Susan J. Decker

Characterization of cortical secretory vesicles from the sea urchin egg

In order to evaluate the potential role of cortical vesicle exocytosis in modifying the egg surface at fertilization, we have begun characterization of the cortical secretory vesicle. Earlier reports (N. K. Detering, G. L. Decker, E. D. Schmell, and W. Lennarz, 1977, J. Cell Biol. 75, 899-914) have described the isolation of an egg cell surface complex which consists of the egg plasma membrane and cortical secretory vesicles. We have now developed a method of dissassembling the cell surface complex and isolating the cortical vesicles. The very low levels of contaminating plasma membrane in this preparation allow the meaningful comparison of plasma membrane and cortical vesicle proteins and lipids. The cortical vesicles were found to be rich in high-molecular-weight PAS-positive proteins. The majority of these glycoconjugates were solubilized by hypotonic lysis of the cortical vesicles and probably represent proteins sequestered inside the intact vesicles. The fatty acid composition of the cortical vesicles was found to be unusually high in arachidonic acid. The fatty acid composition of the cortical vesicles was closely similar to that of the plasma membrane; however, the cortical vesicles were substantially higher in cholesterol content.

Malignant granular cell tumor metastatic to the orbit

OBJECTIVE Malignant granular cell tumor is a rare type of soft tissue sarcoma. To our knowledge, ocular (eyelid) involvement has been described in only two cases. Herein, we report the clinicopathologic features of an unusual case of malignant granular cell tumor metastatic to the orbit. DESIGN Observational case report. METHODS Retrospective review of the medical record and the histopathologic and electron microscopic findings and review of the literature. RESULTS A 72-year-old man with biopsy-proven granular cell tumor in the cervical region was initially seen with proptosis and motility disturbance. A magnetic resonance imaging scan showed a large intraconal mass, and biopsy of the orbital mass revealed granular cell tumor. Histopathologic examination of the primary neck tumor and the orbital mass revealed increased nuclear atypia and pleomorphism in the consecutive lesions. The morphologic impression of granular cell tumor was also supported by the immunohistochemical demonstration of S-100 protein expression and ultrastructural findings typical of granular cell tumor. Six months after the orbital involvement, systemic workup revealed multiple apparent bony and lung metastases. CONCLUSIONS We report the first malignant granular cell tumor metastatic to the orbit and suggest the inclusion of this tumor in the differential diagnosis of metastatic orbital lesions.

Immunoblastic malignant lymphoma in dolphins: Histologic, ultrastructural, and immunohistochemical features

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APO2.7 defines a shared apoptotic–necrotic pathway in a breast tumor hypoxia model†

A breast tumor hypoxia model used to simulate conditions which may exist within an enlarging tumor was examined using documented methods for identifying mechanisms of cell death and compared to the mitochondrial membrane-specific APO2.7 antigen expression. Hypoxic conditions were induced by holding cell pellets of MDA-MB-175-VII breast carcinoma cells in tightly capped centrifuge tubes for up to 10 days. Cells were harvested at 1.5, 3, 4.5, 6, 12, 18, and 24 h, and each 24 h thereafter to 10 days. APO2.7 was monitored in unprocessed cells (no permeabilization prior to staining) for all time points and processed cells (permeabilized prior to staining) for only the first 24 h. Cell viability probes trypan blue and anti-tubulin antibody showed a rapid increase in staining over the first 24 h, as did the phosphatidylserine-specific annexin V and DNA fragmentation by flow cytometry (range of 60-81% positive staining). Light scatter changes indicative of cell death were also quite remarkable. APO2.7 staining never exceeded 42% of the cell pellet over the 10 days of testing compared to greater than 95% staining for all other methods tested. When APO2.7 antigen expression was examined with respect to depth in the cell pellet, it was apparent that cells deeper in the pellet expressed APO2.7 more rapidly; however, fewer cells stained and cells showed fewer apoptotic features on an ultrastructural level than cells at the cell media interface. The study indicates that the anti-APO2.7 antibody may be able to discern apoptotic and incomplete apoptotic cells from necrotic MDA-MB breast cancer cells, traversing a heterogeneous pathway to cell death induced by hypoxia.

Preparation of plasma membranes from fertilized sea urchin eggs

A new method is presented for preparation of highly purified plasma membranes from fertilized sea urchin eggs. The purified plasma membranes are in vesicle form and are highly enriched in ouabain inhibitable, Na+/K+ ATPase activity. Analysis of membrane proteins by sodium dodecyl sulfate-gel electrophoresis indicates that several high-molecular-weight proteins characteristic of plasma membranes from unfertilized eggs are absent in plasma membranes from fertilized eggs.

Non-apoptotic desquamation of cells from corneal epithelium: Putative role for Muc4/sialomucin complex in cell release and survival

Muc4/sialomucin complex (SMC), a large heterodimeric mucin composed of an extracellular mucin subunit ASGP‐1 and a transmembrane subunit ASGP‐2, is present at the rat ocular surface localized mainly to the most superficial layers of the epithelia. To investigate corneal homeostasis and the functions of Muc4/SMC at the ocular surface, we developed a corneal epithelial cell culture system from corneal explants, from which migrating cells formed an epithelial sheet resembling the native epithelium with regard to microanatomy, expression of characteristic markers, cell migration, and Muc4/SMC expression. Cells migrating from the explants expressed smooth muscle actin. Proliferation was detected only on the edge of epithelial sheet in the immature epithelium and throughout the sheet in confluent cultures. Microscopy revealed that the epithelial sheet was formed from four to six layers of cells expressing keratin 3 and Muc4/SMC in forms identical to those expressed at ocular surface in vivo. Electron microscopy showed cells in various morphological states in the process of releasing from the surface of the multilayer (desquamating). Surprisingly, few of these cells showed evidence of apoptosis, either by morphological or DNA fragmentation analyses. These results suggest a new model for desquamation from stratified epithelia, in which desquamation and apoptosis are independent and sequential processes. Desquamating cells also exhibit a high level of Muc4/SMC. Since Muc4/SMC has been shown to be a potent anti‐adhesive and a repressor of apoptosis, we propose that it plays a role in the non‐apoptotic desquamation process.

Trabecular meshwork alteration and intraocular pressure change following pulsed near-infrared laser trabeculoplasty in cats.

BACKGROUND AND OBJECTIVE To comparatively assess the safety and variation in intraocular pressure (IOP) of two pulsed near-infrared lasers (titanium:sapphire and alexandrite) for laser trabeculoplasty versus conventional blue-green argon laser trabeculoplasty in an animal model. MATERIALS AND METHODS The left eyes of 15 healthy cats received a 180 degree laser trabeculoplasty treatment: 5 with a titanium:sapphire laser, 5 with an alexandrite laser, and 5 with an argon laser. Preoperatively and postoperatively, all animals underwent tonometry, gonioscopy, and slit-lamp examination. The cats were observed up to 12 weeks. Scanning electron microscopy and histologic examination were performed to evaluate potential alterations in the trabecular meshwork structure. RESULTS IOP at 1 hour, 1 day, and 1 week following treatment was remarkably lower, irrespective of the laser source used. Following treatment with both near-infrared lasers, gonioscopy showed depigmentation underneath the area of the treated trabecular meshwork and histologic evaluation showed a decrease in pigment density. On scanning electron microscopy, damage to the trabecular meshwork structure could not be detected after treatment with near-infrared lasers. CONCLUSIONS Near-infrared laser trabeculoplasty was found to be effective to temporarily lower IOP in cats. The lasers selectively altered pigment-containing cells, avoiding structural damage of the trabecular meshwork anatomy.