Tryptophan Raman/457.9-nm-excited fluorescence of intact guinea pig lenses in aging and ultraviolet light.

Normal age-matched guinea pig lenses were compared with those exposed to (1) long-term ultraviolet (UV) light (9 months, 353-nm peak) in vivo, and (2) short-term UV light (3.5 hr, 325 nm) in vitro from a helium-cadmium laser. Tryptophan Raman and 457.9-nm-excited fluorescence profiles along the visual axis (VA) were obtained by taking 21 (for tryptophan) or 11 (for fluorescence) successive spectra for each intact lens using the Raman optical dissection technique. To indicate the extent of UV exposure, fluorescence spectra were obtained along the VA (excitation/emission = 457.9/497 nm); these spectra indicated that the major alteration by UV was in the nucleus with the least in the posterior cortex. Normal aging lenses had no apparent change in the tryptophan profile between 3 days and 12 months. The UV-irradiated lenses also showed no appreciable difference from the normal aging patterns. These results indicate that there is no detectable tryptophan photolysis in the intact guinea pig lens by longwave UV light.

Anterior vs posterior intraocular melanoma. Metastatic differences in a murine model.

OBJECTIVE: To study differences in metastatic rate of anterior vs posterior ocular melanoma in a murine model. METHODS: Thirty-eight 12-week-old C57BL6 mice were inoculated into the anterior chamber (AC) or posterior compartment (PC) of the right eye with 5 x 10(5) tissue culture Queens melanoma cells per 5 microL. The right eye was enucleated 14 days after inoculation, and the animal was killed 28 days after inoculation and a necropsy was performed. The eye was evaluated for the presence of melanoma and the mean of the 10 largest nucleoli in tumor cells. Eyes with orbital invasion of the melanoma were excluded. The number of metastases was determined at necropsy. RESULTS: Melanomas grew in 30 of 38 eyes. After exclusion of 9 eyes with orbital invasion of tumor, melanoma was found in 12 mice in the AC group and 9 mice in the PC group. The metastatic rate was significantly lower for AC tumors (33%) than for PC tumors (89%) (P = .02). All AC tumors that metastasized to lungs also metastasized to ipsilateral cervical lymph nodes, and no PC tumors metastasized to ipsilateral cervical lymph nodes. The median number of pulmonary metastases per tumor was significantly smaller for AC tumors than for PC tumors (P = .01) There was a median of 0 metastases (range, 0-14) per AC tumor and 4 metastases (range, 0-38) per PC tumor. CONCLUSION: Posterior ocular melanoma metastasizes more often than anterior ocular melanoma in this murine model.

Murine model of anterior and posterior ocular melanoma.

Iris melanomas are less likely to metastasize than ciliary body or choroidal melanomas. In order to study this difference in metastatic rate, we developed a murine model of anterior chamber (AC) and posterior compartment (PC) melanoma. Eighteen C57BL6 mice were inoculated in the AC (n = 6) or PC (n = 12) with B16F10 melanoma cells and eleven mice were inoculated in the AC (n = 3) or PC (n = 8) with Queens melanoma cells. The animals were sacrificed at 12 to 14 days post inoculation and histologically examined. Results were that 8 of 9 AC tumors and 10 of 20 PC tumors grew. One PC tumor metastasized to the lungs. This model may be used to study anterior versus posterior ocular melanoma differences.

The Emory mouse cataract: changes in the beta and gamma-crystallins during aging and cataractogenesis as revealed by isoelectric focusing of the native soluble proteins.

Isoelectric focusing on ultra-thin polyacrylamide gels was employed to study alterations in the distribution patterns of soluble beta and gamma-crystallins during the development of cataract in the Emory (EM) mouse. These alterations were compared with corresponding changes occurring in clear control lenses of the same age from the cataract-resistant (CR) strain. At two months of age both strains gave similar patterns. At four and fourteen months the EM lens showed depressed beta peaks compared to the CR lens. At fourteen months the EM lens had reduced gamma 2 and gamma 5 compared with the CR lens. At twenty-four months the EM lens had all gamma's markedly reduced, especially gamma 5, with the complete disappearance of gamma 4 which was however still present in the CR lens at twenty-six months. At twenty-four months the EM lens had a relatively large amount of beta 5 but no beta 5a or beta 5b; the latter two were present in the CR lens surrounding beta 5 which was less prominent than in the EM lens. The decrease in gamma's in the twenty-four months EM lens was approximately equalled by an apparent increase in beta's. Of the above changes only the decrease in beta's at four months can be considered precataractous and perhaps a primary change in cataractogenesis. The later changes in both beta and gamma may be secondary although they are certainly at least associated with cataractogenesis. The invariable decrease in gamma with age is noticeably accelerated in cataractogenesis.

Raman spectroscopic evaluation of aging and long-wave UV exposure in the guinea pig lens: a possible model for human aging.

The laser Raman optical dissection technique makes it possible to study individual points of minute volumes (2 X 10(-3) microliter) in the intact living lens in vitro. This technique was used to measure the sulfhydryl and disulfide content of 21 distinct points along the visual axis of the guinea-pig lens after aging and long-wave ultraviolet exposure (9-month duration in vivo). To facilitate comparison between different lenses, data was compiled as the intensity ratio of sulfhydryl (2580 cm-1) to a protein reference signal (2731 cm-1) or disulfide (508 cm-1) to phenylalanine (622 cm-1). These 21 ratios for each experiment were plotted as a function of the distance of the point from the nuclear center of the lens to give a visual axis profile. From these profiles we have found that the loss of sulfhydryl can be accelerated in the guinea-pig lens by in vivo ultraviolet exposure (353 nm peak from an incoherent source) for nine months. There is also a subsequent uniform increase in the disulfide content across the visual axis after UV exposure suggesting a direct sulfhydryl to disulfide conversion in the guinea-pig lens.

A biomechanical evaluation of rabbit corneas in M-K and Dexsol.

We compared the average force required to separate normal corneas at a 50% stromal depth, with the force required to separate corneas stored for 2, 5, 7, 10, and 14 days in either McCarey-Kaufman or Dexsol corneal storage medium. The required interlamellar separation force was calculated by standardizing the width of the test strips. The average required force for 35 fresh rabbit corneas (no storage) was 9.1 +/- 1.5 g/mm. There was no significant change in the required separation force after storage in either medium for up to 14 days.

Distribution of two metabolically related fluorophors in human lens measured by laser microprobe.

Automated Raman microprobe spectrometry revealed the distribution of a major fluorophor, 3-OH-L-kynurenine-O-beta-glucoside, in human lenses from 0.38 to 71 yr. A three-dimensional perspective grid map with fluorescence intensity as the third dimension shows maximum fluorescence in the infant lens nucleus. At 12 yr the fluorescence peak is broadened and a toroid-shaped maximum occurs also in the outer cortex, creating a toroid-shaped minimum between the two maxima. By 71 yr the nuclear maximum is lower but a new (green) fluorophor (excitation 488 nm: emission 530 nm) has appeared as a toroidal maximum in the same location as the blue minimum, suggesting the conversion of the blue fluorophor to the unidentified green fluorophor.