LIPID ADJUVANTS AND ANTIGENS EMBEDDED IN THE MYCOBACTERIAL CELL ENVELOPE

The mycobacterial granuloma is one of the most complex and anatomically de ned immunological lesions in medicine, composed of concentric rings of necrosis, myeloid palisade, peripheral lymphocytes and polymorphonuclear phagocytes. In vaccine design, mycobacterial cell walls, also known as Freund’s adjuvant, have been widely used to stimulate T-cell responses in experiments over the last century. These two widely known immunological phenomena indicate that mycobacteria normally generate immune responses that are strong and differ from other bacterial or viral pathogens. Over the past decade, these immunological observations have been systematically investigated at the cellular and molecular levels, by separating the cell wall into its molecular components, deleting lipid synthase genes and studying mammalian receptors that mediate speci c responses to lipid, phospholipid and glycolipid components of the cell wall. Here, we review recent advances in understanding key immunostimulatory lipids, such as trehalosedimycolate (TDM) and phosphatidylinositol mannosides (PIM), as well as key host receptors including cell surface lectins (mannose receptor, DC-SIGN, Mincle), Toll-like receptors (TLRs) and CD1 proteins. In addition, we discuss new experimental approaches for broadly dissecting the host cell wall to determine the role of individual lipids in immunopathogenesis. INTRODUCTION The remarkably potent and broadly activating immunological properties of the mycobacterial cell wall have been known for more than a century. After Freund’s complete cell wall preparation failed to induce protective immunity in humans (1), its adjuvant properties were recognized. Complete Freund’s adjuvant is a complex mixture of natural cell wall components that is one of the most widely used adjuvants in immunological research. Freund’s adjuvant activates macrophages, promotes dendritic cell maturation and, when used in combination with peptides or proteins, generates MHC-restricted T cell responses (2). Although widely used in animal research, Freund’s adjuvant cannot be administered to humans because CHAPTER 2A.2 LIPID ADJUVANTS AND ANTIGENS EMBEDDED IN THE MYCOBACTERIAL CELL ENVELOPE Chapter02A.2. indd 125 11/02/14 1:06 PM 126 SECTION 2 MAIN PLAYERS the individual bioactive molecules are unknown. However, by discovering the individual molecules that activate mammalian responses and the key receptors that mediate responses, it is becoming possible to rationally develop adjuvants derived from mycobacterial cell wall components and subunit vaccines. Rapid advances in mycobacterial genetics provide methods to pinpoint the key lipid synthases and transporters with non-redundant roles in pathogenesis. This review highlights lipidic adjuvants and antigens in mycobacteria that activate host cells via lectins (Mincle, Dectin 1, mannose receptor), Toll-like receptors (TLR1, TLR2, TLR6, TLR 9), or T cell receptors via CD1. In addition, we discuss lipid modulators of the immune response that act by unknown mechanisms. MYCOBACTERIAL CELL WALL ORGANIZATION Mycobacteria elaborate what might be nature’s most complex lipidic cell envelope, which consists of ve major layers (Figure 2A.2.1). The cytoplasmic membrane is a phospholipid bilayer composed of anionic phospholipids such as cardiolipin, phosphatidylethanolamine and phosphatidylinositol, which resemble molecules found in other species, but are distinguished by a genus-speci c, branched C19 fatty acid known as tuberculostearic acid. The inner phospholipid membrane also contains PIM species, which have been reported to escape the inner membranes to varying degrees. In contrast, the more distal layers of peptidoglycan, arabinogalactan, mycolic acid membrane, as well as the carbohydrate-enriched capsular material, are Actinobacteria-speci c structures composed of lipids found only in mycobacteria and related species. In fact, browsing through recently discovered lipidomic databases (3, 4), it is clear that more than ninety percent of the subclasses of mycobacterial lipids are molecularly distinct from those of humans and other types of bacteria (Table 2A.2.1) (5). This basic observation has several general implications for the role of mycobacterial lipids in mammalian immune responses. First of all, the general lack of overlap in the classes of mycobacterial lipids with those found in other Gram-positive and Gram-negative bacteria provides a potential explanation for the relatively speci c nature of the mycobacterial granuloma as contrasted with classical purulent and septic responses to other bacteria. Thus, understanding the immune response requires direct study of natural mycobacterial pathogens, like MTB, rather than inferences from other bacteria, which have fundamentally different cell wall structures. Secondly, the lack of substantial overlap among lipids found in mycobacteria and mammals means that they contain a broad range of lipids that are foreign to the human immune system. In particular, outer membranes that Chapter02A.2. indd 126 11/02/14 1:06 PM 127 CHAPTER 2A.2 LIPID ADJUVANTS AND ANTIGENS EMBEDDED IN THE MYCOBACTERIAL CELL ENVELOPE