Phagocytosis and related processes like endocytosis and exocytosis allow cells to import and export materials across their plasma membranes. Phagocytosis specifically refers to the ingestion and destruction of large particles by phagocytes like macrophages and neutrophils. This involves receptors recognizing particles, engulfment within a phagosome, and fusion with lysosomes to form a phagolysosome where digestion occurs through oxidative and non-oxidative mechanisms. Careful regulation of phagocytosis, inflammation, and clearance of dead cells is important for immune function and tissue homeostasis.

1. EXOCYTOSIS AND
ENDOCYTOSIS

2. Exocytosis and Endocytosis:
Transporting Material Across the
Plasma Membrane
• Two methods (unique to eukaryotes) for
transporting materials across the plasma
membrane are
- Exocytosis, the process by which secretory vesicles
release their contents outside the cell
- Endocytosis, the process by which cells internalize
external materials

3. Exocytosis Releases Intracellular
Molecules Outside the Cell
• In exocytosis, proteins in a vesicle are released to
the exterior of the cell as the vesicle fuses with the
plasma membrane
• Animal cells secrete hormones, mucus, milk
proteins, and digestive enzymes this way
• Plant and fungal cells secrete enzyme and
structural proteins for the cell wall

4. The process of exocytosis
• Vesicles containing products for secretion move
to the cell surface (1)
• The membrane of the vesicle fuses with the
plasma membrane (2)
• Fusion with the plasma membrane discharges the
contents of the vesicle (3)
• The membrane of the vesicle becomes part of the
cell membrane (4)

5. Figure 12-12A

6. Orientation of membrane
• When the vesicle fuses with the plasma membrane
- The lumenal (inner) membrane of the vesicle
becomes part of the outer surface of the plasma
membrane
- So, glycolipids and glycoproteins that were formed
in the ER and Golgi lumens will face the
extracellular space

7. Figure 12-12B

8. Mechanism of exocytosis
• The mechanism of the movement of exocytotic
vesicles to the cell surface is not clear
• Evidence points to the involvement of
microtubules in vesicle movement
• Vesicle movement stops when cells are treated
with colchicine, a microtubule assembly inhibitor

9. The Role of Calcium in Triggering
Exocytosis
• Fusion of regulated secretory vesicles with
the plasma membrane is generally triggered
by an extracellular signal
• In most cases a hormone or neurotransmitter
binds receptors on the cell surface and
triggers a second messenger inside the cell
• A transient elevation in Ca2+ appears to be
an essential step in the signaling cascade

10. Polarized Secretion
• In many cases, exocytosis of specific proteins is
limited to a specific surface of the cell
• For example, intestinal cells secrete digestive
enzymes only on the side of the cell that faces
into the intestine
• This is called polarized secretion

11. Endocytosis Imports Extracellular
Molecules by Forming Vesicles from
the Plasma Membrane
• Most eukaryotic cells carry out one or more
forms of endocytosis for uptake of extracellular
material
• A small segment of the plasma membrane folds
inward (1)
• Then it pinches off to form an endocytic vesicle
containing ingested substances or particles (2-4)

12. Figure 12-13

13. Membrane flow
• Endocytosis and exocytosis have opposite
effects, in terms of membrane flow
• Exocytosis adds lipids and proteins to the plasma
membrane, whereas endocytosis removes them
• The steady-state composition of the plasma
membrane results from a balance between the
two processes

14. Phagocytosis
• The ingestion of large particles up to and
including whole cells or microorganisms is called
phagocytosis
• For many unicellular organisms it is a means of
acquiring food
• For more complex organisms, it is usually
restricted to specialized cells called phagocytes

15. Phagocytes in immune function
• In humans, two types of white blood cells use
phagocytosis as a means of defense
• Neutrophils and macrophages engulf and digest
foreign materials or invasive microorganisms found
in the bloodstream or injured tissues
• Macrophages are also scavengers, ingesting
cellular debris and damaged cells

16. Phagocytosis
• Phagocytosis, defined as the cellular uptake of
particulates (>0.5 m) within a plasma-membrane
envelope
• E’lie Metchnikoff (1845–1916) made his seminal
studies in the 1880s and extensively explored
the role of phagocytosis is cellular immunity
• Was awarded the Nobel Prize, which he shared
in 1908 with Paul Ehrlich

17. Phagocytosis
• Phagocytosis is a specific form of endocytosis by
which cells internalise solid matter, including
microbial pathogens.
• Professional phagocytes include monocytes,
macrophages, neutrophils, dendritic cells,
osteoclasts, and eosinophils.
• These cells are in charge of eliminating
microorganisms and of presenting them to cells of
the adaptive immune system.

