On the role of type IX collagen in the extracellular matrix of cartilage: type IX collagen is localized to intersections of collagen fibrils.

The tissue distribution of type II and type IX collagen in 17-d-old chicken embryo was studied by immunofluorescence using polyclonal antibodies against type II collagen and a peptic fragment of type IX collagen (HMW), respectively. Both proteins were found only in cartilage where they were co-distributed. They occurred uniformly throughout the extracellular matrix, i.e., without distinction between pericellular, territorial, and interterritorial matrices. Tissues that undergo endochondral bone formation contained type IX collagen, whereas periosteal and membranous bones were negative. The thin collagenous fibrils in cartilage consisted of type II collagen as determined by immunoelectron microscopy. Type IX collagen was associated with the fibrils but essentially was restricted to intersections of the fibrils. These observations suggested that type IX collagen contributes to the stabilization of the network of thin fibers of the extracellular matrix of cartilage by interactions of its triple helical domains with several fibrils at or close to their intersections.

Morphological studies on V2 carcinoma invasion and tumor-associated connective tissue changes in the rabbit mesentery.

The mesentery is a duplicature of the peritoneum consisting of loose connective tissue and covered on both sides by mesothelium. Rabbit V2 carcinoma cells implanted i.p. adhere to the mesenteric surface between contracted mesothelial cells. While invasion from these sites sets in, progressive changes of the connective tissue, reflecting fibroblast stimulation, become apparent and comprise multiplication of connective tissue cells, transformation of fibrocytes into fibroblasts, and enhanced production of fibrillar and nonfibrillar constituents of the extracellular matrix. Tumor invasion into this increasingly dense tissue proceeds in 2 ways. (a) Single cells penetrate into and locomote within the interior, where they divide and give rise to nodules which become surrounded by zones of tissue destruction. (b) Proliferation of surface-attached tumor cells results in the formation of nodules which, preceded by zones of tissue damage, extend into the interior. While evidence for lytic effects in the microenvironment of single tumor cells is lacking, degradation of the fibrillar extracellular matrix is regularly found around tumor nodules and indicates a collective lytic action achieved by tumor cells and, possibly, host cells. These morphological findings are discussed in relation to published bio- and histochemical data on spread of the V2 carcinoma.

Interactions between normal epithelial and squamous carcinoma cells in monolayer culture.

Confrontations of rings of adult human oral mucosa epithelial cells enclosing islands of similar normal epithelium, fibroblasts, and cells of three established lines of human squamous carcinoma in monolayer culture were investigated by phase and reflection microscopy and by time lapse cinematography. Measurements of the dimensions of the rings and islands of cells revealed that, while normal epithelial rings confronted with normal epithelium or fibroblasts migrated continuously inwards, similar rings confronting islands of the carcinomas retreated progressively outwards from the tumor islands. The persistence of substantial cell-free space between the epithelium and tumor cells indicated that the outwards migration of the epithelial rings was not solely due to proliferation of the tumor cells. The tumor-induced migration of normal epithelium in monolayer culture may reflect the response of normal epithelium to carcinoma cells in certain in vivo situations.

Use of the avidin-biotin-peroxidase complex (ABC) method for the localization of rabbit cathepsin B in cells and tissues.

Visualization of rabbit cathepsin B was achieved utilizing monospecific sheep antibodies and the avidin-biotin-peroxidase complex (ABC) method. This technique was applied to stain 1) paraffin sections of the liver, 2) fixed fibroblasts from tissue culture, and 3) fixed mesenteries. Cathepsin B was found to be localized within cells of the lining of the liver sinusoids (most probably Kupffer cells), in perinuclear granules of cultured fibroblasts, and within histiocytes of the mesentery. The results demonstrate that the method permits precise and highly sensitive localization of cathepsin B within cells and tissues. Compared to fluorescent staining of cathepsin B, the ABC method has the advantage that routine paraffin sections can be stained, and that all the orthodox histological staining procedures can still be carried out.

The V2 carcinoma of the rabbit as an integrated model of tumor invasion.

The V2 carcinoma, established from skin carcinomas of cottontail rabbits and transplantable in all strains of domestic rabbits, is a paradigm of invasiveness attainable by squamous cell carcinoma. The main mechanisms contributing to this potential are the pressure of incessant cell proliferation, the capacity of the tumor to grow in compact as well as in dissociated formation, the synthesis of proteinases (chiefly cathepsin B and collagenases) by the tumor cells, and the latter's migratory activity. In addition, the V2 carcinoma elicits a large spectrum of host reactions which favor partly the organism, partly the tumor and thus create the complexity of the invasion phenomenon.

The spread of cancer in the organism. Facts and problems.

In this review, cancer is conceived as an alteration of the suface-monitored social behavior of cells. Apart from impaired growth controls, loss of residency (tissue affiliation) is the most important consequence of this homeostatic disorder. It results in local spread (penetration) which is initiated by locomotive and/or desctructive activities of the neoplastic cells. Access of cancer elements to the circulation possibly leads to distant spread (metastasis). Penetration and metastasis largely depend upon reactions of the organism, which are of an ill-understood, ambiguous nature favoring both the tumor and the host.

Penetration of an ascitic reticulum cell sarcoma of the golden hampster into the body wall and through the diaphragm. A scanning electron microscopic (SEM) study.

