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Biomedical subjects

J Hassell

Publications and source records attributed to J Hassell.

At least 19 recordsLinked to original sources
PubMed1998

Proteoglycan distribution during healing of corneal stromal wounds in chick.

Proteoglycan distribution during corneal stromal healing in growing corneas of young chicks were histologically and immunohistochemically analysed. Single linear incisions to produce partial-thickness wounds were made in the corneas of 5 day old chicks. The corneas were harvested at different times after wounding and processed for either histochemical analyses using periodic acid-Schiff's reaction (PAS) or for indirect immunofluorescence analyses of lumican, keratocan, keratan sulfate, perlecan and laminin. Linear corneal stromal incisions were completely covered by migrated stratified epithelium by day 2 post wounding and resulted in a gaping wound with a thinner stroma. New stromal scar tissue formed between the epithelium and the original stroma that resulted in partial restoration of stromal thickness. During the first two to three weeks of healing, the stromal tissue filling the depression formed from the gaping wound, was hypercellular and PAS positive, indicating significantly higher levels of glycoprotein content but no new Bowman's membrane was formed. By four weeks, the scar tissue occupied a 2-3 mm wide region. Immunofluorescence analyses indicated that other major differences in the healing and normally growing stroma were the increased synthesis and deposition of perlecan and laminin. No differences were evident in the immunofluorescence for keratocan or keratan sulfate in the scar tissue, but the scar tissue did contain markedly decreased levels of lumican. Thus, the regulation of proteoglycan and glycoprotein synthesis is altered in the keratocytes that are recruited to the wounded regions in the growing corneal stroma of post-hatched young chicks. While synthesis and deposition of adhesive molecules including laminin and perlecan are elevated, the synthesis of one of the keratan sulfate proteoglycans, lumican, is reduced in the scar tissue as compared to the normally growing stroma.

Animals↗
PubMed1998

A Rho-associated protein kinase: differentially distributed in limbal and corneal epithelia.

PURPOSE: The authors have developed monoclonal antibodies (mAbs) to characterize the sequential biochemical changes in corneal epithelial cells after they differentiate from stem cells, located in the limbus, and migrate centripetally to follow the pathway of terminal differentiation. The purpose of this study was to identify a protein (recognized by mAb HE1/11F) with increased expression associated with the transition of the limbal epithelium to corneal epithelium. METHODS: The distribution and identification of the protein(s) were performed using an indirect immunohistochemical staining technique and a western blot analysis, respectively. A rabbit corneal epithelial cDNA library, constructed in the Uni-Zap XR vector, was screened with mAb HE1/11F to select cDNA clones expressing polypeptide(s) recognized by this mAb. Additional overlapping cDNA clones were obtained from a primer extension cDNA library to determine the sequence of the complete open reading frame encoding the protein recognized by mAb HE1/11F. RESULTS: Rabbit corneal epithelium exhibited strong immunostaining with mAb HE1/11F, however, the limbal epithelial cells stained weakly. HE1/11F recognized 160-kDa (HEBM1) and 100-kDa (HEBM2) polypeptides in the corneal epithelial extracts. The amino acid sequence of the protein deduced from the nucleotide sequence of the cDNA exhibited a close homology to that of a RhoA (Ras-related small GTPase)-associated serine-threonine kinase (ROCK-I or Rho-associated coiled-coil kinase). A 160-kDa RhoA-binding polypeptide with a molecular mass similar to that of HEBM1 and ROCK-I was detected in the corneal epithelial extracts. These findings strongly suggested that HEBM1 was rabbit ROCK-I. The identity of HEBM1 was further confirmed from the reactivity of mAb HE1/11F with ROCK-I immunoprecipitated from rabbit corneal epithelial extracts using anti-ROCK-I antibodies. CONCLUSIONS: The increased expression of a protein identified as ROCK-I from cDNA analyses is associated with rabbit corneal epithelial differentiation and transition from the limbal to corneal surface. Therefore, a RhoA signaling pathway is likely to be associated with corneal epithelial differentiation (maturation). A close homology among the cDNA sequences of rabbit, mouse, rat, and human ROCK-I indicates that this RhoA-associated kinase is a well-conserved protein.

Amino Acid Sequence↗
PubMed1997

Non-glycosaminoglycan bearing domains of perlecan and aggrecan influence the utilization of sites for heparan and chondroitin sulfate synthesis.