18. Phagocytosis
• In addition, fibroblasts, epithelial cells, and
endothelial cells can also perform phagocytosis.
• These nonprofessional phagocytes cannot ingest
microorganisms but are important in eliminating
apoptotic bodies

19. Phagocytosis
• In these cells, phagocytosis is a mechanism
by which microorganisms can be contained,
killed and processed for antigen presentation
and represents a vital facet of the innate
immune response to pathogens, and plays an
essential role in initiating the adaptive
immune response.

20. Phagocytosis
• Phagocytes must recognize a large number of
different particles that could potentially be
ingested, including all sorts of pathogens and also
apoptotic cells.
• This recognition is achieved thanks to a variety of
discrete receptors that distinguish the particle as a
target and then initiate a signaling cascade that
promotes phagocytosis.
• Receptors on the plasma membrane of
phagocytes can be divided into nonopsonic or
opsonic receptors.

21. Phagocytosis
• Nonopsonic receptors can recognize directly
molecular groups on the surface of the phagocytic
targets called pathogen associated molecular
patterns (PAMPS). Among these receptors, called
pattern recognition receptors (PRRs), there are
lectin-like recognition molecules, such as CD169
and CD33; also related C-type lectins, such as
Dectin-2, Mincle, or DNGR-1; scavenger receptors
; and Dectin-1, which is a receptor for fungal beta-
glucan.

22. Phagocytosis
• Interestingly, toll-like receptors (TLRs) are
detectors for foreign particles, but they do not
function as phagocytic receptors. However, TLRs
often collaborate with other nonopsonic receptors
to stimulate ingestion

23. Phagocytosis
• Opsonic receptors recognize host-derived
opsonins that bind to foreign particles and target
them for ingestion.
• Opsonins include antibodies, complement,
fibronectin, mannose-binding lectin, and milk fat
globulin (lactadherin).
• The best characterized and maybe most important
opsonic phagocytic receptors are the Fc receptors
(FcR) and the complement receptors (CR)

24. Phagocytosis
•
Human phagocytic
receptors and their ligands.
Receptor Ligands Reference(s)
Pattern-recognition
receptors
Dectin-1 Polysaccharides of some yeast cells [29]
Mannose receptor Mannan [30]
CD14 Lipopolysaccharide-binding protein [31]
Scavenger receptor A Lipopolysaccharide, lipoteichoic acid [32, 33]
CD36
Plasmodium falciparum-infected
erythrocytes
[40]
MARCO Bacteria [41]
Opsonic receptors
FcγRI (CD64) IgG1 = IgG3 > IgG4 [42]
FcγRIIa (CD32a) IgG3 ≥ IgG1 = IgG2 [42]
FcγRIIIa (CD16a) IgG [42]

25. Phagocytosis
• The cell membrane then extends around the
target, eventually enveloping it and pinching-
off to form a discreet phagosome.
• This vesicle can mature and acidify through
fusion with late endosomes and lysosomes to
form a phagolysosome, in which
degradation of the contents can occur via the
action of lysosomal hydrolases.

26. The Process of Phagocytosis
~A series of complex steps allowing phagocytes
to engulf and destroy invading
microorganisms.
~Most pathogens have evolved an ability to
evade one or more of the steps (resistance).

27. © 2012 Pearson Education, Inc.
Step 1
• Chemotaxis- Phagocytic cells are recruited to
site of infection or tissue damage by chemical
stimuli (chemoattractants).

28. © 2012 Pearson Education, Inc.

29. Step 2
• Recognition & Attachment- Receptors located
on outside of phagocyte recognize and bind
(directly or indirectly).
~Direct binding-receptors recognize and bind to
patterns of compounds found on invaders
~Indirect binding-particle is opsonized, coating
particle with antibody substance for easier
ingestion

30. Source: Undetermined

31. Step 3
• Engulfment-Phagocytic cell engulfs invader, forming a
membrane-bound vacuole called a phagosome.
~Cytoskeleton of phagocyte rearranges to form armlike extensions
(pseudopods) that surround material being engulfed.

32. Step 4
• Phagolysosome Maturation
• The phagosome changes its membrane
composition and its contents, to turn into a
phagolysosome, a vesicle that can destroy the
particle ingested.
• This transformation is known as phagosome
maturation and consists of successive fusion and
fission interactions between the new phagosome
and early endosomes, late endosomes, and finally
lysosomes.

33. Step 4
• At the end, the mature phagosome, also called
phagolysosome, has a different membrane
composition, which allows it to contain a very acidic
and degradative environment

34. Step 5
• Destruction & Digestion-Oxygen consumption
increases, sugars metabolized (aerobic
respiration), highly toxic oxygen products
produced (superoxide, hydrogen peroxide,
singlet oxygen, hydroxyl radicals).
~As available O2 in phagolysosome is
consumed metabolic pathway switches to
fermentation, producing lactic acid and
lowering pH.
~Enzymes degrade peptidoglycan of the
bacterial cell walls, and other parts of the cell.

