Reinterpretation of the ultrastructure of cartilage matrix.

The epiphyseal cartilage from new-born mouse was treated with collagenase in two ways: either before fixation or after glutaraldehyde fixation. The electron dense granules of the matrix were not seen in the micrographs of cartilage treated with collagenase before fixation. It is concluded that collagen plays a definite role in the formation of the granules at the time of tissue fixation and that the granules are fixation artifacts.     

Biochemical and autoradiographical distribution of hyaluronic acid in calf rib cartilage.

In calf rib cartilage, about one half of total hyaluronate is soluble with guanidinium hydrochloride, the other half only after collagenase treatment. Evidence is presented for its pericellular and intracellular distribution.     

Biochemical and autoradiographical distribution of hyaluronic acid in calf rib cartilage

Summary In calf rib cartilage, about one half of total hyaluronate is soluble with guanidinium hydrochloride, the other half only after collagenase treatment. Evidence is presented for its pericellular and intracellular distribution.Acknowledgment. This work was supported by grants from the Deutsche Forschungsgemeinschaft (Kl 193/10, Mo 183/5).     

Annexin V interactions with collagen

Annexin V was originally identified as a collagen-binding protein called anchorin CII and was isolated from chondrocyte membranes by affinity chromatography on native type II collagen. The binding of annexin V to native collagen type II is stable at physiological ionic strength when annexin V is reconstituted in liposomes. The binding to native collagen types II and X, and to some extent to type I as well, was confirmed using recombinant annexin V. A physiological role for annexin V interactions with extracellular collagen is consistent with the localization of annexin V on the outer cell surface of chondrocytes, microvilli of hypertrophic chondrocytes, fibroblasts and osteoblasts. A breakthrough in our understanding of the function of annexin V was made with the discovery of its calcium channel activity. At least one of several putative functions of annexin V became obvious from studies on matrix vesicles derived from calcifying cartilage. It was found that calcium uptake by matrix vesicles depend on collagen type II and type X binding to annexin V in the vesicles and was lost when collagens were digested with collagenase; calcium influx was reconstituted after adding back native collagen II or V. These findings indicate that annexin V plays a major role in matrix vesicle-initiated cartilage calcification as a collagen-regulated calcium channel.     

Calcium and disease: molecular determinants of calcium crystal deposition diseases

Deposition of basic calcium phosphate (hydroxyapatite, octacalcium phosphate and tricalcium phosphate) (BCP) and crystalline calcium pyrophosphate dihydrate (CPPD) is associated with a variety of aging-related pathologies, including osteoarthritis, cartilage degeneration and pseudogout. These diseases of calcium deposition serve as some of the best-studied examples of how calcium-regulated changes in gene expression can directly lead to pathogenic consequences. Tissue damage can result when crystals stimulate cells to release matrix-degrading molecules or secrete cytokines that stimulate the release of matrix-degrading molecules. Exposure of cultured cells to crystals induces expression of cellular proto-oncogenes such as c-fos, c-myc and c-jun, by a calcium-dependent mechanism, and this response can be blocked by a potential therapeutic compound, phosphocitrate. Activation of the c-fos and c-jun genes is directly involved in expression of metalloproteinases such as collagenase and stromelysin, suggesting that crystal-mediated activation of these genes is directly involved in pathogenesis. In this review recent advances in the molecular mechanisms responsible for crystal-mediated cell activation are discussed.     

Suppression by the cyclohexanetrione Ro 31-0521 of retinoic acid-induced teratogenicity

The cyclohexanetrione Ro 31-0521, which stimulates prostaglandin synthesis, inhibited retinoic acid-induced cartilage degradation in vitro and suppressed the congenital forelimb malformations in rats treated with retinoic acid on day 13 of gestation in a dose-dependent manner.     

The response of costal cartilage to changes in hormonal environment

Résumé 1^o L'hypophysectomie diminue l'incorporation de S³⁵ dans le cartilage costal de jeunes rats. L'hydrocortisone renforce l'action de l'hypophysectomie. Au contraire, le propionate de testostérone stimule la fixation de S³⁵ dans le cartilage costal. Un traitement à l'hormone de croissance donne und réponse qualitativement identique à celle obtenue avec le propionate de testostérone. 2^o Le propionate de testostérone, l'hydrocortisone et l'hormone de croissance n'influencent pas l'incorporation de radiosulfate chez le rat normal.     

