Value of electron microscopy in kidney biopsy diagnosis.

Kidney biopsy reports given during 2003 were collected from the authors' pathology database. A total of 111 biopsies were performed. Five tumor samples were not studied with electron microscopy (EM). Of the remaining 106 biopsies, 85 were studied with EM. EM was not performed in 10/24 transplant biopsies, or in 11/82 cases of suspected primary kidney disease. The role of EM was evaluated by grouping the samples in 3 categories: (1) EM was essential for diagnosis, (2) EM contributed to the interpretation and cleared uncertainties, and (3) EM had no influence on the diagnostic process. In transplant biopsies EM influenced the final diagnosis in 86% of cases (category 2). In biopsies performed for primary kidney disease EM was essential for diagnosis in 18.3% clearly contributed in 53.5%, and had no influence on the final diagnosis in 28.2% of cases. The study suggests that the importance of EM has not decreased during the last few years. Because only about 25% of the EM reports did not have any influence on the diagnostic process, it is recommended that kidney biopsy protocols should include EM in all biopsy cases, or at least tissue should be reserved for EM studies of all cases. Because of the influence of EM on the diagnostic process the need for EM in pathology training should be emphasized.

Modulation of peripheral blood mononuclear cell activation status during Salmonella-triggered reactive arthritis.

OBJECTIVE: To determine the activation status of mononuclear cells in the peripheral circulation during the acute phase and the recovery phase of Salmonella-triggered reactive arthritis (ReA). METHODS: Peripheral blood mononuclear cells (PBMC) were obtained from 8 patients with Salmonella infection (4 with ReA and 4 without) and were studied by reverse transcription-polymerase chain reaction for messenger RNA (mRNA) of proinflammatory and antiinflammatory cytokines, by flow cytometry (FC) for cell surface activation and adhesion molecules, by immunofluorescence (IF) microscopy for bacterial antigens, and by FC, IF, and DNA fragmentation on gel for signs of apoptosis. RESULTS: During the acute phase of the infection, PBMC were activated in all patients, as characterized by high levels of expression of CD14, CD11b, and CD11c on monocytes. In the patients with ReA, PBMC also had the capacity to produce interleukin-1beta (IL-1beta), IL-6, IL-8, IL-10, and tumor necrosis factor alpha. During the amelioration of disease, monocyte activation was decreased in all patients. A complete down-regulation of CD14 was detected only in the patients with ReA, whereas the expression of CD14 in the patients without ReA was positive and was similar to that in healthy controls. In addition, cytokine mRNA levels decreased regardless of the presence of Salmonella antigens in blood cells in all 4 patients with ReA. CONCLUSION: High levels of expression of some activation and adhesion molecules and elevated levels of mRNA for certain cytokines that are predominantly produced by monocytes were found in PBMC from patients with acute Salmonella-triggered ReA, which suggests that these cells are activated. On the other hand, complete down-regulation of CD14 and a marked decrease in the cytokine production capacity during amelioration of the disease suggest that suppression of PBMC activity might be involved in recovery from ReA.

Enterobacterial infection modulates major histocompatibility complex class I expression on mononuclear cells.

Major histocompatibility complex (MHC) class I expression is reduced in several viral infections, but it is not known whether the same happens during infections caused by intracellular enterobacteria. In this study, the expression of MHC class I antigens on peripheral blood mononuclear cells (PBMC) from 16 patients with Salmonella, Yersinia, or Klebsiella infection was investigated. During or after the acute infection, the expression of MHC class I antigens was markedly decreased in eight patients, all with genotype HLA-B27, and six out of eight with reactive arthritis (ReA). A significant decrease of monomorphic MHC class I was found in three patients, of HLA-B27 in eight (P<0.05) and of HLA-A2 in two. However, patients negative for the HLA-B27 genotype, or healthy HLA-B27-positive individuals, did not have a significant decrease of MHC class I antigens. During the decreased expression on the cell surface, intracellular retention of MHC class I antigens was observed, whereas HLA-B27 mRNA levels did not vary significantly. This is the first evidence that enterobacterial infection may down-regulate expression of MHC class I molecules in vivo and that down-regulation is predominant in patients with the HLA-B27 genotype.

