Apoptosis in cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy.

To test the hypothesis that an apoptotic process plays a role in the pathogenesis of cerebral lesions in cerebral autosomal-dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), we examined samples from frontal, temporal, insular, and occipital regions, basal ganglia, and cerebellum from 4 patients with CADASIL, 2 with Binswanger disease, and 3 controls. Apoptotic cells were identified using in situ end labeling and activated caspase 3 immunostaining. Immunolabeling for Notch3, the beta-amyloid protein precursor, and phosphorylated neurofilament protein was performed on successive sections. Apoptosis of vascular cells was markedly increased in status cribrosus in CADASIL, both in basal ganglia and subcortical white matter, suggesting that concomitantly with Notch3 deposition it may play a causative role in the dilatation of Virchow-Robin spaces. Neuronal apoptosis was found in CADASIL, mostly in cortical layers 3 and 5. Its severity correlated semiquantitatively with the extent of ischemic lesions and axonal damage in the underlying white matter. It was more severe in demented patients. Only occasional apoptotic neurons were found in the Binswanger cases and none in the controls. This supports the view that neuronal apoptosis may contribute to cortical atrophy and cognitive impairment in patients with CADASIL and that it may, at least partly, result from axonal damage in the underlying white matter.

Corneal epithelial cultures generated from organ-cultured limbal tissue: factors influencing epithelial cell growth.

PURPOSE: To explore the in vitro proliferative potential of human limbal epithelial cells after 31 degrees C organ-culture storage and to investigate putative factors influencing it. METHODS: 185 cultures of limbal explants were carried-out either from full-thickness explants (n = 102) or from enzymatically dissociated cells (n = 83) seeded on a feeder layer of human keratocytes. Epithelial outgrowth was assessed by phase contrast microscopy using a computerized image analysis software. Cell phenotype was evaluated by transmission electron microscopy and immunocytology. Univariate and multivariate analysis were performed to determine factors influencing epithelial growth in culture. RESULTS: An epithelial outgrowth of 100 square mm or more was observed in 52% of cultures, (average growth area: 440 +/- 256 mm at three weeks). Corneal epithelial phenotype was confirmed by transmission electron microscopy, and cytokeratin pattern. Cytokeratine 19, deltaNp63, nestin and vimentin positive staining revealed undifferentiated epithelial cells in both explant and cell suspension cultures at three weeks. Short death to cornea retrieval time (p < 0.03) and female donors (p < 0.01) were associated with higher cell growth. Enzymatic treatment of explants by trypsin, but not dispase, decreased cell proliferation at two (p < 0.03) and three weeks (p < 0.04). Donor age, duration of corneal storage, and source of the explant did not influence the cell growth. CONCLUSION: Organ-culture conditions can preserve limbal cell mitotic potential if limbal tissue is excised early after circulatory arrest. Human keratocytes can be used as a feeder layer allowing epithelial cells to maintain poorly differentiated phenotype in culture. Further investigations are needed to explain the influence of the donor sex on epithelial cell growth in culture.

Cellular and molecular tunnels surrounding the forebrain commissures of human fetuses.

Glial cells and extracellular matrix (ECM) molecules surround developing fiber tracts and are implicated in axonal pathfinding. These and other molecules are produced by these strategically located glial cells and have been shown to influence axonal growth across the midline in rodents. We searched for similar cellular and molecular structures surrounding the telencephalic commissures of fetal human brains. Paraffin-embedded brain sections were immunostained for glial fibrillary acidic protein (GFAP) and vimentin (VN) to identify glial cells; for microtubule-associated protein-2 (MAP-2) and neuronal nuclear protein (NeuN) to document neurons; for neurofilament (NF) to identify axons; and for chondroitin sulfate (CS), tenascin (TN), and fibronectin (FN) to show the ECM. As in rodents, three cellular clusters surrounding the corpus callosum were identified by their expression of GFAP and VN (but not MAP-2 or NeuN) from 13 to at least 18 weeks postovulation (wpo): the glial wedge, the glia of the indusium griseum, and the midline sling. CS and TN (but not FN) were expressed pericellularly in these cell groups. The anterior commissure was surrounded by a GFAP+/VN+ glial tunnel from 12 wpo, with TN expression seen between the GFAP+ cell bodies. The fimbria showed GFAP+/VN+ cells at its lateral and medial borders from 12 wpo, with pericellular expression of CS. The fornix showed GFAP+ cells somewhat later (16 wpo). Because these structures are similar to those described for rodents, we concluded that the axon guiding mechanisms postulated for commissural formation in nonhuman mammals may also be operant in the developing human brain.

