TY - JOUR
T1 - Characterizing molecular diffusion in the lens capsule
AU - Danysh, Brian P.
AU - Patel, Tapan P.
AU - Czymmek, Kirk J.
AU - Edwards, David A.
AU - Wang, Liyun
AU - Pande, Jayanti
AU - Duncan, Melinda K.
N1 - Funding Information:
This work was supported by National Eye Institute Grant EY015279 to MKD, a Sigma-Xi/National Academy of Sciences research award and a GK-12 fellowship both awarded to BPD, a Beckman Young Scholars Fellowship to TPP, National Eye Institute grant EY010535 to JP, NIH COBRE program grant P20RR016459 , NIAMS grant AR054385 and University of Delaware Research Foundation grants to LW. INBRE program grant P20 RR16472 supported the University of Delaware Bio-Imaging Center. The Authors would also like to thank Chuong Huynh, Larry Scipioni, and Arno Merkle (Carl Zeiss SMT, Inc., Peabody, MA) for their assistance with the helium ion microscopy.
PY - 2010/4
Y1 - 2010/4
N2 - The lens capsule compartmentalizes the cells of the avascular lens from other ocular tissues. Small molecules required for lens cell metabolism, such as glucose, salts, and waste products, freely pass through the capsule. However, the lens capsule is selectively permeable to proteins such as growth hormones and substrate carriers which are required for proper lens growth and development. We used fluorescence recovery after photobleaching (FRAP) to characterize the diffusional behavior of various sized dextrans (3, 10, 40, 150, and 250. kDa) and proteins endogenous to the lens environment (EGF, γD-crystallin, BSA, transferrin, ceruloplasmin, and IgG) within the capsules of whole living lenses. We found that proteins had dramatically different diffusion and partition coefficients as well as capsule matrix binding affinities than similar sized dextrans, but they had comparable permeabilities. We also found ionic interactions between proteins and the capsule matrix significantly influence permeability and binding affinity, while hydrophobic interactions had less of an effect. The removal of a single anionic residue from the surface of a protein, γD-crystallin [E107A], significantly altered its permeability and matrix binding affinity in the capsule. Our data indicated that permeabilities and binding affinities in the lens capsule varied between individual proteins and cannot be predicted by isoelectric points or molecular size alone.
AB - The lens capsule compartmentalizes the cells of the avascular lens from other ocular tissues. Small molecules required for lens cell metabolism, such as glucose, salts, and waste products, freely pass through the capsule. However, the lens capsule is selectively permeable to proteins such as growth hormones and substrate carriers which are required for proper lens growth and development. We used fluorescence recovery after photobleaching (FRAP) to characterize the diffusional behavior of various sized dextrans (3, 10, 40, 150, and 250. kDa) and proteins endogenous to the lens environment (EGF, γD-crystallin, BSA, transferrin, ceruloplasmin, and IgG) within the capsules of whole living lenses. We found that proteins had dramatically different diffusion and partition coefficients as well as capsule matrix binding affinities than similar sized dextrans, but they had comparable permeabilities. We also found ionic interactions between proteins and the capsule matrix significantly influence permeability and binding affinity, while hydrophobic interactions had less of an effect. The removal of a single anionic residue from the surface of a protein, γD-crystallin [E107A], significantly altered its permeability and matrix binding affinity in the capsule. Our data indicated that permeabilities and binding affinities in the lens capsule varied between individual proteins and cannot be predicted by isoelectric points or molecular size alone.
KW - Basement membrane
KW - Binding affinity
KW - Diffusion coefficient
KW - FRAP
KW - Lens capsule
KW - Partition coefficient
KW - Permeabilitiy
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U2 - 10.1016/j.matbio.2009.12.004
DO - 10.1016/j.matbio.2009.12.004
M3 - Article
C2 - 20026402
AN - SCOPUS:77950860248
SN - 0945-053X
VL - 29
SP - 228
EP - 236
JO - Matrix Biology
JF - Matrix Biology
IS - 3
ER -