Defining the physiologically normal coating and pathological deposit: an analysis of sulfur-containing moieties and pellicle thickness on hydrogel contact lenses.

PURPOSE: Historically, biochemical studies of the interaction between tears and hydrogel contact lenses have not been coordinated with the study of the morphological ultrastructure of the phenomena. Moreover, terms that have distinct and different meanings--pellicle, coating, deposit, and biofilm--have been used interchangeably and often incorrectly when applied within the context of the general field of contact lens biotechnology to describe the tear-polymer interaction. We describe our elucidation of morphological and elemental characteristics of the normal pellicle that forms on the lens surface and urge standard use of the word "pellicle" to specify this entity. METHODS: Fourteen worn hydrogel lenses (8 Group 1 and 6 Group 4 lenses according to the FDA classification) were rinsed, quartered, and fixed or dried, depending on the analysis to be performed. Scanning electron microscopy (SEM) was used to examine the morphology of the pellicle and quantify its thickness. X-ray analysis was used to detect elements associated with the anterior, central, and posterior portions of the lenses and their relative distribution. RESULTS: A distinctive morphological pellicle ranging from 0.1 to 8.6 microns was present on 12 of the 14 lenses. The pellicle was thicker on the Group 4 lenses than on the Group 1 lenses (P < 0.003). However, the pellicle on Group 1 lenses became thicker with increasing lens age (P < 0.02), but not as thick as on Group 4 lenses. Morphologically distinct lipid or jelly bump deposits were observed at the surface of both lenses from a single patient wearing 2 week old Group 4 lenses. Eleven lenses had sulfur-bearing tear components on the anterior zone. Sulfur was deposited within the matrix of nine lenses. The sulfur containing moieties were more prevalent on Group 4 lenses (P < 0.002). More sulfur was assayed on older lenses (P < 0.004). The anterior lens zone had more sulfur-bearing tear components than did the posterior or center zones (P < 0.05). CONCLUSIONS: The physiologically normal pellicle is a distinct morphological entity covering the anterior lens surface. Abnormal deposits such as the discrete microgel region, known as jelly bumps, are not part of the physiologically normal pellicle at the anterior lens surface and have the potential to induce pathology. Sulfur-containing moieties within the matrix may represent the breakdown of large proteins and mucoproteins or intact proteins, as well as contaminants such as cosmetics and environmental pollutants. It is also possible that entire small proteins, such as lysozyme, impregnate the matrix. The moieties that become entrapped within the matrix or rigidly adhere to the matrix should be considered true deposits.

Ciliary protein conservation during development in the ciliated protozoan, Oxytricha.

The ciliated protozoan Oxytricha fallax possesses multiple highly localized clusters of basal bodies and cilia, all of which are broken down and rebuilt during prefission morphogenesis-with one major exception. The adoral zone of membranelles (AZM) of the ciliate oral apparatus contains approximately 1,500-2,000 basal bodies and cilia, and it is the only compound ciliary structure that is passed morphologically intact to one daughter cell at each cell division. By labeling all proteins in cells, and then picking the one daughter cell possessing the original labeled AZM, we could then evaluate whether or not the ciliary proteins of the AZM were diluted (i.e., either by degradation to constituent amino acids or by subunit exchange) during cell division. Autoradiographic analysis demonstrated that the label was highly conserved in the AZM (i.e., we saw no evidence of turnover), and electrophoretic data illustrate that at least one of the proteins of the AZM is tubulin. We, therefore, conclude that for at least some of the ciliary and basal body proteins of Oxytricha fallax, AZM morphological conservation is essentially equivalent to molecular conservation.

Effects of cystment on cells of Oxytricha fallax possessing supernumerary dorsal bristle rows.

Cells of Oxytricha fallax possessing cytotactically inherited supernumerary dorsal bristle rows can redevelop those dorsal supernumerary rows after cystment, even though supernumerary ventral cortical structures are permanently lost through cystment. Previous work has demonstrated: (1) that cystement involves a complete dedifferentiation of all ciliary structures-all cilia, basal bodies, microtubules and fibres; and (2) that all ventral ciliary structures arise from a single ciliary primordium during excystement. These observations suggest the following conclusions. (1) The information for the redevelopment of supernumerary dorsal bristle rows during excystment is associated with some ultrastructurally unidentifiable molecular structure of the cyst cortex. (2) Cytotactic information for the development of cortical patterns is retained in at last two locations in the resting cyst; one location specifies the site of development of the ventral ciliature whereas the other specifies the location and pattern of the dorsal ciliature.

Cytogeometrical determination of ciliary pattern formation in the hypotrich ciliate Stylonychia mytilus. II. Stability and field regulation.

