pathology review 2

1.     What is the mechanism that leads to the manifestation of “tumor” or swelling at the site of inflammation?

Increased vascular permeability. Edema.  (exudate)

In response to injury, inflammatory soluble mediators (like histamine, bradykinin, and cytokines) are released, leading to increased vascular permeability. This allows plasma proteins and fluid to escape from the blood vessels into the interstitial space, contributing to local edema. Some mediators also generate pain.

Process:
1. Recognition of tissue damage and/or pathogens – DAMPs (danger molecules – host cell origin) and
PAMPs (pathogen molecules – microorganism origin)
2. Release of soluble mediators from cells or generation from circulation = cytokines, chemokines, and
vasoactive molecules
 a. Cytokines IL-1b or TNFa → activates cells to release vasoactive molecules
 b. Activation of complement cascade → generation of C5a and C3a, called “anaphylotoxins”
 because they are vasoactive
c. Generation of prostaglandins and leukotrienes → vasoactive
d. Release of histamine (cell-derived, mostly mast cells) → activated by cytokines and
complement
e. Bradykinins (from serum or tissue, activated by Factor XII of coagulation cascade)
3. Endothelial cells respond to vasoactive molecules

a. Retraction of endothelial cells to form intercellular gaps → vascular permeability and leakage
of protein rich fluid into extravascular space
i. = Edema (Tumor)
b. Relaxation of vascular smooth muscle → vasodilation
i. Slowing of blood flow, increased blood flow at the site of inflammation
ii. = Redness (Rubor); Calor = (Heat)
c. Pain mediators (Dolor) = prostaglandins, bradykinins
2. Contrast this mechanism from other primary mechanisms of edema/effusion formation (i.e. increased
hydrostatic pressure or decreased oncotic pressure)

2.     Contrast this mechanism from other primary mechanisms of edema/effusion formation (i.e. increased hydrostatic pressure or decreased oncotic pressure).

Increased hydrostatic pressure: elevated pressure in capillaries results from conditions like heart failure or venous obstruction, it pushes fluid out of vessels into surrounding tissues. Fluid is transudate (protein-poor).

Decreased oncotic pressure: determined by plasma proteins (albium), nephrotic syndrome, liver cirrhosis, malnutrition) fluid leaks out of vascular space. Formation of transudate- more generalized. Can affect areas like the legs.

Comparison: fluid from increased vascular permeability is exudate, protein rich. Vascular permeability changes are direct result of injury and inflammation, hydrostatic and oncotic pressure are systemic.

3.     To alleviate the inflammation (and the pain), you take an Advil or two. What is the target of Advil?

Advil targets the eicosanoid pathway, it is nonselective, it will target both cox-1 and cox-2.

Eicosanoids: Lipid mediators of inflammation, 5 types of phospholipid-derived molecules. Originate from endothelial cells, platelets and innate leukocytes upon activation or injury.

1.     Prostaglandins

2.     Leukotriens

3.     Thromboxans

4.     lipoxins

4.     How does this target inhibition by Advil work to interfere with inflammation?

Advil inhibits the induction of cox 1 and cox 2. Cox 1 is an enzyme responsible for the inflammatory response made by prostaglandins. The Advil targets the production of prostaglandin and, therefore, targets both cox 1 and cox 2.  inhibits the formation of prostaglandins and thromboxanes – the key
lipid mediators which mediate the vascular response and the cardinal signs of inflammation.

Prostaglandins: promote inflammation, travel to the dedicated blood supply, and tell vessels to dilate. More blood gets to areas and more white blood cells to fight infection. Increases vascular permeability. Stimulate nociception. Creates pain and changes the internal thermostat through the thalamus. Inhibit platelet aggregation.  Can also stimulate platelet aggregation.

You know it is likely that bacteria were introduced into the wound. Answer the questions below that address the responses and immune cell types involved in preventing establishment of a bacterial infection.

5.     This species of bacteria has never been “seen” before by your immune system. Which 2 of the 3 complement activation pathways are likely to contribute to complement-mediated elimination of this bacteria?

1.     Alternative Pathway: This pathway can be activated spontaneously on microbial surfaces, even in the absence of antibodies. It is crucial for the immediate response to new pathogens. The alternative pathway is continuously activated at low levels (tick-over), and when complement proteins interact with the surface of the bacteria, it leads to the formation of the C3 convertase, resulting in opsonization and lysis of the bacteria.

2.     Lectin Pathway: This pathway is activated when mannose-binding lectin (MBL) or other lectins bind to specific carbohydrates on the surface of the bacteria. Since this is a new species, the pattern recognition receptors in the lectin pathway can recognize common carbohydrate structures found on many bacteria, facilitating complement activation.

