoprotein functions as either a homotypic or heterotypic adhesion molecule contributing to cell migration, inflammatory processes and wound healing. We have recently shown that PECAM-1 is expressed on a variety of hemopoietic progenitor cell types and on stromal macrophages from human bone marrow. In this review, we discuss the possible functional significance of this molecule for both hemopoietic cell differentiation and for mature cells.

Synthesis and assembly of virus-like particles of human papillomaviruses type 6 and type 16 in fission yeast Schizosaccharomyces pombe.

We have synthesized capsid proteins of human papillomavirus types 6 (HPV 6) and 16 (HPV 16) in fission yeast Schizosaccharomyces pombe and produced virus-like particles (VLP). The capsid proteins were localized in the nucleus by indirect immunofluorescence and cell fractionation analyses. The VLP were produced in both yeast clones synthesizing L1 alone and L/L2 and purified by sulfato-cellulofine chromatography. Electron microscopic examination showed that these VLP were similar in structure to native HPV particles. Two HPV 16 L1 variants (16 B27L1 and 16 T3L1), isolated from benign cervical samples, produced many more (68- and 14-fold) VLP than the prototype L1 (16 PL1) derived from cervical carcinoma. Coexpression of the HPV 6 L2 protein with 6 L1 and 16 B27L1 proteins increased the production level of the VLP four- and twofold, respectively. The L2 was not detected in the VLP purified with sulfato-cellulofine column, although the L2 was purified in the same fraction containing HPV 6 and 16 B27-VLP by size-fractionation using Sepharose column. Interaction between 6 L2 and 6/16 L1 proteins was not detected by the coimmunoprecipitation assays with either L1 or L2 antibodies. These results suggest that the L2 is not incorporated into the VLP synthesized in yeast.

CD66 identifies the biliary glycoprotein (BGP) adhesion molecule: cloning, expression, and adhesion functions of the BGPc splice variant.

Within the hematopoietic lineage, the monoclonal antibody (MoAb) CD66 reacts with cells of the granulocyte lineage, but not with the majority of progenitor cells from human bone marrow. Our previous studies have shown that CD66 binds specifically to at least three carcinoembryonic antigen (CEA) superfamily members, ie, CEA itself, nonspecific cross-reacting antigen (NCA), and CGM1, but not to CGM6 (NCA-95). In this report, we show that CD66 will also identify the biliary glycoproteins (BGP). A full-length cDNA for the BGPc molecule (a cytoplasmic splice variant of BGPa) was isolated by expression cloning using the CD66 MoAbs. This protein has an identical extracellular and transmembrane sequence to BGPa with one N-terminal IgV like domain, three IgC-like extracellular domains (A1, B1, and A2), plus a transmembrane domain, but the cytoplasmic domain is spliced by 53 nucleotides. Reverse transcriptase-polymerase chain reaction experiments show that this splice variant can be detected in colonic carcinoma cell lines, in primary colonic adenocarcinomas, and in myeloid and B-cell lines to varying degrees. Quantitative analyses of BGPc RNA expression by RNase protection indicate that abundant levels occur only in the colonic, but not in the hematopoietic, cell lines tested. Studies presented here show that BGPc mediates homotypic adhesion and suggest that the cytoplasmic splicing does not alter the initial homotypic adhesion properties of BGPa.

Histological methods for assessing myelin sheaths and axons in human nerve trunks.

Although there are many histological techniques for assessing myelin sheaths and axons in paraffin embedded or frozen sections of the peripheral nervous system, modern approaches usually use plastic embedded material. Although plastic embedding is superior for small cutaneous branches, this method has limited value for histological assessment of nerve trunks. We report three methods which together yield a comprehensive approach for thorough and detailed investigation of human nerve trunks. The rapid osmication method permitted assessment of myelinated nerve fibers from frozen sections at operation, thus providing the surgeon with guidance on the extent of nerve resection. The modification presented here resulted in permanent slides, allowing comparison of results with those of the other two procedures. The new osmium-hematoxylin technique could be performed on paraffin embedded nerves. Paraffin, unlike plastic, permitted the study of the whole cross sectional area of the nerve in single sections. Moreover, the sharp image of the myelin permitted computerized morphometry. The significantly modified axonal silver impregnation technique was performed on frozen sections mounted on glass slides, as opposed to the time-consuming impregnation of free-floating sections. The latter technique had a high success rate and permitted semiquantitative assessment of axons in nerve trunks. These methods can be performed in any routine histology laboratory and resulted in greater accuracy compared to conventional methods.

