Protein array characterization of bioactive proteins secreted by immortalized human corneal epithelium in response to pseudomonas constituents.
OBJECTIVE
To use protein arrays to delineate the spectrum of angiogenic bioactive protein modulators that might be secreted and up-regulated by the corneal epithelium in response to killed bacterial products.


METHODS
Immortalized human corneal epithelial cells were grown in culture, serum starved, and exposed to heat-killed Pseudomonas aeruginosa in a dose-dependent manner. The resultant culture medium was screened by antibody arrays for 43 proteins that can modulate angiogenesis and immune and inflammatory processes. Parallel analysis was carried out on tears recovered in the open and closed eye phases (OTF and CTF) of the diurnal cycle.


RESULTS
Array analysis reveals that the immortalized cells constitutively secrete several proteins and up-regulate the secretion of IL-6, IL-8, and GRO in response to killed bacteria. Also evident was the emergence of a strong signal for GM-CSF and moderate/weak signals for MCP-1, MMP-9, Leptin, and INF gamma in a dose-dependent manner. Several of these proteins, including IL-6, IL-8, GRO, MMP-9, TIMP-1, and MCP-1, accumulate in the CTF. Other proteins are unique to tear fluid.


CONCLUSIONS
Nine proteins were identified that are secreted by epithelium in response to killed bacteria that contribute to the innate and adaptive defense system through potentiating PMN and macrophage recruitment, activation, and opsonization in a cooperative manner. The vast majority of these proteins are angiogenic modulators, perhaps contributing to the imbalance between angiogenic and angiostatic processes and risk of corneal vascularization.
Sack
Robert
R
Department of Biological Sciences, SUNY Optometry, New York, New York 10036, USA. rsack@sunyopt.edu
Sathe
Sonal
S
Beaton
Ann R
AR
McNamara
Nancy
N
Fleiszig
Suzanne
S
Ni
Minjian
M
D043924:Angiogenesis Modulating Agents / Q000378:metabolism; D002460:Cell Line; D002462:Cell Membrane; D016207:Cytokines / Q000378:metabolism; D019573:Epithelium, Corneal / Q000378:metabolism* / Q000382:microbiology / Q000473:pathology; D005136:Eye Proteins / Q000378:metabolism*; D006801:Humans; D007113:Immunity, Innate; D040081:Protein Array Analysis / Q000379:methods*; D011550:Pseudomonas aeruginosa / Q000502:physiology*; D013666:Tears / Q000378:metabolism; D015854:Up-Regulation
D043924:Angiogenesis Modulating Agents; D016207:Cytokines; D005136:Eye Proteins