In vivo downregulation of innate and adaptive immune responses in corneal allograft rejection by HC-HA/PTX3 complex purified from amniotic membrane

Heavy chain-hyaluronic acid (HC-HA)/PTX3 purified from human amniotic membrane (AM) was previously observed to suppress inflammatory responses in vitro. We now examine whether HC-HA/PTX3 is able to exert a similar effect in vivo, using murine models for keratitis and corneal allograft rejection. The...

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Bibliographic Details
Published in:Investigative ophthalmology & visual science 2014-03, Vol.55 (3), p.1647-1656
Main Authors: He, Hua, Tan, Yaohong, Duffort, Stephanie, Perez, Victor L, Tseng, Scheffer C G
Format: Article
Language:English
Subjects:
Acute-Phase Proteins
Adaptive Immunity - physiology
Allografts
Amnion - cytology
Amnion - immunology
Animals
C-Reactive Protein - pharmacology
CD4-Positive T-Lymphocytes - immunology
Cells, Cultured
Corneal Transplantation
Cytokines - biosynthesis
Cytokines - drug effects
Disease Models, Animal
Enzyme-Linked Immunosorbent Assay
Female
Flow Cytometry
Graft Rejection - immunology
Graft Rejection - metabolism
Graft Rejection - pathology
Humans
Hyaluronic Acid - pharmacology
Immunity, Innate - physiology
Mice
Mice, Inbred BALB C
Mice, Inbred C57BL
Serum Amyloid P-Component - pharmacology
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Summary:Heavy chain-hyaluronic acid (HC-HA)/PTX3 purified from human amniotic membrane (AM) was previously observed to suppress inflammatory responses in vitro. We now examine whether HC-HA/PTX3 is able to exert a similar effect in vivo, using murine models for keratitis and corneal allograft rejection. The in vitro effect of HC-HA/PTX3 was tested using OTII ovalbumin (OVA) transgenic, purified CD4(+) T cells, or IFN-γ/lipopolysaccharide (LPS)-stimulated RAW264.7 cells. Cytokine production was measured by ELISA, while cell surface markers and cell proliferation were determined by flow cytometry. In vivo effects of HC-HA/PTX3 were analyzed by quantifying the recruitment of enhanced green fluorescence-labeled macrophages and by measuring the expression of arginase 1 (Arg-1), IL-10, and IL-12 in LPS-induced keratitis in the macrophage Fas-induced apoptosis (Mafia) mouse. The effect of corneal allograft survival in a complete major histocompatibility complex (MHC) mismatched mouse model was assessed by grading corneal opacification. In vitro studies demonstrated that HC-HA/PTX3 significantly enhanced the expansion of FOXP3 T cells and suppressed cell proliferation and protein expression of IFN-γ, IL-2, CD25, and CD69 in activated CD4(+) T cells. Furthermore, immobilized HC-HA/PTX3 significantly upregulated IL-10 gene expression but downregulated that of IL-12 and IL-23 in activated RAW264.7 cells. Finally, in vivo subconjunctival injection of HC-HA/PTX3 significantly prolonged corneal allograft survival, suppressed macrophage infiltration, and promoted M2 polarization by upregulating Arg-1 and IL-10 but downregulating IL-12. HC-HA/PTX3 can suppress inflammatory responses in vivo by modulating both innate and adaptive immunity of macrophages and CD4(+) T cells.
ISSN:1552-5783
0146-0404
1552-5783
DOI:10.1167/iovs.13-13094