Optimized Ki-67 staining in murine cells: a tool to determine cell proliferation.

The reliable analysis of the cell cycle status has become increasingly relevant for scientific and clinical work, especially for the determination of tumor cell growth. One established method to characterize the proliferation activity of cells is the analysis of the Ki-67 protein. Ki-67 is expressed in the nucleus during the whole cell cycle except for the G0 phase. Several different protocols exist for the examination of the Ki-67 protein in tissue and cell culture, but most of them are defined for human cells. For the analysis of the Ki-67 protein in murine tissue and cell culture there is a variety of protocols existing which recommend different fixation and permeabilization reagents or special kits. In this study, we established a reliable protocol for Ki-67 staining in murine cells and tissue based on PFA fixation, which can be used not only for flow cytometry but also for immunofluorescence microscopy analysis. We tested our protocol successfully with three different Ki-67 anti-mouse antibodies in cell culture, regenerating liver tissue and mouse melanoma tumor to demonstrate the general applicability.

Arteriovenous fistula presenting as a tumor of the upper eyelid.

We report, to our knowledge for the first time, a case of arteriovenous fistula presenting as a tumor of the upper eyelid in an 80-year-old patient. Surgical excision through an anterior approach brought excellent cosmetic and functional results.

Beta 2-adrenergic receptors on peripheral blood mononuclear cells in patients with rheumatic diseases.

In order to investigate the influence of the autonomic nervous system on the immune response in rheumatic diseases such as rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE), we determined the number and the dissociation constants of beta 2-adrenergic receptors (beta 2R) on peripheral blood mononuclear cells (PBMC) in patients with RA characterized by either low or high disease activity, patients with SLE, and healthy age-matched controls by means of a radioligand binding assay with [125I]iodocyanopindolol. The number of beta 2R was significantly decreased in all three patient groups as compared with the healthy controls. In SLE patients, a significant negative correlation between beta 2R and the anti-ds-DNA antibody titre was observed (r = -0.57, P < 0.01). These data demonstrate the close correlation between the number of beta 2R on PBMC and the extent of disease activity in patients with RA and SLE and suggest an involvement of the autonomic nervous system in the pathogenesis of rheumatic disorders.

Decreased density of beta-adrenergic receptors on peripheral blood mononuclear cells in patients with rheumatoid arthritis.

There is growing evidence that the autonomic nervous system influences the immune response by activation and modulation of beta 2-adrenergic receptors (beta 2R) on immunocompetent cells. However, little is known about its pathogenetic role in autoimmune disease. Therefore, the number and the dissociation constants (beta 2R-KKD) of beta 2R on peripheral blood mononuclear cells (PBMNC) were determined in 21 patients with rheumatoid arthritis (RA) and 9 healthy age and sex matched controls [healthy donors (HD)] by a radioligand binding assay with [125I]iodocyanopindolol. The density of beta 2R (x +/- SEM = 2,120 +/- 103 binding sites/cell) and the beta 2R-KD (x +/- SEM = 6.8 +/- 0.8 pM) were significantly decreased in patients with RA compared to HD (number of beta 2R: 3,960 +/- 372, beta 2R-KD: 16.7 +/- 3.6, p less than 0.01). In RA, the number of beta 2R was significantly lower in patients with high systemic disease activity (n = 11, number of beta 2R: 1,850 +/- 134) than in patients with low inflammatory activity (n = 10, number of beta 2R: 2,418 +/- 95, p less than 0.004). In addition, there were significant negative correlations between the beta 2R density and an arbitrary systemic activity score (r = -0.74, p less than 0.001), the Ritchie index (r = 0.77, p less than 0.001), and various variables of disease activity (r = 0.59 to -0.78, p less than 0.005), respectively. Our data demonstrate the close interplay between the systemic inflammatory activity and the beta 2R characteristics in patients with RA. Our results provide further evidence that the immune response may be influenced by the sympathetic nervous system.