Cell: matrix adhesion and hemidesmosomes
In collaboration with Ilene Gipson at the Schepen’s Eye Research Institute and Harvard Medical School, Dr. Stepp and colleagues showed that the α6β4 integrin was an integral membrane component of hemidesmosomes. This was an important discovery for a number of reasons. While the integral membrane components of the desmosomes (desmoglein, desmocoliln) and adherens junctions (E-cadherin) had begun to be characterized, none of the integral membrane components of hemidesmosomes had been identified. Hemidesmosomes mediate cell substrate adhesion between epithelia and basement membrane via keratin intermediate filaments. Dr. Stepp was a post-doctoral fellow in the laboratory of Richard Hynes at MIT in the Cancer Center from 1986-1988 during which The Hynes lab named and characterized the first integrin. Integrins were thought to mediate cell adhesion to substrates via actin microfilaments. Arriving at Harvard and Schepen’s Eye Research Institute with knowledge of integrins, Dr. Stepp observed that α6 integrin was restricted to the basal aspect of the basal cells where hemidesmosomes were located. It’s localization was disrupted during migration and it became restricted to the basal aspect of the basement membrane once again when cell migration ceased and hemidesmosomes were reassembled. Collaborating with Ilene Gipson, Dr. Stepp demonstrated that α6 integrin was present in hemidesmosomes. A paper describing those results was published in PNAS in 1990. It was the first paper to demonstrate an integral membrane component of the hemidesmosome and was the first demonstration in vivo of an integrin within an adhesion complex with known biological relevance. The demonstration of integrins as components of hemidesmosomes helped to launch interest in integrin research.
- Stepp, M.A., Spurr-Michaud, S.J., Tisdale, A., Ellwell, J., and Gipson, I.K.1990. α6ß4 Integrin Heterodimer is a Component of Hemidesmosomes. Proc. Natl. Acad. Sci. USA, 87, 8970-8974. PMID: 2247472. Link
- Giancotti, F.G., Stepp, M.A., Suzuki, S., Engvall, E., Ruoslahti, E. 1992. Proteolytic processing of endogenous and recombinant β4 integrin subunit. J. Cell Biol. 118, 951-959. PMID: 1500432. Link
- Gipson, I.K., Spurr-Michaud, S., Tisdale, A., Elwell, J., Stepp, M.A. 1993. Redistribution of the hemidesmosome components α6β4 integrin and bullous pemphigoid antigens during epithelial wound healing. Exp. Cell Res. 207, 86-98. PMID: 8319775. Link
Development of Models for the Study of Recurrent Epithelial Erosion and Limbal Stem Cell Deficiency
The Stepp Lab developed and characterized a mouse model that continues to advance understanding of corneal homeostasis and wound healing. The model is used to study recurrent epithelial erosions and the model was optimized for the study of limbal stem cell deficiency. It has been adopted and modified by others and is being used in research labs to help advance treatment options for those suffering from corneal pathology. The model was first described in a paper published in 2004 and numerous additional papers from the Stepp Lab and others using this model have followed. Prior to the description of the mouse recurrent erosion model, the only way to study recurrent erosions was using genetically susceptible dog breeds.
- Pal-Ghosh, S., Pajoohesh-Ganji, A., Brown, M., Stepp, M.A. 2004. A mouse model for the study of recurrent corneal epithelial erosions: α9β1 integrin implicated in progression of the disease. Invest. Ophthalmol. Vis. Sci. 45, 1775-1788. PMID: 15161840. Link
- Pal-Ghosh, S., Blanco, T., Tadvalkar, G., Pajoohesh-Ganji, A., Parthasarathy, A., Zieske, J.D., Stepp, M.A. 2011. MMP9 cleavage of the β4 integrin ectodomain leads to recurrent epithelial erosions in mice. J. Cell Sci. 124, 2666-2675. PMID: 21750188. Link
- Stepp, M.A., Zieske, J.D., Trinkaus-Randall, V., Kyne, B.M., Pal-Ghosh, S., Tadvalkar, G., Pajoohesh-Ganji, A. 2014. Wounding the cornea to learn how it heals. Exp. Eye. Res. 121, 178-193. PMID: 24607489. Link
Adult Corneal Epithelial Stem Cells and Corneal Goblet Cells
When the mouse cornea is wounded near the limbus, migration of a population of stem cells and associated niche cells, referred to as compound niches, move onto the corneal surface. Over time they expand in number. Because they express simple epithelial keratins and contain goblet cells, their expansion leads to a condition similar to limbal stem cell deficiency. Conjunctival goblet cells do not migrate onto the cornea nor give rise to the goblet cells that expand on the cornea in this model. Wounds that induce goblet cell expansion do not disrupt the limbal epithelium and its population of progenitor cells. The source of the goblet cells is either the progenitor cells that make up the compound niche or the corneal epithelium itself. In this model, the progenitor cells that give rise to corneal epithelium also generate goblet cells but they only do so when injuries near the limbus occur. The signals that induce differentiation of progenitor cells into goblet cells are not known.
