Human nasal septal cartilage: analysis of intracellular enzyme activities, glycogen content, cell density and clonal proliferation of septal chondrocytes of healthy adults and acromegalic patients.

The human septal cartilage is of ectodermal origin and contributes to midfacial growth and development. Acromegaly is an endocrine disease due to growth hormone (Gh) excess originating from a somatotrophic adenoma of the pituitary gland. Excessive Gh levels lead to high insulin-like growth factor I (IGF I) concentrations, which are known to stimulate cartilage growth in vivo and in vitro. One of the salient clinical pictures is coarsening of the midface and enlargement of the septal cartilage. Septal cartilage was obtained from 8 acromegalic patients during transnasal hypophysectomy and from 10 healthy adults during septoplasty to analyse the following aspects of cartilage biochemistry, metabolism and growth. 1. Intracellular glycogen, the major source of energy of chondrocytes, was determined enzymatically and found to be drastically reduced in acromegaly. 2. Several intracellular enzymes, related to biomatrix degradation, showed a strict local pattern of distribution. Cathepsin B activity, a neutral proteinase degrading both the helical and nonhelical region of the collagen molecule was significantly increased in acromegaly, whereas alkaline phosphatase activity, an enzyme related to mineralization of the cartilage at the chondroosseous junction was depressed in acromegaly. 3. The cell density in some areas of the septal cartilage was increased in acromegaly, whereas the clonal proliferation rate of its chondrocytes in response to serum and growth factors was decreased. Chondrocytes both of healthy adults and acromegalic patients could be effectively stimulated by insulin-like growth factor I and II and to a lesser extent by epidermal growth factor.

[Organization of the stroma in the bony nasal septum and its homologues in representative skulls]. / Organizacja zrebu kostnego przegrody nosa i jej homologów u przedstawicieli czaszkowców.

In literature the notion and systematization of osseous nasal septum complex in man and other mammals are not fully and precisely verified. The examinations of the processes of ossification in ++bony cranium and nasal septum were being made in birds, fishes, reptiles and mammals. The skulls were macerated and the specimens were stained by use of alizarin. The radio- and photography were used for documentation. The results proved the opinion that all the osseous elements of nasal septum in mammals derived from the interorbital septum and axially palatal complex in fishes, reptiles and birds.

[Concentration of trace elements in the inferior turbinates and in the nasal septum of the human]. / Konzentration der Spurenelemente in der unteren Nasenmuschel und im Septum des Menschen.

The trace elements in 47 inferior turbinates (concha nasalis inferior) and in the nasal septum were examined by means of synchrotron radiation, spectroscopy of characteristic x-rays, and "sub-micron elemental mapping with the Oxford scanning proton microprobe", and the results compared. Both first methods displayed an age-dependent reduction of Zn and an increase of Pb concentration. In some specimens the As concentration was very high. The disposition of all elements except Zn, As and Fe was uniform. Zn and As were concentrated on the surface of the septum and Fe in the region of vascular channels.

The fate of homologous nasal septal cartilages in tympanoplasty.

We have used homologous nasal septal cartilage for tympanoplasty for the last 8 years and obtained satisfactory results. In order to demonstrate the fate of homograft cartilage implanted into the middle ear, mucopolysaccharides have been studied by means of enzyme digestion. The matrix of normal septal cartilage was divided into three regions: 1) pericellular region; chondroitin sulfate B, 2) distal interstitial region; hyaluronic acid and chondroitin sulfates, 3) peripheral interstitial region; collagen. In preserved cartilage, chondroitin sulfate B was lacked out, but hyaluronic acid and collagen remained intact though the amount of mucopolysaccharides diminished slightly when compared with normal septal cartilage. Homograft cartilages evidenced depletion of mucopolysaccharides. Homograft cartilages should be used for the purposes mentioned, though not as material for columella, nor for reconstruction of large bone defect.

Isolation and characterization of high-buoyant-density proteoglycans from bovine femoral-head cartilage.

