Week 1 - Wednesday

Question 1

Where is the spleen located?

Answer 1

The spleen resides in the left upper quadrant, retroperitoneal, along ribs 9, 10 and 11.

Question 2

An immunoglobulin is isolated and identified. It is found at low levels in serum, but plays a major role in fighting helminthic infections. What is the likely class of this immunoglobulin?

Answer 2

IgE:

Before binding to antigen, IgE binds tightly to Fc receptors on basophils and mast cells — thus, it is the least common Ig in serum.

Once bound to the Fc receptors on basophils and mast cells, binding of cognate antigen (e.g., allergens) by IgE triggers release of cytokines and other mediators involved in the allergic response — this is a Type 1HSR (hypersensitivity reaction).

During an infection with invasive helminths (e.g., Strongyloides, Trichinella, Ascaris, Necator, Ancylostoma), IgE mediates an ADCC (antibody-dependent cell-mediated cytotoxicity) response:

IgE binds to the surface of the helminth → eosinophils (which have Fc receptors for IgE, aka FcεRs) bind to the free Fc “tail” of the IgE, triggering eosinophil degranulation → release of cytotoxic mediators (e.g., lytic enzymes, perforin, granzymes, tumor necrosis factor) to try to kill the helminth.

Serum levels of IgE tend to rise during infections with invasive helminths (e.g., Strongyloides, Trichinella, Ascaris, Necator, Ancylostoma).

Question 3

What is the pathway blood travels through in the spleen?

Answer 3

Blood is filtered through the spleen via the following route: splenic artery → trabecular arteries → central arteries (surrounded by white pulp) → sinusoids (in red pulp) → red pulp veins → trabecular veins → splenic vein (notice how blood flows from inside the spleen towards the capsule, i.e. white pulp to red pulp)

Question 4

With respect to immunology, define the term: “mature naïve lymphocytes”.

Answer 4

Definition:
“Mature naïve lymphocytes” = immunocompetent B- and T-cells which have already completed the selection process in the bone marrow and thymus, respectively, but have not yet been stimulated by foreign antigen.

“Mature” = immunocompetent = capable of mounting an immune response following antigenic stimulation

“Naïve” = not yet exposed to cognate antigen

Question 5

What is the arterial supply of the spleen? What makes this unusual?

Answer 5

The spleen is derived, around the 6th week of development, from mesenchymal cells of the dorsal mesentery. Despite its mesenchymal origin, the spleen still shares blood supply with organs of the foregut. Specifically, the spleen receives blood from the splenic artery, the largest branch of the celiac trunk.

A posterior perforation of the stomach, typically due to an ulcer, may lead to perforation of the splenic artery.

Question 6

What type of immunity does the complement system represent?

Answer 6

Innate immunity: includes many cellular and humoral devices, from nonspecific barriers (e.g., skin) to recognition and destruction of specific pathogens (e.g., complement-mediated opsonization of microorganisms for subsequent phagocytosis).

Innate immunity is maintained by a vast repertoire of serum proteins that recognize general classes of invading agents and which constantly sample the local environment. This provides for a fast immune response; however the proteins do not exhibit any memory in the case of recurrent infection.

The innate immune system involves:

• NK (natural killer) cells
• Phagocytes, neutrophils
• Dendritic cells
• Complement

Question 7

An ambitious college student is studying T-cell activation. On the topic of lymphocyte activation, which of the following is incorrectly paired?

AMHC-II and TCR

BB20 and CD 28

CMHC-I and TCR

DAPC and T helper cell

EB7 and CD 28

Answer 7

BB20 and CD 28

The first signal in T helper cell activation is foreign antigen presented on the MHC-II on the APC and recognized by the TCR of the T helper cell. The second signal is a costimulatory signal involving interaction of B7 from the APC-antigen presenting cell with CD28 on the T helper cell. Activation of the cytotoxic T cell involves the first signal, which involves endogenously synthesized (viral or self) proteins that are presented on MHC-I of the virus infected cell and recognized by TCR on the T cytotoxic cell. B20 is an associated marker on B cells and is not directly involved with T cell activation.

