Phagocytes and Phagocytosis
Phagocytes and Phagocytosis
- One of the most ancient forms of innate immunity is phagocytosis. It is a defensive reaction entailing the capture and digestion of foreign particles (e.g. bacteria or remnants of disintegrated cells) by phagocytes.
- Metchnikoff first discovered in 1882 that amoeboid cells of the mesoderm in starfishes were able to engulf and digest various foreign particles.
- Metchnikoff subdivided the cells capable of phagocytosis into microphages and macrophages.
- Microphages include granular leukocytes: neutrophils, eosinophils and basophils. Neutrophils and Eosinophils possess a marked ability for phagocytosis. Basophils are non- phagocytic in nature.
- Macrophages comprise a large set of specialized cells. They are consolidated into the mononuclear phagocyte system. It includes mobile cells (blood monocytes, phagocytes of the lymph nodes and spleen, connective tissue hystiocytes, etc.) or non-mobile elements (the resident macrophages of the lymphatic tissue and spleen, endotheliocytes of the blood vessels, liver resident macrophages or Kupffer stellate cells, mesangial macrophages, alveolar macrophages, glial macrophages, osteoclasts, etc.).
- Myeloid monocyte precursors differentiate in the bone marrow into promonocytes and then into mature monocytes. The latter comes into the blood flow. This circulating cell population migrates throughout the capillary wall into tissues and organs and transforms into resident macrophages.
- Molecular differentiation markers of macrophages are CD14, CD16 (FcγRIII), CD32 (FcγRII), CD64 (FcγRI), receptors to C3b-component of complement (CD35) and cytokine receptors (to IL-2, IL-4, γ-IFN, etc.).
- Macrophages possess 3 main biological functions: phagocytosis, antigen presentation, and cytokine secretion.
The process of phagocytosis consists of 5 main stages.
- The first phase is chemotaxis stage. Many microbial agents produce chemotactic agents that attract phagocytic cells.
- Chemotaxis deficiency may account for enhanced susceptibility to certain infections; these defects may be acquired or inherited.
- This stage involves phagocyte approaching the microbe by means of positive chemotaxis.
- Under the influence of microbial products activation of phagocytes occurs. It leads to a change in the cellular actin contractility, thus conferring amoeboid motility to phagocytes.
- There is a tremendous number of different chemoattractive agents known as chemokines (more than 100 substances).
- One of the most important chemokines is the above-mentioned IL-8, C5a-complement component and others, which attract macrophages and neutrophils toward the focus of inflammation.
- Also, some bacteria produce chemical substances such as LPS, which can attract leukocytes.
- At the second stage adsorption of the microorganisms to the surface of the phagocyte takes place (adhesion stage). This event is mediated by several recognition mechanisms. Tight binding is maintained by the specific interaction of phagocyte receptors (e.g., from Toll-like receptor family) with pathogen-associated molecular patterns.
- Target recognition often involves carbohydrate elements and lectins both of phagocyte and bacterial origin.
- The above-mentioned variant of microbial inactivation is known as non-immune phagocytosis.
- It may develop in the absence of serum antibodies or complement. Such “surface phagocytosis” occurs early in the infectious process before antibodies would become available.
- Immune phagocytosis evolves with the help of immune recognition.
- Phagocytosis is much more efficient in the presence of specific antibodies or complement components like C3b (so-called opsonins) that cover the surface of bacteria, thus facilitating their ingestion by phagocytes.
- Opsonization can occur by three basic mechanisms: by antibodies alone, by immune complex or by complement fragments (mainly via C3b and C5a components).
- Macrophages have membrane receptors to Fc portion of antibody and to C3b and C5a components of the complement system. These receptors stimulate the phagocytosis of antibody-coated particles.
- The third stage of phagocytosis is named ingestion (or engulfment) stage. The attached bacteria activate the ingestion phase by stimulating intracellular actin contraction with the formation of pseudopodia enwrapping the object of phagocytosis.
- Bacteria become completely encased within a vacuole (endosome or phagosome). In 1 minute the lysosomal granules fuse with the phagosome resulting in phagolysosome formation, and the lysosomal contents expel nearby the engulfed microorganism. This is followed by the activation of a great number of microbicidal mechanisms.
- In the fourth phase intracellular digestion of the engulfed microbes is activated (digestion stage).
- Ingestion of foreign particles (e.g., microorganisms) triggers several effects of phagocytic cells.
- There is a tremendous increase in activity of the hexose monophosphate shunt that generates NADPH.
- Primary key reaction in phagocytosis is catalyzed by phagocyte NADPH oxidase, which initiates the formation of multiple reactive oxygen species (ROS). This process is known as respiratory burst – the major microbicidal mechanism in phagocytes.
- The main ROS agents are superoxide anion (O2-), hydroxyl radicals (•OH) and singlet oxygen.
- Superoxide anion undergoes conversion into hydrogen peroxide under the influence of superoxide dismutase, and subsequently to hydroxyl radicals (•OH).
- All of these products have the outstanding chemical reactivity making them powerful microbicidal agents. Hydroxyl radical is one of the most reactive chemicals known.
- Furthermore, the combination of peroxide and halide ions forms a potent halogenating system capable of killing both bacteria and viruses. Hydrogen peroxide and the halogenated compounds are more stable than free radicals. They diffuse further and subvert the microbials outside the cell.
- Reactive nitrogen species (nitric oxide NO and peroxynitrite) act through the inactivation of certain microbial electron transport enzymes and also by the production of (•OH) radicals.
- Activation of reactive nitrogen species is termed as a nitrosative burst or nitrosative stress. It plays a significant role in the elimination of certain pathogens, e.g., Mycobacterium tuberculosis.
- Also, the family of cationic proteins and peptides, known as defensins, attacks the bacteria inside the phagolysosome. They are about of 3.5-4 kDa of molecular weight, being rich in arginine.
- These antibiotic peptides act as biocides against a broad spectrum of gram-positive and gram-negative bacteria, fungi and a number of enveloped viruses.
- Further damage of bacterial structures is caused by neutral proteinases (such as cathepsin G) and by other endosomal hydrolytic enzymes like hyaluronidase or nucleases. Lysozyme and lactoferrin are also potent microbicidal factors that are oxygen-independent and can function under anaerobic conditions.
- Low pH within phagosome facilitates microbial degradation.
- Finally, the killed microbial bodies are digested by hydrolytic enzymes, and the phagocytized microbes become completely disintegrated (complete phagocytosis).
- Besides complete phagocytosis, incomplete phagocytosis occurs in certain bacterial infections (gonorrhoea, legionellosis, leishmaniasis, tuberculosis, leprosy, etc.)
- In those cases, microorganisms are engulfed by phagocytes but don’t lose their viability, and even may reproduce.
- The mechanisms of microbial survival within phagocytes are supported by capsule production (like Klebsiella pneumonia), or by the block of phagosome-lysosomal fusion (e.g., Legionella pneumophila) or by the chemically resistant structure of the microbial body (e.g., the presence of highly stable lipids and waxes in M. tuberculosis).
- The last fifth phase of phagocytosis is the release of degradation products, where the non-digested microbial remnants are discharged outside from the cell.