Proteoglycans were extracted from bovine (15-18 months old) femoral-head cartilage. The heterogeneity of the A1D1 proteoglycan fraction was examined by gel chromatography, sedimentation velocity, sucrose rate-zonal centrifugation and CS2SO4 isopycnic centrifugation. In all cases polydisperse but unimodal distributions were obtained. Chemical analysis of the preparation yielded a galactosamine/glucosamine molar ratio of 7:1, and 13C n.m.r. spectroscopy showed that the chondroitin sulphate comprised equal proportions of the 4- and 6-sulphate isomers. Gel chromatography of a papain and Pronase digest of the proteoglycan indicated that the chondroitin sulphate chains had a Mn of approx. 10500. The mean buoyant density of the proteoglycan in pure CS2SO4 was 1.46 g/ml. Physical characterization of the proteoglycan preparation in 4M-guanidine hydrochloride, pH 7.4, by using conventional light-scattering gave a radius of gyration of 42 nm and a Mw of 0.96 X 10(6). Quasi-elastic light-scattering in the same solvent yielded a translational diffusion coefficient, D020, of 5.41 X 10(-8) cm2 X S-1, and ultracentrifugation gave a sedimentation coefficient, S020, of 12.0S. Thus from sedimentation-diffusion studies a Mw of 1.36 X 10(6) was calculated. The possible origins for the differences in the two molecular-weight estimates are discussed. It is concluded that the high-buoyant-density proteoglycans from bovine articular cartilage are significantly smaller than those from bovine nasal septum, and that this is largely due to the smaller size of their chondroitin sulphate chains.

Histochemical properties of cartilage proteoglycans.

Proteoglycan interaction with alcian blue at different concentrations of magnesium chloride was studied both in vitro and in histological sections of paraffin-embedded tissues. Our experiments indicate that a) proteoglycans with different contents of chondroitin sulfate and keratan sulfate, prepared under nondegradative conditions, are not distinguishable on the basis of the critical electrolyte concentrations at which staining is abolished; b) the state of aggregation of proteoglycans only very slightly affects the alcian blue affinity of the macromolecules at different concentrations of magnesium chloride; c) the interaction of proteoglycans with other components of the connective tissue matrix is an important factor in determining the strength of binding of alcian blue to the polyanionic macromolecules in histological sections. These factors should be considered in interpreting histochemical data obtained by staining tissue sections with alcian blue at different concentrations of magnesium chloride. Proteoglycans, like glycosaminoglycans, are only weakly periodic acid-Schiff-positive.

A comparison of the proteoglycans produced by rabbit articular chondrocytes in monolayer and spinner culture and those of bovine nasal cartilage.

The structural and immunological properties of the glycosaminoglycans and the core proteins of bovine nasal cartilage proteoglycan and the proteoglycans produced by rabbit articular chondrocytes in spinner and monolayer culture were compared. Culture medium with 35SO4- or 3H-serine-labeled proteoglycan was mixed with bovine nasal cartilage 4M guanidine-HCl extract and digested with trypsin. The proteoglycan fragments were then isolated by DEAE-cellulose chromatography and fractionated by dissociative CsCl density gradient centrifugation. Approximately 90% of the 35SO4 incorporated into proteoglycan by the cultured chondrocytes was in chondroitin sulfates and about 5% in keratan sulfate. Although there was considerable overlap in the Sepharose 4B elution of the tryptic proteoglycan fragments of highest buoyant density, some monolayer-produced proteoglycan fragments eluted earlier and some spinner-produced proteoglycan fragments eluted later than the proteoglycan fragments from bovine nasal cartilage. These differences in apparent fragment size could relate to differences in glycosaminoglycan chain length, since the glycosaminoglycans released by treatment with alkali from monolayer-produced proteoglycan in part eluted from Sepharose 4B earlier and those from spinner-produced proteoglycan in part eluted later than the chondroitin sulfate chains released from bovine cartilage proteoglycan. After digestion with chondroitinase ABC, 3H-serine-labeled high density tryptic proteoglycan fragments from monolayer and spinner culture yielded Sepharose 6B elution profiles which were similar to each other but did not coincide with the peaks of carbazole reactivity found with similarly treated fragments of bovine nasal cartilage proteoglycan. Cross-reactivity was demonstrated by radioimmunoautography between bovine cartilage and rabbit chondrocyte proteoglycan fragments restricted to gradient fractions of low buoyant density, but immunological cross-reactivity was not found for the antigens associated with the keratan sulfate-rich and chondroitin sulfate-bearing tryptic fragments of bovine nasal cartilage proteoglycan. These studies indicate that the proteoglycan core proteins produced by rabbit articular chondrocytes in monolayer and spinner culture are, in part, different from the core protein of bovine nasal cartilage proteoglycan and that the three proteoglycans differ in the length of some of their chondroitin sulfate chains.