SEM studies on infiltration of the ascitic form of the hamster reticulum cell sarcoma HaTu 25 into the ventral body wall and through the diaphragm were performed during 6 consecutive days after intraperitoneal transplantation. The findings allow an interpretation of the course of events based on 3 main stages: 1) Contraction of mesothelial cells with partial exposure of the submesothelial stratum. 2) Preferential attachment of tumor cells to these denuded areas. 3) Advance of tumor cells within defects gradually extening from the submesothelial stratum of the musculature. These stages were more pronounced and took a more rapid course at the peritoneal side of the diaphragm than at the body wall. At the pleural side of the diaphragm the appearance of single tumor cells within widened intercellular spaces of the mesothelium was recorded prior to the onset of penetration at the peritoneal surface. The rapid migration of tumor cells through the diaphragm as well as the particularly intensive tumor infiltration into this organ is thought to be connected with the mechanism of intravasation of tumor cells into the lymphatic plexus of the diaphragm. During the whole sequence of events, HaTu 25 cells were found to have maintained their spherical configuration and characteristic surface architecture. Apparently, growth pressure is of minor or no importance in this spacial mode of tumor penetration, rather the action of proteolytic enzymes elaborated by the tumor cells has to be taken into consideration.

In vitro motility of cells from human epidermoid carcinomas. A study by phase-contrast and reflection-contrast cinematography.

The motile behavior of six cell lines derived from human squamous carcinomas (two from the larynx, four from the tongue) was studied by cinematography under phase- and reflection-contrast illumination. The recorded cell activities consist in spreading, stationary and translocation motility, and aggregate formation. Within this common pattern, quantitative modifications ("sub-pattern") are stable properties of the individual cells lines. Such modifications are particularly evident with regard to the dynamic texture of the aggregates which ranges from loose, netlike structures to compact islands with smooth borders. Accordingly, the intensity of cell traffic within and around the aggregates varies considerably. It is discussed to what extent the in vitro motility of the carcinoma cell populations reflects their behavior in the organism and thus the significance of cell movements for invasion.

Release of proteinases by cultures of human cell lines derived from squamous carcinomas of the tongue and larynx.

Seven human cell lines derived from squamous carcinomas of the tongue and larynx were examined for their ability to produce and secrete proteinases. All cell lines were able to release into the culture medium cysteine proteinase and plasminogen activator-like activities. All lines differed from each other in the amount of enzymes secreted, in the kinetics of the secretion, in the quality of the enzymes produced an in the intracellular pool of these activities. These features constitute potential criteria for classifying the biochemical behavior of the tumor cell lines and for explaining their different ability to invade soft tissues and bone.

Cell adhesion: a general appraisal.

Most data on cell adhesion relate to in vitro conditions; for this reason the subject of this review is adhesion of cells to plane inorganic substrata. Adhesion is conceived of as a process requiring energy and comprising distinct steps, most notably the secretion of attachment proteins, the build-up of attachment sites, and the attachment site--induced organization of the cytoskeleton. The grip and stick concept (Rees et al. 1977) is a very adequate interpretation of this course of events. Agents and functions involved in the stages of adhesions are briefly outlined and a few possible extrapolations to cell adhesion in vivo are indicated.

Intermediate-sized filaments in leukemia cells.

Electron microscopic studies on human acute leukemias have shown that leukemic populations contain spherical and polarized cells in various proportions. As recorded by time-lapse cinematography, the two cell configurations represent different functional states: resting cells are completely spherical, locomotive cells are polarized with a conspicuous extension posteriorly. In 9 out of 12 cases of acute myeloid leukemia the two cell configurations were found to coincide with a different pattern of intermediate-sized filaments (ISF). Most spherical myeloblasts possessed large bundles of ISF (a minority had small bundles), whereas polarized myeloblasts showed small groups or single filaments. A similar correlation between cell shape and arrangement of ISF was observed in a transplantable undifferentiated rat leukemia. Two concepts can be distinguished with regard to the role of fibrillar structures in leukemic myeloblasts: thick bundles of ISF either represent a pathological state or have a functional significance. A tentative interpretation of our own results provides some arguments in favor of a disaggregation-reaggregation cycle of thick ISF bundles, whereas a pathological ("end stage") nature of these structures appears less likely.

Comparative morphology of cells located on glass and in the loose connective tissue: a study by scanning electron microscopy.

Most of the studies dealing with cellular shape, surface configuration, and motility are carried out in vitro on plane substrata. During the past years, the direct transfer of results obtained under these conditions to the cellular behavior displayed in the living organism, has been increasingly challenged. For this reason we have investigated the above mentioned functions of different cell classes localized on glass and in the loose connective tissue. The cells utilized were: fibroblasts and macrophages from normal rat and rabbit mesenteries, V2 rabbit carcinoma cells and L5222 rat leukemia cells. The combination of time-lapse cinematography and scanning electron microscopy revealed that motility and surface features are the same, irrespective of the immediate surrounding. Cellular shape and attachment, on the other hand, are dependent on the substrate. Fibroblasts, macrophages and cells of epithelial origin, including carcinoma cells, flatten on glass, but have a rounded configuration in the tissue. The flat leading lamellae displayed during locomotion on glass, are not evident in cells migrating through tissues. What regards attachment devices, extensively studied on glass, their formation and position within a tissue are, at present, a matter of speculation. Although it can be assumed that a similar process is operable in vivo and in vitro, clarification rests upon the use of ultrahistochemical techniques.