Journal of Clinical Apheresis 7: 163-168 (1992)

Basic Principles of Immunology (Immune Response) for Hemapheresis Practitioners Janice M. Sigmon Therapeutic Plasma Exchange Program, Nephrology Service, Walter Reed Army Medical Center, Washington, D.C. Advances in medical knowledge and technology have identified the essential elements involved in the human immune system, their relationships and interactions, and offer more advanced concepts in the design, function, and maintenance of immune response. Immune response begins with the earliest progenitor cell and transfers its legacy of protection through white blood cells and the complement system. A basic understanding of immunology is essential to the hemapheresis practitioner as new treatment regimens requiring hemapheresis interventions de0 1992 Wiley-Liss. Inc. velop.

Key words: stem cells, lymphocyte activation, complement, immunoglobulins

INTRODUCTION

IMMUNITY

The dynamics of blood that enable the body to fight infection and protect itself from the constant challenge of environmental microbes and viruses is more commonly known as the immune system. The development of Acquired Immune Deficiency Syndrome (AIDS) in the 1980s demanded the rapid expansion of medical knowledge and technology, which has shed new light on the multifaceted relationships and interactions of the immune system and their subsequent effect on the body. The fact that white blood cells (WBCs) play a major role in this process is not new knowledge. However, the mechanism by which these cells interact is just beginning to be elucidated. A basic understanding of the WBC differential and the function of these cells is essential to understanding the mechanism of immune response.

Immunity is the resistance to infection conveyed by the combined forces of the white blood cells: granulocytes, monocytes, and lymphocytes (Fig. 2). Natural immunity is conveyed at birth and is not enhanced by repeated exposure. It is a general immunity which does not require a stimulus to develop. Examples of natural immunity include gastrointestinal bacteria, which assist in the digestive process; ABO blood group antibodies; intact skin, which prevents against infection; stomach and vaginal pH, which kills bacteria and fungi; and cilia, which filter epithelial surfaces in the nose and trachea. Acquired immunity is stimulated by, and developed after, exposure to a foreign substance. The foreign substance can be a protein, drug, carbohydrate, etc. The foreign molecule is called an antigen or an immunogen, and is capable of stimulating an immune response. Although smaller immunogens are capable of stimulating an immune response, an immunogen is usually greater than 40,000 Daltons in molecular weight. Uniquely specific, the foreign antigen is “fingerprinted and booked” by “memory” T-lymphocytes so that the antigen can be identified if it is seen again. The substance produced in response to the foreign antigen is called an antibody or an immunoglobulin,

ONTOGENY OF BLOOD

At conception, parental chromosomes, one set from each parent, convey the genetic instruction for the development of the human fetus. The production of pluripotential stem cells, the progenitor blood cells, occurs in the earliest stages of fetal development. These pluripotential stem cells multiply and differentiate into 2 cell lines: hematopoietic stem cells, which become the red blood cell, platelet, and granulocyte/ monocyte precursors; and lymphopoietic stem cells, which become the B-lymphocyte and T-lymphocyte precursors (Fig. I ) . At the precursor phase, the cells have matured and have become committed colony-forming units of discrete blood cell types. 0 1992 Wiley-Liss, Inc.

Address reprint requests to Janice M Sigmon, M A , MT(ASCP) SBB, Nephrology Service, Bldg 2. Room 4903. Wdlter Reed Army Medicdl Center, Wdshington, D C 20307-5001 The opinions and assertions contained herein are the privdte views of the author and are not to be construed as official or d5 reflecting the views of the Department of the Army or the Depdrtment of Defense

Sigmon

164

/

Hematopoietic Stem C e l l CD34*

Committed Erythroid Precursor

Pluripotential Stem Cell CD34+

/ \ \

Ly m p hop oi e t ic Stem C e l l CD349

Committed

Committed Platelet Precursor

\

Granulocy te/Monocyte Precursor

A Erythrocyte Progenitors

Megakar yocyle Progenitors

Granulocyte

Monocyte

Progenitors

Progenitors

(Mar row)

(Marrow)

(Marrow)

(Marrow)

Red Blood C e l l (RBC)

Platelet

/ \ Committed R Cell P re cur6 or

I Pre-R Cell (Marrow)

I

Mature B Cell

Corn mit ted T Cell Precursor

I Pre-T Cell (Thymus)

I

Mature T Cell

Eosinophil Basophil

adk Fig. 1 . Blood cell lineage from stem cell to mature, discrete cell type [1,2]. Thc precursor cells of all lineages appear to possess the CD34 antigen. while the committed colony-forming cells are negativc for the CD34 antigen. This distinguishing characteristic is often utilized in bone marrow purging 131.

