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Biomedical subjects

M R Hernandez

Publications and source records attributed to M R Hernandez.

At least 55 records · Page 3Linked to original sources
PubMed1994

Localization of collagen type III mRNA in normal human optic nerve heads.

Previous studies using immunocytochemistry and electron microscopy have localized collagen type III (CIII) in the human optic nerve head (ONH). CIII is present in the core of the cribriform plates of the lamina cribrosa and increases with age. In this study, in situ hybridization was used to localize mRNA for CIII in specific cells in human fetal and adult ONH to determine its cellular origin and to provide temporal and regional information on the synthesis of CIII in the ONH. Human ONH from donors with no history of eye disease (16-24 weeks gestation and 6-75 years) were fixed, embedded and sectioned serially. Sections were hybridized with antisense riboprobe for CIII and controls with sense riboprobe and processed for ISH. Immunoperoxidase staining with antibodies against GFAP and von Willebrand factor was used to detect astrocytes and vascular endothelial cells respectively. Fetal ONH: CIII mRNA was localized in most blood vessels throughout the ONH in sections hybridized with antisense probe. The label was localized to endothelial cells of small vessels. Endothelial cells of central retinal vessels were not labeled. In the lamina cribrosa, CIII mRNA was localized to small vessels, but not to astrocytes. Fibroblasts of the peripapillary sclera were labelled with antisense probe. Young-Adult ONH: In the cribriform plates, lamina cribrosa cells hybridized the probe at all ages. Few astrocytes hybridized the probe. Some endothelial cells of small vessels were also labeled. In the insertion region, most vessels were labeled in young ONH, but very few, if any, in old adults. The apparent intensity of hybridization signal associated with blood vessels in adult ONH is markedly reduced with age. No hybridization was observed inside of nerve bundles consistent with the absence of mRNA inside axon and CIII in this location. Localization of mRNA for CIII to blood vessels in fetal lamina cribrosa suggests a vascular origin for CIII in the plates, and perhaps, for lamina cribrosa cells. In young and old lamina cribrosa, localization of CIII mRNA suggests lifelong synthesis of this protein in agreement with age-related increase of CIII in this tissue.

Adolescent↗
PubMed1993

Collagen type I mRNA levels in cultured human lamina cribrosa cells: effects of elevated hydrostatic pressure.

In primary open-angle glaucoma, elevated intraocular pressure is associated with distortion and reorganization of the connective tissue plates of the lamina cribrosa. We have previously postulated that the resident cells of the lamina cribrosa may respond to elevated intraocular pressure by altering the biosynthesis or degradation of extracellular matrix. To determine the response of lamina cribrosa cells to increased pressure, we have compared cultures of human lamina cribrosa cells, from five individuals, maintained under control and pressurized conditions in vitro. Cells from third to fifth passage cultures of human lamina cribrosa subjected to elevated hydrostatic pressure (50 mm Hg) for 7 days changed shape from flat and polygonal to elongated, synthesized and secreted increased amounts of collagen type I as shown by immunofluorescent localization, and exhibited increased mRNA levels of collagen type I (199 +/- 36% of control) (mean +/- S.D.), as determined by slot-blot hybridization. In contrast, beta-actin mRNA levels were unchanged, indicating that the effects of elevated pressure are probably relatively selective. Our data indicate that elevated pressure increases the synthesis of collagen Type I by human lamina cribrosa cells in vitro. In vivo, lamina cribrosa cells may react to changes in their environment by modulating, specifically, changes in the mRNA levels, production, and secretion of extracellular matrix macromolecules. The relationship of these changes in extracellular matrix to those observed in glaucoma remains to be determined.

Adolescent↗
PubMed1993

Glaucoma: changes in extracellular matrix in the optic nerve head.

Primary open angle glaucoma (POAG), the most common form of glaucoma, is characterized by irreversible loss of axons from the optic nerve. The site of damage to the axons is at the level of the lamina cribrosa in the optic nerve head. It has been hypothesized that structural and biochemical abnormalities in the extracellular matrix (ECM) of the lamina cribrosa underlie the progressive compressive and remodelling of this connective tissue in glaucoma. In this review, we present evidence of specific changes in collagen and elastic fibers, major ECM components in the lamina cribrosa of glaucomatous eyes.

Collagen↗
PubMed1992

Ultrastructural immunocytochemical analysis of elastin in the human lamina cribrosa. Changes in elastic fibers in primary open-angle glaucoma.

