Antimicrobial and cytotoxic activities of cells

BY

ADEDEJI EBENEZER

15/SCI 03/1001

Postgraduate Biochemistry Programme

Department of Chemical Sciences
College of Sciences
Afe Babalola University

Ado-Ekiti, Ekiti State

Introduction

Phagocytes are a class of cells which are capable of ingestion and destruction of
microorganisms and viruses that are responsible for inciting an inflammatory
response. Phagocytosis of microbes plays an essential role in the health of the
immune system. Once pathogens invade the tissues, neutrophils or
polymorphonuclear leukocytes (PMNs), and then local and blood-borne
macrophages, are recruited to the site of inflammation. PMNs have their origin in
multi-potential stem cells in the bone marrow. They differentiate in the marrow
and are released in a mature form, containing a full complement of bactericidal
agents. They are short-lived cells which constitute 30-70% of the circulating white
blood cells (leukocytes). PMNs have cytoplasmic granules that are formed from
the Golgi apparatus. These membranous granules are called lysosomes and contain
the various bactericidal and digestive enzymes which can destroy bacterial cells
after engulfment. The contents of lysosomes include lysozyme, cationic proteins,
acid hydrolases, proteases, peroxidase and lactoferrin (Alberts et al., 2002). PMNs
also contain large stores of glycogen; since they derive most of their metabolic
energy from glycolysis, they can function efficiently in anaerobic environments.

Macrophages (also called mononuclear phagocytes) also arise from bone marrow
stem cells which give rise to promonocytes which develop into monocytes that are
released into the blood stream (Alberts et al., 2002). Monocytes make up 3-7% of
the circulating white blood cells and are actively phagocytic and bactericidal
(Alberts et al., 2002). Within 2 days or so, the blood stream monocytes (sometimes
called wondering macrophages) emigrate into the tissues where they settle down,
enlarge and become fixed macrophages (tissue histocytes), which also have
phagocytic potential. Macrophages are more active in phagocytosis than
monocytes and develop many more granules containing hydrolytic enzymes. New
macrophages can develop by cell division under inflammatory stimuli, but most
macrophages are matured blood monocytes. PMNs play a more important role in
the acute stages of an infection, while macrophages are principally involved in
chronic types of infections (Alberts et al., 2002). PMNs circulate in the blood
stream, and during an acute inflammatory response they migrate through the
endothelial cell junctions as part of the inflammatory exudate. They migrate to the
focus of the infection and ingest the foreign agents. PMNs which have become
engorged with bacteria usually die and largely make up the material of pus.
Macrophages, which are also attracted to the area during an inflammatory
response, are slower to arrive and become increasingly involved in chronic
infections. By 10 to 30 minutes after ingestion, many pathogenic and
nonpathogenic bacteria are killed followed by lysis and digestion of the bacteria by
lysosomal enzymes. The microbicidal activities of phagocytes are complex with
metabolic products, as well as lysosomal constituents, responsible for this diversity
in action. These activities differ to some extent in PMNs, monocytes and
macrophages. The microbicidal activities of phagocytes are usually divided into
oxygen-dependent and oxygen-independent events:

Oxygen-independent activity

Lysosomal granules contain a variety of extremely alkaline proteins that strongly

inhibit bacteria, yeasts and even some viruses (Alberts et al., 2002). A few
molecules of any one of these cationic proteins appear able to inactivate a bacterial
cell by damage to their permeability barriers, but their exact modes of action are
not known (Alberts et al., 2002). The lysosomal granules of neutrophils contain
lactoferrin, an extremely powerful iron-chelating agent, which withholds potential
iron needed for bacterial growth. The pH of the phagolysosome may be as low as
4.0 due to accumulation of lactic acid, which is sufficiently acidic to prevent the
growth of most pathogens. This acidic environment apparently optimizes the
activity of many degradative lysosomal enzymes including lysozyme,
glycosylases, phospholipases, and nucleases (Alberts et al., 2002).

Oxygen-dependent activity

Liganding of Fc receptors (on neutrophils, monocytes or macrophages) and

mannose receptors (on macrophages) increases their O2 uptake, called the
respiratory burst or "oxygen burst" (Alberts et al., 2002). These receptors activate a
membrane-bound NADPH oxidase that reduces O2 to O2- (superoxide). Superoxide
can be reduced to OH (hydroxyl radical) or dismutated to H2O2 (hydrogen
peroxide) by superoxide dismutase. O2, OH, and H2O2 are activated oxygen species
that are potent oxidizing agents in biological systems which adversely affect a
number of cellular structures including membranes and nucleic acids (Alberts et
al., 2002). Furthermore, at least in the case of PMNs, these reactive oxygen
intermediates can act in concert with a lysosomal enzyme called myeloperoxidase
(MPO).

MPO is one of the lysosomal enzymes of PMNs which is released into the
phagocytic vacuole during fusion to form the phagolysosome. MPO uses H2O2
generated during the respiratory burst to catalyze halogenation (mainly
chlorination) of microbes contained within the phagolysosome (Alberts et al.,
2002). Such halogenations are a potent mechanism for killing cells. When the
NADPH oxidase and myeloperoxidase systems are operating in concert, a series of
reactions leading to lethal oxygenation and halogenation of engulfed microbes
occurs (Alberts et al., 2002).

Cytotoxicity

Cytotoxic activity is generally carried out by CD8+ T lymphocytes (CTLs). Lytic

granules containing granzymes and perforins are released against target cell. The
perforins lyse cells by the formation of pores in the lipid bilayer. The pores consist
of polymers of perforin, which have a cylindrical structure that is lipophilic on the
outside and hydrophilic down a hollow center with an inner diameter of 16 nm
(Alberts et al., 2002). The granzymes are proteases, so although they have a role in
triggering apoptosis in the target cell, they cannot act directly to fragment the
DNA. Rather, they must activate an enzyme, or more probably an enzyme cascade,
in the target cell. Granzyme B can cleave the ubiquitous cellular enzyme CPP-32,
which is believed to have a key role in programmed cell death in all cells. CPP-32
is a caspase and activates a nuclease, called caspase-activated deoxyribonuclease
or CAD, by cleaving an inhibitory protein (ICAD) that binds to and inactivates
CAD (Alberts et al., 2002). This enzyme is believed to be the final effector of
DNA degradation in apoptosis.

Another mechanism of cell lysis exists for effector CTLs and Th1 cells. It involves
the binding of Fas in the target cell membrane by the Fas ligand, which is present
in the membranes of activated cytotoxic T cells and Th1 cells. Ligation of Fas
leads to activation of caspases, which induce apoptosis in the target cell (Alberts et
al., 2002).
Reference

Alberts, B., Johnson, A., Lewis, J., Raff, M., Roberts, K. and Walter, P. (2002). Molecular Biology of the
Cell 4th ed. New York: Garland Science.