A kinetic study of DNA and basic protein metabolism during spermatogenesis in the sand crab, Emerita analoga.

Cytochemical and radioautographic techniques define and confirm a staging scheme for developing spermatids of the decapod crab, Emerita analoga. Quantitative photometric data demonstrate that developing spermatids lose a significant proportion of their nuclear proteins, as evidenced by diminishing binding of fluorodinitrobenzene. Photometric results also show that much (but not all) of the spermatid nuclear protein loss is in somatic-type histone, as evidenced by a dramatic fall in the histone/DNA ratio of these cells during a period in which nuclear DNA content remains constant. By the end of spermiogenesis, the sperm nuclear histone and protamine content is approximately zero, whereas some nonbasic protein persists. Loss of spermatid nuclear somatic-type histone is not accompanied by synthesis of gamete-type histone (e.g. protamine or arginine-rich histone), showing that the processes of displacement and synthesis of nuclear basic proteins during histone transition are not subject to obligatory coupling. Labeling studies suggest that nonbasic acrosomal proteins (presumably partly enzymes) are synthesized in the cytoplasm, after which they move into the acrosome. Stainable basic proteins accumulate in the acrosome during precisely the period of nuclear somatic histone loss, suggesting nuclear-cytoplasmic transfer.

Matrix loss and synthesis following a single impact load on articular cartilage in vitro.

Articular cartilage biopsies were subjected to a single impact load and the metabolic response of the chondrocytes investigated using radiolabelled precursors for protein ([3H]leucine) and glycosaminoglycan ([35S]sulfate). The severity of the impact was controlled by using different masses and drop heights in a purpose built drop tower. Loss of matrix components was studied by prelabelling prior to loading, the possible repair response by pulse labelling at defined intervals after loading. There was an increase in the loss of both labels from the tissue with increasing severity of impact though the patterns of loss were different. Only 25%-40% of the sulfate was lost over a two week period and the loss increased with the severity of impact. This contrasted with 60% of the leucine being lost over the same period independently of loading. In addition to the loss of synthetic activity caused by cell death, there was a suppression of incorporation immediately following loading. This eventually recovered and increased above control values but the recovery time appeared to depend on the severity of the impact. These results provide preliminary evidence for a repair response.

Soluble antigen extracts used as blocking agents to obtain specificity in serotyping of Streptococcus mutans.

Specific immunofluorescent (IF) conjugates were prepared for Streptococcus mutans serotypes d and g. Serotype specificity was obtained by blocking the cross-reacting antibodies with soluble antigen extracted from cells of the cross-reacting strains. Soluble antigen extracts are particularly useful in obtaining specificity with quantities of conjugates that are too small to be efficiently subjected to adsorption procedures. They also can be used advantageously to block traces of cross-reactivity that frequently remain with conjugates that have been subjected to conventional adsorption procedures. S. mutans isolates from dental plaque samples were identified as belonging to serotypes d or g on the basis of IF staining with conjugates that were made type-specific by blocking the cross-reacting antibodies with appropriate soluble antigen extracts.

Biocompatibility of particles of GORE-TEX cruciate ligament prosthesis: an investigation both in vitro and in vivo.

This research was to determine the biocompatibility of simulated wear particles from a GORE-TEX cruciate ligament prosthesis using in vitro and in vivo techniques. Exposure of normal human synovial fibroblasts and primary mouse peritoneal macrophages to the particles revealed no cytotoxicity, as determined by lactate dehydrogenase release, but beta-N-acetyl-D-glucosaminidase was released from macrophages at high concentrations, showing inflammatory potential. This was not seen when the particles were injected into mouse knees, where no adverse reactions were observed. These simulated wear particles were shown to be biocompatible using these in vitro and in vivo systems.

Biocompatibility of diamond-like carbon coating.

Diamond-like carbon coating is an inert, impervious hydrocarbon coating with properties suitable for use in the biomedical field, particularly in orthopaedic implants. Mouse peritoneal macrophages and mouse fibroblasts were grown on tissue culture plates treated with diamond-like carbon and the biocompatibility assessed both biochemically and morphologically. This investigation showed that diamond-like carbon coating caused no adverse effects on cells in culture and therefore merits further investigation as a coating for biomedical use.