3.     classical pathway would typically require prior exposure to the bacteria and the formation of antibodies, so it is less likely to be involved in the immediate response to a previously unencountered pathogen. Thus, the alternative and lectin pathways are the primary contributors to complement-mediated elimination in this scenario.

6.     What is the first innate immune cell to respond to this infection and which chemokine is responsible for the specific recruitment of this cell?

Neutrophils: most numerous cell in innate immune response. The first wave of cells to enter inflamed tissue. Primarily driven by IL-8 (CXCL8).

Phagocytosis: pathogens are engulfed in the phagosome which acquires degradative properties by maturation.

The questions below pertain to the cellular interactions and sequence of events in the lymph node that drained this infected wound which ultimately led to an adaptive immune response that eliminated this bacterium.

7.     What cell type is most responsible for antigen presentation of bacterial peptides (proteins) to establish an adaptive immune response?

Dendritic cells: 3^rd class of phagocytic cells (granulocytes, macrophages). Dendritic cells engulf and degrade pathogens. The function is not for pathogen clearance, it is for antigen presentation and activation of T cells. Dendritic cells link innate and adaptive immunity by responding to PAMPS while immature(induce migration, enhance antigen processing, up-regulate co-stimulatory mol) the cells mature and are no longer phagocytic, they express high levels of antigen on MHC mol.

8.     Which antigen presentation molecule will be most likely used by this cell type to establish the adaptive immune response?

MHC molecules.

9.     List the “three signals” this cell type will provide to a naïve T cell in your lymph node to establish this adaptive immune response

Signal 1: MHC-peptide complex on surface of dendritic cell binding to the T cell receptor
Signal 2: Costimulation – CD80/86 molecules on surface of dendritic cell binding to CD28 molecule on
surface of T cell → leads to production of IL-2

Signal 3: Local cytokine milieu that drives T cell polarization → these cytokines come from DCs and other
innate immune cells in response to PRR activation by PAMPs

10.  In the adaptive immune response, both T cells and B cells are likely to contribute. What class of immunoglobulin (antibody) will be most likely produced by B cells in response to this bacterial infection?

IgM because it is always first until class switching occurs, then IgG.

IgG: most common type of antibody in blood, only antibody that can cross placenta

IgA: secreted at mucosal sites, neutralizes pathogens in these lumens (GI, resperitory)

IgM: first antibody in response to antigen

11.  What CD4 T cell-associated molecule is most critical to providing “help” to the B cells to increase the amount and affinity of this immunoglobulin for this bacterium?

CD40L (CD40 ligand). This is expressed on the surface of CD4+ helper T cells and binds CD40 on the surface
of B cells

12.  In response to this bacterial infection, what are the expected innate cell-derived cytokine(s), CD4 T-helper cell types, and T cell derived cytokine(s)?

Innate cytokines = CXCL8, IL12, TNFalpha, IL-1, IL-6
CD4+ helper T cell type = Th1 cells; their principal product is IFN gamma = activates macrophages to enhance
phagocytosis, pathogen killing/clearance activities among many other functions.

13.  List alternative receptors your immune system would use to recognize this pathogen if it were a virus and not a bacterium.

We are looking for other pattern recognition receptors (PRRs). For viruses, think of intracellular or
endosomal located PRRs: TLRs 3,7,8, and 9, RIG-I receptors

14.  If this were a viral infection, what would be the expected innate cell-derived cytokine(s), T cell response, and T cell derived cytokine(s)?

Innate cytokines = Type I interferons (IFNa and IFNb), IL-12, IFN gamma (NK cell derived)
T cell response = CD4+ Th1 cells and CD8+ cytotoxic lymphocytes and their cytokine products of IL-2, IFN
gamma

15.  What other innate immune cell type would also be critical to eliminating this viral infection? List the ways this cell can become activated and three mechanisms this cell uses to kill virally infected cells

Natural Killer (NK) cells
Activation occurs in response to: 1) Downregulation/loss of MHC class I expression; 2) Upregulation
of stress-induced ligands on infected cells; 3) Type I interferons
Mechanisms of killing: 1) Release of cytoplasmic granules containing perforin and granzyme →
osmotic lysis/induction of apoptosis; 2) Antibody-dependent cell mediated cytotoxicity (ADCC) →
antibody bound to cell surface antigen (e.g. of a tumor/transformed cell) then binds Fc receptor on
NK cell → end result is perforin/granzyme release; 3) IFN gamma – has direct cytotoxic and anti-
proliferative effects on cells

16.  If this were a parasitic infection, what would be the expected innate immune cells involved, T cell response, and T cell derived cytokine(s)?