Phosphate ester groups in proteoglycans from bovine nasal cartilage.

1. Proteoglycan subunits isolated by standard procedures from bovine nasal cartilage, previously incubated in the presence of [32P]phosphate contain [32]-phosphate ester groups as a regular structural component. 2. Contamination of the proteoglycan subunit with 32P-labeled nucleic acids could be excluded by repeated cesium chloride density gradient centrifugation under associative and dissociative conditions, lanthanum chloride precipitation, gel filtration and by the resistance of the proteoglycan subunit associated 32P to phosphoric diester hydrolases. 3. The [32P]phosphate ester groups are associated to the chondroitin sulfate peptide fraction obtained by proteolytic digestion of the proteoglycan subunit molecule. Degradation of the chondroitin sulfate peptide by chondroitinase ABC resulted in a 32P-labelled oligosaccharide peptide fraction, that contains xylose, galactose, glucuronic acid and inorganic phosphate in a molar ratio 1 : 2 : 1 : 0.12. 4. 32P radioactivity is released as inorganic phosphate by treatment of the 32P-labelled oligosaccharide peptide with acid phosphatase or alkali.

Selective aggregation of proteoglycans with hyaluronic acid.

Conditions were established to separate proteoglycan aggregate (AH1) from a bovine nasal septum extract by associative rate zonal sedimentation on a NaCl gradient. The AH1 has a higher protein content than the mixed aggregate-monomer (A1) isolated by conventional associative CsCl density gradient centrifugation from a portion of the same extract. The same associative rate zonal conditions separated the A1 fraction into aggregated AH1 containing hyaluronic acid and nonaggregated proteoglycan monomer (N1) essentially free of hyaluronic acid. The AH1 fraction is richer in protein and keratin sulfate than is N1. Dissociative rate zonal sedimentation of A1 under conditions which totally sedimented most of the disaggregated monomer (AH1-D1) and the nonaggregated monomer N1 separated a less sedimentable protein and keratan sulfate-rich proteoglycan monomer (AH1-D2). Chromatography on Sepharose 2B under dissociative conditions demonstrated that the nonaggregated N1 monomer is intermediate in size between the disaggregated monomers AH1-D1 and AH1-D2. N1 has a buoyant density higher than AH1 and is practically equivalent to AH1-D1. All are dense fractions so that separation by CsCl density gradient equilibration is not feasible.

Comparative studies on human and bovine nasal cartilage proteoglycan complex components.

Human and bovine nasal proteoglycan complex components were prepared in parallel. Some molecular properties of human and bovine proteoglycan subunits (PGS) were compared. Two major human "link-like proteins" have been characterized and purified; their compositions and molecular weights were very similar to those observed for the previously described bovine "link proteins".

Investigation of molecular motion of proteoglycans in cartilage by 13C magnetic resonance.

13C nmr spectral parameters were measured for intact bovine nasal cartilage tissue, the purified proteoglycan aggregate, and chondroitin 4-sulfate. A comparison of integrated intensities obtained for four different samples of fresh tissue with an ethylene glycol standard indicated that at least 80% of the total glycosaminoglycan carbons in the tissue contributed to the spectrum. This result was confirmed by intensity measurements obtained at 56 degrees on fresh tissue and at 37 degrees after extensive papain digestion of fresh tissue. Spin lattice relaxation times and nuclear Overhauser enhancements were analyzed in terms of the following models of molecular motion: (a) single correlation time; (b) log X2 distribution of correlation times; and (c) anisotropic motion. The analysis indicates that the segmental motions of glycosaminoglycan chains are characterized by a broad distribution of correlation times centered at about 50 ns. Slow motion contributions to glycosaminoglycan line widths were reduced by dipolar decoupling (gammaH2/2pi = 65 kHz). Collagen intensity was observed in dipolar decoupled spectra, but not in scalar decoupled spectra of intact tissue, showing that the type II collagen in cartilage undergoes anisotropic motion like the type I collagen in tendon. Only glycosaminoglycan resonances were observed in spectra of a solution of proteoglycan aggregate before and after chondroitinase digestion. After subsequent digestion with papain, protein resonances were observed. These results suggest that the protein portions of the proteoglycan aggregate structure, in contrast with the glycosaminoglycan chains, have restricted backbone mobility and consequently a defined backbone structure.