Biocompatibility of particulate polymethylmethacrylate bone cements: a comparative study in vitro and in vivo.

The biocompatibility of particles of four different commercial polymethylmethacrylate (PMMA) bone cements was evaluated by exposing human synovial fibroblasts and mouse peritoneal macrophages to particles of cement. Cell integrity and inflammatory potential were assessed using enzyme release and microscopical examination. Results suggested the occurrence of cell damage in both cell types and macrophage studies indicated inflammatory potential. The response in vivo was investigated by intra-articular injection of the particles into mouse knee joints. Clinical and histological evaluation was performed over 2-52 wk. Particles of all four cements were well tolerated in the joints.

Preparation of antiserums for use in the fluorescent antibody identification of certain plaque bacteria.

The interaction of many factors involved in FA antiserum production determines the quality of each antiserum or conjugate prepared. Performance and physicochemical characteristics of 58 FA antiserums to S mutans currently in use suggested that many of these conjugates could have been improved by using the methods and evaluation procedures described in this symposium. Deficiencies in the conjugates included low titers, incomplete fractionation, inadequately labeled antibody, fluorochromed albumin, free fluorescein and cross-reactions. Titers ranged from 1:1 up to 1:4,000. Titers for S mutans serotype c conjugates were uniformly low. S mutans serotype c conjugates were prepared from antiserums produced using a modified immunization schedule. The schedule used both viable and killed whole cells and gave FA titers as high as 1:1,000 (adjusted to 10 mg protein/ml). Procedures presented in this paper and the report by Pittman and co-workers27 should permit direct FA serum titers of up to 1:1,000 for each of the recognized serotypes of S mutans. The availability of highly specific antiserums with adequate titers will advance the use of the FA technique to identify microorganisms directly in specimens, such as dental plaque.29,30 This technique could be used to study such phenomena as the transmission of microorganisms from person to person and the establishment of the oral flora. It could also be used in epidemiological studies and perhaps to monitor the effect of a therapeutic agent on microbial composition of dental plaque.

Application of fluorescent antibody methods in the analysis of plaque samples.

Direct FA staining for S mutans serotypes may be performed on smears made from plaque or strain isolates and on colonies attached to agar plates of black membrane filters. Staining with single conjugates directed to S mutans serotypes (a to e) as well as with polyvalent a-, b-, d-, f-fluorescein label and c-, e-rhodamine label conjugates indicate that serotype c is the most common. This is in agreement with many reports by other investigators. Cross-reactions with many S mutans conjugates occurred with organisms resembling Lancefield groups C and G streptococci.

Prevalence of Streptococcus mutans serotypes, Actinomyces, and other bacteria in the plaque of children.

Selected microbial components in dental plaque were determined for children in Biddeford, Maine and Colombia, South America. Using cultural methods, Streptococcus mutans was detected in 51.4% of the Colombian children and 63.3% of the Maine children. Serotype c was predominant in both populations. The greatest difference between the two groups occurred with serotypes d and g which were present in 25% of the Colombian children with S. mutans and were not detected in the Maine children. In the specimens examined with specific FA conjugates. Actinomyces was the predominant genus, present in all individuals and comprising an average of 52% of all cells.

Plaque, caries, periodontal diseases, and acculturation among Yanomamö Indians, Venezuela.

The number of DM and d teeth and surfaces was recorded for 220 Yanomamö Indians from three groups of villages with different degrees of contact with Western culture. Specimens of plaque were taken from the teeth, transported in a holding solution, cultured, and examined for specific oral streptococci. In addition, the periodontal health and oral hygiene of one group of villagers were assessed using the Russell PI and the Greene & Vermillions OHIS. Caries experience among the Yanomamö was shown to be positively associated with exposure to Western culture. S. mutans was recovered with about the same frequency from specimens taken from the teeth of Indians living at all three village locations. However, the presence of S. mutans alone did not account for the disparity in dental caries scores. The examinees had abundant and persistent accumulations of soft deposits on their teeth accompanied by markedly inflamed gingival tissues. However, periodontal pockets and loss of appreciable amounts of bone did not appear as early in life nor were they as severe as reported for some other populations which practice little oral hygiene. Those disparities in the distribution of plaque-induced oral diseases between Western populations and the Yanomamö warrant further study.

