Bence-Jones Protein in Urine

Bence-Jones protein is a type of abnormal protein present in the urine of some 35 to 65 per cent of patients with multiple myeloma. This disease, myelomatosis, is characterized by a large increase in the number of plasma cells, which are probably of an abnormal type. Associated with the plasma cell proliferation is a moderate to large increase in the globulin fraction of serum, the total globulin level at times reaching values of 11 to 12 gm./100 ml. Generally, the increased globulin behaves electrophoretically as gamma globulin. On occasions, a peak, the so-called "M" peak, is seen between the beta and gamma globulin fractions, and even more rarely, in the alpha globulin region. Recent studies have shown that gamma globulins are made up of four peptide chains linked together as a unit. The inner two are the longer and are referred to as H (heavy) chains; the outer two are shorter and are called L (light) chains. In myelomatosis, there appears to exist a considerably increased and unbalanced production of the two forms, so that serum contains a high level of abnormal gamma globulins. These may be either the normal type of gamma globulin, designated as 7S or IgG, or the less common IgA type. The excess of short L forms is excreted and is detected in the urine in the form of those abnormal proteins first detected by Bence-Jones some 100 years ago.

Normal proteins, such as the common albumins and globulins, when heated, do not coagulate and precipitate out of solution until the temperature reaches 56 degree to 70 degree. Bence-Jones proteins coagulate at a much lower temperature, 40 degree to 60 degree of Celcius,; furthermore, unlike normal proteins, the precipitated Bence-Jones proteins redissolve as the temperature of the solution is increased to between 85 degree and 100 degree of Celcius., and often reprecipitate as the temperature is again decreased to 45 degree to 50 degree of Celcius. Because of this peculiar behavior on heating, Bence-Jones proteins have been referred to as pyroglobulins. The Bence-Jones proteins have a molecular weight of the order of 45,000, roughly one-fourth that of complete gamma globulin molecules.

Urine Protein

All urines contain some protein. The protein excreted by healthy individuals is of the order of 50 to 100 or perhaps 150 mg./24 hr. After considerable muscular exertion the value may be as high as 250 mg. Random specimens will contain under 10 mg./100 ml., although in overnight specimens the level may be as high as 15 to 20 mg./100 ml. Proteinuria is said to be present whenever the urinary protein output is greater than that reflected in these normal values. Not all proteinuria is clinically significant, but persistent abnormal levels of protein in the urine is an indicator of the presence of kidney and urinary tract disease. Thus, the detection of urinary protein and the quantitative assessment of the degree of proteinuria are very important laboratory procedures.

In general, the urinary proteins reflect those present in plasma, along with special proteins of renal tubular origin, and those exuded from the tissues that line the genito-urinary tract. The glomerular filtrate contains small amounts of all plasma protein fractions, but most of the filtered protein is reabsorbed in transit through the kidney tubules. Some of the globulins present in urine are of smaller size than their electrophoretic counterparts in serum. The new techniques of immunoelectrophoresis and antibody-antigen diffusion in agar are rapidly changing our knowledge of urine proteins. The old concept that albumin was the only protein present in any significant amount, even in pathological urines, is no longer valid, and therefore the term albuminuria should no longer be used. Some 40 to 60 per cent of the proteins present in normal urines are mucins (glycoproteins) originating from the linings of the lower  genitourinary tract, and are usually of no clinical significance.

Proteinuria is often a manifestation of primary renal disease, although transient proteinuria may occur with fevers, thyroid disorders, and in heart disease, in the absence of renal disease. Proteinuria may be evident very early in the course of various renal disease states. With such conditions as pyelonephritis, reflecting bacterial infection in the kidney, and acute glomerulonephritis, often associated with recent streptococcal infections, the degree of proteinuria is slight, usually amounting to less than 2 gm. per day. In chronic glomerulonephritis and in the nephrotic syndrome, including lipoid nephrosis, and in some forms of hypertensive vascular disease (nephrosclerosis), protein loss may vary from a few grams to as much as 20 to 30 gm. per day. Proteinuria is encountered in certain other disease entities when and if they give rise to kidney lesions. Among these are lupus erythematosus, amyloidosis, toxemia of pregnancy, septicemia, and certain forms of drug and chemical poisoning. In multiple myeloma, not only does one find a modest increase in protein output, but in 40 to 60 per cent of the cases, the urine contains a group of abnormal proteins referred to as Bence-Jones proteins.

In healthy individuals, transitory elevations in urine protein output are encountered after intense exercise or work, and after exposure to cold. Orthostatic proteinuria is benign condition in which protein excretion is normal when the patient is lying down (prone), but is mildly elevated when the patient walks or stands erect for any period of time. Persons subject to orthostatic proteinuria often are embarrassed during medical examinations for insurance because their urines are found to be positive, when tested for protein after the applicants had been on their feet and active for a good part of the day. On rechecking the urine in the morning after a night's rest, the urine is usually found to be negative for protein. 

Plasma and Serum Proteins

CLINICAL SIGNIFICANCE

Blood is made up of particulate cell forms suspended in a fluid medium called plasma, a very complicated mixture of inorganic, and simple and complex organic materials dissolved in water. If the blood is collected without anticoagulant, in vessels with siliconized or non polar surfaces, the fluid separating is referred to as native plasma. This approximates very closely the plasma actually present in circulating blood. It is the practice, however, to use such anticoagulants as oxalate, citrate, EDTA, and heparin to prepare specimens of plasma for study or analysis. This differs from native plasma because it is modified by the presence of the added chemicals and by the partial loss of calcium bound by oxalate, if this is used. If blood is permitted to clot, the fluid separating is referred to as serum. It lacks the protein fibrinogen present in plasma, the fibrinogen having been transformed into insoluble fibrin in the clotting process. The fibrinogen constitutes only some 3 to 6 per cent of the total plasma proteins. It is satisfactory and much more convenient to use serum rather than plasma in clinical chemical studies.

