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Technical Program TRACK 3

Methods: SU-8 photoresist spun-coated on a silicon wafer was patterned face, however, poses major technical challenges to in vitro investigation
with desired features (circular holes 500 µm in diameter with 1200 µm cen- of its physiology and pathology using traditional cell culture models. As a
ter-to-center spacing) using conventional photolithography. Polydimethylsi- result, research in this area has relied heavily on expensive and time-con-
loxane (PDMS) was then cast over the SU-8 pattern to form positive impres- suming ex vivo or in vivo animal studies that often fail to model biological re-
sions of the desired features, and circular “buttons” were bore out using an sponses in humans. These critical drawbacks of exiting models have greatly
industrial punch. These buttons were then attached one-by-one to the pillars limited our fundamental understanding and hampered the development of
of a larger PDMS structure whose pillars matched the wells of an industry new therapeutic approaches to ocular surface diseases. To address this se-
standard 96-well plate. This multi-part PDMS stamp then served as a mask rious lack of physiological model systems, we have developed a microengi-
to protect desired circular regions of adsorbed collagen (with dimensions neered organ-on-a-chip model of the human eye that replicates 3D architec-
noted above) from ablation by oxygen plasma. The iHeps (Cellular Dynamics ture, physiological functionality, and dynamic mechanical microenvironment
International, Madison, WI) selectively attached to the collagen islands and of the ocular surface in the human eye.
then murine embryonic 3T3-J2 fibroblasts were seeded in the remaining
bare areas to create iMPCCs. Live markers in iMPCCs were measured using As the first step towards the development of this eye model, we generated
published protocols. In addition, ~1-week stabilized co-cultures were treated the porous 3D shell scaffolds by microengineering planar polystyrene cell
with up to 100×Cmax (maximum plasma concentration) of a drug three times culture scaffolds to create a curvature similar to that of the cornea in vivo.
over six days. Liver functions (i.e. albumin, urea) in drug-treated wells were Throughout prolonged cell culture, the 3D shell scaffolds retained the orig-
compared to solvent-only control wells. A drug that reduced at least one of inal curvature and maintained its structural stability. These scaffolds were
the measured functions to below 50% of controls was declared toxic, where- then incorporated into our eye-on-a-chip system by sandwiching them be-
as a drug that did not reduce any of the functions to below 50% of controls tween upper and lower PDMS slabs that contained a circular chamber and a
was declared non-toxic. microfluidic channel, respectively. To recreate a stromal layer in the cornea,
the sandwiched scaffold was impregnated with human primary keratocytes
Results and Discussion: Several major liver-specific mRNA transcripts (i.e. suspended in collagen precursor. We also recapitulated the unique concen-
HNF-4α, HNF-6) were up-regulated in iMPCCs relative to control cultures. tric patterns of the corneal and conjunctival epithelia using our novel 3D cell
Albumin and urea production by iMPCCs both reached steady-state levels patterning techniques that enabled precise positioning and deposition of
of 5-6 µg/hr/million cells by the first week of culture and remained stable human corneal and conjunctival epithelial cells at the central and peripheral
for 3 more weeks. Activities of several cytochrome P450 (CYP) enzymes regions of the scaffold surface, respectively. Furthermore, simulation of eye
were also maintained in iMPCCs for 4 weeks, reaching up to 73% of levels blinking was accomplished by integrating a biomimetic hydrogel eyelid with
observed in stable PHH cultures. Furthermore, iMPCCs were able to detect our microengineered cell culture model. The hydrogel eyelid was linked to
21 of 37 hepatotoxic drugs (65% sensitivity) and 10 of 10 non-toxic drugs a computer-controlled miniature motor and electromechanically actuated
(100% specificity). These results were remarkably similar to outcomes seen to slide along the convex scaffold surface to mimic the eyelid movements.
