Orcein staining and the identification of polytene chromosomes.

Acetic orcein staining of polytene chromosomes was introduced in 1941 shortly after the initial studies on aceto-carmine-stained chromosomes by Bridges (2) and has remained a standard method of preparation. Orcein dye can be purchased in both its natural form as extracted from two species of lichens, Rocella tinctoria and Lecanora parella, and a synthetic form. The mechanism of staining is not clearly understood because the stain itself is a variety of phenazones, which may interact at an acid pH with negatively charged groups or possibly interact hydrophobically with chromatin.

Transformation of MC3T3-E1 cells following stress and transfection with pSV2neo plasmid.

A continuous cell line, MC3T3-E1 cells, originally derived from murine calvaria bones, loses its osteogenic properties as a result of extended passage number under stress conditions. These aged/stressed MC3T3-S cells, although nontumorigenic, do not display some of the osteogenic properties characteristic of the MC3T3-E1 cells. Altered properties include low expression of alkaline phosphatase, diminished collagen synthesis and inability to form mineralized nodules in vitro.

Effect of methyl mercaptan on synthesis and degradation of collagen.

Measurements of the conversion of [14C]-proline to [14C]-hydroxyproline were employed to assess the effect of methyl mercaptan on intra- and extracellular metabolism of collagenous proteins in human gingival fibroblast cultures. Following a 30-min pulse, 10 ng of methyl mercaptan per ml of 95% air/5% CO2 head-space suppressed collagen synthesis by 39% and increased the intracellular degradation of newly synthesized collagen from 26% to 42%. Parallel cultures assayed for proline transport demonstrated a 29% inhibition of [14C]-proline uptake.

Stimulation of enzyme and cytokine production by methyl mercaptan in human gingival fibroblast and monocyte cell cultures.

The volatile sulphur compound methyl mercaptan (CH3SH) is a by-product of protein metabolism and a principal component of oral malodour. This investigation examines the effect of CH3SH on the enzymatic activities of cathepsins B and G and elastase, and on the production by human gingival fibroblasts of two key factors, prostaglandin E (PGE) and cAMP, of the PGE2-cAMP-dependent pathway, which may contribute to the increased production of collagenase and tissue destruction in human periodontal disease. The results demonstrate that CH3SH alone, or in combination with interleukin-1 (IL-1) or lipopolysaccharide, can significantly enhance the secretion of PGE2, cAMP and procollagenase by human gingival fibroblasts.

Evaluation of a model for post-partum arthritis and the role of oestrogen in prevention of MRL-lpr associated rheumatic conditions.

Sixty-eight percent of female MRL-lpr mice developed a post-partum exacerbation of their mild spontaneous arthritis within 30 days of parturition. The flare became evident between 5 and 15 days after delivery. Histologically it was characterized by a significant increase of subsynovial inflammation and synovial hyperplasia without changes in the level of cartilage and bone erosion.

Lpr and MRL background gene involvement in the control of adjuvant enhanced arthritis in MRL-lpr mice.

The MRL-lpr mouse strain develops a mild spontaneous arthritis which can be enhanced by the intradermal injection of complete Freund's adjuvant (CFA). In this study we examined the requirement of the lpr gene and background MRL genes on CFA-enhanced murine arthritis. MRL+, MRL-lpr, AKR/J, and B6-lpr mice (experimental) and B6 mice (control) were injected intradermally with CFA containing M.

Acquisition of resistance to 6-azauridine through DNA amplification in neoplastic but not normal osteoblasts.

In this communication, we have characterized the resistance to AZUrd in tumorigenic mouse C3H-OS osteosarcoma cells and non-tumorigenic MC3T3-E1 osteoblast cells. DNA and RNA blot analysis showed a 30-fold increase in UMP synthase specific DNA and a 10-fold increase in mRNA, respectively, in resistant versus non-resistant C3H-OS cells. No corresponding increases in either UMP synthetase DNA or mRNA were evident in resistant MC3T3-E1 osteoblasts.