Some 92 to 93 per cent of plasma or serum is solvent water; of the 7 to 8 per cent of total solutes present, the proteins occur in the greatest concentration, roughly 6.8 to 8.8 (6.5 to 8.5) gm./100 ml. of plasma (serum) water. Because of the large molecular weight of proteins, however, their molar concentration calculates to about 0.80 to 1.10 mM/L. In clinical work, the concentration of proteins is given in terms of grams per 100 ml. of serum volume. It would be more precise to express it in terms of serum water. The latter value is 1.07 times the serum volume figure.

Plasma proteins serve a number of different functions in the organism. They play a nutritive role, inasmuch as they constitute a portion of the amino acid pool of the body; thus, the proteins are a form of storage amino acids. If needed, these proteins can be broken down in the liver to produce amino acids for use in building other proteins. Alternatively, they can be deaminated to give keto acids that can be mobilized to provide caloric energy or be transformed into carbohydrates and lipids. Plasma proteins also act as transport agents: many vital metabolites, metal ions, hormones, and lipids are transported about the body, bound to and carried by certain specific proteins. Some proteins have important special functions of their own, such as enzymes, the immune antibodies among the globulins, and the various proteins associated with blood coagulation.

Another important function of proteins is a physicochemical one. The plasma proteins, being large, colloidal molecules, are nondiffusible; i.e., they cannot move through the thin capillary wall membranes as can most other blood solutes. They are thus entrapped in the vascular system and exert a colloidal osmotic pressure, which serves to maintain a normal blood volume, and a normal water content in the interstitial fluid and the tissues. The albumin fraction is most important in maintaining this normal colloidal osmotic or oncotic pressure in blood. If the albumin falls to low levels, water will leave the blood vessels and enter the extracellular fluid and the tissues, thus producing edema.

The maintenance of the acid-base balance in blood also involves the plasma proteins. As amphoteric compounds, they function as buffers to minimize sudden, gross changes in the pH of the blood.

Despite their presence together, the various proteins of plasma do not originate from the same source. The liver is the main organ for the synthesis of albumins and alpha and beta globulins, and perhaps some nonimmune gamma globulins; among these these proteins are included the blood clotting and transport proteins. The cells of the reticuloendothelial system (spleen, bone marrow, lymph nodes) serve as the source of the antibody, immune gamma globulins and perhaps some beta globulins.

The level of total serum proteins found in healthy young and middle-aged adults is 6.0 to 8.2 gm./100 ml. of serum. In plasma, fibrinogen increases this value by an additional 0.2 to 0.4 gm./100 ml. A diurnal variation of 0.5 gm./100 ml. reflects small changes in the ratio of vascular to nonvascular fluid in the course of daily activity. In disease states both the total protein and the ratio of the individual protein fractions may change independently of one another. In states of dehydration, total protein may increase some 10 to 15 per cent, the rise being reflected in all protein fractions. Dehydration may result either from a decrease in water intake, as occurs in frank water deprivation (thirst), or from excessive water loss, as occurs in severe vomiting, diarrhea, Addison's disease, and diabetic acidosis. The absolute quantity of serum proteins is unaltered, but the concentration is increased because of the decreased volume of solvent water. In multiple myeloma, the total protein may increase to over 10 gm./100 ml., the increase being almost entirely due to the presence of markedly elevated levels of myeloma proteins (abnormal forms of gamma globulins). The quantities of other proteins are essentially unaltered.

Hypoproteinemia, characterized by total protein levels below 6.0 gm./100 ml., is encountered in many unrelated disease states. In the nephrotic syndrome large masses of albumin may be lost in the urine as a result of leakage of the albumin molecules through the damaged kidney. In salt retention syndromes, water is held back to dilute out the retained salt, resulting in the dilution of all protein fractions. Large quantities of proteins are lost in patients with severe burns, extensive bleeding, or open wounds. Water is replaced by the body more rapidly than is protein, effecting a decreased total protein concentration. A long period of low intake or deficient absorption of protein may affect the level and composition of serum proteins, as in sprue and in other forms of intestinal malabsorption, as well as in acute protein starvation (kwashiorkor). In these conditions the liver has inadequate raw material to synthesize serum proteins to replace those lost in the normal turnover (wear and tear) of proteins and amino acids.

In general, changes in total proteins may occur in one, several, or all fractions. It is also possible for significant changes to occur in different directions in different fractions, without changes in the total protein concentration.

The physician may occasionally request only a value for total protein in serum; however, more commonly he will ask for total protein, the values for the albumin, and total globulin fractions, and for the albumin-globulin (A/G) ratio. In healthy young and middle-aged adults, the albumin may vary from 3.8 to 4.7 gm./100 ml., and the total globulins from 2.3 to 3.5 gm./100 ml. The range found for the albumin-globulin ratio is 1.1 to 1.8, averaging 1.5. The globulins are usually separated by a salt fractionation procedure, but may be estimated from the sum of electrophoretically separated fractions. Albumins can be estimated separately by virtue of their ability to bind certain dyes such as methyl orange and hydroxyazobenzoic acid. These techniques will be discussed later.