with PHHs using the same set of drugs (70% sensitivity, 100% specificity). We successfully demonstrated spreading of artificial tear fluid and surface
On the other hand, conventional cultures of iHeps with their reduced func- wetting of scaffold during the hydrogel movements. The motor was pro-
tional capacities showed a 38% reduction in sensitivity. iMPCCs were able grammed to simulate physiological blink scenarios of the human eye by pre-
to distinguish the relative toxicity of structural drug analogs (toxic/non-toxic cisely controlling kinematics and patterns of the eyelid movements.
drug pairs: troglitazone/rosiglitazone, tolcapone/entacapone) and displayed
known bioactivation-mediated mechanisms of acetaminophen toxicity. In Our on-going studies focus on leveraging this model system to develop a
conclusion, controlling interactions between iHeps and embryonic fibro- microengineered human disease model of dry eyes. We aim to demonstrate
blasts using microfabrication tools can significantly mature and sustain liver the capability of our human blinking “eye-on-a-chip” to recapitulate the
functions in the iHeps for at least 4 weeks. Proof-of-concept drug toxicity central disease mechanisms in the dry eye disease, which are the ocular
studies show that iMPCCs could be useful for an initial toxicity screen of a surface tissue damages caused by tear hyperosmolarity as a result of exces-
large drug library in industry standard 96-well plates. In the future, iMPCCs sive evaporation and/or insufficient secretion of the tear fluid.
could be used in human-on-a-chip platforms being designed to assess
multi-organ responses to pharmaceuticals. Lastly, the development of a li- We believe that our human blinking eye-on-a-chip system offers the promise
brary of patient-specific iHeps could enable personalized drug screening. to address critical technical barriers to progress ocular biology and clinical
ophthalmology by providing a novel experimental platform that enables rep-
4:40pm A human blinking “eye-on-a-chip” lication, visualization, and analysis of key biological processes in the human
eye. Moreover, this approach may enable the development of human dis-
ease models that represent cost-effective and more predictable alternatives
Technical Presentation. NEMB2016-6051 to conventional animal models for the identification and development of
new therapeutic approaches.
Jeongyun Seo, Woo Yul Byun, Andrea Frank, Department of Bio-
engineering, University of Pennsylvania, Philadelphia, PA, United 5:00pm Microfluidic Hydrogel System Provides Improved In Vi-
States, Giacomina Massaro-Giordano, Vivian Lee, Vatinee Bun- tro Platform for Blood Brain Barrier Studies
ya, Scheie Eye Institute, Department of Ophthalmology, Perelman
School of Medicine at the University of, Philadelphia, PA, United Technical Presentation. NEMB2016-6104
States, Dan Dongeun Huh, Department of Bioengineering, Univer-
sity of Pennsylvania, Philadelphia, PA, United States Candice Hovell, Yoshitaka Sei, Cole Weiler, Georgia Institute of
Technology, Atlanta, GA, United States, Gilda Barabino, City Col-
The ocular surface is a central anatomical and functional unit of the eye that lege of New York, New York, NY, United States, Lakeshia Taite,
protects the ocular system from external environment. As a principal protec-
tive barrier in this unit, the cornea consists of epithelium and a sub-epithelial Texas A&M University College of Veterinary Medicine & Biomedical
collagen-rich stromal tissue containing keratocytes. At the circumferential Sciences, College Station, GA, United States, YongTae Kim, Geor-
margin of the cornea, the corneal epithelium grades into the conjunctival gia Institute of Technology, Atlanta, GA, United States
epithelium lined with goblet cells that are responsible for producing mucus
into the tear fluid. The ocular surface is also under the constant influence of The development of new drug candidates is a timely and costly process.
dynamic microenvironment created by spontaneous eye blinking-induced Central nervous system (CNS) drugs are particularly challenging to develop
eyelid movements and concomitant spreading of the tear fluid that permits because of the presence of the blood brain barrier (BBB). The BBB main-
hydration and lubrication of the cornea and conjunctiva. tains the homeostatic environment of the brain by restricting the passage of
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molecules from systemic circulation via a variety of specialized endothelial
This structural, functional, and environmental complexity of the ocular sur- processes. These specializations arise from exposure to chemical and me-