Physical Barriers

Physical barriers are most obvious in the arthropods. Tough chitinous exoskeletons can protect arthropods against all types of attackers. The horseshoe crab (Limulus polyphemus) not only has a hard exoskeleton but also can protect itself against bacteria in polluted water by secreting a specialized glycoprotein through pores in the carapace. On contact with endotoxins, this glycoprotein coagulates, sealing the pores and immobilizing any invading bacteria. Likewise, if bacteria enter horseshoe crab hemolymph, clotting factors are activated by lipopolysaccharides (LPSs) and result in local clot formation that traps invaders. Other invertebrates such as the coelenterates, annelids, mollusks, and echinoderms secrete masses of sticky mucus when attacked, thus immobilizing potential invaders. This mucus may contain antimicrobial peptides.

Innate Immunity

Invertebrates use three major innate defense subsystems: phagocytosis by blood or body cavity cells; protease cascades that lead to fluid clotting, melanin formation, and opsonization; and the production of a wide variety of antimicrobial peptides. Their initial defensive response is to use rapidly attacking phagocytic cells. This can be highly efficient, killing more than 99% of the invading bacteria. The few surviving bacteria, however, are somewhat more resistant to phagocytic cells than normal. These survivors induce the second stage of the response, the production of potent antibacterial peptides that wipe out any surviving bacteria and ensure that none survive to permit the development of resistance.

Prophenoloxidase (proPO)-Activating System

This system, found in arthropod hemolymph, consists of multiple enzymes that, when activated, generate a cascade of proteases leading to the production of the inert polymeric pigment melanin (Figure 40-2). The system is activated by the interaction of bacterial and fungal LPSs, peptidoglycans, and glucans with hemocytes. Activation also occurs through cuticular and hemolymph proteases. The proPO system generates phenoloxidase, a sticky enzyme that binds to foreign surfaces. This enzyme acts on tyrosine and dopamine to generate melanin and deposit it around inflammatory sites. Melanin polymer is deposited in the tissues surrounding invaders to form an impermeable barrier that blocks their nutrient uptake. Oxidizing agents and other antimicrobial molecules are also generated during melanin synthesis.

RNA Interference

The intracellular RNA interference pathway (RNAi) is a gene-silencing system that appears to have evolved to prevent viruses from replicating within infected cells. It is especially important as a defense system in invertebrates (Figure 40-3). RNA normally occurs only in a single-stranded (ss) form. Long segments of double-stranded RNA (dsRNA) are not present in healthy eukaryotic cells, but they do occur if a cell is infected by RNA viruses. When a virus-infected cell produces dsRNA, it is rapidly degraded into many short fragments by an enzyme called dicer. These fragments, or small-interfering RNAs (siRNAs) are then stabilized by a protein complex called the RNA-induced silencing complex (RISC). Half of these siRNAs will be complementary to the viral messenger RNAs (mRNAs) and as a result can serve as templates and bind them. Once these viral mRNAs have been captured by binding to the RISC complex, they are degraded rapidly, and viral replication effectively blocked.

Adaptive Immunity

Invertebrates do not make antibodies. The ability to mount adaptive immune responses arose with the jawed vertebrates. Nevertheless, proteins belonging to the immunoglobulin superfamily have been detected in arthropods, echinoderms, and mollusks as well as in protochordates. Some of these proteins can bind specifically to foreign molecules. In insects, there is a protein member of the immunoglobulin superfamily called Dscam that can be extensively diversified by alternate splicing. Isoforms of Dscam are expressed in immune tissues and secreted as soluble proteins into the hemolymph. Drosophila species have the potential to express more than 18,000 isoforms of this molecule. Individual hemocytes may express 14 to 50 forms of Dscam that can bind to bacteria and enhance their phagocytosis. It is not known how self-recognition by Dscam is prevented.

Graft Rejection

Invertebrates can reject allografts and xenografts. For example, cell-mediated allograft rejection occurs in sponges, coelenterates, annelids, and echinoderms. When two identical sponge colonies are placed side by side and made to grow in contact with each other, no reaction occurs. If, however, sponges from two different colonies are made to grow in contact, local destruction of tissue occurs along the area of contact as each sponge attempts to destroy the other.

Annelids such as earthworms can reject both allografts and xenografts. The rejection of xenografts (from other species of earthworms) takes about 20 days. Cells invade the graft, and the grafted tissue turns white, swells, becomes edematous, and eventually dies. If the recipient worms are grafted with a second piece of skin from the same donor, the second graft is rejected faster than the first. This ability to reject second grafts rapidly may be adoptively transferred by coelomocytes from sensitized animals.

Immunological “Big Bang”

The adaptive immune system depends on possession of two key antigen receptor systems, the TCR and the B cell receptor (BCR). Both require the rearrangement of V, D, and J gene segments to form functional, antigen-binding receptors. Invertebrates and cyclostomes cannot rearrange these genes, but cartilaginous and bony fish can. Sometime during the 100 million years between the divergence of jawless and jawed vertebrates and the emergence of cartilaginous and bony fish, about 450 Mya, the enzymatic machinery needed for the recombination of V gene segments emerged. The mechanism of this sudden appearance is unknown. It has been suggested, however, that a transposon carrying the precursors of the recombinase-activating genes RAG1 and RAG2 (most likely a bacterial integrase) was successfully inserted into an immunoglobulin superfamily V-like gene within the germline of the early jawed vertebrates (Figure 40-5). As a result, the immunoglobulin gene could be expressed only after splicing mediated by the RAG enzymes. Thus emerged, in a major evolutionary leap, the ability to generate antigen-binding sites and functional immunoglobulins. This, for the first time, permitted animals to respond specifically to previously encountered antigens. The advantages of this new “improved” system were such that it is now a feature of all jawed vertebrates. This did not, of course, result in discarding of the innate immune defenses. Lectins, the complement system, and the NK cell system remain essential components of vertebrate immunity. It is also important to point out that adaptive immunity did not confer invincibility to infectious agents. It simply made life more difficult for them and conferred an incremental selective advantage on animals with such defenses. The selective advantage of adaptive immunity came with an attendant cost—the potential for autoimmune disease.

Adaptive Immunity

Both cartilaginous and bony fish can mount adaptive immune responses and have a complete set of lymphoid organs except for a bone marrow (Figure 40-6). They have a thymus located just above the pharynx that arises from the first gill arches. In immature fish, small pores lead from the pharynx to the thymus, suggesting that it may be stimulated directly by antigens in the surrounding water. Thymectomy in fish can lead to prolongation of allograft survival and reduced antibody responses. Antibodies or antigen-binding cells may be detected in the thymus during an immune response, suggesting that it contains both T-like and B-like cells. Although the thymus may involute in response to hormones or season, age involution is inconsistent, and the thymus may be found in many older fish.