- Pajoohesh-Ganji, A., Pal-Ghosh, S., Tadvalkar, G., Stepp, M.A. 2012. Corneal goblet cells and their niche: implications for corneal stem cell deficiency. Stem Cells 30, 2032-2043. PMID: 22821715. Link
- Pal-Ghosh, S., Tadvalkar, G., Jurjus, R.A., Zieske, J.D., Stepp, M.A. 2008. BALB/c and C57BL6 mouse strains vary in their ability to heal corneal epithelial debridement wounds. Exp. Eye Res. 87, 478-486. PMID: 18809399. Link
The Heparan Sulfate Proteoglycan Syndecan-1 Modulates Corneal Epithelial Cell Adhesion, Migration, and Sensory Nerve Targeting
Syndecan-1 is a membrane associated heparan sulfate proteoglycan that mediates many of the functions thought to be mediated by the binding of growth factors, matrix proteins, and membrane receptors with so called “heparin binding sites.” The syndecan-1 null mouse was generated by Merton Benfield’s lab at Children’s Hospital in Boston. It had a skin wound healing phenotype that could not be easily characterized due to it’s complexity. Along with several of Dr. Bernfield’s lab members, Dr. Stepp demonstrated that the mouse was null for syndecan-1 expression and that it had a wound healing phenotype that involved reduced corneal and epidermal cell migration due to alterations in integrin expression. The Stepp Lab has published several papers over the years related to syndecan-1 including one on the syndecan-1 null skin cancer phenotype in collaboration with Dr. Stuart Yuspa at the NCI. A review was recently written by the Stepp Lab focusing on syndecan-1 and its role in wound healing. Currently, the Stepp Lab is looking at the role of syndecan-1 in mediating corneal sensory nerve reinnervation after wounding.
- Stepp, M.A., Gibson, H.E., Gala, P.H., Sta. Iglesia, D.D., Pajoohesh-Ganji, A., Pal-Ghosh, S., Brown, M., Aquino, C., Schwartz, A.M., Goldberger, O., Hinkes, M.T., Bernfield, M. 2002. Defects in keratinocyte activation during wound healing in the syndecan-1-deficient mouse. J. Cell Sci. 115, 4517-4531. PMID: 12414997. Link
- Stepp, M.A., Liu, Y., Pal-Ghosh, S., Jurjus, R.A., Tadvalkar, G., Sekaran, A., LoSicco, K., Jiang, L., Larsen, M., Li, L., Yuspa, S.H. 2007. Reduced migration, altered matrix and enhanced TGFβ1 signaling are signatures of mouse keratinocytes lacking Sdc1. J. Cell Sci. 120, 2851-2863. PMID: 17666434. Link
- Stepp, M.A., Pal-Ghosh, S., Tadvalkar, G., Rajjoub, L., Jurjus, R.A., Gerde M., Ryscavage, A., Cataisson, C., Shukla, A., Yuspa, S.H. 2010. Loss of syndecan-1 is associated with malignant conversion in skin carcinogenesis. Mol. Carcinogenesis 49, 363-373. PMID: 20082322. Link
- Stepp, M,A., Pal-Ghosh, S., Tadvalkar, G., Pajoohesh-Ganji, A. 2015. Syndecan-1 and Its Expanding List of Contacts. Adv. Wound Care 4, 235-249. PMID: 25945286. Link
The Intraepithelial Corneal Nerves
The eye is innervated by neurons derived from both the central nervous system and peripheral nervous system. While much research has been done over the years on the retinal nerves, the corneal nerves are less well characterized. The ophthalmic branch of the trigeminal ganglion contains somas of neurons that innervate the cornea. These peripheral nervous system derived nerve fibers provide sensory functions for the cornea and are referred to as intraepithelial corneal nerves and consist of subbasal nerves and intraepithelial nerve terminals. These nerves project for several millimeters within the human corneal epithelium; the nearest Schwann cells are millimeters away in the corneal stroma. When the trigeminal ganglion is severed, the corneal epithelial cells stop proliferating, undergo apoptosis or senescence, and erosions form proving that the nerves provide trophic factors to corneal epithelial cells. The Stepp Lab has shown that during homeostasis, corneal epithelial cells function as glial cells to support the intraepithelial corneal nerves. Thus, the corneal sensory axons and the cells of the corneal epithelium are co-dependent. Corneal epithelial cells activate in response to injury via mechanisms similar to those induced in Schwann cells during Wallarian Degeneration. They phagocytize distal axon fragments within hours of intraepithelial corneal nerve crush wounds. During aging, the proteins, lipids, and mitochondria within the intraepithelial corneal nerves become damaged in a process exacerbated by UV light and are shed. Intraepithelial corneal nerves shed their aged and damaged termini and continuously elongate to maintain their density. The Stepp Lab is using the cornea to study PNS sensory nerve recovery after injury and in response to aging.
- Pajoohesh-Ganji, A., Pal-Ghosh, S., Tadvalkar, G., Kyne, B.M., Saban, D.R., Stepp, M.A. 2015. Partial denervation of sub-basal axons persists following debridement wounds to the mouse cornea. Lab. Invest. 95: 1305-1318. PMID: 26280222. Link
- Pal-Ghosh, S., Pajoohesh-Ganji, A., Tadvalkar, G., Kyne, B.M., Guo, X., Zieske, J.D., Stepp, M.A. 2016. Topical Mitomycin-C enhances subbasal nerve regeneration and reduces erosion frequency in the debridement wounded mouse cornea. Exp. Eye. Res. 146: 361-369. PMID: 26332224. Link
- Stepp, M.A., Tadvalkar, G., Hakh, R., Pal-Ghosh, S. 2017. Corneal epithelial cells function as surrogate Schwann cells for their sensory nerves. GLIA 65: 851-863. PMID: 27878997. Link