Perlecan and aggrecan are proteoglycans that receive primarily heparan sulfate and chondroitin sulfate side chains, respectively. Their large multidomained core proteins have little or no homology to each other and their glycosaminoglycan (GAG) attachment sites are restricted to certain domains only. We examined the involvement of the non-GAG bearing domains in designating the GAG type added to the GAG attachment domain by preparing cDNA constructs that expressed perlecan/aggrecan chimeras as recombinant products in COS-7 cells and then determining the size and GAG composition of the recombinant products. The results showed that domain I of perlecan receives primarily (73-81%) heparan sulfate when coupled with domain II and III of perlecan, but when coupled with the G3 domain of aggrecan, it receives primarily (59-63%) chondroitin sulfate. Furthermore, the chondroitin sulfate attachment region of aggrecan received GAG side chains more readily when coupled to the G3 domain of aggrecan than when coupled to domains II and III of perlecan. The GAG side chains on all these recombinant products were small and similar in size. These findings indicate that the utilization of attachment sites for heparan and chondroitin sulfate or the sulfation of these GAGs can be influenced, in part, by non-GAG bearing domains.

Aggrecans↗
PubMed1996

Expression pattern of collagen IX and potential role in the segmentation of the peripheral nervous system.

Segmentation of the peripheral nervous system of vertebrates requires guidance cues located in the adjacent somitic mesoderm. Recent experiments suggest that inhibitory molecules in the posterior somite may influence segmentation by restricting the outgrowth of axons and the migration of neural crest cells to the anterior somite. A potential candidate for an inhibitory molecule is collagen IX, a chondroitin sulfate proteoglycan made by sclerotome cells of the somite and by the notochord. Immunohistochemical localization of collagen IX demonstrated that its expression in the posterior sclerotome of the somite correlates with axon outgrowth and neural crest cell migration through the anterior sclerotome. In vitro, sensory neurites on fibronectin, and motor neurites on basal lamina extract, avoid regions which contain substrate-bound collagen IX. This effect can be abolished by chondroitinase treatment, suggesting that the glycosaminoglycan component of the molecule is responsible for this activity. Further, collagen IX elicits a similar avoidance behavior by neural crest cells in vitro. These data suggest that collagen IX contributes to the segmentation of the peripheral nervous system in vivo.

Animals↗
PubMed1989

The need for enhancers in gene expression first appears during mouse development with formation of the zygotic nucleus.

Microinjection of the firefly luciferase gene coupled to a thymidine kinase (tk) promoter provided a quantitative assay to evaluate the requirements for gene expression in individual mouse oocytes and embryos. Polyoma virus (PyV) enhancers had no effect on the level of gene expression or competition for transcription factors as long as the DNA remained either in the oocyte germinal vesicle or the pronuclei of one-cell embryos. Expression of injected genes could be observed in pronuclei because the signal that normally triggers zygotic gene expression in two-cell embryos still occurred in one-cell embryos arrested in S phase. However, when the tk promoter was injected into zygotic nuclei of two-cell embryos, enhancers increased the number of embryos that expressed luciferase as well as the level of luciferase activity per embryo. PyV enhancer mutation F101, selected for growth in mouse embryonal carcinoma F9 cells, stimulated expression in developing two-cell embryos about seven times better than the wild-type PyV enhancer and competed effectively for factors required for transcription. These results were consistent with the fact that enhancers are required to activate the PyV origin of DNA replication in developing two-cell embryos but not in one-cell embryos. The maximum levels of gene expression in oocytes, one-cell embryos, and developing two-cell embryos (1:67:21) were inversely related to the extent of chromatin assembly, but the need for enhancers was independent of chromatin assembly. Therefore, it appears that the need for enhancers to activate promoters or origins of replication results from some negative regulatory factor that first appears as a component of zygotic nuclear structure.

Animals↗
PubMed1988

The presence of heparan sulfate proteoglycans in the neuritic plaques and congophilic angiopathy in Alzheimer's disease.

Two immunocytochemical probes were used to specifically identify and localize heparan sulphate proteoglycans (HSPGs) in 17 cases of Alzheimer's disease (AD). A monoclonal (HK-102) and an affinity-purified polyclonal antibody, each recognizing specific domains on the protein core of a basement membrane-derived HSPG, localized HSPGs to the amyloid fibrils present in neuritic plaques (NPs) and congophilic angiopathy (CA) in the brains of Alzheimer's patients, with weak to no immunostaining in neurofibrillary tangles from the same tissues. HSPGs were also demonstrated in "primitive plaques," suggesting that their accumulation takes place during early stages of plaque development. Immunolocalization of HSPGs to subsets of astrocytes and neuronal cells, particularly those in close proximity to NPs and CA, suggested possible involvement of these two cell types in deposition of HS-PGs into the amyloidotic lesions. The current study not only identifies a new component (HSPGs) present in the amyloid deposits of NPs and CA but also suggests that astrocytes, neurons, or both may be involved in its deposition at these sites.

Alzheimer Disease↗
PubMed1987

Influence of a reconstituted basement membrane and its components on casein gene expression and secretion in mouse mammary epithelial cells.

