Activation Of Dll4/Notch Signaling And Hypoxia-Inducible Factor-1 Alpha Facilitates Lymphangiogenesis In Lacrimal Glands In Dry Eye

PurposeBy using hypoxia-inducible factor-1 alpha conditional knockout (HIF-1 alpha CKO) mice and a dry eye (DE) mouse model, we aimed to determine the role played by delta-like ligand 4 (Dll4)/Notch signaling and HIF-1 alpha in the lymphangiogenesis of lacrimal glands (LGs).MethodsC57BL/6 mice were housed in a controlled-environment chamber for DE induction. During DE induction, the expression level of Dll4/Notch signaling and lymphangiogenesis in LGs was measured by quantitative RT-PCR, immunoblot, and immunofluorescence staining. Next, lymphangiogenesis was measured after Dll4/Notch signal inhibition by anti-Dll4 antibody or.-secretase inhibitor. Using HIF-1 alpha CKO mice, the expression of Dll4/Notch signaling and lymphangiogenesis in LGs of DE-induced HIF-1 alpha CKO mice were assessed. Additionally, the infiltration of CD45(+) cells in LGs was assessed by immunohistochemical (IHC) staining and flow cytometry for each condition.ResultsDE significantly upregulated Dll4/Notch and lymphangiogenesis in LGs. Inhibition of Dll4/Notch significantly suppressed lymphangiogenesis in LGs. Compared to wild-type (WT) mice, DE induced HIF-1 alpha CKO mice showed markedly low levels of Dll4/Notch and lymphangiogenesis. Inhibition of lymphangiogenesis by Dll4/Notch suppression resulted in increased CD45+ cell infiltration in LGs. Likewise, CD45+ cells infiltrated more in the LGs of HIF-1 alpha CKO DE mice than in non-DE HIF-1 alpha CKO mice.ConclusionsDll4/Notch signaling and HIF-1a are closely related to lymphangiogenesis in DE-induced LGs. Lymphangiogenesis stimulated by Dll4/Notch and HIF-1 alpha may play a role in protecting LGs from DE-induced inflammation by aiding the clearance of immune cells from LGs.