Date Presented

Spring 5-2026

Document Type

Thesis

First Advisor

Dr. David O. Freier

Second Advisor

Dr. Allison B. Jablonski

Third Advisor

Dr. Price Blair

Abstract

Macrophages are a type of white blood cell that have important functional roles in the innate immune system, the body’s first line of defense against invading pathogens. Macrophages possess two phenotypic forms, M1 and M2, which are functionally and metabolically different and respond to different stimuli. M1 macrophages respond to bacterial products, such as lipopolysaccharide (LPS), by producing pro-inflammatory mediators. Furthermore, M2 macrophages respond to 8 Bromo cAMP (8Br) by producing tissue repair and wound healing properties. Second messenger signaling through cAMP plays an important role in the resolution of inflammation and inflammatory mediators. Metabolism plays a key role in macrophage polarization, resulting in distinct metabolic pathways and functional outputs. Understanding metabolic activity and cAMP signaling pathways in macrophages is important to evaluate their functional role in inflammatory responses and resolution. A key metabolic distinction in macrophage phenotypes is the metabolism of L-Arginine. L-Arginine is the substrate for two different enzymes in macrophages: inducible nitric oxide synthase (iNOS) and arginase. Arginase activity can be measured in RAW 264.7 murine macrophages following 8Br and LPS stimulation. The RAW 264.7 murine macrophage cell line is a standard model for evaluating pro-inflammatory stimuli. It is known that two different isoforms of arginase are expressed in macrophages. 8Br (0 - 1000μM) stimulated arginase activity in a dose-response manner in both the presence and absence of 100 ng/mL LPS. Furthermore, 8Br (100μM) combined with 1 ng/mL LPS shows decreased Nitric Oxide (NO) response. Future experimentation will aim to examine which isoform is stimulated (via LPS and 8Br) and measurable in our systems. Establishing clear protocols for measuring arginase activity can further extend an understanding of the metabolic activity underlying macrophage polarization.

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