Pathology MOD 1: Serum Proteins PDF

PATHOLOGY 08/20/2024. MOD 1: SERUM PROTEINS Dr. Daniel Abraham...

PATHOLOGY 08/20/2024. MOD 1: SERUM PROTEINS Dr. Daniel Abraham G. Gonzales Trans Group/s: __ PART 1: SERUM PROTEINS II. SERUM VS. PLASMA SERUM PLASMA Liquid part of blood Separates after centrifugation Blood is allowed to clot or Anticoagulant is added to coagulate FIRST blood FIRST Contains: Contains: Serum Protein Classification. ○ Antibodies ○ Serum ○ Antigens components A. SERUM ELECTROPHORESIS ○ Electrolytes (antibodies, etc.) How are proteins separated in the first place? ○ Proteins ○ Coagulation ○ Through serum protein electrophoresis (SPEP), factors not strictly a need to know for the lecture, but important to note to understand how and why NO coagulation factors protein separate the way they do What happens during SPEP? Coagulation factors such as Addition of ○ Several samples of serums are placed in gel fibrinogen are NOT present anticoagulant results in attached to two electrodes of opposite charges because they are used up the presence of ○ A current then passes through the gel which and stay with the solid coagulation factors after causes the proteins to move toward a particular components of blood after separation electrode based on the proteins electric charge and clotting size III. SERUM PROTEINS Serum proteins appear in varying forms and locations throughout the serum. Some may freely circulate throughout the serum or others may act as carriers, electrolytes, or ions. Once isolated from the serum, these proteins can be classified in different ways. ○ One classification is albumin and globulins Albumin: comprises the majority of serum proteins Globulin: refers to the rest of the serum proteins other than albumin Further subdivided into alpha-1, alpha-2, beta, and gamma (as seen in the figure below) ○ Another way of classifying these proteins is based on how easily detectable they are in electrophoresis Major serum proteins Minor serum proteins Serum Protein Electrophoresis Example. Note the separation of the different columns These columns represent the different types of serum proteins, each of these columns or bands contain different proteins discussed in the previous slides. Bands can be thicker or thinner depending on the amount of protein in that particular section Pathology - Mod # Topic Title 1 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.  ○ Large amount of albumin contains thickest strand, ○ The opposite occurs instead in a negative acute whereas areas with few proteins are relatively phase reactant, so, it decreases when in an lighter and thinner. inflammatory state 1. CLINICAL SIGNIFICANCE 1.1 Peripheral Edema Decreased levels of albumin Albumin also serves role in maintaining oncotic pressure Decreased levels will cause fluids to leak out of the capillaries and into the surrounding tissues causing edema 1.2 Analbuminemia Congenital absence of albumin Peripheral edema is NOT present here since the body has compensatory mechanism for this state 1.3 Bisalbuminemia Refers to the presence of a variant of albumin Serum Electrophoresis Result Example. alongside the normal albumin This results in two albumin bands which is present IV. MAJOR SERUM PROTEINS upon electrophoresis Proteins that are readily resolved and detected on electrophoresis gels. 1.3 Glycosylated Albumin Up to 25% of albumin can become glycosylated A. PREALBUMIN during hyperglycemia referred to as the glycosylated A fraction that migrates in the position faster than albumin albumin towards the positive electrode Due to its sensitivity, this is used clinically as another Mostly produced in the liver but other organs as well way to assess diabetic control It is rich in tryptophan and has a short 2-day half-life This is a helpful alternative especially in diabetics with compared to other major serum proteins hemolytic anemias where HbA1c or glycosylated It has tetrameric structure with each monomer able to hemoglobin is not easily obtained or is unreliable. bind to a molecule of thyroxine (T4), hence, it is also Fructosamine is a similar measurement used in known as Thyroxine-binding prealbumin (TBPA) or patients with protein losing nephropathy instead. Transthyretin (TTR) It also plays a role in vitamin A metabolism 1.4 Albuminuria Presence of albumin the urine 1. CLINICAL SIGNIFICANCE Trace amounts of albumin in