The elastic fiber consists of several components: a central core of alpha-elastin and a microfibrillar sheath containing three components: fibrillin, microfibril-associated glycoprotein, and a 35-kD protein with amine oxidase activity. Elastin is a major component of the elastic fibers of the extracellular matrix (ECM) of the lamina cribrosa, and elastic fibers undergo marked changes in primary open-angle glaucoma (POAG). These changes, as demonstrated previously, include loss and fragmentation of elastic fibers at the bottom of the glaucomatous cup and disorganization in the peripheral walls of the cup. The author characterized the changes in elastic fibers with age and in POAG at the ultrastructural level, using colloidal gold immunostaining and anti-human alpha-elastin antibody. In fetal eyes, there was no detectable elastin in the ECM of the lamina cribrosa. In infant eyes, elastin was present in microfibrillar aggregates in the core of the plates. In young adults, thin elastic fibers were present that ran longitudinally in the core of the plates. With age, elastic fibers become thicker, tubular, and surrounded by densely packed collagen fibers. In mild POAG, tubular elastic fibers no longer were identifiable. Fragments of elastic fibers and microfibrillar aggregates stained positively for elastin suggested new synthesis of elastin that was not organized into tubular elastic fibers. In advanced POAG, masses of nonfibrillar elastin-positive material had a spotted appearance. Throughout the cribriform plates, there was a loss of collagen fibers, proliferation of basement membranes, and bundles of elastin-negative microfibrils not associated with collagen or elastic fibers. The progression of marked changes in elastic fibers and the disorganization of the ECM of the lamina cribrosa was associated with the loss of function and continuous remodeling of the optic nerve head in POAG.

Adolescent↗
PubMed1991

Localization of collagen types I and IV mRNAs in human optic nerve head by in situ hybridization.

Using in situ hybridization, individual cells expressing mRNAs for collagen types I and IV were localized in fixed-tissue sections of adult and fetal human optic nerve heads. Astroglial cells lining the cribriform plates and cells inside the cribriform plates of the lamina cribrosa had mRNA for collagen type IV. Cells in the glial columns, pial septa, and vascular wall also contained mRNA collagen type IV. Collagen type I mRNA was expressed by cells of the cribriform plates of the lamina cribrosa of adults. Few cells in the glial columns, pial septa, and blood vessels had mRNA for collagen type I. Scleral fibroblasts contained mRNA for collagen type I. These results indicated that the expression of mRNA for both collagen types I and IV paralleled the localization of these extracellular matrix proteins in the optic nerve head and suggested that both collagen types were synthesized in this tissue throughout life.

Adolescent↗
PubMed1990

Changes in the extracellular matrix of the human optic nerve head in primary open-angle glaucoma.

Using immunofluorescent staining, we were able to characterize the changes in composition and distribution of the macromolecules making up the extracellular matrix of the lamina cribrosa of the glaucomatous human optic nerve head. In tissue adjacent to the glaucomatous cups, there was marked disorganization and loss of fibers of elastin within the cores of the cribriform plates. Collagen type VI, normally sparse, increased in quantity considerably throughout the lamina cribrosa in glaucomatous eyes with all degrees of damage. Collagen type IV and other basement membrane macromolecules appeared to extend into nerve bundles, presumably filling in spaces previously occupied by nerves. There was no appreciable change in the postlaminar region, which indicates the specificity of the extracellular matrix changes in the lamina cribrosa. Our results indicate that changes in the extracellular matrix play an important role in the progression of the glaucomatous process and may be a causative agent of the disease.

Aged↗
PubMed1989

Age-related changes in the extracellular matrix of the human optic nerve head.

By using immunofluorescent staining we were able to characterize the age-related changes in the macromolecules making up the extracellular matrix of the lamina cribrosa of the human optic nerve head. As the cores of the cribriform plates enlarged with age, there were age-related increases in the apparent density of collagen types I and III and elastin that constituted the connective tissue support of the nerve bundles. Collagen type IV coated the cribriform plates as basement membranes and was also present within the cores as a fine filamentous network, which increased in density with age as the cribriform plates expanded. As this tissue ages, individual differences leading to more or less of a particular macromolecule of the extracellular matrix may alter the support function of the lamina cribrosa and influence the degeneration of the optic nerve associated with glaucoma.

Aging↗
PubMed1988

Cell culture of the human lamina cribrosa.

The extracellular matrix of the lamina cribrosa may be important in the changes in the optic nerve head associated with glaucoma. To investigate the cell biology of this tissue, human lamina cribrosa was explanted in tissue culture and two cell types grown from this tissue were characterized. The most common cell type obtained was a large, flat, polygonal cell which was negative for glial fibrillar acidic protein (GFAP) and could be serially subcultured. This cell type synthesized collagens type III and type IV, fibronectin and elastin. Much less commonly grown was a cell type with conspicuous long processes and which was positive for GFAP. This presumed astrocyte synthesized collagen type IV and fibronectin. Fibroblastic cells were not obtained from this tissue but were easily grown from sclera. The cells that we have cultured from the human lamina cribrosa may produce the extracellular matrix present in the cribriform plates of this tissue and be important in the glaucomatous process.

Adult↗
PubMed1987

Extracellular matrix of the human lamina cribrosa.