Cytotoxic (CD8+) T cell activation:

First signal: Antigen presentation via MHC-I of a virus-infected cell. This is recognized by TCR on the cytotoxic T cell. The CD8 co-receptor stabilizes this interaction.

Second signal: IL-2 secreted by Th cells binds IL-2R on CD8+ cells → activation to kill the virus infected cell.

Question 8

What are the two major components of the white pulp?

Answer 8

The white pulp is a major center for production and development of lymphocytes. Within the white pulp, exists the periarteriolar lymphatic sheath (PALS) and lymphoid nodules called Malpighian corpuscles (not to be confused with Malpighian renal corpuscles).

The periarterial lymphatic sheath (PALS) surrounds the central arteriole in the white pulp and is composed of T cells. Dendritic cells may also be found in the PALS area, functioning as APCs for their neighboring T cells.

The lymphoid nodules contain germinal centers, which are primarily composed of B cells.

Question 9

What cells are B cells derived from? What type of B cell is released into the circulation from the bone marrow?

Answer 9

Lymphoid stem cell → pro-B cell (progenitor) → pre-B cell (cytoplasmic μ) → immature B cell (surface IgM), which is released into the circulation and is called a mature (naïve) B cell when it simultaneously expresses both surface IgM+ and IgD+ (via alternative splicing).

Question 10

Can you describe 3 general steps involved in the T cell-dependent activation of a naïve B cell?

Answer 10

T cell-dependent activation of a mature (naïve) B cell (surface IgM+ and IgD+) involves 3 steps:

1) B cell surface immunoglobulins (B cell receptors) are bound and cross-linked by their specific cognate antigen → clustering of B cell receptors triggers endocytosis of receptor-antigen complexes into the B cell cytoplasm
2) Endocytosed antigen is dissociated from the B-cell receptor and then cross-linked toMHC class II peptides → antigen-MHC II complexes are then inserted into the B cell membrane and displayed on the B cell surface 3) TH (Helper T cells) recognize this MHC-complexed antigen and provide a co-stimulatory signal.

The co-stimulatory signal is required for activation of B cells and differentiation into plasma cells.

• B7 protein on B cell interacts with CD28 on Th cell → activates Th cell
• CD40 protein on B cell interacts with CD40L (CD40 ligand) on Th cell → activates B cell.

Question 11

Describe the paracortex of a typical lymph node.

Answer 11

Paracortex: T-cell rich area that lies between the lymph node cortex and medulla.

Paracortex = site where antigen is presented (by APCs) to T helper cells and some B cells → stimulated B and T cells travel to the cortex to activate the primary follicles, forming secondary follicles with germinal centers

Paracortical size / cellularity varies depending on underlying pathology — for example:

1. In immunocompetent patients, certain types of infections stimulate lymphocyte proliferation → ↑ number of circulating lymphocytes → hyperplastic, larger-than-normalparacortex due to massive lymphocyte influx
2. Thymic aplasia or hypoplasia (e.g., DiGeorge syndrome) → ↓ number of circulating lymphocytes → hypoplastic, smaller-than-normal paracortex due to lack of lymphocyte influx

Question 12

What is a role of the spleen during fetal development that is usually pathogenic in an adult?

Answer 12

During development, the spleen is responsible for fetal erythropoiesis between the second to seventh months (the liver and bone marrow are also partially responsible for hematopoiesis during this time as well). Splenic erythropoiesis after birth is usually pathogenic.

Question 13

What are secondary lymphoid organs? Do they facilitate a process that is antigen-dependent or antigen-independent?

Answer 13

Secondary lymphoid organs: sites of antigenic stimulation of mature naïve lymphocytes → lymphocyte activation. This process is clearly antigen-dependent (i.e., stimulation byforeign antigen is required).

Normally, all secondary lymphoid organs have lymphoid follicles.