35. Step 6
• Exocytosis-membrane-bound vesicle
containing digested material fuses with the
plasma membrane. Material is expelled to
the external environment.

36. Phagocytosis of bacteria

37. Intracellular Killing
• There are two mechanisms of killing:
• Oxygen dependent killing
• Oxygen independent killing

38. Intracellular Killing and
Digestion
• Several minutes after phagolysosome formation,
the first detectable effect on the microorganism is
the loss of the ability to reproduce.
• Inhibition of macromolecular synthesis occurs
sometime later and many pathogenic and non-
pathogenic bacteria are dead 10 to 30 minutes
after ingestion.
• The mechanisms phagocytes use to carry out this
killing are diverse and complex.

39. Intracellular Killing and
Digestion
• Oxygen-dependent mechanisms
• Binding of Fc receptors on neutrophils,
monocytes and macrophages (also binding of
mannose receptors on macrophages) causes
an increase in oxygen uptake by the
phagocyte called the respiratory burst.
• This influx of oxygen leads to creation of
highly reactive small molecules that damage
the biomolecules of the pathogen.
•

40. Intracellular Killing and
Digestion
• Oxygen-dependent mechanisms
• NADPH oxidase reduces O2 to O2
- (superoxide).
Superoxide can further decay to hydroxide radical
(OH.) or be converted into hydrogen peroxide
(H2O2) by the enzyme superoxide dismutase.

41. Intracellular Killing and
Digestion
• Oxygen-dependent mechanisms
• In neutrophils, these oxygen species can act in
concert with the enzyme myeloperoxidase to form
hypochlorous acid (HOCl) from H2O2 and chloride
ion (Cl-). HOCl then reacts with a second molecule
of H2O2 to form singlet oxygen (1O2), another
reactive oxygen species.

42. Intracellular Killing and
Digestion
• Oxygen-dependent mechanisms
• Macrophages in some mammalian species
catalyze the production of nitric oxide (NO) by the
enzyme nitric oxide synthase. NO is toxic to
bacteria and directly inhibits viral replication.

43. Intracellular Killing and
Digestion
• Oxygen-dependent mechanisms
• It may also combine with other oxygen species to
form highly reactive peroxynitrate radicals. All of
these toxic oxygen species are potent oxidizers
and attack many targets in the pathogen.
• At high enough levels, reactive oxygen species
overwhelm the protective mechanisms of the
microbes, leading to their death..

44. Intracellular Killing and
Digestion
• Oxygen-independent mechanisms
• The pH of the phagolysosome can be as low
as 4.0 and this alone can inhibit the growth of
many types of microorganisms, enhances the
activity of lysozyme, glycosylases,
phospholipases and nucleases present in the
phagolysosome that degrade various parts of
the microbe.

45. Intracellular Killing and
Digestion
• Oxygen-independent mechanisms
• A variety of extremely basic proteins present
in lysosomal granules strongly inhibit
bacteria, yeast and even some viruses.
• The phagolysosome of neutrophils also
contains lactoferrin, an extremely powerful
iron-chelating agent that sequesters most of
the iron present, potentially inhibiting bacterial
growth.

46. Summary of killing
mechanisms

47. Macrophage activation
• Characteristics
~Toll-like receptors-allow them to sense dangerous
materials.
~Produce pro-inflammatory cytokines (M1), alerting
other cells in the immune system.
~Activated macrophages-increases killing power with
assistance from certain T cells. This cooperation
between innate and adaptive host defenses induces
production of nitric oxide and oxygen radicals,
helping to destroy microbes.

48. ~ If activated macrophages fail to destroy
microbes and chronic infection occurs, large
numbers can fuse together forming giant cells.
~Granulomas- concentrated groups of
macrophages, T cells, giant cells. Contain
organisms and material that can’t be destroyed
by walling off and retaining the debris to
prevent infection of more cells. Granulomas
are commonly part of the disease process in
TB, histoplasmosis, and other diseases.

49. Efferocytosis
• Efferocytosis is the guarding mechanism to
remove dying/dead cells from tissues during
growth and remodeling and is executed
primarily by tissue MΦ
• During infection, large numbers of cells
involved in host defense succumb to cell
death. These cells have to be removed to
limit tissue damage and inflammation.