Photodynamic therapy; a comparison with other immunomodulatory treatments of adjuvant-enhanced arthritis in MRL-lpr mice.

Although numerous experimental immunomodulatory regimens have been reported to be effective in the treatment of rheumatoid arthritis, they also produce undesirable side effects. An alternative specific modality of localized treatment is photodynamic therapy (PDT). In this study we treated 13-week-old MRL-lpr mice whose spontaneous arthritis was enhanced by intradermal injection of Freund's complete adjuvant (FCA).

Complete Freund's adjuvant induces an earlier and more severe arthritis in MRL-lpr mice.

A study was performed on the effect of CFA on the spontaneous arthritis of MRL-lpr mice. The development of swelling and erythema was monitored for 1 mo after injecting 13- to 14-wk-old mice intradermally with CFA, at which time the histopathology of the joints and serologic responses to extracellular matrix proteins were investigated. In a series of six experiments, 67 to 82% of mice showed early clinical evidence of arthritis in contrast to the low percentage observed in control animals.

Effect of volatile thiol compounds on protein metabolism by human gingival fibroblasts.

The levels of volatile sulphur compounds (VSC) in periodontal pockets and mouth air have been found to correlate with severity of the disease process. The purpose of this study was to examine the influence of hydrogen sulphide and methyl mercaptan on protein metabolism of human gingival fibroblasts. The incorporation of labelled amino acids into protein was used to evaluate effects on total protein content.

Modulation of human gingival fibroblast cell metabolism by methyl mercaptan.

Methyl mercaptan (CH3SH) is a malodorous compound whose levels are elevated in mouth and crevicular air of individuals with active periodontal disease. Since it may play a role in the disease process, its effects were evaluated using human gingival fibroblast cultures and viable porcine unkeratinized oral mucosal tissue sections. Results showed that the protein content of CH3SH-exposed cell cultures pulsed with [14C]-labelled glycine and proline was decreased by approximately 25%.

Characterization of volatile sulphur compounds production at individual gingival crevicular sites in humans.

The present investigation describes a method for collection and analysis of volatile sulphur compounds (VSC) from gingival crevicular sites in humans. Tenax-GC trapping devices were used to adsorb and concentrate VSC from crevicular air at -55 degrees C, which were then thermally desorbed at 120 degrees C. Gas chromatographic (GC) analyses were performed using a Tracor 550 GC equipped with a flame-photometric detector and a Teflon column packed with 5% polyphenyl ether and 0.

Antibodies to extracellular matrix proteins in the sera of MRL-lpr mice.

Autoimmune MRL-lpr mice develop a spontaneous arthritis displaying similar articular and extra-articular features to rheumatoid arthritis in humans. In this study we used an ELISA assay to evaluate the serological responses of MRL-lpr mice to select extracellular matrix proteins associated with the joint. Significant levels of antibodies to collagens types I, II, III, IV, and V were demonstrated starting between 17 and 20 weeks of age.

Gas chromatographic method for trapping and detection of volatile organic compounds from human mouth air.

The present investigation describes a convenient method for collection and analysis of volatile organic compounds from 25 ml mouth air samples. Tenax-GC trapping devices coated with Teflon are used to adsorb and concentrate volatile organic compounds in mouth air at -20 degrees C, which are then thermally desorbed at 140 degrees C. Gas chromatography (GC) analyses are performed using an aluminum column coated with Teflon and packed with 2% poly-MPE on 80/100 mesh Tenax-GC, and employing a flame ionization detector.

Conservatism of sites of tRNA loci among the linkage groups of several Drosophila species.

The sites of seven tRNA genes (Arg-2, Lys-2, Ser-2b, Ser-7, Thr-3, Thr-4, Val-3b) were studied by in situ hybridization. 125I-labeled tRNA probes from Drosophila melanogaster were hybridized to spreads of polytene chromosomes prepared from four Drosophila species representing different evolutionary lineages (D. melanogaster, Drosophila hydei, Drosophila pseudoobscura, and Drosophila virilis).