Lymphocytes: (25

Circulate throughout the body Identify ‘foreign‘ substances Provide ‘memory’ against foreign substances Secrete ‘cytokines’ to summons other lymphs

- 40%)

Monocytes: (0 - 5%)

Reside in tissues Become Macrophages, Phagocytes, or Mast Cells

Monocyte

- Macrophages:

Stimulate inflammation. fever Synthesize and secrete complement Possess receptor for Fc portion O f lg Express Class I and II MHC Antigens

Granulocytes: (40

Basophil

- 65%)

Circulate throughout the body Provide Chemotaxis

@

Eosinophil

Neutrophil

Fig. 2. White blood cell morphology is readily apparent when a thin smear of blood is stained with Wright’s stain and viewed under a microscopc to obtain a differential.

which will “search and destroy” the foreign antigen before it can cause infection, complement activation, transfusion reaction, or other adverse manifestation (Table I). With each subsequent exposure the concentration and potency of the antibody is multiplied. THE ROLE OF THE WHITE BLOOD CELLS IN IMMUNE RESPONSE

The monocyte-macrophage is the body’s alarm system. Monocytes leave the bloodstream for the tissues, particularly the spleen, liver, peritoneum, lymph nodes, tonsils, and gastrointestinal tract. These cells process foreign material, splitting it and digesting it to obtain useful information about the foreign substance. When activated, the monocyte-macrophage can signal the need for lymphocyte involvement by secreting chemical substances such as interferon and lymphokines. These substances activate CD4’ T-helper lymphocytes and eventually result in an inflammatory process. At the same time, they increase their own metabolism by secreting interleukin- 1 (IL- 1) and increase their cell-surface receptors by secreting interleukin-2 (IL-2). At times, the

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165

TABLE I. Manifestations of Immune Response Immunoglobulins (produced by plasma cells) Complement activation Graft vs. host disease (GVHD) Monokines (produced by phagocytes) Type 1 interferon Interleukin- 1 Tumor necrosis factor (TNF) Colony Stimulating Factors (CSF) (produced by phagocytes and lymphocytes) Granulocyte-macrophage C S F (GM-CSF) Monocyte-macrophage C S F (M-CSF) Granulocyte CSF (G-CSF) Interleukin-7

monocyte-macrophage becomes phagocytic when activated. It circulates like Pac-Manzc, engulfing and removing bacteria and other foreign protein. The lymphocyte is the body's police force. Lymphocytes circulate throughout the body and identify the monocyte-processed foreign substances. Once the identification is made, other lymphocytes may be summoned to assist in the elimination of the foreign substances. The lymphocyte has an extended longevity, some living up to 3 and more years in the body. These long-lived cells provide the memory bank that stores the identities of previously fingerprinted foreign material. There are 2 kinds of lymphocytes: B-lymphocytes and T-lymphocytes (Fig. 3). B-lymphocytes make up 10-15% of the circulating lymphocyte population and provide humoral immunity. B-lymphocytes recognize surface markers of foreign antigens on transfused red cells and transplanted tissues, i.e., Rh, Fya, K, Jka, etc. or human leukocyte antigens (HLA). Some B-lymphocytes are lymphopoietic stem cells which have been programmed in the marrow to enter the bloodstream as ptasma cells and to synthesize and secrete specific immunoglobulin when activated. The plasma cells are also programmed to expand or reproduce so that sufficient quantities of immunoglobulin are produced to eliminate the foreign antigen. They deteriorate rapidly when their task is complete. T-lymphocytes provide cellular immunity, which includes the killing of intracellular pathogens, delayed hypersensitivity skin reactions, graft-versus-host disease (GVHD), and tumor rejection. T-lymphocytes are lymphopoietic stem cells that are processed by the thymus and are capable of providing a variety of functions, i.e., helper, killer, suppressor, etc., which are required in monocyte-macrophage and B-lymphocyte activities. Modern technology in the form of flow cytometry allows the different T-lymphocytes to be classified according to function and surface marker (antigen). These

Ly mp ho D o iet ic

Committed B Cell Precursor

ComGitted T Cell Precursor

I

I

Pre-B Cell (Marrow)

Pre-T Cell (Thymus)

I

Mature B-Lymphocyte

Humoral Immunity Fig. 3.