We used immunoperoxidase staining and double immunofluorescent staining to demonstrate the macromolecular components of the extracellular matrix of the lamina cribrosa from young human donors. The cribriform plates were made up of a core of elastin fibers with a sparse, patchy distribution of collagen type III. The plates were coated with collagen type IV and laminin; these basement membrane components were presumably made by the astrocytes that were distributed on the surfaces of the plates. The insertion of the lamina cribrosa in the sclera was made up of concentric, circumferential elastin fibers that surrounded the lamina cribrosa and were continuous with the elastin in the cribriform plates. Astrocytic processes extended into the bundles of elastin fibers, whereas the basement membrane components extended into the sclera. The mechanical properties of the macromolecules of the extracellular matrix of the lamina cribrosa may make this tissue compliant and sensitive to intraocular pressure. Perhaps individual differences in the macromolecular components of this tissue contribute to the glaucomatous changes in the optic nerve head.

Adolescent↗
PubMed1987

Human trabecular meshwork cells in culture: morphology and extracellular matrix components.

This study demonstrates the presence of laminin and collagen type IV in the extracellular space of human trabecular meshwork (HTM) cells in culture and its absence in cultures of fibroblasts from sclera adjacent to the outflow pathway. These basement membrane components can be detected by standard immunohistochemical techniques. Positive staining for these macromolecules is found only after the HTM cells have reached confluence. The presence of laminin and human collagen type IV in the early passages can serve as additional criteria for identification of HTM cells in culture. These cells provide a useful experimental system for studying the effects of drugs and other factors on the synthesis of components of the extracellular matrix.

Adolescent↗
PubMed1986

Extracellular matrix of the human optic nerve head.

Double-antibody immunofluorescent studies of sectioned human optic nerve head indicated the marked presence of collagen type IV and laminin in the extracellular matrix of the lamina cribrosa. These macromolecules were layered transversely across the nerve fascicles and appeared to constitute the cribriform plates. Relatively little collagen types III and I were present in the extracellular matrix of this tissue and fibronectin was not detected in appreciable amounts. These results indicated that the lamina cribrosa contains a specialized extracellular matrix of the central nervous system made up of plates of material resembling basement membrane. The major macromolecular components of the lamina cribrosa do not resemble those of sclera.

Adult↗
PubMed1986

Potentiation of collagen synthesis in explants of the rabbit eye by 5 beta-dihydrocortisol.

The biologic effect of 5 beta-dihydrocortisol on collagen synthesis was evaluated. The metabolite was found to potentiate subthreshold levels of dexamethasone in increasing 3H-proline incorporation in cells of the outflow region of the rabbit. Digestion of the tissue with highly purified collagenase indicated that the 3H-proline was incorporated into collagen type protein. This study demonstrates another biologic activity of 5 beta-dihydrocortisol, a metabolite found to accumulate in cells cultured from trabeculectomy specimens from patients with primary open angle glaucoma.

Animals↗
PubMed1985

The effect of dexamethasone on the synthesis of collagen in normal human trabecular meshwork explants.

This study demonstrates that the trabecular meshwork cells of the human eye incorporate 3H-Proline into collagen during in vitro incubation. Addition of dexamethasone to the incubation mixture produced a marked decrease in this incorporation. Dexamethasone was active at 10(-8) M and higher concentrations. The specificity of the hormone effect was demonstrated by its inability to alter 3H-leucine incorporation in these cells. These results indicate that dexamethasone decreases the synthesis of collagen, a major component of the extracellular matrix, in the trabecular meshwork.

Collagen↗
PubMed1983

Glucocorticoid target cells in human outflow pathway: autopsy and surgical specimens.

Target cells for glucocorticoids were found in trabeculectomy specimens obtained surgically from humans with glaucoma (primary and secondary) and from nonglaucomatous autopsy eyes using an autoradiographic technique. Specific nuclear localization of 3H-dexamethasone was found in cells of the trabecular meshwork, scleral fibroblasts, and in the endothelial lining of both Schlemm's canal and the outflow vessels. Glucocorticoids may alter the outflow facility by a direct effect on the metabolism of these cells. The autoradiographic method is suitable for studying competitors of glucocorticoid binding in small surgical specimens of human tissue involved in the outflow of aqueous humor.

Aqueous Humor↗
PubMed1983

The effect of dexamethasone on the in vitro incorporation of precursors of extracellular matrix components in the outflow pathway region of the rabbit eye.

The present study demonstrates that the cells of the outflow pathway region incorporate precursors of the extracellular matrix components during in vitro incubation. Addition of dexamethasone (10(-7) M) to the incubation mixture led to changes in precursor incorporation that were precursor and cell specific. The steroid significantly decreased the incorporation of 3H-glucosamine and increased the incorporation of 3H-proline in the outflow pathway cells and in the fibroblasts of the adjacent sclera. The specificity of the hormone effect was demonstrated by its inability to alter 3H-leucine incorporation. By contrast, in iridial fibroblasts dexamethasone caused a decreased incorporation of 3H-glucosamine while not affecting the incorporation of 3H-proline. Our results suggest that dexamethasone causes an increase in collagen synthesis in the target cells most closely related to the outflow pathway. Further, the decreased incorporation of 3H-glucosamine with dexamethasone suggests decreased synthesis of glycosaminoglycans, glycoproteins or possibly glycolipids in these same cells.

Animals↗