Th2 cells, eosinophils, basophils, mast cells; IL-4, IL-5, IL-13, IgE

With appropriate clearance of the bacteria, the injury was able to resolve without substantial damage to the tissue. The questions below pertain to the sequence of events that led to the resolution of the inflammatory response and healing of the injury.

17.  Which innate immune cell is primarily responsible for resolution of inflammation and promotion of healing?

Macrophage

18.  What is the specific “polarization” state of this innate immune cell which helps to promote resolution of inflammation and healing? What are the molecules that would promote this polarization versus one of sustained inflammation?

M2/alternatively activated/anti-inflammatory macrophages - Immune suppressive/Anti-inflammatory
cytokines or Th-2 derived cytokines (TGFb and IL10); engulfment of apoptotic cells; absence of Th1
promoting stimuli like LPS/other PAMPs, IL-12 etc. → M2 macrophages mediate promotion of fibrosis,
angiogenesis/tissue repair

19.  What specific CD4 T cell subset is also effective at suppressing inflammatory responses? What is the transcription factor that drives differentiation of naïve T cells into this subset?

Regulatory T cells;FoxP3

20.  With this healing response, you are likely to develop a scar at the site of injury. What is the name of this process and what is the scar composed of?

Fibrosis/granulation tissue → collagens and blood vessels

Scenario 2: Early into a month-long backpacking trip through India, you find yourself on public transportation in the company of individuals with a productive cough. You don’t think much of it and expect that if this is transmissible, you may be sick within the next few days. However, you feel perfectly healthy and continue on with the remainder of your trip then return home. After describing this scenario at your next physical exam visit, your physician decides to test you for tuberculosis using the tuberculin skin test (TST). The nurse administers an injection of “purified protein derivative (PPD)” into the dermis (below the skin) of your right forearm and tells you to return 48 hours later to have the test evaluated. After 48 hours, there is swelling at the injection site on your arm, and when you return to the physician’s office, the TST test is interpreted as positive.

21.  What form of immune response is the TST test measuring (i.e. innate vs adaptive, cell-mediated vs. antibody)?

The TST test is measuring an adaptive/antigen-specific cell-mediated immune response to the proteins in the PPD

22.  What form of hypersensitivity is being evaluated by the TST test?

The TST test measures a type IV hypersensitivity, also known as Delayed Type Hypersensitivity (DTH)

23.  What major form of CD4 T-helper cell is involved in this response and which T cell derived cytokine is critical to this response?

Th1 cells and IFN gamma

24.  Using arrows between each step in the process, diagram the critical immune events occurring between initial exposure and infection with the Mycobacterium tuberculosis organism, followed by skin testing and the manifestation of swelling at the PPD injection site.

Initial exposure:
1. Aerosol exposure → inhalation of Mtb organism into the lung
2. Phagocytosis of the bacteria and pattern recognition by a sentinel cell = Alveolar macrophage in the
lung
3. Recruitment of antigen presenting cells (APCs) to the site of infection
4. Processing of the bacteria by the APC → presentation of antigen peptides by MHCII on the surface
5. Migration to the draining lymph node
6. Presentation and interaction of MHCII and B7 molecules of the APC with TCR and CD28 on CD4 T cells
a. Some presentation by MHCI will occur → activation of CD8 T cells
7. Proliferation of CD4 T cells and generation of T cell memory
Weeks later:
8. Injection of PPD into skin
9. Acquisition and presentation by APC
10. Activation of effector CD4 T cells from the memory cell population
11. Production of IFNg at the skin site
12. Recruitment of innate leukocytes and additional T cells to the skin site
13. Swelling from cellular response → Positive PPD test

25. How does the adaptive immunity in this hypersensitivity differ from the immunity developed in the
first scenario above?

The first scenario describes an innate inflammatory reaction and subsequent priming of naïve T and B cells
leading to the generation of an adaptive/antigen-specific immune response.
This TST test positive reaction requires that an antigen-specific memory T cell population was already
generated. The memory response must be “awakened” to become an effector response after recognizing
peptides (PPD proteins) on antigen presenting cells. The activated T cells then produce IFNg which activates
innate leukocytes and the inflammatory response recruits more leukocytes to the skin, leading to a swelling
that is the result of cellular response (as opposed to immediate hypersensitivity (type I) for which the
swelling is mediated heavily by the vascular response)

Definitions:

Circulation:

-       Perfusion: the passage of fluid through the circulatory system to an organ or tissue, usually referring to delivery of blood to cap bed in tissue

