case study [chemistry] Questions:

1. What are this patient’s most striking laboratory results and

An Elderly Male Without how do you explain them?
2. What is this patient’s most likely diagnosis?
a Beta-Band on Serum 3. What is the pathogenesis of primary causes of low or ab-
Lipoprotein Electrophoresis sent LDL-C?
4. What are the clinical manifestations in individuals with
Chakshu Gupta, MD, James J. Maciejko, MS, PhD
this condition?
Departments of Pathology and Internal Medicine, St. John Hospital
5. What methods are available for quantifying the principal
and Medical Center, Detroit, MI
analyte involved in this patient’s condition?
DOI: 10.1309/X3LYTDQQJ5J8QU8R

Possible Answers:

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Patient 1. Low density lipoprotein cholesterol (LDL-C) was unde-
69-year-old African-American male. tectable by calculation and by lipoprotein electrophoresis of
our patient’s serum. The undetectable LDL-C by calculation
Chief Complaint using the Friedewald formula prompted lipoprotein
Patient presented to the emergency room at a local hospital electrophoresis of our patient’s serum.
with a nosebleed. The emergency room physician ordered The absence or extremely low concentration of LDL-C in
routine laboratory tests, including a lipid panel, as part of the the plasma can be the result of primary or secondary causes.
work-up on this patient. Primary causes of low or absent LDL-C include
abetalipoproteinemia and hypobetalipoproteinemia. Abetal-
Past Medical History ipoproteinemia is a rare autosomal recessive disorder in
There was no history of headache, double vision, skin rash, which plasma cholesterol, triglyceride, and total phospho-
trauma, or bleeding disorder. He claimed a history of hyper- lipid concentrations are extremely low.1-4 In this disorder,
tension, but had not been compliant with his antihypertensive both the phospholipid and fatty acid composition of the
medication. plasma lipoproteins are abnormal. Screening tests for abetal-
ipoproteinemia include a lipid panel, followed by lipoprotein
Physical Examination electrophoresis. The absence of a beta-lipoprotein band on
Significant only for elevated blood pressure (190/93 mmHg) electrophoresis indicates extremely low or absent plasma
and nosebleed. LDL-C. Definitive diagnosis of abetalipoproteinemia
depends on immunochemical demonstration of the absence,
Family History or markedly reduced concentration, of apolipoprotein B (apo
Unremarkable/not known B), the principal apolipoprotein of LDL-C.1
Hypobetalipoproteinemia is unrelated to abetalipoproteine-
Principal Laboratory Findings mia and is inherited as an autosomal dominant trait.1-3 In this
See [T1] and [I1]. disorder, total cholesterol concentration is low, triglyceride

T1
Principal Laboratory Findings

Reference Values*
Test Patient’s Results Desirable or Near or Above Borderline High Very
Optimal Optimal High High

Appearance of serum Slight turbidity Clear – – – –

TC, mg/dL 88 <200 – 200-239 >240 –
TG, mg/dL 153 <150 – 150-199 200-499 >500
LDL-C,† mg/dL Und <100 100-129 130-159 160-189 >190
280 HDL-C,‡ mg/dL 64 <40, Low; ∃60, High
Male Female CHD Risk
TC:HDL-C ratio 1.4 3.4 3.3 = average
5.0 4.4 average
9.6 7.1 2x average
24.0 11.0 3x average
*According to current guidelines of the National Cholesterol Education Program (NCEP).7 †Calculated according to the Friedewald formula: LDL-C = TC - (HDL-C) - (0.2 x TG). ‡High

levels of HDL-C and low levels of LDL-C are desirable because of the reduced risk of CHD in individuals with this combination of lipoprotein results.
TC, total cholesterol; TGs, triglycerides; LDL-C, low density lipoprotein cholesterol; HDL-C, high density lipoprotein cholesterol; Und, undetectable; CHD, coronary heart disease