Monocytes that have ingested Yersinia enterocolitica serotype O:3 acquire enhanced capacity to bind to nonstimulated vascular endothelial cells via P-selectin.

Reactive arthritis is usually a self-limiting polyarthritis which develops after certain gastrointestinal or urogenital infections. Microbial antigens found in the inflamed joints are thought to play a key role in the development of this disease. It is not known how antigens of the pathogenic organisms migrate from the mucosal tissues into the joints. The data presented here show that mononuclear phagocytes which mediate the dissemination of several intracellular pathogens acquire an enhanced capacity to bind to nonstimulated vascular endothelial cells after phagocytosis of Yersinia enterocolitica O:3, one of the causative organisms of reactive arthritis. The increased binding to previously nonstimulated endothelial cells was mediated by P-selectin, whose translocation to the endothelial cell surface was induced by monocytes with intracellular Yersinia bacteria. These results suggest that mononuclear phagocytes may be responsible for the dissemination of bacterial antigens and the initiation of the joint inflammation in reactive arthritis.

Infectious agents as triggers of reactive arthritis.

Reactive arthritis was originally defined as a sterile joint inflammation after infection elsewhere in the body, but this view has been challenged in the past decade since different antigens and DNA and RNA of various triggering microbes have been shown to exist at the sites of inflammation in the joints. It has been suggested that microbial antigens, or intact pathogens, are important for the pathogenesis of reactive arthritis, at least in the early phase of the disease, but the exact mechanism of how the pathogens contribute to the development of this usually self-limiting polyarthritis has not been discovered. This article reviews the theories on the role of infectious agents as triggers of reactive arthritis.

Yersinia enterocolitica serotype O:3 alters the expression of serologic HLA-B27 epitopes on human monocytes.

The expression of serologic HLA-B27 epitopes on leukocytes of patients with reactive arthritis or ankylosing spondylitis has been shown to be modified in the course of the disease. The purpose of this work was to study whether phagocytosis of arthritis-triggering microbes in vitro alters the expression of HLA-B27 molecules on human antigen-presenting cells and to characterize the underlying mechanisms. Human monocytes and HLA-B27- or HLA-A2-transfected human U-937 cells were exposed to Yersinia enterocolitica serotype O:3. The expression of different epitopes of HLA-B27 was monitored by using immunofluorescence, and their synthesis was determined by quantitative immunoprecipitation. Our results show that phagocytosis of Y. enterocolitica serotype O:3 changed the expression of serological HLA-B27 epitopes. This was due to the reduced synthesis of HLA-B27 molecules. The expression of especially the epitopes which depend on the presence of peptides in the antigen-binding groove was changed. The expression of the ME1 epitope, which has been shown to be important for T-cell recognition in patients with reactive arthritis, was decreased. Down-regulation of epitopes important for the T-cell recognition may impair the elimination of arthritis-triggering microbes and lead to persistent infection. In addition, Y. enterocolitica serotype O:3 seemed to alter the repertoire of peptides presented by the HLA-B27 molecules on human monocytes. This may have a role in the pathogenesis of reactive arthritis via an autoimmune mechanism.

False-negative serological HLA-B27 typing results may be due to altered antigenic epitopes and can be detected by polymerase chain reaction.

Serological typing with the microlymphocytotoxicity test (MLCT) and flow cytometry (FC) using HLA-B27 antisera is commonly used for the determination of HLA-B27. However, in some patients tested more than once, negative results have turned out to be positive at following investigations. We retested by polymerase chain reaction (PCR) samples from 20 randomly selected patients with reactive arthritis or Reiter's syndrome who had now been followed for 20 yr. Ten of the patients were originally tested to be HLA-B27 positive and 10 HLA-B27 negative by the MLCT. All 10 serologically HLA-B27 positive individuals were also positive in the PCR. However, 2/10 patients interpreted as being HLA-B27 negative were positive by PCR. At this time, the same two patients were also positive in the routine MLCT and FC using four different monoclonal antibodies against HLA-B27. PCR is superior to serological techniques to determine HLA-B27 positivity unequivocally, since it is based on the detection of HLA-B27 gene sequences.

Expression of MHC class II molecules on human monocytes is regulated independently from each other after phagocytosis of bacteria.