Corneal keloid: clinical, ultrasonographic, and ultrastructural characteristics.

A 70-year-old man was referred to us with a 2-year, progressive, painless decrease in visual acuity in the right eye. Ocular history included extraction of a traumatic cataract with a transclerally fixated posterior chamber intraocular lens. Slitlamp examination showed a raised, white, vascularized mass covering the cornea. The lesion was removed by superficial lamellar keratectomy. Light microscopy examination confirmed the diagnosis of corneal keloid. These uncommon lesions usually develop in adults after corneal traumas, surgery, or inflammatory processes. They have also been described in children with Lowe's syndrome, Rubinstein-Taybi syndrome, and other ocular developmental disorders.

Candida parapsilosis keratitis.

PURPOSE: To present clinical, microbiologic, and histopathologic features of keratitis due to Candida parapsilosis. METHODS: Clinicomicrobiologic evaluation of four patients (four eyes) with culture-proven C. parapsilosis keratitis. The patients were evaluated for symptoms, visual acuity, clinical observations, microbiologic examination of corneal scrapings, and pathologic examination of corneal buttons. RESULTS: Three cases were observed after penetrating keratoplasty, and one case occurred after inhalation of corticosteroids. Clinical presentation of C. parapsilosis keratitis showed a great diversity. There was one case of crystalline keratopathy and three cases of suppurative corneal infiltrate. Histopathology of corneal buttons showed interlamellar accumulations of yeast. Medical treatment included topical amphotericin B and systemic triazoles. Penetrating keratoplasty was required in three patients. CONCLUSION: Risk factors for C. parapsilosis keratitis may include corticosteroid use and prior corneal transplantation. The prognosis of C. parapsilosis keratitis with antifungal and surgical therapy may vary from good visual outcome to intraocular extension with phthisis bulbi.

[Amniotic membrane transplantation for the treatment severe acanthamoeba keratitis]. / Intérêt de la greffe de membrane amniotique dans le traitement des kératites amibiennes sévères.

BACKGROUND: It is now possible to treat ocular surface disorders by means of amniotic membrane transplantation. We performed a study to determine the efficacy of this technique in the treatment of severe Acanthamoeba keratitis. METHODS: We studied six patients with severe, painful, nonhealing Acanthamoeba keratitis who underwent one or two amniotic membrane transplantation procedures between February 2001 and January 2003. Histopathological analysis of the corneal buttons was performed in four cases. RESULTS: Eight amniotic membrane transplantation procedures were performed. The mean length of follow-up was 14 (range 3-21) months. The mean interval between institution of medical treatment and the procedure was 3.6 months. All patients had progressive stromal lesions caused by an inflammatory reaction. Complete reepithelialization occurred in four cases, and partial healing in two cases. Ocular inflammation and tissue destruction were decreased in all cases, pain was lessened in five cases, and corneal neovascularization was decreased in four cases. No postoperative complications were observed. Amniotic membrane was observed under dysplastic corneal epithelium on histologic examination. INTERPRETATION: Amniotic membrane transplantation may be a safe and effective treatment of severe Acanthamoeba keratitis, particularly during the inflammation phase. It may permit penetrating keratoplasty to be delayed.