the urine is considered as abnormal (beware in urinalysis results) It is used mainly as a marker for nutritional status Measurement of microalbumin to urine is the standard Its production is heavily dependent on adequate care of management for: nutrition or any alterations in hepatic function such ○ Diabetes mellitus as in liver disease ○ Early detection of complications (mainly diabetic Therefore, it is a better early indicator of any changes nephropathy) in nutrition compared to other serum proteins Able to enter the CSF more easily due to its compact size which results in the presence of the distinct prealbumin band Other forms of serum proteins are also present only in the CSF If you see a prealbumin band upon electrophoresis of an unknown fluid, it is most likely to contain or be CSF. B. ALBUMIN The single most abundant protein in the plasma Serve as a general transport or carrier protein, as well as mobile repository of amino acids for incorporation into other proteins Has a half-life of 17 days Elevated infrequently during dehydration and artifactually during prolonged application of tourniquet Decreased values can be seen if there is impaired Albuminuria. synthesis by the liver or if there are losses such as in asichies or protein losing nephropathies C. α1-ANTITRYPSIN (AAT) Since it decreases during illness, it is also referred to as The major of the two alpha-1 globulins “negative acute phase reactant” Coded by SERPINA1 gene on chromosome 14 ○ Acute phase reactant: markers in the serum that increase acutely during inflammation; Pathology - Mod 1 Serum Proteins 2 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.  Main function: prevent inappropriate severe Haptoglobin-hemoglobin complexes → transported biochemical responses to inflammation wherein it and recycled into iron and bilirubin counteracts trypsin and neutralize proteases released by leukocytes 1. CLINICAL SIGNIFICANCE Elevated values → stress, infection acute inflammation, tissue necrosis ○ Acute phase reactant Decreased values → after hemolytic episodes (especially massive hemolysis) ○ Transfusion reactions ○ Decrease is due to the use of haptoglobin in forming complexes and cleared in the circulation ○ Decreased in liver disease or in individuals with congenital deficiencies F. β-LIPOPROTEIN Under the beta-globulins Low density lipoprotein Position and appearance during electrophoresis is sensitive to the recent ingestion of fatty food especially in hypercholesterolemia or increased cholesterol levels in the blood → greater staining of beta-lipoprotein α1-ANTITRYPSIN. 1. CLINICAL SIGNIFICANCE AAT deficiency → pulmonary emphysema or liver cirrhosis Elevated values are expected whenever acute inflammation occurs Enzyme is considered a nonspecific acute phase reactant D. α₂-MACROGLOBULIN (AMG) The largest major non-immunoglobulin protein in plasma Levels in the serum are similar in AAT ○ Higher in women due to estrogen Binds to proteases and inactivates it β-Lipoprotein. G. TRANSFERRIN Major beta-globulin Serves to transport ferric ions from intracellular or mucosal stores to bone marrow Normally, ranges from 200-400 mg/dL Measured as iron-binding capacity or IBC α2-Macroglobulin. 1. CLINICAL SIGNIFICANCE Rises 10x more in nephrotic syndrome Elevated in early diabetic nephropathy E. HAPTOGLOBIN Major protein migrating in the alpha-2 globulin region Half live of about 4 days Transferrin. Functions to preserve body iron in protein stores by binding to hemoglobin during lysis of RBC 1. CLINICAL SIGNIFICANCE Increase 2x in short-term iron deficiency Pathology - Mod 1 Serum Proteins 3 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.  Increased during acute phase response and 1.1 Hemochromatosis pregnancy or use of contraceptive medications Hereditary disorder resulting in cirrhosis, diabetes, Decreased during extensive coagulation due to being cardiomyopathy, arthritis, and other endocrine disorder used up due to toxic effects of excess-free iron 1.2 Antibacterial Effect Transferrin has an antibacterial effect due to its ability to remove iron from bacteria that require it for growth 1.3 Atransferrinemia Congenital deficiency of transferrin Presents with microcytic anemia and iron overload 1.4 Protein-losing Nephropathy Transferrin along with iron are lost from the circulation and into the urine 2. NOTABLE VARIANTS Fibrinogen. 