In vivo matrix-guided human mesenchymal stem cells

Microfracture of subchondral bone results in intrinsic repair of cartilage defects. Stem or progenitor cells from bone marrow have been proposed to be involved in this regenerative process. Here, we demonstrate for the first time that mesenchymal stem (MS) cells can in fact be recovered from matrix material saturated with cells from bone marrow after microfracture. This also introduces a new technique for MS cell isolation during arthroscopic treatment. MS cells were phenotyped using specific cell surface antibodies. Differentiation of the MS cells into the adipogenic, chondrogenic and osteogenic lineage could be demonstrated by cultivation of MS cells as a monolayer, as micromass bodies or mesenchymal microspheres. This study demonstrates that MS cells can be attracted to a cartilage defect by guidance of a collagenous matrix after perforating subchondral bone. Protocols for application of MS cells in restoration of cartilage tissue include an initial invasive biopsy to obtain the MS cells and time-wasting in vitro proliferation and possibly differentiation of the cells before implantation. The new technique already includes attraction of MS cells to sites of cartilage defects and therefore may overcome the necessity of in vitro proliferation and differentiation of MS cells prior to transplantation. Received 3 November 2005; received after revision 15 December 2005; accepted 4 January 2006     

Platelet heterogeneity and dense tubular system changes on activation

Platelets impregnated with heavy metals appeared as 3 distinct morphological types: 'reticular' cells with a polygonal dense tubular network and stained granules, dark metallophilic cells, and pale metallophobic cells with microvesicles and non-staining granules. On stimulation, type 1 cells decreased while type 3 cells increased, suggesting that with activation dense tubules break up into microvesicles and granules become metallophobic. In the type 2 cells a different functional mechanism may exist.     

Ultrastructural changes in the neural lobe of the rat pituitary induced by reserpine treatment.

The doses of reserpine, which are generally used to deplete amines from the nervous tissue caused marked ultrastructural changes in the neural lobes of reserpine treated rats. The findings suggested depletion of neurosecretory granules, increased lysosomal activity and changes in the pituicytes.     

Noncollagenous, nonproteoglycan macromolecules of cartilage

Extracellular matrix comprises approximately 90% of cartilage, with collagens and proteoglycans making up the bulk of the tissue. In recent years, several abundant cartilage proteins that are neither collagens nor proteoglycans have been characterized in detail. The putative roles of these proteins range from involvement in matrix organization or matrix-cell signaling (PRELP, chondroadherin, cartilage oligomeric protein and cartilage matrix protein) through to molecules that are likely to be involved with modulation of the chondrocyte phenotype (CD-RAP, CDMPs, chondromodulin and pleiotrophin). Other molecules, such as the cartilage-derived C-type lectin and cartilage intermediate layer protein have no role as yet. Due to the difficulties associated with experimentally manipulating a tissue that is 90% extracellular matrix in a manner that can be readily transferred to the whole organism, many of these molecules have been focused on by a surprisingly small number of researchers. This review focuses on newly discovered proteins and glycoproteins in cartilage, with a bias towards those that have structural roles or that are unique to cartilage. Received 7 January 1999; accepted 11 March 1999     

A preliminary report on the fine structure of Tripneustes esculentus eggs.

The fine structure of Tripneustes esculentus eggs was studied with the aid of an electron microscope. Cells obtained from this West Indies sea urchin showed cortical granules, mitochondria forming a rosette around lipid granules, endoplasmic reticula, Golgi apparatus and annulated lamellae. These structures appear identical to those seen in eggs of the Atlantic sea urchin: Arbacia punctulata.     

A simple enzymatic method for isolation of hepatocytes

Summary A new method is described for isolating hepatocytes by an enzymatic procedure: 0.05% bacterial collagenase in calcium-free Hanks' solution is injected into the liver tissue which is removed aseptically. The time required to prepare culture and risk of bacterial and fungal contamination is greatly reduced.     

Effect of bromocriptine on the "prolactin-like cell" of Nectophrynoides occidentalis (Amphibia, viviparous Anura): optical and electronic microscopical studies]

Bromocriptin has an effect on prolactin-like cells of Nectophryno?des occidentalis. 16 days after delivery, these cells are fully active in normal animals. Retention of secretory granules and myelinic formations have been observed in the prolactin-like cells of females treated by CB 154.     

