Macrophages & Neutrophils

• Macrophages


  Activated macrophages produce cytokines
• IL-1 & TNF


  • Activate endothelial cells lining the blood vessels
  • Increase the expression of adhesion molecules and neutrophil chemo-attractants such as IL-8
  • Early response (4-96 hours)
• IL-8


  • Recruit neutrophils from the blood into tissues
  • Enhance the binding of neutrophils to adhesion molecules
  • Early response (4-96 hours)
• IL-6 & IL-12


  • Activate lymphocytes
  • Induces maturation of T-cells
  • Increases antibody production by B-cells
  • Adaptive response
• Early Host Response


  1. Macrophages are stimulated by soluble products of bacteria

  2. Activated macrophage directs the early host response to infection which is to bring in another phagocyte, the neutrophil, from the blood to the site of infection

  3. Neutrophils are present in the blood in large numbers and have a short half life of only 6 hours, so many are made and lost each day

  4. The macrophage and the neutrophil play an important role in this inflammatory response that occurs 4 - 96 hours after exposure to infection

  5. The macrophage synthesises and releases cytokines, which are chemical signals that communicate between cells

  6. The cytokines stimulate neutrophil recruitment from the blood
• Systemic Effects


  • Cytokines also have long range systemic effects when they enter the bloodstream
  • IL-1, TNF and IL-6 are endogenous pyrogens, ie they increase body temperature, which decreases bacterial replication. They also stimulate the acute phase response, ie the synthesis of proteins in the liver that increase activation of complement
  • IL-6 and IL-12 also enhance the adaptive immune response
• The Early Response


  1. Neutrophil recruitment and inflammation

  2. Neutrophils leave the blood vessels during early inflammation response

  3. During inflammatory response, vessels dilate bringing more blood to the area

  4. The permeability of the blood vessel wall increases and leads to accumulation of fluid in the tissue causing swelling and pain

  5. Inflammation is characterised by redness (rubor), heat (calor), pain (dolor) & swelling (tumor)

  6. Neutrophils are able to interact with the blood vessel wall and start to migrate

  7. The movement of cells across the blood vessel wall is carefully regulated by cytokines produced by the macrophage, which orchestrate this response
• Migration of Neutrophils – Step 1 – Rolling Adhesion


  1. Recognition by endothelial cell surface selectin molecules (E-Selectin) of carbohydrate residues on the neutrophil surface

  2. Carbohydrate residues belong to glycoproteins on neutrophil surface

  3. Weak and reversible interaction that causes the neutrophil to slow down and roll along endothelial surface

  4. It also enhances the interaction of adhesion molecules in the 2nd step

  5. E-Selectin expression is increased by TNF which is released by activated macrophages at tissue sites of infection

  6. Neutrophil recruitment occurs at tissue specific sites
• Migration of Neutrophils – Step 2 – Adhesion Molecules

  1. Adhesion molecules such as lymphocyte function associated antigen-1 (LFA-1) on neutrophils bind to the intracellular adhesion molecule-1 (ICAM-1) on the endothelial

  2. Chemokines are a family of proteins that attract white blood cells

  3. IL-8 is a chemokine and a very potent attractant for neutrophils

  4. IL-8 made by endothelial cells is bound to the cell surface and interacts with specific receptors on the neutrophil to direct the movement of neutrophils towards site of infection

  5. When IL-8 binds to its receptor, the binding between adhesion molecules o the neutrophil and endothelium is made stronger and the neutrophils stop rolling

  6. Expression of both ICAM-1 and IL-8 on endothelium is increased by TNF
• Migration of Neutrophils – Step 3 – Diapedesis

  1. Neutrophils respond to concentration gradients of IL-8 which acts as a chemoattractant and move across the blood vessel wall into the tissue where IL-8 is present at higher concentrations than in the blood

  2. The neutrophil squeezes between endothelial cells and penetrates the basement membrane to reach the site of infection
• Migration of Neutrophils – Step 4

  1. The migration of neutrophils through the tissue matrix under the influence of concentration gradients of IL-8

  2. Migration involves the interaction of the neutrophil with components of the matrix and the neutrophil releases proteolytic enzymes that degrade the matrix and increase rate of migration

  3. At site of infection neutrophil phagocytoses and kills bacteria, supporting role of macrophage
• Neutrophil Recruitment Through Tissue

  1. Neutrophils migrate into tissues by a process called haptotaxis, that is, they move up a gradient of chemoattractants that are bound to the tissue matrix or stroma

  2. These chemoattractants are produced by bacteria, and by stromal cells in the tissue matrix such as fibroblasts and epithelial cells

  3. The cells crawl over the surface of the matrix towards high concentrations of chemoattractants

  4. The stromal cells release chemokines such as IL-8, and also two lipid mediators leukotriene B4 (LTB4) and platelet activating factor (PAF)

  5. The neutrophils initially respond to these chemoattractants, but their ultimate goal is the bacterial infection

  6. The bacteria release peptides with a formyl group attached (formyl peptides) that are not made by human cells, and they activate complement

  7. The neutrophils move towards the bacteria as they are directed by concentration gradients of bacterial peptides, and activated complement, in this case complement component C5a

  8. Once in the vicinity of the bacteria they are able to undertake the tasks of bacterial phagocytosis and killiing
• Phagocytosis & Bacterial Killing

  • The neutrophil has a multi-lobed nucleus, and is often called a polymorphonuclear cell (PMN)
  • The cytoplasm is packed with many dense granules that fall into 3 categories; the primary (azurophil) granules, the secondary (specific) granules and the tertiary granules
  • Each of these contain different anti-microbial proteins
  • In addition there are small secretory vesicles that because they contain plasma proteins such as albumin are thought to be formed by endocytosis
  • Neutrophil phagocytosis involves two different receptor classes, those that recognise activated complement such as CR1 and CR3 and those that recognise immunoglobulins (FcgR)
  • Complement components opsonise bacteria, and in addition immunoglobulins (or antibodies) also opsonise bacteria
  • Once bound, the target bacteria are ingested into a phagosome
  • Once in the phagosome, primary and secondary granules fuse with the phagosome and release their contents on to the bacteria to kill them
• Proteases


  Elastase, cathepsin G and proteinase 3, enzymatically breakdown components of bacterial cell walls, making them susceptible to peptide antibiotics such as the defensins
• Defensins


  Small peptides that have antimicrobial properties because they are highly basic (positively charged) peptides that act by inducing the permeability of microbial membranes, and act on both gram negative and gram positive bacteria
• BPI


  Bactericidal/permeability increasing protein binds endotoxin (LPS) and is therefore toxic to gram negative bacteria
• Reactive oxygen


  • The NADPH oxidase enzyme transfers electrons to molecular oxygen and generates highly reactive free radicals called superoxides, that leads to the generation of a range of antimicrobials
  • First superoxide dismutase (SOD) converts superoxide to hydrogen peroxide (H2O2), and in the presence of normal concentrations of chloride ions and the enzyme myeloperoxidase (MPO) generates hypochlorous acid (the main ingredient of household bleach) which is a very effective antimicrobial