Secondary lymphoid organs include:

1. Lymph nodes
2. Spleen
3. MALT (mucosal-associated lymphoid tissue) — e.g., tonsils, adenoid glands, Peyer’s patches

Question 14

Where does positive and negative selection of T-cells occur in the thymus?

Answer 14

The Thymus is the major site of T-Cell development

Functions to convert hematopoietic progenitor cells → immature thymocytes → T-cells

The entire organ is enclosed by an investing capsule. Just deep to the capsule is the cortex. The medulla is the thymic center.

Formed by fusion of the ventral wings of the third branchial pouch

Cortex: dense with immature T-cells. It is the predominant site of positive selection (selecting functional T-cells).

Medulla: pale-colored because it contains mature T-cells, epithelial reticular cells, andHassall’s corpuscles (remains of epithelial tubes which grow out from the third branchial pouches).

It is the predominant site of negative selection (elimination of autoreactive T cells).

Positive selection induces apoptosis of cells that bind too weakly while negative selection induces apoptosis of cells that bind too strongly. Cells with high-medium binding survive. Binding refers to the ability of the T-cell receptors to bind to either MHCclass I/II or peptide molecules.

Question 15

What are the two major functions of the spleen (in general)?

Answer 15

The spleen is a secondary lymphoid organ with roles in blood filtration and microbial defense.

Question 16

Describe the medulla of a typical lymph node.

Answer 16

Medulla: consists of large blood vessels, medullary cords, and medullary sinuses

Medullary cords are rich with antibody-producing plasma cells

These antibodies exit the lymph node via cortical sinuses, which drain into the medullary sinuses, then into the efferent lymphatics

Medullary sinuses are endothelial-lined lymphatic spaces that collect lymph destined for the efferent lymphatics. They contain macrophages and reticular cells.

Question 17

Describe the medulla of a typical lymph node.

Answer 17

Medulla: consists of large blood vessels, medullary cords, and medullary sinuses

Medullary cords are rich with antibody-producing plasma cells

These antibodies exit the lymph node via cortical sinuses, which drain into the medullary sinuses, then into the efferent lymphatics

Medullary sinuses are endothelial-lined lymphatic spaces that collect lymph destined for the efferent lymphatics. They contain macrophages and reticular cells.

Question 18

Each B cell can only make Ig (immunoglobulin) specific to one antigen — what determines each B cell’s idiotype?

Answer 18

Each B cell’s antigen specificity (idiotype) is determined by the three-dimensional shape formed by the N-terminal of heavy and light chains (at the tips of the “Y”).

B cells that are reactive to self antigens undergo clonal deletion in the bone marrow or are inactivated in the periphery (clonal anergy).

Question 19

Through what specialized type of venules do lymphocytes enter lymph nodes?

Answer 19

HEVs (high endothelial (postcapillary) venules) transport T- and B-cells from the bloodstream to the lymph node paracortex:

In the bloodstream, L-selectins on mature naïve lymphocytes weakly bind to addressins on endothelium →

This weak interaction allows stronger integrin-mediated binding and subsequent diapedesis →

Lymphocytes leave the blood and are carried by HEVs into the paracortical area of lymph nodes.

Guided by chemokine gradients, B-cells migrate to the cortex while T cells primarily remain local within the paracortex.

Question 20

What are the two main components of the red pulp of the spleen?

Answer 20

The red pulp is composed of the splenic cords of billroth and splenic (venous) sinusoids. The splenic sinuses are engorged with blood, giving this area its red color.

Sinusoids are marked by gaps in the endothelium that assist in the filtration of blood. Abnormal and/or aged red blood cells become trapped as they attempt to squeeze through the narrow spaces of the sinusoids. The trapped cells are subsequently removed by splenic macrophages.

Macrophages lining the splenic cords of billroth in the red pulp sample sinusoidal blood by extending processes through gaps in the sinusoidal endothelium and basement membrane. Here, they function to remove old and damaged red blood cells as well as encapsulated bacteria (recall SHiNE SKiS mnemonic for encapsulated bacteria – S. pneumonia, H. influenza type B, N. meningitides, E. coli, Salmonella, K. pneumonia, Group B Strep).