Early skin responses of hibernating and nonhibernating ground squirrels to topical applications of DMBA

Skin patches on hibernating and nonhibernating ground squirrels were treated with multiple topical applications of the carcinogen 7,12-dimethylbenza(a)anthracene. Nonhibernators showed blistering, peeling, drying, hair loss, increased vascularization and hyperpigmentation in proportion to DMBA concentration. The latter was apparently due to a) an increased number of dermal and epidermal melanocytes and b) the appearance of melanocytes with large coarse cytoplasmic granules. Notably, hibernators remained free of gross skin changes and were histologically similar to untreated controls.     

Avantages et inconvénients de la fixation par perfusion du tissu nerveux à la lumière d'une donnée de cytochimie

Summary It has been extensively demonstrated that perfusion is the best fixation procedure for providing good morphological evidence ofstructures, especially in the case of nervous tissue. But it might be questioned if it should also be preferred whencytochemical data have to be obtained or compared.As a preliminary attempt to answer this question, lipoproteic inclusions in nerve cells of vegetative ganglia of adult cat have been considered. These occur in about 25% of cells after immersion in fixative, whereas if perfusion is used, a relation appears between the amount of saline perfused before the fixative fluid and the percentage of cells containing inclusions: this falls to 10% after 300 ml saline, to 1% after 800 ml. In conclusion, though structural artefacts have been avoided and fixation appears excellent, perfusion is significantly responsible for a definite cytochemical alteration.     

Aggrecan, aging and assembly in articular cartilage

The primary function of articular cartilage to act as a self-renewing, low frictional material that can distribute load efficiently at joints is critically dependent upon the composition and organisation of the extracellular matrix. Aggrecan is a major component of the extracellular matrix, forming high molecular weight aggregates necessary for the hydration of cartilage and to meet its weight-bearing mechanical demands. Aggregate assembly is a highly ordered process requiring the formation of a ternary complex between aggrecan, link protein and hyaluronan. There is extensive age-associated heterogeneity in the structure and molecular stoichiometry of these components in adult human articular cartilage, resulting in diverse populations of complexes with a range of stabilities that have implications for cartilage mechanobiology and integrity. Recent findings have demonstrated that aggrecan can form ligands with other matrix proteins. These findings provide new insights into mechanisms for aggregate assembly and functional protein networks in different cartilage compartments with maturation and aging.     

Peroxidase activity of neutrophil granules in two cases of congenital myeloperoxidase deficiency]

The neutrophils and monocytes of two patients with hereditary myeloperoxidase deficiency lacked myeloperoxidase activity as determined by light and electron microscopic cytochemical staining. Using Graham-Karnovsky media, azurophils of neutrophils were devoid of peroxidase whereas all eosinophilic and basophilic granules exhibited normal peroxidase activity. After incubation in alkaline diaminobenzidine media which stains the catalase of microperoxisomes, some small granules were seen to be strongly stained in both immature and mature neutrophils. These small granules were distinct from all other neutrophilic granules which lacked a positive reaction. Only, in the presence of cyanide or aminotriazole, peroxidatic activity was also detected in some ellipsoid azurophils. This observation suggests that these substances activated an oxidase whose nature is discussed.     

Collagen treated with (+)-catechin becomes resistant to the action of mammalian collagenase

Treatment of radioactively labeled guinea-pig skin soluble collagen or calf skin collagen with the flavonoid (+)-catechin makes the collagen resistant to the action of mammalian collagenase but not to the action of bacterial collagenase. Complete resistance to the action of the mammalian enzyme may be achieved by incubating 0.6 mg of collagen (dry weight) with 0.1 mM (+)-catechin, followed by dialysis to remove the unbound flavonoid. Since incubation of the mammalian enzyme with (+)-catechin does not inhibit its activity, it is postulated that (+)-catechin binds tightly to collagen and modifies its structure sufficiently to make it resistant to enzyme degradation.     

Platelet heterogeneity and dense tubular system changes on activation

Summary Platelets impregnated with heavy metals appeared as 3 distinct morphological types: reticular cells with a polygonal dense tubular network and stained granules, dark metallophilic cells, and pale metallophobic cells with microvesicles and non-staining granules. On stimulation, type 1 cells decreased while type 3 cells increased, suggesting that with activation dense tubules break up into microvesicles and granules become metallophobic. In the type 2 cells a different functional mechanism may exist.