laboratorymedicine> april 2003> number 4> volume 34 ©

levels are usually in the low-normal range, and high density
lipoprotein cholesterol (HDL-C), while variable, is generally
normal. The LDL-C level in homozygous individuals is
below the 10th percentile for age and gender, and below the
50th percentile in heterozygous individuals. In addition, a
faint beta-band will be observed on lipoprotein electrophore-
sis of the patient’s serum and minimal LDL-C concentrations
will be detected by immunochemical or ultracentrifugal
analysis.
Secondary causes are far more common than primary
causes and include systemic diseases, malignancies, anti-
LDL-C antibodies, and laboratory errors. Systemic diseases
such as malnutrition, alcoholism, gastrointestinal disease
(malabsorption states), and lipid storage diseases can result in
markedly reduced total and LDL-C levels.1-4 Lipid storage
diseases such as Gaucher’s and Niemann-Pick disease acceler-
ate lipoprotein catabolism which markedly decreases plasma
concentrations of LDL-C.4 Malignancies, in which tumor
cells up-regulate LDL-C receptors, markedly reduce the
plasma LDL-C concentration. Hematopoietic malignancies
(eg, acute and chronic myelogenous leukemias, myelodyspla-
sia with myeloid metaplasia, and multiple myeloma) have
been reported to be associated with very low HDL-C and
LDL-C levels.1,4 Therefore, a work-up to exclude any occult
malignancy (especially hematopoietic) should be undertaken
in patients with extremely low or undetectable LDL-C, if clin-
ically indicated. Several autoimmune disorders (eg, rheuma-
toid arthritis and systemic lupus erythematosis) and
dysglobulinemias produce anti-LDL antibodies which reduce
the plasma LDL-C concentration.1 It is postulated that faster
catabolism of antibody-bound LDL-C in vivo and cryoprecipi-
tation of immune complexes in vitro are the mechanisms ex-
plaining the reduction in LDL-C in these disorders. Although
laboratory analytical errors occur less frequently with in-
creasing automation and technological advancements in the
medical laboratory, pre-analytical errors in patient preparation,
collection techniques, and sample handling are potential
sources of erroneously low LDL-C values. Repeating the tests
comprising the lipid panel or redrawing the patient and testing
a fresh specimen can potentially resolve such errors.
2. Most likely diagnosis: Absence of LDL-C secondary to
systemic disease or malignancy. Unfortunately, our patient
had a fragmented history and was lost to follow-up. Never-
theless, from his known history and examination during his
emergency room visit, he did not have any overt clinical
signs or symptoms of either abetalipoproteinemia or familial
hypobetalipoproteinemia. Thus, it is more likely that the ab-
sence of LDL-C in our patient’s plasma occurred secondary
to a systemic disease (eg, malnutrition) or an unidentified un-
derlying malignancy.
3. Pathogenesis of primary causes of low or markedly
reduced LDL-C concentration: Abetalipoproteinemia is an
autosomal, recessively inherited disorder characterized by
© laboratorymedicine> april 2003> number 4> volume 34
+
(α)
(pre-β)
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(β)
0
Patient Control —
[I1] The patient’s serum lipoprotein electrophoresis pattern demonstrates
the absence of a beta-lipoprotein band (arrow) compared to a normal
control sample demonstrating the usual location of alpha (α), pre-β, and
beta (β) lipoprotein bands from the anode (+, positively-charged
electrode) to the cathode (-, negatively-charged electrode). 0, origin or
point-of-application of serum.
extremely low total cholesterol and triglyceride levels (<30
mg/dL), and the complete absence of apo B-containing
lipoproteins (ie, chylomicrons, very low density lipoproteins
(VLDL), LDL, lipoprotein “little a” [Lp(a)]). Moreover, a
rare variant of abetalipoproteinemia exists in which, in con-
trast to classical abetalipoproteinemia, triglycerides are ab-
sorbed in the intestine and chylomicrons are formed. The
normal serum triglyceride levels that occur in individuals
with this variant form of abetalipoproteinemia accounts for
its name, “normotriglyceridemic abetalipoproteinemia.”5
The defect in abetalipoproteinemia appears to be in intes-
tinal and hepatic apo B metabolism. Apo B-100 (hepatic ori-
gin) and B-48 (intestinal origin) are degraded shortly after
transcription due to defects in microsomal transport proteins
with resulting lack of both chylomicron and VLDL forma-
tion.1,2,4 Consequently, lipids accumulate in the liver and in-
testine (enterocytes). The intestinal lipid accumulation leads
to severe lipid malabsorption. While apo B-containing
lipoproteins (ie, LDL, VLDL, and chylomicrons) are absent
in the plasma of affected individuals, both the apo B gene 281
and its products are normal. In patients with normotriglyceri-
demic abetalipoproteinemia, there is selective inhibition of
the hepatic synthesis of apo B-100; however, the intestinal
synthesis of apo B-48 is apparently not affected.5 Familial hy-
pobetalipoproteinemia is inherited as an autosomal dominant
disorder, in which mutations in the apo B gene lead to either a
regulatory or local defect in the production of apo B.2,3
 Apolipoprotein B levels are low to absent, depending on
whether the individual is heterozygous or homozygous for
the defective gene. Lipoproteins with beta or pre-beta mobil-
ity are decreased or absent after lipoprotein electrophoresis
of the patient’s serum.
4. The clinical manifestations of abetalipoproteinemia in
childhood or early adolescence include 5 basic features: 1)
fat malabsorption, 2) hypolipidemia (undetectable plasma
VLDL and LDL-cholesterol levels), 3) acanthocytosis, 4) re-
tinitis pigmentosa, and 5) ataxic neuropathic disease.2,3
Moreover, dietary fat in the form of chylomicrons is not pro-
between them.3 In heterozygous individuals, hematologic and
gastrointestinal manifestations are mild, including fewer
acanthocytes, limited fat malabsorption, and minor deficien-
cies of fat-soluble vitamins. Both homozygous and heterozy-
gous individuals can show neuromuscular and ophthalmic
manifestations, including ataxia, diminished reflexes, and re-
tinitis pigmentosa.
Methods for precisely quantifying the principal analyte,
LDL-C, (ie, “8-quantitation”) associated with our patient’s
condition include both indirect and direct methods. Indirect
methods assume that the total cholesterol concentration is
equal to the sum of all cholesterol-containing lipoproteins in