Bcl-2 protein localizes to the chromosomes of mitotic nuclei and is correlated with the cell cycle in cultured epithelial cell lines.

bcl-2 gene expression confers a survival advantage by preventing cells from entering apoptosis. In contrast to the previously described cytoplasmic localization of Bcl-2 in epithelial cells in vivo, in this study we have demonstrated, in a series of human epithelial cell lines, that Bcl-2 also localizes to mitotic nuclei. Both immunocytochemical and immunoelectron microscopical examinations localize this protein to nuclei and in particular to chromosomes. Nuclear Bcl-2 expression in these cell lines is correlated with the cell cycle. There is relatively strong expression during mitosis, most intense during prophase and metaphase, declining in telophase and then the protein becomes undetectable soon after separation of the two daughter cells. The expression and distribution of Bcl-2 is influenced by treatment with excessive thymidine. These results indicate that Bcl-2 may protect the cells from apoptosis occurring during mitosis and suggest a possible role for the protein in cell immortalization.

Regeneration at the predilective damage sites of nerve trunks in treated leprosy.

Superficially located large and medium sized mixed peripheral limb nerves in active leprosy have previously been shown to have well-recognized fusiform swellings. It is generally agreed that these are the sites of predilective nerve involvement where the severest degeneration and fibrosis occur. A semiquantitative histopathological study on one of these sites, the flexor retinaculum region of the posterior tibial nerve, has been carried out on 14 treated leprosy patients who suffered from total sensory loss to the foot for between 2 and 40 years. The following observations were made: (1) large-scale nerve regeneration was present as characterized by numerous Schwann cells and unmyelinated axons which formed regeneration clusters; (2) thick myelinated axons were either absent or present only in very low numbers; (3) the intraneural fibrosis was usually not severe; (4) the presence of active inflammation probably interfered with nerve regeneration; (5) it appeared that this regeneration started shortly after the onset of therapy and persisted for decades; (6) lepromatous cases were characterized by evenly distributed pathology, whereas borderline tuberculoid cases had an unevenly distributed pathology; (7) the massive nerve regeneration observed was functionally ineffective--these findings indicate that the total nerve damage may affect the more peripheral nerve branches.

An immunoelectron microscopical study of the expression of class II major histocompatibility complex during chronic relapsing experimental allergic encephalomyelitis in Biozzi AB/H mice.

Immunoelectron microscopical techniques have been used to study class II major histocompatibility complex (MHC) expression by cells in the spinal cords of Biozzi AB/H mice with chronic relapsing experimental allergic encephalomyelitis. Throughout the course of disease both astrocytes and endothelia failed to express significant levels of class II MHC antigens. The major central nervous system resident cell types found to express class II MHC antigens were the perivascular microglia, with infiltrating macrophages and some lymphocytes being strongly positive.

Mycobacteria in nerve trunks of long-term treated leprosy patients.

Mycobacteria were present in 4 out of 8 mixed peripheral nerve trunks from patients (3 BT and 1 BL) treated with DDS and/or MDT for periods ranging from 21 months to 8 years. Most of the bacilli appeared to be 'whole'. Nerve destruction with areas of granulomatous infiltration appeared more active than expected. Possible reasons for a continued presence of bacilli in treated nerves and its implications in 'relapse' are discussed.

Expression of vascular addressins and ICAM-1 by endothelial cells in the spinal cord during chronic relapsing experimental allergic encephalomyelitis in the Biozzi AB/H mouse.