When primary mouse mammary epithelial cells are cultured on plastic, they rapidly lose their ability to synthesize and secrete most milk proteins even in the presence of lactogenic hormones, whereas cells cultured on released type I collagen gels show greatly enhanced mRNA levels and secretion rates of beta-casein and of some other milk proteins. We show here that culture on a reconstituted basement membrane from Engelbreth-Holm-Swarm tumor (EHS) allows greater than 90% of cells to produce high levels of beta-casein. By comparison, 30-40% of cells on released type 1 gels and only 2-10% of cells on plastic express beta-casein after 6 days in culture. Because only 40% of cells from late pregnant gland produced beta-casein before culture, the EHS matrix can both induce and maintain an increased level of casein gene expression. Individual basal lamina components were also evaluated. Type IV collagen and fibronectin had little effect on morphology and beta-casein mRNA levels. In contrast, both laminin and heparan sulfate proteoglycan increased beta-casein mRNA levels (1.5- to 4-fold and 2- to 8-fold, respectively). However, for heparan sulfate proteoglycan, increased message was not accompanied by increased secretion of beta-casein. Profound morphological differences were evident between cells cultured on plastic and on EHS matrix, the latter cells forming ducts, ductules, and lumina and resembling secretory alveoli. These results emphasize the vital role of the extracellular matrix in receiving and integrating structural and functional signals that can direct specific gene expression in differentiated tissues.

Animals↗
PubMed1975

Female homosexuals' concepts of self, men, and women.

Twenty-four female homosexuals and 24 female heterosexuals matched for age and education were given the Draw-A-Person Test (DAP) and, on the Gough-Heilbrun Adjective Check List (ACL), were asked to describe themselves, men in general, and women in general. Results indicated that homosexuals, relative to the heterosuxual controls: (a) sexualized their drawings (p smaller than .02) and embellished the female figure more (p smaller than .05); (b) showed no evidence of confusion in gender identity on the DAP or ACL; (c) scored sifnigicantly higher on Autonomy, Change, and Exhibition on the ACL; and (d) scored significantly lower on Abasement, Defensiveness, Deference, Endurance, Order, Personal Adjustment and Self-control on the ACL. It was concluded that the female homosexual may be more independent, changeable, and sexually preoccupied and less well adjusted than her heterosexual counterpart.

Adult↗
PubMed1975

Group/audiovisual instruction for patients with diabetes. Learning achievements and time economics.

Patients with diabetes receiving instruction in group classes utilizing audiovisual teaching techniques achieved significantly higher post-test scores than those taught individually in the traditional bedside manner. In addition to the significantly greater learning, the dietitian's time was reduced by 100 per cent, based on classes of eight patients. The study was conducted by randomly allocating forty-five qualified patients to control or experimental groups, pre-testing them using a questionaire-interview, giving bedside or class dietary instruction, and post-testing, using the same questionaire. Various personal characteristcs of the participants, as well as data relating to the diabetic state, were also examined for possible relationships to test scores and to one another.

Achievement↗
PubMedDate not supplied

Localization by immunofluorescent microscopy of several collagen types and of a basement membrane proteoglycan in rat glomerular epithelial and mesangial cell cultures.

Confluent cultures of rat glomerular epithelial and mesangial cells were studied by immunofluorescent microscopy, using affinity-purified antibodies directed against collagen of type I-V or an antiserum directed against a basement membrane (BM) proteoglycan. The epithelial cells were stained by antibodies directed against type I, IV and V collagen, whereas the mesangial cells were stained by all the antibodies directed against the different collagenous antigens tested. Therefore, only mesangial cells contained antigenic determinants of type III collagen. On the contrary, both cell types possessed BM proteoglycan antigens. The data suggest that rat glomerular epithelial and mesangial cells may be implicated in the biosynthesis of different components of normal (and pathological) glomerular BM.

Animals↗
PubMedDate not supplied

Expression and structure of cartilage proteins.

Cartilage has unique physical characteristics attributable to the presence of an unusually high content of proteoglycan embedded in the network of collagen fibrils. Advances in understanding the structure of these components and how their synthesis is regulated have been greatly assisted by the application of molecular biology. For example, an immortalized rat chondrocyte cell line was obtained by infection with a recombinant retrovirus encoding the myc gene product. Several positive and negative DNA regulatory elements of the collagen II gene have been identified that appear to be important in the regulation of this gene in chondrocytes. The complete primary structure of the cartilage proteoglycan (aggrecan) core protein deduced from cDNA sequence displays a complex multidomain structure including numerous repeats of Ser-Gly sequences and sequence homologies with link protein and animal lectins. Such studies advance our understanding of normal morphogenetic events and lay the groundwork for determining the basis of molecular and genetic defects.

Animals↗