Induced and natural break sites in the chromosomes of Hawaiian Drosophila.

gamma-Irradiation of a laboratory strain of the Hawaiian species Drosophila heteroneura yielded 310 breaks in the five major acrocentric polytene chromosomes. Their map positions conform to the Poisson distribution, unlike most of the 436 natural breaks mapped in 105 closely related species endemic to Hawaii. Genome element E is longer and has more induced breaks than the others.

The influence of aging on lysosomal acid DNase of the rat submandibular gland.

Lysosomal and cytoplasmic fractions were prepared from rat submandibular glands for investigation of the release of lysosomal acid DNase in relation to aging. It was found that the acid DNase activity ratio for cytoplasmic/lysosomal fractions in rats aged 27 months was higher than that in three-month-old rats. The release of acid DNase from the lysosomal fraction by shaking was markedly increased in the fraction from the older animals.

Improved high-performance liquid chromatography method for quantitation of proline and hydroxyproline in biological materials.

Numerous high-performance liquid chromatography systems have been described for the determination of hydroxyproline (Hyp) and proline (Pro) levels in biological materials. These methods are generally complicated and have shortcomings in applicability due to poor separation, low sensitivity or derivatization-associated problems. The large number of chemical components present in biological samples further complicates the analysis of Hyp which usually occurs in extremely low concentrations.

Radiochromatographic analysis of H2S-collagen complexes in rat-tail type I collagen.

Suspensions of 15 mg lyophilized type I, acid-soluble, rat-tail tendon collagen in 0.02 M tris/0.134 M NaCl (pH 7.

Sulfur uptake by type I collagen from methyl mercaptan/dimethyl disulfide air mixtures.

Type I acid-soluble collagen, suspended in 0.02 M Tris/0.13 M NaCl buffer (pH 7.

Effect of hydrogen sulfide and methyl mercaptan on the permeability of oral mucosa.

Hydrogen sulfide (H2S) and methyl mercaptan (CH3SH) are the volatile sulfur compounds (VSC) that were investigated for a possible role in the etiology of periodontal disease. The results show that the permeability of porcine non-keratinized sublingual mucosa is increased by up to 75% or 103% following exposure to H2S and CH3SH, respectively. The effect may be attributed to VSC reaction with tissue components resulting in alteration in the integrity of the tissue barrier.

Volatiles of exogenous origin from the human oral cavity.

The volatiles found in the headspace above male and female saliva were examined by combined gas chromatography--mass spectrometry. This had led to the identification of a number of constituents of exogenous origin. The most likely source of these products are atmospheric and water pollutants as well as food stuffs and cosmetic products.

Quantitative differences in volatiles from healthy mouths and mouths with periodontitis.

It has been suggested that orally derived volatile aromatic amines may be of possible diagnostic use and may contribute to the etiology or pathogenesis of periodontal disease. Using gas chromatography and gas chromatography/mass spectrometry, we identified and quantitated pyridine and picolines in the headspace of incubated whole saliva from healthy and diseased subjects. The oral health of subjects was evaluated by four standard oral-health indices.

Characterization of volatile sulphur production by pathogenic and non-pathogenic strains of oral Bacteroides.

Marked differences were observed in intermediate sulphur metabolism between non-pathogenic strains of Bacteroides melaninogenicus var melaninogenicus (CP-) and pathogenic Bacteroides melaninogenicus asaccharolyticus (CP+). The CP+ strains, which produced collagenase and protease and caused formation of abscesses when injected subcutaneously into groins of guinea pigs, produced copious amounts of volatile sulphur compounds (VSC) which consisted predominantly of CH3SH and (CH3S)2. Hydrogen sulphide occurred in considerably lesser amounts.