The expression of adhesion molecules on central nervous system (CNS) endothelia was examined during chronic relapsing experimental allergic encephalomyelitis (CREAE) in the Biozzi AB/H mouse. Active disease episodes (acute and relapse) were associated with the up-regulation of MALA-2, the murine homologue of intercellular adhesion molecule-1 (ICAM-1), on CNS endothelia and the infiltration of ICAM-1-positive mononuclear cells. In addition, the high endothelial venule (HEV)-associated MECA-325 antigen was evident in perivascular lesions, particularly in relapsing disease. The peripheral lymph node HEV-associated vascular addressin defined by MECA-79 antibody was not detectable in the CNS during CREAE. However, the mucosal HEV addressin was evident in lesions, which ultrastructurally was found to be expressed on the surface of endothelial cells by immunoelectron microscopy. The expression of adhesion molecules, such as ICAM-1, may provide a means by which both the initial neuroantigen-specific and the subsequent antigen-non specific cells extravasate into the CNS. Such infiltration may induce the expression of the vascular addressins which may then provide a means of site-selective cellular recruitment leading to disease progression.

Major histocompatibility complex class II antigen expression in nerves in leprosy; an immunoelectronmicroscopical study.

A technique for immunoelectronmicroscopy has been used to investigate major histocompatibility class II expression in leprosy nerves. In normal nerves, endothelial cells and occasional endoneural cells (not Schwann cells) were constitutively class II positive. In both paucibacillary and multibacillary leprosy nerve biopsies, infiltrating leukocytes were positive but class II-positive Schwann cells were not seen. These observations indicate that Schwann cells may not be involved in presenting Mycobacterium leprae antigens to T cells in leprosy. This conflicts with evidence from in vitro studies, but may be explained by the fact that in vivo Schwann cells are surrounded by basement membranes and are closely associated with axons.

Denatured muscle grafts for nerve repair. An experimental model of nerve damage in leprosy.

About 20% of patients with leprosy develop localised granulomatous lesions in peripheral nerves. We report experiments in guinea-pigs in which freeze-thawed autogenous muscle grafts were used for the treatment of such mycobacterial granulomas. Granulomas were induced in guinea-pig tibial nerves and the animals were left for 7 to 100 days in order to assess maximal damage. The local area of nerve damage was then excised and the gap filled with denatured muscle grafts. Clinical assessment after periods up to 150 days showed good sensory and motor recovery which correlated well with the histological findings. The muscle graft technique may be of value for the treatment of chronic nerve lesions in selected cases of leprosy.

Induction of chronic relapsing experimental allergic encephalomyelitis in Biozzi mice.

Experimental allergic encephalomyelitis (EAE) was induced in Biozzi AB/H (antibody high) mice by sensitization with spinal cord homogenate in adjuvant. Biozzi AB/H mice were highly susceptible to EAE induction and followed a chronic relapsing pattern of disease. Disease episodes were characterized by mononuclear infiltration of the central nervous system, with demyelination being particularly evident in relapse. The cellular infiltrates, which were associated with immunoglobulin deposition, consisted of macrophages and primarily CD4-positive T lymphocytes. However, similarly treated Biozzi AB/L (antibody low) mice were markedly less susceptible to EAE induction than AB/H mice. Thus, Biozzi mice should prove valuable for the study of chronic relapsing EAE.

The repair of large peripheral nerves using skeletal muscle autografts: a comparison with cable grafts in the sheep femoral nerve.

Recovery of morphological characteristics was compared in the femoral nerves of sheep at varying times up to 10 months after nerve repair. Groups of sheep receiving coaxially aligned freeze-thawed skeletal muscle autografts were compared with those receiving three-strand cable grafts made from autogenous cutaneous nerve. At all times after implantation more nerve fibres could be counted distal to the muscle grafts than distal to cable grafts. Indices of nerve fibre maturation were indistinguishable between the two groups at 10 months. The results are discussed in relation to the possible use of the technique for repairing large mixed nerves.

An immunoelectronmicroscopical study of the expression of major histocompatibility complex (MHC) class II antigens in guinea pig sciatic nerves following induction of intraneural mycobacterial granulomas.

A guinea pig model of nerve damage in leprosy has been used to investigate the expression of major histocompatibility complex (MHC) class II antigens in granulomatous lesions in nerves. Using an immunoelectronmicroscopical technique, infiltrating mononuclear cells and endoneural fibroblast-like cells are shown to be class II-positive in the experimental neural lesions. Schwann cells are not class II-positive under these conditions, although at the light microscope level Schwann cell-like cells appear to be positively stained. This illustrates the value of immunoelectronmicroscopy in the investigation of cell surface proteins in situ as compared with conventional light immunohistochemistry.