1

Mature T-Ly m p ho cy te

Cellular Immunity

Differentiation of lymphopoietic prccurbors

surface markers are designated as CD for clusters of differentiation. All pluripotential stem cells have a surface marker differentiated as CD34' antigen. As the pluripotential stem cell becomes committed and matures into a specific cell type, the CD34' antigen is lost and the characteristic antigen of the cell type becomes the dominant surface marker. Helper T-lymphocytes (formerly designated as T-4 lymphocytes) have a specific CD4' antigen. These lymphocytes produce cytokines, which activate either B-lymphocytes to become plasma cells and secrete antibody or macrophages to secrete interferon and lymphokines to produce inflammation (Fig. 4). Suppressor/ cytotoxic T-lymphocytes (formerly designated as T-8 lymphocytes) have a specific CD8' antigen. These cells process the foreign antigen and destroy it by cell lysis. Suppressor T-lymphocytes also serve to terminate the immune response function by secreting inhibitors when the immune response is no longer needed. The normal ratio of CD4' to CD8' lymphocytes is 1 .O-2.8. Alteration of this ratio can result in immunosuppression. T-lymphocytes mediate immunity at the major histocompatibility complex (MHC; Fig. 5). The CD4' lymphocytes code for the Class 11 antigens (HLA-D and -Dr) of the MHC. The CD8+ lymphocytes code for the Class I antigens (HLA-A, -B, and -C) of the MHC. A mismatch at the MHC can result in graft-versus-host disease (GVHD).

166

Sigmon Monocyte-Macrophage Processes Antlgen and Secretes: 1. I n t e r l e u k l n - 1 2. I n t e r l e u k l n - 2 3. L y m p h o k l n e s : Interferon BCGF

@

Monocyte-Macrophage

Q

Mature B-Lymph8 Become Plasma C e l l s

Antigen

1.

Activated T - H e l p e r C D - 4 * Lymphs Secrete: Cytoklnes

A c t i v a t i o n Phase:

2.

C o g n i t i v e Phase:

Processing Information, Differentiating Specific Protein and Proliferating

Blndlng of t h e Forelgn Antlgen by M o n o c y t e - M a c r o p h a g e S u r f a c e R e c e p t o r s

A n t l b o d y Tagged Antigen E n g u l f e d by P h a g o c y t e

I na c t i v a t e d Plasma Cells Deteriorate

@

@

T

- S uCppD -re8 +a s o r s

A n t l b o d y Tagged Ant l g e n Plasma C e l l s Produce S p e c l f l c A n t i b o d y

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@ Memory

@@

T-Lymph8

@ 3. E f f e c t o r Phase: D e s t r u c t i o n of t h e Foreign A n t i g e n

4.

Memory T-bymphs

Suppressor Phase: I n h i b i t i o n of I m m u n e R e s p o n s e

Fig. 4. Immune response as mediated by a monocyte-macrophage. The phases of activity are similar regardless of the specific cellular or humordl activity. The endpoint of the immune response will vary according to the particular white cells involved.

IMMUNOGLO BULI N (Ig) PRODUCT10N

Chromosome #6

Antibodies or immunoglobulins are protein molecules (Fig. 6) produced by plasma cells in response to a foreign antigen. There are 5 classes of immunoglobulins, which differ in size, concentration, and function within the plasma portion of blood (Fig. 7). The immune response mechanism is illustrated in Figure 8. COMPLEME NT ACT1VAT1ON

Complement is a family of 1 1 different enzymatic proteins which circulate in the plasma and mediate the immune response. When activated, the proteins come together (like matching pieces of a puzzle), attaching to the initiating molecule. If the last complement components attach, the molecule is lysed. If the cascade does not come to completion, the complement-tagged molecule

6 L i

Centromere

B c

/A

i

Complement Components

Fig. 5 . The major histocompatibility complex (MHC) is a group of genes located on the short arm of Chromosome 6. These genes code for the human leukocyte antigens (HLA). as well as the complement components.

Immunology for the HP

167

Immune Response F(ab): Antibody Binding Site

Light Chain-

Secondary Response (Anamnestid Primary Response

Heavy Chain-

FC Macrophage Binding Site

Fig. 6. The immunoglobulin molccule. The variable regions (shaded) on the F(ab) portion convey the antibody specificity according to the molecular configuration. The constant region (unshaded) contains the Fc portion, which signals for other immunologic activity. i.e., complement activation and/or fixation, binding with white blood cells, etc. 10

14

30 60

120

2

7

14

30 6 0 1 2 0

Days

@& IgM

IgG Smallest monomer 75% of all immunoglobulin Found in extravascular spaces Capable of binding complement

IgA

4

Pentamer

g41b

15% of all immunoglobulin Has a 'J' Chain Mostly intravascular Excellent binder of complement

Monomer, Dimer. or Trimer 10% of all immunoglobulin Has 'secretory' piece Found in bodily secretions Prevent entry of foreign substances into the body

IgE

%P

Monomer

Monomer

~ 1 %of all immunoglobulin

(1% of all immunoglobulin

Found on 8-lymphs when they are bound to antigen

Seen in histamine reactions Mostly on surface of basophile and mast cells

IgD No known function

Fig. 7 . Characteristics of the 5 classes of immunoglobulin (Ig) molecules.