Reactive arthritis is usually self-limiting polyarthritis which develops in HLA-B27 positive individuals after certain gastrointestinal or urogenital infections. The pathogenesis of reactive arthritis is unknown but T cells seem to have a crucial role. Most of the antigen-specific T cells isolated from the synovial fluid have been MHC class II restricted. The role of antigen presentation in the pathogenesis of reactive arthritis has been studied relatively little. In this work the authors studied the effect of arthritis-triggering bacterium (Yersinia enterocolitica O:3) on the expression of MHC class II molecules on human monocytes and found that the expression of different MHC class II molecules was regulated independently from each other in half of the individuals after certain incubation periods. In these cases the expression of HLA-DP was parallel to the expression of HLA-DQ, while HLA-DR expression went to the opposite direction or did not change at all. No difference between HLA-B27 negative and HLA-B27 positive healthy individuals was seen. The authors conclude that independent regulation of the expression of different MHC class II antigens on antigen-presenting cells is a more common phenomenon than usually thought.

Yersinia lipopolysaccharide is modified by human monocytes.

Reactive arthritis is usually self-limiting polyarthritis, which develops after certain gastrointestinal or urogenital tract infections, mostly in susceptible HLA B27-positive individuals. In the pathogenesis of this arthritis, it is probably important that structures of the causative bacteria are found in the affected joints. The structure found in the synovial fluid phagocytes of the patients with reactive arthritis after Yersinia, Salmonella, and Shigella infections has always been lipopolysaccharide (LPS) of the causative bacteria. It has been in a highly processed form but still immunoreactive. To follow the degradation process of LPS, we fed peripheral blood monocytes of healthy blood donors with heat-killed Yersinia enterocolitica O:3 bacteria in vitro and monitored the fate of LPS by immunofluorescence and immunoblotting methods. Heat-killed bacteria were used since Y. enterocolitica O:3 bacteria are able to live inside monocytes in vitro and dividing intracellular bacteria would have made it impossible to monitor the degradation process of LPS with these methods. Both the core region and the O-polysaccharide chain of LPS persisted in cytoplasmic vacuoles and on plasma membrane of monocytes through the 7-day follow-up time. Migration properties of processed LPS in sodium dodecyl sulfate-polyacrylamide gel electrophoresis suggested structural modifications of LPS. We also demonstrated that core epitopes appearing on the surface of Yersinia-fed monocytes on day 4 of incubation were processed intracellularly, suggesting that LPS-containing phagocytes are a constant source of membrane-active LPS in their microenvironment as well as in the joints of arthritic patients.

Intracellular pathogens and professional phagocytes in reactive arthritis.

Reactive arthritis is a postinfectious complication which develops after certain infections, mostly gastrointestinal or urogenital. Antigenic structures of the causative microbes, but no live organisms, have been demonstrated in inflamed joints. The host factors as well as the microbial antigens responsible for the initiation of the arthritic process are unknown. The pathogenesis of reactive arthritis is discussed here with special reference to the intracellular life of the causative microbes and to monocytes/macrophages, which may be involved in early events of the arthritic process as well as in maintenance of the autoimmune type of responses.

Nurse cells of the bursa of Fabricius: do they exist?

Cell-cell interactions in B lymphocyte development have so far been incompletely characterized, mostly due to lack of a special organ for B cell maturation in the mammalian species. Certain well-known lymphostromal interactions in the thymus have raised the question whether similar interactions with nurse cells would also operate in the development of B cells. We have tested this hypothesis in the chicken bursa of Fabricius, an organ specific for the B cell maturation. To identify possible nurse cells, with viable lymphocytes enclosed, the cells in the bursa of Fabricius were dispersed with collagenase and trypsin. Light and electron microscopic examination of bursa cell suspensions showed four types of aggregates, identified by low magnification light microscopy as potential nurse cell-like complexes. Electron microscopy revealed that all aggregates consisted of epithelial cells, and complexes of epithelial cells with lymphocytes enclosed were not observed. These findings indicate that interactions similar to those seen in the avian and mammalian thymus between epithelial nurse cells and T lymphocytes are not a part of the avian B cell differentiation process.