Lymphatic pumping and lymphatic transport of antigen to lymph node

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Lymphatic vessels play essential roles in maintaining fluid balance and facilitating immune defense. While the heart beats to drive blood circulation, lymph transport primarily relies on lymphatic vessel pumping. Lymphatic pumping is driven by lymphatic smooth muscle cell (SMC) contraction. The function of lymphatic pumping is to propel lymph and cells transport to the draining lymph node (LN). When antigen and dendritic cells reach the draining lymph node (LN) through afferent lymphatic vessels, they must exit the vessels and migrate deep into the LN to activate T cell responses. However, it is not clear how changes in lymph flow may affect antigen delivery and antigen presentation in LN. In my previous studies, I have developed an intravital imaging model to study lymphatic pumping in mice. Using this model, I demonstrated that nitric oxide (NO) is required to maintain lymphatic pumping in physiological condition via the activation of the endothelial-derived nitric oxide synthase (eNOS). In contrast, the inducible nitric oxide synthase (iNOS) is responsible for the inhibition of lymphatic pumping in oxazolone skin sensitization induced inflammation (OX-inflammation). We hypothesize that NO regulates lymphatic pumping, which consequently modulates lymph flow velocity and flow pattern, and affect antigen delivery in lymph node. ***OBJECTIVES***Objective 1. To determine how NO regulates lymphatic SMC to control lymphatic pumping. We will study if NO directly signals on SMCs to cause SMC relaxation or via indirectly interrupt SMC and lymphatic endothelial cell (LEC) interaction. We will use eNOS-/- mice, or broad NOS inhibitor L-NMMA or iNOS-/- mice to block NOSs and study how lymphatic SMCs are regulated in physiological and OX-inflammation. ***Objective 2. To determine how lymphatic pumping controls the lymph flow velocity and flow pattern. We will measure the lymph flow in wild type, eNOS-/- and iNOS-/- mice in physiological and inflammatory conditions. We will study how lymph flow regulated LEC gene expression with cell culture using flow chamber to manipulate lymph flow velocity and flow pattern in variant conditions. ***Objective 3. To determine how lymphatic pumping manipulates antigen delivery to the draining LN. Free form of small antigens travel along the LN conduit system and are quickly captured by lymph node resident dendritic cells (DCs). We will use fluorescently labeled ovabulmin as small antigen to measure the antigen delivery in the draining LN when lymphatic pumping is impaired in variant conditions. We will study how these changes will alter the efficiency of antigen capture by LN DCs and initiation of T cell responses. ***The long-term goal of my studies will address fundamental questions of lymphatic pumping and lymphatic transport of antigen to the draining lymph node in order to efficiently present antigen and generate immune defense.**