2.1 Carbohydrate-deficient Transferrin Found in persons who engage in heavy alcohol 1. CLINICAL SIGNIFICANCE consumption Used as a specific marker for chronic alcohol abuse 1.1 Dysfibrogenemia Disease wherein the presence of an abnormal 2.2 Asialotransferrin hereditary variant of fibrinogen results in impaired Found in CSF, eye, and ear clotting, an increased tendency to bleed or bruise Purpose: evidence of presence of CSF in cases of head (hemorrhagic diathesis), or an increased tendency to trauma with nasal drainage or in fistula fluid after certain develop thrombosis. otologic or ear procedures Similar to prealbumin, the presence of Asialotransferin 1.2 Congenital Afibrogenemia upon electrophoresis of the fluid may indicate CSF Fibrinogen is absent, although the condition is not as severe as hemophilia in terms of arthrosis or joint H. COMPLEMENT bleeding Separate fraction of beta-globulin Consists mainly of C3 component PART 2: MINOR SERUM PROTEINS Values are increased during acute phase response due to its status as an acute phase reactant Decreased values occurs in autoimmune diseases V. MINOR SERUM PROTEINS Activated and used to form immune complexes and Proteins in the serum that are not usually detected by subsequently deposited within tissues standard protein electrophoresis due to their low levels A. CERULOPLASMIN A copper binding protein located in the a2-globulin band Contains most of the copper in the plasma It exhibits ferroxidase activity, which is important in iron metabolism Migrates along with the a2-globulins Complement. 1. CLINICAL SIGNIFICANCE Used as convenient marker for assessing disease activity in rheumatic disorders: lupus and arthritis I. FIBRINOGEN Present only in the plasma and not serum Most abundant of the coagulation factors Forms the fibrin clot Located in beta-globulin band Ceruloplasmin in the a2-globulin band. Pathology - Mod 1 Serum Proteins 4 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.  Wilson’s Disease: rare genetic disease that results in Decreased values are seen due to: impaired hepatic excretion of copper into the bile, ○ Non-specific urinary losses resulting in toxic deposition of copper in the tissues. ○ Liver failure (decreased synthesis) ○ Severe when not treated ○ Porphyrias, rhabdomyolysis, chronic ○ Diagnostic criteria: neuromuscular disease Low serum ceruloplasmin levels ○ The most profound decrease is seen during Liver disease intravascular hemolysis Neurologic signs Hemopexin serves as a substitute for when Keyser-Fleisher rings in the cornea due to haptoglobin cannot bind to more hemoglobin deposition of copper in the descemet’s Used as an additional aid to diagnose hemolysis membrane earlier Increased copper concentrations in urine and upon liver biopsy Hemopexin in the B-globulin band. Wilson’s Disease. D. a1-ACID GLYCOPROTEIN or OROSOMUCOID a1-globulin that serves as a binding agent for B. Gc-GLOBULIN progesterone aka Vitamin D-binding protein (DBP) ○ Also assists the transport and metabolism of Migrates along with the a1-globulins progesterone Decreased values occur in severe liver disease and Binds to lidocaine to keep it in active circulation trauma patients, who develop organ dysfunction and ○ May be used for monitoring lidocaine levels sepsis Elevated levels during pregnancy Congenital absence of Gc-globulin may result in Considered an acute phase reactant impaired Vitamin D transport and may be lethal ○ Nephrotic syndrome may also result in losses of DBP, some of which may be complexed with vitamin D a1-Acid Glycoprotein in the a1-globulin band. E. C-REACTIVE PROTEIN Gamma globulin present in the serum of patients with different disorders, except pneumococcal infections Gc-Globulin in the a1-globulin band. A general scavenger molecule that rises strikingly when there is tissue necrosis C. HEMOPEXIN or B-GLOBULIN Clinical significance: B-globulin which, similar to haptoglobin, serves to ○ Serves as a highly sensitive acute-phase preserve the body’s iron stores and limits toxicity of reactant free heme ○ Used as a rapid presumptive test to differentiate ○ Binds heme released by degraded hemoglobin via bacterial and viral infections its very strong affinity, which is then is cleared Bacterial infection commonly presents with from circulation by hepatocytes high CRP Pathology - Mod 1 Serum Proteins 5 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.  ○ Used by rheumatologists to monitor the progression/remission of certain autoimmune diseases ○ Higher than normal CRP levels are linked to a greater risk of cerebrovascular stroke or myocardial infarction Normal electrophoretogram. A normal electrophoretogram shows: ○ Nearly invisible prealbumin ○ Large albumin peak ○ Small a1 peak ○ Board a2 peak ○ Bimodal beta peak (sometimes) ○ Broad gamma peak CRP in the gamma-globulin band. VI. PATTERNS OF PROTEIN ABNORMALITIES Bimodal Beta Peak. B. ABNORMAL 1. ACUTE PHASE OR IMMEDIATE RESPONSE Acute Phase or Immediate Response. Electrophoretogram (top), agarose gel electrophoresis (middle), and the corresponding proteins per band (bottom). Albumin is slightly decreased. The a2 band is elevated due to increased levels of The columns in the gel represent the different types of haptoglobin or other proteins, such as a-1 antitrypsin. proteins ○ The bands will be thicker or thinner depending on 2. DELAYED RESPONSE the amount of protein in that column ○ The density of the bands correspond to the height of the peak on the electrophoretogram ○ Ex. a large amount of protein in a specific band (eg. albumin) results in a thicker and denser band on gel and a higher peak in the electrophoretogram Meanwhile, fewer proteins in a column result in a lighter band on gel and a lower peak on the electrophoretogram. A. NORMAL Delayed Response. Extension or continuation of the acute response Pathology - Mod 1 Serum Proteins 6 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.  Greater decrease in albumin compared to the acute response 4. NEPHROTIC SYNDROME Further increase in a2 band Broadening of the gamma region due to a polyclonal increase in immunoglobulins 3. LIVER CIRRHOSIS Nephrotic Syndrome. Nephrotic syndrome and other protein using Liver Cirrhosis. nephropathies result in decreased albumin, a1, B1, and gamma globulins Liver cirrhosis results in decreased albumin due to ○ a2 macroglobulin and B-lipoprotein are decreased synthesis by the liver and increased loss increased. There is also the presence of beta-gamma bridging ○ Occurs due to a polyclonal increase in immunoglobulins (several or all are increased) 5. HYPOGAMMAGLOBULINEMIA ○ These immunoglobulins are located throughout the beta to the gamma columns ○ Because of the increase in proteins, the corresponding column in the agarose gel band will also be more dense or thickened. Subsequently results in the increased spike which bridges the beta to the gamma column. Hypogammaglobulinemia. Shows nearly to completely absent gamma fractions Occurs in: ○ Neonates ○ Congenital immunodeficiency states ○ Adults with lymphoreticular disorders ○ Post-chemotherapy states ○ Hypoproteinemic states 6. MONOCLONAL GAMMOPATHY Immunoglobulins (highlighted in yellow) throughout the beta and gamma columns. Monoclonal Gammopathy. Recognition of this pattern is most important and widespread clinical application of serum protein electrophoresis Paraprotein: abnormal protein secreted by a monoclonal proliferation of plasma cells, especially in cases of multiple myeloma ○ aka M spike, M protein, or immunoprotein In contrast to polyclonal gammopathy, this presents as a Beta-gamma bridging in agarose gel. relatively thin spike on electrophoretogram. Pathology - Mod 1 Serum Proteins 7 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.  VII. SUMMARY There are several different proteins in the serum that are classified into major and minor, and have different functions Several normal or abnormal states may influence the amount of proteins in the serum Depending on the amount of proteins, this results in different appearances seen on electrophoresis and electrophoretogram Polyclonal Gammopathy in Liver Cirrhosis. In polyclonal gammopathy (in the case of liver cirrhosis), multiple immunoglobulins are increased, ranging from the beta to the gamma column However, in monoclonal gammopathy, there is an increase of only one immunoglobulin, which corresponds to homogenous and needle-shaped elevation compared to the other gamma or beta elevations discussed previously. Examples of Monoclonal Gammopathy in Agarose Gel. Homogenous and needle-shaped elevation. Pathology - Mod 1 Serum Proteins 8 of 8 The use of trans, practice questions, and evals ratio must be used discreetly and social media/public exposure of the aforementioned shall be strictly prohibited.