-       Ischemia: restriction of blood flow to tissue (poor profusion)

-       Hemorrhage: loss of blood from vasculature

-       Petechia: pinpoint hemorrhage, can be up to 1mm

-       Ecchymosis: larger hemorrhage up to a few cm

-       Purpura: petechia and ecchymoses on mucus membranes

-       Hemo or Hema: forms used to describe hemorrhage

-       Hyperemia: active increased blood flow

-       Congestion: passive, reduced outflow of blood

Heart anatomy:

-       4 chambers that perform the pumping function, heart is enclosed in the pericardium

-       Chamber walls have 3 layer architecture like vessels (epicardium, myocardium, endocardium)

-       Myocardium: myocytes, contains actin and myosin filaments in sarcomeres

-       Intercalated discs: coordinate contraction, intercellular junctions that facilitate mechanical and electrical coupling

-       Nutrients and O2 delivered via coronary, perfusion occurs during diastole

Edema and Effusions:

-       Edema and effusions develop when normal physiological homeostasis (small net movement of fluid into interstitium returned via lymphatic circ) is perturbed (the capacity of the lymphatic drainage system is exceeded)

-       Interstitium: the space between cells in tissues

-       Edema: excessive movement of fluid out of vessels and accumulation in tissues

-       Effusion: excessive movement of fluid out of vessels and accumulation in the body cavity

Mechanisms of edema formation:

-       Increased hydrostatic pressure: increase in fluid pressure within vasculature

-       Decreased plasma osmotic pressure: loss of ability of plasma proteins to retain/attract water

-       Lymphatic obstruction: decreased outflow of fluid from the interstitium

Edema and Effusions fluid characterization:

-       Transudate: non-inflammatory, low protein, and nucleated cell count with clear colorless fluids.

-       Exudates: inflammatory, results from increased vascular permeability, high protein and cell counts, infectious or noninfectious but from irritants

-       Chylous effusion: from trauma or obstruction or thoracic duct, usually in thorax, milky white from high lymph and fat content

-       Neoplastic effusion: tumor cells

-       Hemorrhagic: trauma or neoplasia

Inflammation:

Process:

Stimulus is recognized -> cells send signal for help -> vessels open for invited cells -> cells enter tissue through opening and contribute to response.

5 R’s of inflammation:

1.     Recognition of injurious event

2.     Recruitment of leukocytes

3.     Removal of agent

4.     Regulation of inflammatory response

5.     repair of tissue damage

Responses of acute inflammation:

-       vasodilation, increased vascular permeability, smooth muscle constriction, and immune cell recruitment.

-       Consequences of acute inflammation: redness, swelling, heat, pain, fever, nausea, tissue damage

Immune diseases:

-       B cells - generate antibodies = Large proteins that recognize specific
sequences or conformation on proteins

-        CD4 “helper” T cells - Recognize specific foreign proteins then function
to assist innate cells and adaptive cells with the immune response

-       CD8 “cytotoxic” T cells - Remove cells with intracellular infections, such
as virus

-       Hypersensitivity: release and production of inflammatory mediators caused by a hyperactive immune response to specific molecules. Requires adaptive immunity against spec mol. B cell production of specific antibody, CD4 helper or CD8 cytotoxic t cells. Antibody-dependent 1-4. T cell dependent 4.

-       Type I: antibody-dependent, dependent on generation of IgE antibody, IgE bound to Fc-E receptor.  immediate development of pathologic response. Ex. Allergy, anaphylaxis

o   Exudate, contains many eosinophils, responds to chemokines from mast cell and macrophage

-       Mast Cells: allergic and hypersensitive reactions, purple-colored basophilic granules. Mucosal. Promote acute inflammation, activation causes granule exocytosis, high expression of surface IgE

-       Eosinophil granules: activate mast cell-derived mediators, cytoplasmic granules contain: Major basic protein, arylsulfatase, histaminase, peroxidase, matrix metalloproteinase

-       Anaphylaxis: hyper-acute severe systemic response, low BP, resppiritory distress, itchiness, GI upset

-       Type 2: antibody dependent, IgG or IgM, bound antibodies mediate destruction of target, leads to cytoxicity (death of target). Increases or decreases cellular function. Autoimmune disease.

-       Type 3: antibody dependent, IgM or IgG, related to concentration of antigen, exogenos or autoantigens, critical component is formation of immune complexes. Kidney, joints, lung, eye

Type 4: delayed type, antibody independent, CD4 and CD8 T-cells, cause of many chronic immune diorders, chronic inflamation, destruction of autoimmune target