Inge L. Eestermans

Depletion of macrophages and disappearance of postcapillary high endothelial venules in lymph nodes deprived of afferent lymphatic vessels.

SummaryThe afferent lymphatic vessels of rat popliteal lymph nodes were interrupted, and the histological alterations in the lymph nodes occurring 1 to 14 weeks after operation were studied. One week after operation the number of macrophages was considerably reduced and continued to decrease during the subsequent time periods studied. A 6 weeks most macrophages had disappeared. Simultaneously the immunological activity diminished and had completely disappeared 8 weeks after operation. Three weeks after operation the endothelial cells of the postcapillary high endothelial venules had flattened, and the number of immigrating lymphocytes was greatly reduced. Subsequently the lymph nodes became depleted of both macrophages and lymphocytes, leaving only the reticuloendothelial framework.

Ontogeny of milky spots in the human greater omentum: An immunochemical study

Milky spots in the human greater omentum are preformed specific accumulations of primarily macrophages within the stroma of the greater omentum. To obtain a better understanding of milky spots in the human greater omentum, the development and the earliest forms of milky spots in the human greater omentum were studied, with special attention to the macrophage population.

Phosphatase cytochemistry with cerium as trapping agent

SummaryLead is prevalently replaced by cerium as trapping agent in phosphatase cytochemistry to prevent nonspecific precipitation. Recently, substrate specific but artefactual lcad precipitates have been described in the nuclear envelope (NE) and rough endoplasmic reticulum (RER) due to a local matrix effect. In the present study a verification was carried out of the localization of acid phosphatase and glucose-6-phosphatase in the NE and RER of rat peritoneal macrophages and hepatocytes respectively with cerium. It appeared that precipitates of cerium phosphate in NE and RER of peritoneal macrophages do not represent sites of acid phosphatase activity but are due to the matrix effect. However, in rat hepatocytes these organelles demonstrate true reactive sites for glucose-6-phosphatase.

Novel isolation and purification method permitting functional cytotoxicity studies of macrophages from milky spots in the greater omentum

Milky spots in the greater omentum are well organized perivascular infiltrates of leukocytes which are probably involved in the clearance of tumor cells from the peritoneal cavity. In milky spots, macrophages are the predominant cell type forming a distinct population of cells. To investigate whether these macrophages have a function in the control of metastatic spread in the peritoneal cavity, a novel isolation and purification method was developed in order to study the functional cytotoxicity of macrophages from milky spots in the greater omentum against tumor cells in vitro. In order to obtain a cell suspension, greater omenta of unstimulated healthy male WAG/RIJ rats were incubated in collagenase/DNase suspension and filtered. Subsequently, macrophages were isolated and purified using flow cytometry by sorting unstained cells on the basis of size and internal complexity. Macrophages and other cells were identified by routine May-Grünwald-Giemsa staining and by immunophenotyping with the specific macrophage monoclonal antibody ED 1. Furthermore, macrophage subtypes were characterized by ultrastructural analysis. Functional cytotoxicity of the isolated macrophages was assayed against the syngeneic CC 531 tumor cell line in a colorimetric MTT assay. From three greater omenta of healthy rats 1.16 +/- 0.16 x 10(6) macrophages were isolated with a purity of 83 +/- 2% and a viability of > or = 96%. The macrophages were of the exudate (monocytic), exudate-resident and resident cell type and were in equal proportions. The contaminating cells were mainly mesothelial. A maximum cytotoxicity of approximately 30% was reached with the macrophage fraction at an effector-to-target ratio of 10. Furthermore, it was established that the mesothelial cells did not exhibit cytotoxicity.

Electron dense granules and the role of buffers: artefacts from fixation with glutaraldehyde and osmium tetroxide

Electron dense granules may appear in tissues after glutaraldehyde prefixation and osmium tetroxide postfixation. In order to determine the conditions under which the granules are formed various vehicles in pre‐ and post‐fixatives were tested on lymph node, thymus and heart.

An ultrastructural study of dopamine-immunoreactive nerve fibres in milky spots of the human greater omentum

An ultrastructural study was performed to examine the presence and distribution of dopamine-immunoreactive nerve fibres in milky spots of the human greater omentum. Non-myelinated nerve fibres were located perivascularly as well as throughout the milky spots. Dopamine immunoreactivity was demonstrated in the nerve fibres and in a portion of the macrophage population. These results demonstrate a discrepancy between human and non-human milky spots.

FALSE LOCALIZATION OF ACID-PHOSPHATASE ACTIVITY IN THE NUCLEAR-ENVELOPE AND ENDOPLASMIC-RETICULUM OF RAT PERITONEAL-MACROPHAGES

Acid phosphatase cytochemistry using lead salt methods was performed on rat peritoneal macrophages obtained by the intraperitoneal injection of dextran five days previously. Lead precipitate was present in the nuclear envelope, the rough endoplasmic reticulum, Golgi apparatus and lysosomes in about 50% of these cells. The formation of reaction product appeared to be substrate-specific and was sensitive to sodium fluoride in all these sites. However, only in the nuclear envelope, the rough endoplasmic reticulum and Golgi apparatus could lead salt precipitation be prevented by (a) omission of the washing procedure following the incubation step, (b) postincubation in a medium containing sodium fluoride, or (c) washing in buffer containing lead salt. It is concluded that precipitation of lead salt does not prove the presence of acid phosphatase activity in these organelles. The formation of precipitate in these sites is probably due to a local matrix effect, facilitated by the persistence of acid phosphatase activity in the lysosomes and a suboptimal trapping efficiency of phosphate ions during the washing procedure which follows in the incubation step.