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duced in the intestine due to premature degradation of apo B-
48 in enterocytes. Fat malabsorption is present from birth,
and affected infants show poor appetite, loose, voluminous
stools, and little weight gain. In addition, most of these pa-
tients have deficiencies of fat-soluble vitamins, especially
vitamin E and/or vitamins A and K. Vitamin D is not
deficient as it is absorbed independently of chylomicrons.
Both vitamins A and K can also be transported across the in-
testines independent of lipoproteins, and clinical deficiency
may not be severe.2,3 Acanthocytes [constituting 50% to
100% of all red blood cells (RBCs)] are found circulating in
the peripheral blood and are thought to form due to maldistri-
bution of lipids between the bilayers of the RBC membrane.3
Affected individuals may also have extensive demyelination,
muscle weakness, loss of deep-tendon reflexes, and retinopa-
thy. The cause of the neuromuscular abnormalities remain
obscure, although progression of these abnormalities can be
inhibited and even reversed with supplementation of large
oral doses of vitamin E.3,4
The clinical presentation of patients with familial hypo-
betalipoproteinemia depends upon zygosity. Patients with
homozygous familial hypobetalipoproteinemia are clinically
indistinguishable from those with abetalipoproteinemia, and
family and genetic studies are required to distinguish
the serum [ie, TC = LDL-C + HDL-C + VLDL-C + interme-
diate-density lipoprotein-cholesterol (IDL-C)]. The most
widely used indirect method for asseesing LDL-C concentra-
tion is the Friedewald equation:
LDL-C = TC – (HDL-C) – (TG/5)
The term, TG/5 (or 0.2 x TG, because 1/5 = 0.2), pro-
vides an estimation of VLDL-C concentration based on the
average ratio of triglyceride to cholesterol in VLDL and the
concentration of IDL-C is assumed to be small compared to
the other cholesterol-containing lipoproteins in human
serum. A different factor for estimating VLDL-C concentra-
tion (ie, TG/2.175) is used when the lipoprotein cholesterol
concentrations are expressed in units of millimoles/liter
(mmol/L). The Friedewald equation will underestimate
LDL-C concentration in serum samples obtained from
whole blood specimens collected from non-fasting individu-
als and when TG concentrations are >400 mg/dL. The equa-
tion will overestimate LDL-C concentration in individuals
with Type III hyperlipoproteinemia because of the accumu-
lation of chylomicron and VLDL remnants in the plasma of
such individuals. Direct methods for quantifying LDL-C
concentration include preparative ultracentrifugation to sep-
arate LDL from other lipoproteins followed by cholesterol
analysis of the LDL fraction; selective precipitation of