Nasal glands in newborn infants.

The entire mucosa of the nasal septum from four newborn infants was removed, stained by the PAS-alcian blue whole-mount method, and the density of mucous glands was determined in 12 different localities. The interindividual median density was 30.7 glands/mm2 and the median count 13,500 glands. The glands were regularly distributed over the entire respiratory region, but with a significant decrease in density in the posterior half. It has been demonstrated that all glands are laid down before birth, that their density decreases with increasing age until it reaches 8.4 glandss/mm2 at an adult age, and that acute or recurrent acute catarrhal or inflammatory changes of the nose do not lead to the newformation of glands.

Isolation and physical characterization of hyaluronic acid prepared from bovine nasal septum by cetylpyridinium chloride precipitation.

Raw extract in 2 m CaCl2 of bovine nasal septum cartilage was eluted from 4 per cent agarose gel to give a "void volume" Fraction v-4, which was indistinguishable in composition and behavior on viscometric and sedimentation analysis from the densest fraction obtained by associative centrifugation in a cesium chloride density gradient. The sulfated proteoglycan was precipitated (Fraction A) by cetylpyridinium chloride from acidic solutions of Fraction v-4 or of dialyzed raw ectract. Neutralization under conditions of low ionic strength precipitated a further small fraction (B), which contained from 0.5 to 1 per cent of the uronic acid in the original extract. Analysis by associative and dissociative density gradient centrifugation demonstrated that Fraction B resembled in effective density known samples of hyaluronic acid from other sources. Gel chromatography of proteolytic digests of Fractions A and B on 6 per cent agarose indicated that cetylpyridinium chloride precipitation essentially separated sulfated proteoglycan (A) from hyaluronic acid (B). A viscosity-average molecular weight of about 5 x 10(5) was estimated for a sample of Fraction B purified in a dissociative (4 M guanidine hydrochloride + CsCl) density gradient. Sedimentation velocity data were consistent with this result. Analysis of hexosamines showed that the sample contained 96 per cent glucosamine, confirming the identification of hyaluronic acid. The proteoglycan fraction (A) resembled "subunits" in its sedimentation behavior.

[Interaction of connective tissue structural glycoprotein with proteoglycans and glycosaminoglycans]. / Vzaimodeistvie strukturnogo glikoprotein soedinitel'noi tkani s proteoglikanami i glikozaminoglikanami

It was shown that structural glycoprotein (SGP) of connective tissue isolated from be cattle heart valves and nasal septum cartilage formed water-soluble complexes with protein-chondroitin-4-sulphate and different heparin fractions. SGP formed no mentioned complexes with hyaluronic acid. Probably this plays an important role in the formation of collagen and elastin fibers.

Endonasal venular permeability in rats exposed to the cold.

Sixteen rats were killed 22 hrs after being injected with a dose of 4 mCi(198)Au mixed with 16 mg of inert colloidal gold/100g body wt. The anterior nasal mucosa of the septum and turbinates in 10 rats which were injected and kept in a temperature of 5-7 degrees C showed mean radioactive counts 2-3 times higher than those recorded in the same areas in 6 control rats kept in ordinary room temperature (22 degrees C). In all sixteen rats the highest counts were obtained in the RES organs, namely the liver, spleen, adrenals and the bone marrow, in descending order. Other tissues revealed no significant radioactivity except for minor counts in the lungs and kidneys. The importance of these findings regarding the localisation of granulomatous diseases and vasculitis in the nose is discussed.