Local cell membrane deformations due to receptor-ligand bonding as seen by reflection microscopy.

Understanding surface receptor clustering and redistribution processes at the cell-matrix contact zone requires detailed knowledge of the spatial integration of these molecules in the architecture of this complex interface. Here we present and discuss critically a procedure to extract such information combining reflection contrast microscopy (RCM) and reflection interference microscopy (RIM). As model system, we used living human umbilical vein endothelial cells (HUVEC) adhering to laminin-coated surfaces and investigated the distribution of the alpha2beta1 (CD29/CD49b) integrin at the contact zone of these cells. First, we applied freeze-fracture electron microscopy to gain information on microscopic details of the alpha2beta1 distribution at the contact zone. Next, we visualized and analyzed the overall lateral distribution of the integrins applying RCM using immunogold-labeling with 10 nm labels and a special silver enhancement technique. We found that RCM can be used to determine the lateral position of the marked receptor molecules to an accuracy of about 100-200 nm, instead of large morphological changes at the contact zone during silver enhancement. Finally, we combined RCM with RIM and analyzed the interference pattern of the contact zone around the label positions. Thus, we were able to detect changes of the average shape of the cell membrane due to receptor-ligand bonding of a size down to the resolution of the techniques.

The expression of substance P and its neurokinin-1 receptor mRNA in the bovine corpus luteum of early developmental stage.

Related to all leukocytes, 90% of eosinophils are recruited into the bovine corpus luteum of early developmental stage. We here describe a simultaneous appearance of substance P (SP)-positive fibre-like structures and the neurokinin-1 (NK-1) receptor mRNA for SP. Substance P was depicted by using indirect immunohistology and immunofluorescence localization. The dot blot analysis confirmed the presence of SP at the protein level. Using nested reverse transcribed-polymerase chain reaction (RT-PCR) analysis, a 358 bp long partial bovine receptor mRNA for SP (NK-1) was sequenced in the spinal cord. The mRNA for SP and for the NK-1 receptor were then detected in the corpus luteum of early developmental stage with RT-PCR and nested RT-PCR. We conclude: The production of SP and the expression of NK-1 receptor mRNA may be involved in the selective recruitment of eosinophils into the bovine corpus luteum of early developmental stage.

Leucocyte proliferation in the bovine corpus luteum.

Leucocytes vary in type and number during the lifespan of a corpus luteum. The aim of this study was to determine whether there is an increase in the number of lymphocytes and macrophages as a result of local proliferation. Bovine corpora lutea were classified into stages of development, secretion and regression. A new double immunolabelling method was established for nuclear Ki-67 antigen (a marker for cell proliferation) and for leucocyte surface antigens (detection of CD2-, CD3-, CD4-, CD8-positive lymphocytes and CD14-positive monocytes). Differential cell counting was performed. Between the stages of development and regression there was an increase in the number of T-lymphocytes and macrophages. The percentage of proliferating leucocytes in relation to the total number of proliferating cells was approximately 20% at the stage of advanced secretion and 70% at late regression. The increase in the number of proliferating leucocytes at late regression was due to CD14-positive macrophages. These macrophages migrated from small blood vessels into the septa of corpora lutea at the early stage of regression. Macrophages showed local proliferation in the late stage of regression when capillaries were no longer present. It is concluded that the physiological involution of the corpus luteum is an inflammatory-like condition, which includes local proliferation of monocytes.

Difference in localization of eosinophils and mast cells in the bovine ovary.

The bovine ovary contains a considerable number of leucocytes which can be located with an antibody against the CD18 molecule. In the present study, subtyping and cell counting were carried out on histological sections stained with Sirius red for eosinophils and with toluidine blue for mast cells. The CD18(+) cells were identified immunohistologically. Eosinophils and mast cells contributed considerably to the CD18(+) pool. The number of eosinophils in the corpus luteum increased rapidly in early development to approximately 90% of the CD18(+) cells, and decreased to 30% during secretion and to 10% during regression. Mast cells were not detectable in the follicles, the corpus luteum and the periphery of the cortex, but were observed in the interstitial cortical stroma and the medulla. The number of mast cells in these regions, which corresponded to 60-76% of the CD18(+) cells, did not change significantly throughout the oestrous cycle. It is concluded that eosinophils are selectively recruited at the periovulatory period and that mast cells are unevenly distributed.

[Sex differences in morphologic aging processes]. / Geschlechtsunterschiede morphologischer Alternsprozesse.

According to Max Bürger (2) it is justified to speak of sexual divergence of biomorphosis. Morphometrical investigations have proved the existence of such sexual differences. The following results were obtained: The first example concerns the increase of fatty tissue in parathyroid glands which we measured on histological slides of 135 human samples by means of the point counting method. The fatty tissue of males was shown to increase continuously with statistic significance, whereas this process stops in females during their generative phase between the 2nd and 5th decade. Evidence for the sexual divergence was also obtained by investigating the fatty tissue between the bundles of muscle fibers of the tongue, however, without being able to statistically verify these results. We found the same sexual differences by measuring the muscle cell regression and connective tissue proliferation in the ciliary muscle of human eyes and tunica muscularis of the small intestine as well. All findings prove that the female is able to stay biologically younger during her generative activity compared to males. After menopause the aging processes speed up in female tissues without making up for the "age lead" of males. It is also worth mentioning that macroscopic anthropometric features do confirm the same fact, e.g., length and breadth of human auricles, ranging in age from 18 to 51 years.

Anatomic basis of micturition and urinary continence. Muscle systems in urinary bladder neck during ageing.

Dorschner et al have described the unique function and form of several different muscle systems of the urinary bladder neck. If these systems have different functional responsibilities, then the muscles must undergo different ageing processes, as stated in the theory of function-dependent ageing. One characteristic of histologic ageing is the change over time in the proportion of muscle cells to connective tissue, a phenomenon we have demonstrated in both the ciliary muscle and in the two muscle systems of the small intestine. Using an SIS-Image Analysing System, we have now measured automatically the ratios of muscle cells to connective tissue in sections from several regions of the urinary bladder neck, taken from 50 male and 15 female cadavers. Our results confirm new functional explanations of the different muscle systems in the bladder neck. The relative volume of muscle cells in both the sphincter trigonalis m. and the dilator urethrae m. diminishes continuously with age. In the ejaculatorius m., however, the volume of muscle cells first increases until beginning at the end of the third decade, it decreases until senescence. As was presumed, the proportion of muscle cells in the detrusor vesicae m. does not decline during the later decades. The volume of muscle cells and fibers in both urethral sphincter muscles, however, decreases with age, beginning in early childhood.