After a splenectomy, there is a significant risk for infection with encapsulated bacteria. Oftentimes patients will be prophylactically treated with antibiotics to prevent such infection. In addition, the following can be observed on a blood smear:

Howell-Jolly bodies in RBCs

Target cells

Thrombocytosis

Question 21

Name 2 main functions of lymph nodes.

Answer 21

Lymph nodes are secondary lymphoid organs that perform 2 main functions:

1. Nonspecific filtration of lymph — macrophages within lymph nodes may trap / destroy malignant cells, microorganisms (e.g., viruses, bacteria, fungi), debris (e.g., anthracotic pigment).
2. Facilitates interaction between lymphocytes and APCs (antigen presenting cells —e.g., macrophages, dendritic cells) → activation of lymphocytes (e.g., B cells, T cells)

Question 22

Where do all T cells originate from?

Answer 22

All T cells originate from hematopoietic stem cells in the bone marrow, however development takes place in the thymus.

The earliest thymocytes found in the thymus are CD4-CD8- → then CD4CD8 → thymocytes finally develop into either:

• CD4CD8-, also known simply as CD4 helper T cells, or Th cells; or
• CD4-CD8, also known simply as CD8 cytotoxic T cells, or CTLs.

Note: only 1-2 percent of T cells leave the thymus; most of the remaining T cells undergoapoptosis.

Question 23

What is the function of the marginal zone of the spleen?

Answer 23

Between the white pulp and the red pulp is a marginal zone composed of antigen presenting cells (APCs) and specialized B cells. The primary role of the marginal zone is to trap antigens from circulation to present them to splenic lymphocytes.

Question 24

An experimenter adds papain to a sample of IgG. Explain what would happen to an individual immunoglobulin and what the products of the reaction would be.

Answer 24

Papain is a proteolytic enzyme often used in experiments. It cleaves the peptide bonds at the hinge region of the antibody → 2 identical Fab fragments (which can bind antigen) and 1 Fc fragment (which can bind complement).

Question 25

B-cells and T-cells are derived from lymphoid stem cells — where are these lymphoid stem cells located?
Where do B-cells mature? Where do T-cells mature?

Answer 25

Although B-cells and T-cells both originate in the bone marrow from lymphoid stem cells, T-cells mature in the thymus whereas B-cells mature in the bone marrow:

Question 26

What are the three major zones of the spleen?

Answer 26

Grossly, the spleen is composed of a capsule and two distinct areas termed red pulp andwhite pulp, separated by the marginal zone (capsule à red pulp à marginal zone à white pulp, from outside in).

Question 27

Describe the cortex of a typical lymph node.

Answer 27

Cortex: composed of lymphoid follicles (mostly B-cells within a loose network of antigen-presenting follicular dendritic cells) as well as reticular cells, macrophages, and some scattered CD4 helper T cells.

Prior to antigenic stimulation, primary follicles are composed of small, resting B cells

In response to antigenic stimulation, secondary follicles form. These contain a pale germinal center (mainly activated, proliferating B cells) and a mantle zone (both antigen-presenting B cells and T helper cells)

Question 28

Can you name three CD cell surface markers found on B cells?

Answer 28

B cell markers: CD 19, CD 20, CD 21

CD 19: co-receptor with CD21; involved in signal transduction during B cell activation

CD 20: function unknown

CD 21 (CR2, C3d receptor): co-receptor with CD19; involved in B-cell activation. Is also the receptor for complement component C3d

EBV (Epstein-Barr virus) gains access into B cells via CD 21 → infectious heterophil-positive mononucleosis

Question 29

Can you describe 4 things that contribute to B cell idiotypic diversity during B cell maturation?