The effect of starch glove powder on joint and other tissues.

The purpose of this research was to establish the possible role of starch glove powder in complications following orthopaedic surgery using in vivo and in vitro techniques. Exposure of primary mouse peritoneal macrophages to starch glove powder caused 10% release of prostaglandin E2 (0.1 mg/ml, 16 h) but no increased release of lactate dehydrogenase, demonstrating that cell integrity had not been compromised. Long-term tissue reaction to starch glove powder was investigated in vivo by injection into mouse knee joints. Over a period of 52 weeks no inflammatory response was elicited, no starch was observed in the regional lymph nodes and none was found in joints after the 8th week. Starch glove powder appeared to be innocuous in the joint and although prostaglandin E2 release was stimulated in vitro, this had no apparent effect on joints in vivo.

Meeting continuing professional education needs.

The interest in the Diploma of Advanced Nursing Studies is indicative of the need and demand for flexible, student-centred learning opportunities that link theory with clinical practice. The programmes use study guides that offer students a sequential or problem-solving interactive vehicle between the student and teacher. Distance learning provides students with access to flexible learning opportunities when their choices are constrained by location and personal circumstances.

Evaluation of excitation light sources for incident immunofluorescence microscopy.

A variety of fluorescent excitation light sources were compared using a standard fluorescein solution or a bacterial conjugate with immunofluorescent microscopy. Quantitative data were obtained with microscope photometric apparatus. Both the quantitative data and comparative conjugate titering suggest that the 450-W xenon arc excited significantly more fluorescence than did the more commonly used 250-W mercury arc or the 100-W halogen lamp. The conjugate could be diluted 4 to 32 times more using the 450-W xenon. Additional advantages of 450-W xenon excitation include sufficient energy of wave lengths between 470 to 490 mm, thus permitting narrow-band excitation resulting in less autofluorescence and the ability to perform fluorescent-antibody procedures without the darkening of ambient room light.

Antibiotic susceptibility of Streptococcus mutans: comparison of serotype profiles.

A total of 82 strains of Streptococcus mutans representing serotypes a through g were tested for susceptibility to erythromycin, penicillin, methicillin, lincomycin, tetracycline, vancomycin, gentamicin, streptomycin, neomycin, kanamycin, bacitracin, and polymyxin B. Strains included stock cultures and isolates from human and animal dental plaque. Minimal inhibitory concentrations were determined by a broth-microdilution procedure. The major differences in antibiotic susceptibility observed among the serotypes resulted with antibiotics which act on the cell surface. Bacitracin was most active against serotype a strains and polymyxin B against serotype b strains. Serotypes a, d, and g were less susceptible than the other serotypes to methicillin.

Comparative recovery of Streptococcus mutans on ten isolation media.

The ability of Streptococcus mutans (Bratthall serotypes a through e) to grow on 10 isolation media was examined. The number and morphology of the colonies were observed to vary on different media. The use of blood-sucrose media consistently produced the highest recoveries. Mitis salivarius agar (MS) and higher recovery values than modified medium 10 (MM10SB), Trypticase-yeast extract-cystine medium (TYC), or MS with 1% tellurite (MST). MST with 40% sucrose (MS40S), MST with 20% sucrose and 0.2 U of bacitracin per ml (MSB), and Carlsson medium with 1% sulfasoxazole (MC), media formulated for the selection of S. mutans, were the most inhibitory for all serotypes. The morphology of several S. mutans strains was atypical on MC and MS40S, making positive identification difficult. Absence of growth of serotype a strains on MSB and serotype d strains on MC were the two major differences observed among the serotypes. Results are discussed in terms of the difficulties in making quantitative determinations from cultural data.