Phagocytosis and killing

The lecture asks: what does opsonin mean? The word comes from the Greek for “to prepare food”. An opsonin coats a microbe and gives the phagocyte a handle to grip. Phagocytes can recognize some microbes directly (mannose and scavenger receptors), but coated microbes are engulfed far more efficiently.

After binding, the cell extends pseudopods around the particle and closes them into a phagosome. Granules and lysosomes fuse with it to form a phagolysosome. Killing happens inside that compartment, so the most toxic products are kept away from the neutrophil's own cytoplasm, until the cell dies or regurgitates its contents and damages the surrounding tissue.

When a neutrophil is activated, its oxygen consumption jumps, not to make ATP but to make oxidants: the respiratory (oxidative) burst. The enzyme is NADPH oxidase (phagocyte oxidase), whose membrane and cytosolic subunits assemble on the phagosome membrane only when the cell is stimulated.

NADPH comes from the hexose monophosphate shunt, whose first enzyme is G6PD; very severe G6PD deficiency can weaken the burst. Iron can also turn H₂O₂ into hydroxyl radical (Fenton reaction), the same chemistry that injures cells in Chapter 1. Myeloperoxidase is stored in the azurophilic (primary) granules and gives pus its green tint.

Neutrophil granules hold the enzymes that finish the job in the phagolysosome: elastase, collagenases, gelatinase, cathepsins, lysozyme and myeloperoxidase. The same contents also escape into the tissue, which is why inflammation damages the host it protects.

Where neutrophils accumulate densely around pyogenic bacteria, these proteases and oxidants digest bacteria, dead neutrophils and the surrounding tissue alike: liquefactive necrosis. The liquid product is pus, and a localized collection of it is an abscess (2.5). Antiproteases in plasma and tissue fluid, such as α₁-antitrypsin and α₂-macroglobulin, normally limit the spread; when they are overwhelmed or deficient, the damage extends (emphysema in α₁-antitrypsin deficiency).

Chédiak–Higashi syndrome (autosomal recessive) is caused by mutations in LYST, a lysosomal trafficking regulator. The lecture calls it a protein-trafficking defect that disturbs microtubule-guided movement, so lysosomes fail to fuse with phagosomes. Lysosome-related organelles instead fuse with each other into giant granules, and every cell type that uses such organelles is affected.

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In chronic granulomatous disease (CGD), NADPH oxidase does not work. About two-thirds of cases are X-linked (the gp91phox subunit, gene CYBB); the rest are autosomal recessive. Neutrophils are recruited normally, engulf normally and form pus, but cannot make superoxide, so their own burst produces no H₂O₂ and no HOCl.

Why catalase matters. Many bacteria generate H₂O₂ themselves. If a catalase-negative organism (for example a streptococcus) is engulfed, the CGD neutrophil can borrow that H₂O₂, and its intact myeloperoxidase turns it into HOCl. Catalase-positive organisms destroy their own H₂O₂ (2 H₂O₂ → 2 H₂O + O₂), leaving the neutrophil nothing to work with. Those organisms survive: **Staphylococcus aureus, Serratia marcescens, Burkholderia cepacia, Nocardia, Aspergillus* and Candida*.

Organisms that survive inside phagocytes keep stimulating T cells and macrophages, so neutrophils are joined by lymphocytes and macrophages that fuse into giant cells: granulomas, often with abscesses, in lymph nodes, lungs, liver, skin and bone. Pneumonia is the most common presentation; Aspergillus is the leading cause of death.

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Testing the burst. In the nitroblue tetrazolium (NBT) test, stimulated neutrophils that make superoxide reduce the yellow dye to dark blue formazan; blue cells mean the burst is intact (a positive test). CGD neutrophils stay unstained. The more sensitive dihydrorhodamine (DHR) flow cytometry test measures oxidation of DHR to fluorescent rhodamine after stimulation with PMA, a protein kinase C agonist: CGD cells do not fluoresce. Mothers who carry X-linked CGD show two populations, one normal and one not, because of X inactivation.

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Myeloperoxidase deficiency (autosomal recessive) is the most common inherited phagocyte defect. The burst runs normally and makes plenty of H₂O₂, which alone kills most bacteria, but no HOCl. Most people never know they have it. The exception is *Candida*: disseminated or persistent candidiasis, especially in people with diabetes, whose neutrophils already work less well.

Five inherited defects, five places where the same pathway can break. For each one you are told only what is molecularly wrong. Predict the blood and the tissue first; the lab then shows where the neutrophil stops and which clinical clue gives it away.