The first regeneration sequence after folding of right fragments of Stylonychia mytilus results in formation of mirror-imaged incomplete doublets illustrating independent determination of polar and lateralaxes. Analysis of the second morphogenetic sequence illustrates that the independent determination of polar and lateral axes is stable through a subsequent cellular reorganization and confirms that cytogeometry participates in determination of ciliary pattern. The morphogenetic inversion of cirral row primordia in this type of fragment is reflected in the structure of the individual cirri. These data not only extend and confirm our earlier study on this type of fragment, but also are consistent with the conclusion derived from data on a different type of mirror-imaged doublet, that global patterning and assembly of ciliature are independently determined.

Analysis of the effects of encystment and excystment on incomplete doublets of Oxytricha fallax.

Singlet cells of Oxytricha fallax possessing supernumerary rows of dorsally located marginal cirri were encysted, excysted, and observed for retention of the supernumerary rows. Without exception, cells lost the supernumerary rows as a result of cystment processes. (Encystment previously has been shown to result in the total resorption of ciliature.) In contrast, supernumerary dorsally located marginal cirral rows develop true-to-type during prefission morphogenesis and thus are inherited from cell to cell. These (and other) observations suggest that at least two levels of cytotactic control of cell patterning are operative on the ciliate cortex; one is dependent upon visible ciliature whereas the other is dependent upon an as yet ultrastructurally unidentifiable molecular architecture.

Patterning and assembly of ciliature are independent processes in hypotrich ciliates.

Mirror-imaged doublets of the hypotrich ciliate Pleurotricha lanceolata were induced and analyzed with respect to the overall patterning (structural asymmetry and polarity) of the individual components of the ciliature. The overall pattern is arranged as a mirror image, but the individual components in the two halves of the doublet show the same organizational asymmetry. These data demonstrate the independence of the mechanisms for this kind of large-scale (global) patterning and control of assembly of the individual ciliary components.

Qualitative and morphometric evaluation of vocal cord lesions produced by the carbon dioxide laser.

The canine larynx was studied to obtain qualitative and quantitative data accurately defining the destructive effects of the carbon dioxide laser on epithelial tissue by administering doses of varied energy to the dorsal surface of the vocal cords. Resultant lesions subsequently were analyzed by light, transmission, and scanning microscopy. Three major morphologic patterns of vocal cord lesions were observed: 1. cup shape, 2. doughnut shape and, 3. complex star shape. The patterns produced in the laryngeal tissue reflect the operative mode (energy distribution within the focused spot of energy) of the laser at the time of surgery. The mode resulting in the doughnut shaped pattern (energy distribution around the periphery of the spot) produced deepest lesions, with minimal thermal conduction laterally, whereas the mode resulting in the cup-shaped pattern (energy distribution uniform throughout the spot) produced at comparable doses shallower lesions with larger diameter. Too few star-shaped patterns were observed to permit meaningful morphometric analysis.

The structure and development of the dorsal bristle complex of Oxytricha fallax and Stylomychia pustulata.

Oxytricha fallax and Stylonychia pustulata possess 6 rows of dorsal bristle units. Each dorsal bristle unit consists of a pair of kinetosomes; the anterior kinetosome has a cilium and the posterior kinetosome a ciliary study. The kinetosome pair, located at the bottom of a cortical pit surrounding the cilium and ciliary stub, is surrounded by an asymmetrical fibrillar mass. Future rows 1-4 are formed from 2 sets of primordia originating within mature dorsal rows 1-3. Rows 5 and 6 originate from the anterior regions of both right marginal cirral primorida. Old dorsal bristle units utilized in formation of primordia are presumably maintained in the new rows of the proter and opisthe; those outside the primordia resorbed. The morphogenetic pattern of the Oxytrichidae is similar to those of the Urostylidae and Holostichidae, but quite different from that of the Euplotidae.

Nuclear gene dosage effects on mitochondrial mass and DNA.

In order to assess the effect of nuclear gene dosage on the regulation of mitochondria we have studied serial sections of a set of isogenic haploid and diploid cells of Saccharomyces cerevisiae, growing exponentially in the absence of catabolite repression, and determined the amount of mitochondrial DNA per cell. Mitochondria accounted for 14% of the cytoplasmic and 12% of the total cellular volume in all cells examined regardless of their ploidy or their apparent stage in the cell cycle. The mean number of mitochondria per cell was 22 in the diploid and 10 in the haploids. The volume distribution appeared unimodal and identical in haploids and diploids. The mitochondrial DNA accounted for 12.6 +/- 1.2% and 13.5 +/- 1.3% of the total cellular DNA in the diploid and haploid populations, respectively. These values correspond to 3.6 x 10(-15) g, 2.2 x 10(9) daltons, or 44 genomes (50 x 10(6) daltons each) per haploid and twice that per diploid cell. On this basis, the average mitochondrion in these cells contains four mitochondrial genomes in both the haploid and the diploid.