may be eliminated by phagocytosis, opsonization, or anaphylaxis. There are 2 pathways of complement activation: the classical pathway and the alternate pathway (Fig. 9). The presence of antigen bound by IgM or IgG antibody can activate the classical pathway of the complement system. Specific red cell antibodies characteristically activate complement, i.e., ABO, Kidd, P,, H, and some M antibodies, etc. The alternate pathway is initiated by microorganisms, i.e., bacteria, fungi, viruses, tumor cells, etc. GRAFT-VERSUS-HOST DISEASE (GVHD)

GVHD can occur if viable donor lymphocytes engraft and multiply, overwhelming an immunosuppressed patient. The immunocompetent donor lymphocytes per-

Fig. 8. The immune response mechanism. The primary iwnnune response t o foreign substances occurs within 10- 14 days of exposure. The initial immunoglobulin is an IgM antibody which convert., t o a potent, long-lasting IgG antibody within 14-28 days after exposure. This IgG antibody reinains in the plasma. although often at an undetectable concentration. The secondtrn immune resporise (~ncrmne.sric~ response) occurs when the body is again challenged by thc same antigen. The second stimulation produces the same IgM response (recalling the first episode), but the conversion to IgG is much faster (3-5 days) and the resulting antibody is much more potcnt. remaining in highest concentrations for about 14 days. The antibody level falls off much more slowly and usually remains detectable in the individual's plasma.

ceive the recipient as foreign and mount an immune response, which can result in fever, rash, diarrhea, bone marrow suppression. and death. GVHD can be prevented by low-dose ( 1,500-5,000 rads) irradiation for 2-4 minutes of blood components required for transfusion and tissue used for transplantation. This will prevent the mitotic (reproductive) process of the immunocompetent donor lymphocytes, thus eliminating their propensity to multiply and overwhelm. IMMUNE RESPONSE IN DISEASE

While the immune response system plays a significant role in the maintenance of health, a normally functioning immune system can present problems in the successful treatment of diseases, requiring blood component transfusion and transplantation. Exposure to blood group and tissue antigens which are not present in the patient often leads to the development of specific antibodies, which reduce the survival of the transfused or transplanted tissues. Once a patient has become sensitized (developed antibody), identification of the specific antibody and

168

Sigmon

-

Classical Pathway Antigen-Antibody Complex Clq

+

Clr

+

+

Alternate Pathway

Ca++

Mg++

Cls

Microorganism: virus, fungi, tumor, etc.

+

, I

8 -

C4b2a

Factor B Factor D

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C3

+

#

CS + p

C3b

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Cell Lysis Fig. 9 . The complement cascade. The interaction of the components depends upon the presence of the pieces. As each reaction occurs, it lays the foundation for the next sequence. The bar over the components designates an activated component complex. The circles denote

complement components released into the plasma when split from the original component. The release of these byproducts can initiate anaphylaxis and hypersensitivity reactions.

search for compatible components often delays treatments and can cause further complications to the patient’s recovery. Examples include patients who are alloimmunized (refractory) to random platelets who subsequently require HLA-matched platelets and patients awaiting bone marrow transplant who become sensitized through transfusion of blood components. The immune system can become dysfunctional and begin to develop antibodies specifically against itself. The aberrant autoimmune process can result in a variety of diseases, i.e., rheumatoid arthritis, myasthenia gravis, systemic lupus erythematosus, Guillain Bane syndrome, Goodpasture’s syndrome, etc. The stimulus for the autoimmune process may be bacterially or virally induced, the result of exposure to toxic agents, a genetic susceptibility, or some other idiopathic event. Treatment for autoimmune diseases includes chemotherapy, steroid therapy, and therapeutic plasma exchange, and often requires a combination of these therapies.

REFERENCES 1. Tregellas W M , Keating LJ: “Immunology.” Arlington. VA:

American Association of Blood Banks. 1985, p 3. 2. Kasprisin CA. Snyder EL: “Bone Marrow Transplantation: A Nursing Perspective. A Workshop.” Arlington, VA: American Association Blood Banks. 1990, p 3. 3. Sacher RA, McCarthy L J , Sibinga CTS: “Processing of Bone Marrow for Transplantation. A Workshop. Arlington, VA: American Association of Blood Banks, 1990, p 49. ”

SELECTED READING Harmening D, Calhoun L, Polesky HF: “Modern Blood Banking and Transfusion Practices,” 2nd ed. Philadelphia: F.A. Davis, 1989, pp 43-64. Roitt I , Brostoff J , Male D: “lmmunology.” St. Louis: CV Mosby C o . , 1989. Walker RH. Hoppe PA, Judd WJ, Ness P. Polesky HF, Rolih SD, Snyder EL. Vengelen-Tyler V, Ward M: “Technical Manual.” 10th ed. Arlington. VA: American Association of Blood Banks, 1990, pp 121-146.