Answer 29

During B-cell maturation, 4 things contribute to idiotypic diversity:

1) Existence of multiple allotypes of V, D, and J genes (inherited from parents)

2) Gene recombination:
VDJ recombination → ↑ diversity of heavy chain variable domains
VJ recombination → ↑ diversity of light chain variable domains

3) N-nucleotide addition: Tdt (terminal deoxynucleotidyl transferase) randomly inserts nucleotides into the DNA sequence at the junctions of V, D, and J segments.
- In B cells, Tdt is only active during recombination of heavy chain gene segments (VDJrecombination), not active during recombination of light chain gene segments (VJ recombination).
- In T cells, Tdt is active during rearrangement of all gene segments in the formation ofTCR (T-cell receptor).

4) Combinatorial diversity: Refers to 2 processes during B cell maturation:

• Variable arrangement of receptor gene segments
• Any heavy chain can associate with any light chain → different heavy chain-light chain combinations form different epitopes

Once VDJ and VJ recombination are complete, allelic exclusion inactivates one chromosome in a homologous pair → ensures that only one Ig idiotype is expressed in each B cell

Question 30

Can you describe two examples of T cell-independent activation of B cells?
- How are these similar? different?

Answer 30

There are two examples of T cell-independent activation of B cells:

Type 1: An antigen called a mitogen (e.g., the highly repetitive surface structure of certain parasites) can bind B cell surface molecules that are not B cell receptors but are nevertheless linked to B cell receptors → clustering of mitogen-bound B cell surface molecules → clustering of B cell receptors linked to the mitogen-bound B cell surface molecules → polyclonal T cell-independent activation of B cells that do notrecognize the inciting mitogen. In other words, a single mitogen can activate many different B cells with many different specificities, none of which are specific for the mitogen!

Type 2: Some non-protein antigens like carbohydrates [e.g., the LPS(lipopolysaccharide) endotoxin in the cell wall of most gram-negative bacteria] have repeating epitopes that can cross-link a large number of B cell receptors at once → clustering of B cell receptors → antigen-specific T cell-independent activation of B cells that recognize the repeated epitope

Question 31

An immunologist isolates several protein fractions from an antibody. The fraction in which she is most interested is neither the heavy chain nor the Fc portion. What protein does this likely represent?

Answer 31

Light Chain

L chains are either κ (kappa) or λ (lambda). Either one can be found in any of the 5 different Ig classes, however each particular Ig molecule only contains either κ or λ.

Light chain is not found in the Fc portion. The N-terminus of the L chain is involved in the antigen binding site.

Question 32

What are the 2 possible fates of a mature (naïve) B cell following antigenic stimulation?

Answer 32

Upon antigenic stimulation → mature (naïve) B cells may become either antibody-producing plasma cells or memory B cells.

Somatic Hypermutation (aka Affinity Maturation) (occurs in B cells only, not in T cells):

• After antigen stimulation → rapid clonal proliferation of B cells (which produce antibodies against one particular antigen) → naturally occurring mutations in this rapid process yield antibodies with varying affinities for the antigen → B cells which produce antibodies with the highest antigen affinity are selected
• Note: this is the only change in idiotype that occurs after antigen stimulation

Following antigen-specific activation, mature (naïve) B cells are able to produce an immunoglobulin isotype other than IgM or IgD (e.g., IgG, IgA, or IgE) by isotype switching, which is accomplished through irreversible genomic recombination of heavy chain constant regions.

Question 33

Which two immunoglobulins bind complement?

Answer 33

The C-terminal region of the Fc portion of IgG and IgM binds complement. The Fc portion of IgA/G/E activates effector cells.

Question 34

What are primary lymphoid organs? Do they facilitate a process that is antigen-dependent or antigen-independent?

Answer 34

Primary lymphoid organs: sites of lymphocyte birth and/or maturation via a process that does not involve stimulation by foreign antigen (i.e., an antigen-independent process) → mature naïve lymphocytes

Normally, primary lymphoid organs do not contain lymphoid follicles.

Primary lymphoid organs include:

1. Bone marrow
2. Thymus