Alon Kahana, M.D., Ph.D.
Research Projects

Dr. Kahana in his labThe orbit is the anatomic structure that contains the eye and all of its associated tissues, including muscles, nerves, blood vessels and connective tissues. It is encased in a complex bony structure that protects the eye and separates it from the brain. Our research focuses on the morphogenesis of the orbit and the potential of regenerative medicine to provide novel therapeutic approaches to orbital diseases. Our lab utilizes zebrafish because of the model’s significant advantages for studying embryologic and regenerative processes.

Orbital tissues have a unique embryologic origin in the neural crest, which is a transient population of stem cells that migrate from the invaginating neural tube to destinations throughout the body. Neural crest-derived cells are responsible for tissues as varied as heart valves, blood vessels, bones, endocrine glands, and the autonomic nervous system. However, there is no other place in the body to which neural crest cells contribute and/or influence more cell types within such a small space than the orbit. The neural crest contributes directly or indirectly to most of the orbital bones, the orbital connective tissues (including muscle pulleys), sclera, corneal stroma and endothelium, trabecular meshwork, sensory nerves, blood vessels, iris, and extraocular muscles.

The lab has two overlapping research interests: (1) regeneration of damaged extraocular muscles, and (2) the role of the neural crest in ocular development and disease.

3-day-old zebrafish embryo
The α-actin::EGFP transgene marks the extraocular and jaw muscles of 3 day-old zebrafish embryos. Fish have 6 extraocular muscles for each eye, just as humans do: 4 rectus muscles, and 2 oblique muscles.

Extraocular muscle regeneration

Binocular vision requires exact control of eye movement, which is accomplished by six pairs of extraocular muscles, which are biologically unique and genetically distinct from other skeletal muscles. Extraocular muscle disorders can cause neurologic vision loss in children (amblyopia), or debilitating double vision in adults. Zebrafish have a unique capacity for regenerating complex tissues and organs, using a combination of resident stem cells and cellular dedifferentiation. Our lab developed a unique model for studying the regenerative response to extraocular muscle damage. We discovered that Zebrafish can regenerate fully functional muscles even after removal of 50-80% of the muscle. Using transcriptome analysis of regenerating muscle cells, we identified several pathways that are important in this process. Interestingly, many of these pathways are also known to be involved in cancer. We are now characterizing the biological mechanisms that underlie extraocular muscle regeneration. Our goal is to utilize regenerative medicine to improve clinical outcomes of extraocular muscle disorders, including congenital strabismus, trauma and thyroid eye disease. This work also has potential to contribute to the broader fields of stem cell biology and regenerative medicine.

Orbital development, disease, and the role of neural crest cells

Diseases such as thyroid-related eye disease may be the result of derangements in neural crest-derived cells that are normally present in the orbit. Craniofacial syndromes such as Apert and Crouzon, as well as anterior segment dysgenesis syndromes of the eye, are also known to involve derangements in neural crest development. Our research explores the biology of neural crest-derived tissues in the orbit by using Zebrafish as a model system. Zebrafish are transparent during embryogenesis, are genetically tractable, and as vertebrates, they are remarkably good models for human biology and genetics. Our research focuses on the signals that control neural crest migration into the eye and orbit, as well as the intricate processes that control neural crest cell fate. The ability of adult orbital tissues to trans-differentiate, as well as the unique biological behavior of a variety of orbital tumors and inflammatory processes, may be related to the neural crest origins of much of the orbit. Our goal is to develop novel diagnostic tools as well as potential treatments for neural crest-related eye diseases such as congenital glaucoma and anterior segment dysgenesis syndromes, as well as for severe orbital diseases such as cancer and thyroid-related eye disease.

Paraffin sections of the head taken from zebrafish embryos & adults reveal remarkable similarity with the human eye and orbit
Paraffin sections of the head taken from zebrafish Paraffin sections of the head taken from zebrafish

Movie 1: Adult zebrafish display a stereotypical optokinetic response (OKR)

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Adult zebrafish display a stereotyped pattern of eye movements, known as an optokinetic reflex (OKR), in response to moving visual stimuli. This response is evoked by placing zebrafish on a stationary pedestal within a rotating optokinetic drum that is marked on the inner wall by alternating black and white stripes. Generation of this response requires (1) vision, and (2) extraocular muscle activity. In the movie, the first 10 seconds reveal left-sided eye pursuits followed by rapid right-sided saccades in response to counter-clockwise rotating stripes. At 11 seconds, drum rotation is changed to clockwise, and the fish responds appropriately by tracking the stripes to the right. This adaptive trait is found in most vertebrates and allows for moving images to be stabilized on the retina. The Kahana lab is using this behavioral assay to assess extraocular muscle function in our extraocular muscle regeneration research.

Movie 2: Rostral Neural Crest Migrates in Dorsal and Ventral Waves Around the Eye to Populate the Frontonasal Process and Periorbital Mesenchyme.

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Multiphoton time-lapse imaging of Tg(sox10::EGFP) from 12 to 30 hpf demonstrated that sox10 expressing cells migrate into the head via waves that migrate dorsal and ventral to the eye to populate the frontonasal process and periorbital mesenchyme.

Movie 3: Thyroid hormone signaling is required for initiation of ventral wave migration of rostral neural crest cells. (Multiphoton time-lapse microscopy)

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Following morpholino knockdown of the thyroid hormone receptor Thraa (which would block thyroid hormone signaling), the ventral wave fails to migrate, while cells of the dorsal wave die following migration.

Movie 4: Exogenous retinoic acid rescues the phenotype of thyroid hormone receptor knockdown

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Time-lapse imaging of Tg(sox10::EGFP) from 12 to 30 hpf of Thraa MO knockdowns treated with 1 nM RA (starting at 12 hpf) demonstrated rescue of migration of the ventral wave of the rostral neural crest.

 

Kahana research facilities

Last Modified: Thursday, 18-Apr-2013 10:18:33 EDT