50. Efferocytosis
• As these cells are also parasitized by
intracellular pathogens, which now lose their
niches to cell death, the need to contain the
infection makes efferocytosis an essential
process during the host response to
intracellular bacteria.
• Removal of senescent and dead cells is
therefore essential to maintain tissue
homeostasis and integrity and to promote
healing

51. Efferocytosis
• Under homeostatic conditions within the
body, macrophages are the prime cells to
clear out apoptotic bodies and remnants
thereof as well as necrotic material

52. Leucocytes and inflammation
• Inflammation is the body’s normal response
to tissue injury and infection with the purpose
of repairing any damage and returning tissue
to a healthy state.
• It is a highly regulated process with both pro
and anti-inflammatory components that work
together to ensure a quick resolution and
restoration

53. Leucocytes and inflammation

54. • The recruitment of leukocytes to the sites of
inflammation and leukocyte-derived inflammatory
mediators contributes to the development of tissue
injury associated with inflammatory diseases.
• The first step in the pathogenesis of inflammatory
conditions is adhesion of circulating leukocytes to
activated vascular endothelial cell in the inflamed
tissues and subsequent transmigration through the
endothelial cells.
Leucocytes and inflammation

55. • During these processes, leukocytes are activated to
secrete a variety of substances such as growth
factors, chemokines and cytokines, complement
components, proteases, nitric oxide, and reactive
oxygen metabolites, which are considered to be one
of the primary sources of the tissue injury.
Leucocytes and inflammation

56. Inhibit adherence: M protein,
capsules
Streptococcus pyogenes, S. pneumoniae
Kill phagocytes: Leukocidins Staphylococcus aureus
Lyse phagocytes: Membrane
attack complex
Listeria monocytogenes
Escape phagosome Shigella
Prevent phagosome-lysosome
fusion
HIV
Survive in phagolysosome Coxiella burnetti
Microbial Evasion of
Phagocytosis

57. Receptor-Mediated Endocytosis
• Cells acquire some substances by receptor-
mediated endocytosis (or clathrin-dependent
endocytosis)
• Cells use receptors on the outer cell surface to
internalize many macromolecules
• Mammalian cells can ingest hormones, growth
factors, serum proteins, enzymes, cholesterol,
antibodies, iron, viruses, bacterial toxins

58. Low-density lipoproteins
• Low-density lipoproteins (LDL) are internalized
by receptor-mediated endocytosis
• The internalization of LDL carries cholesterol into
cells
• The study of hypercholesterolemia and
connection to heart disease led to the discovery
of receptor-mediated endocytosis and a Nobel
Prize for Brown and Goldstein

59. Process of receptor-mediated endocytosis
• Specific molecules (ligands) bind to their
receptors on the outer surface of the cell (1)
• As the receptor-ligand complexes diffuse
laterally they encounter specialized regions
called coated pits, sites for collection and
internalization of these complexes (2)
• In a typical mammalian cell, coated pits occupy
about 20% of the total surface area

60. Figure 12-15

61. Process of receptor-mediated endocytosis
(continued)
• Accumulation of complexes in the pits triggers the
accumulation of additional proteins on the
cytosolic surface of the membrane
• These proteins—adaptor protein, clathrin,
dynamin—induce curvature and invagination of
the pit (3)
• Eventually the pit pinches off (4), forming a
coated vesicle

62. Process of receptor-mediated endocytosis
(continued)
• The clathrin coat is released, leaving an
uncoated vesicle (5)
• Coat proteins and dynamin are recycled to the
plasma membrane and the uncoated vesicle
fuses with an early endosome (6)
• The process is very rapid and coated pits can be
very numerous in cells

63. Figure 12-16

64. Several variations of receptor-mediated
endocytosis
• Epidermal growth factor undergoes endocytosis
and is a signal that stimulates cell division
• As EGF receptors are internalized, the cell
becomes less responsive to EGF,
desensitization
• Defective desensitization through failure to
endocytose the receptor can lead to excess cell
proliferation and possible tumor formation

65. Variations of receptor-mediated
endocytosis
• Receptors may be concentrated in coated pits
independent of ligand binding
• In this case, ligand binding triggers
internalization
• In another variation (e.g., LDL receptors)
receptors are constitutively concentrated and
constitutively internalized independent of ligand
binding

66. After internalization
• Uncoated vesicles fuse with vesicles budding from
the TGN to form early endosomes
• Early endosomes are sites for sorting and recycling
of materials brought into the cell
• Early endosomes continue to acquire lysosomal
proteins from the TGN and mature to form late
endosomes, which then develop into lysosomes

67. Recycling plasma membrane receptors
• Receptors from the plasma membrane are
recycled due to acidification of the early endosome
• The pH gradually lowers as the endosome
matures, facilitated by an ATP-dependent
proton pump
• The lower pH dissociates ligand and receptors,
allowing receptors to be returned to the membrane

68. Clathrin-Independent Endocytosis
• Fluid-phase endocytosis is a type of pinocytosis
for nonspecific internalization of extracellular fluid
• This process does not concentrate the ingested
material, and contents are routed to early
endosomes
• It proceeds fairly constantly and compensates for
membrane segments added by exocytosis