Concentration and Function of Apolipoproteins in Serum Lipoproteins8

Apolipoprotein Concentration (%) in Serum Lipoprotein Component

T2
Apo Chylo VLDL LDL HDL2 HDL3 Function

A-I 33 Trace Trace 65 62 Cholesterol transport from cell membranes; cofactor of LCAT
A-II Trace Trace Trace 10 23 PL-binding properties; structural protein of HDL
A-IV 14 – – ? Trace Unknown
B 5 25 95 3 – B-100: Intracellular formation or transcellular transport of VLDL; LDL
receptor interaction
282 B-48: Intracellular formation or transcellular transport of chylomicrons
and VLDL; LDL receptor interaction?
C 32 55 2 13 5 C-I,-II: cofactor of adipose tissue LPL
D ? ? ? 2 4 Unknown
E 10 15 3 3 1 Receptor interaction with apo B,E receptor cells; inhibitor of adipose
tissue LPL
Other 6 5 5 4 5 –

Apo, apolipoprotein; Chylo, chylomicrons; VLDL, very low density lipoproteins; LDL, low density lipoproteins; HDL2 and HDL3, high density lipoprotein subfractions, as determined by
ultracentrifugation; LCAT, lecithin:cholesterol acyltransferase; PL, phospholipid; LPL, lipoprotein lipase; ?, unknown

laboratorymedicine> april 2003> number 4> volume 34 ©

LDL-C with polyvinyl sulfate or heparin/manganese chloride
(MnCl2) at low pH; and immunochemical methods that use
polyclonal antibodies against apo A-1 and apo E to precipi-
tate lipoproteins rich in these types of apolipoproteins (ie,
VLDL, IDL, and HDL) and leaving behind only the LDL
fraction for quantitation of its cholesterol content.6 The rela-
tive concentration of various apolipoproteins in the serum
lipoproteins and their putative function are shown in T2.
Based on the data presented in T2, the principal apolipopro-
tein constituent of the major lipoproteins present in human
serum are: apo C (VLDL); apo B (LDL); apo A (HDL).
Keywords: lipoprotein, abetalipoproteinemia, Friedewald
equation, familial hypobetalipoproteinemia, low density
lipoprotein cholesterol
1. Naito HK. Coronary artery disease and disorders of lipid metabolism. In:
Clinical Chemistry, Theory, Analysis, Correlation, 3rd ed. St. Louis: Mosby-
Year Book Inc; 1996:661-663.
© laboratorymedicine> april 2003> number 4> volume 34
2. Bachorik PS, Denke MA, Stein EA, et al. Lipids and dyslipoproteinemia. In:
Clinical diagnosis and management by laboratory methods, 20th ed.
Philadelphia: W B Saunders; 2001:235-244.
3. Kane JP, Havel RJ. Disorders of the biogenesis and secretion of lipoproteins
containing the B apolipoproteins. In: The metabolic basis of inherited disease,
6th ed. New York: McGraw-Hill Information Services; 1989:1139-1164.
4. Goldberg IJ, Ginsberg HN. Disorders of lipoprotein metabolism. In: Harrison’s
principles of internal medicine, 14th edition. New York: McGraw-Hill
Companies; 1998:2138-2148.
5. Malloy MJ, Kane JP, Hardman DA, et al. Normotriglyceridemic
abetalipoproteinemia. Absence of the B-100 apolipoprotein. J Clin Invest.
1981;67:1441-1450.
6. Rifai N, Bachorik PS, Albers JJ. Lipids, lipoproteins, and apolipoproteins. In:
Tietz textbook of clinical chemistry, 3rd ed. Philadelphia: W B Saunders;
1999:843-845.
Downloaded from https://academic.oup.com/labmed/article/34/4/280/2504304 by guest on 16 April 2021
7. Expert panel on detection, evaluation, and treatment of high blood cholesterol
in adults. Executive summary of the third report of the National Cholesterol
Education Program (NCEP) expert panel on detection, evaluation, and treatment
of high blood cholesterol in adults (adult treatment panel III). JAMA.
2001;285:2486-2497.
8. Assman G. Lipid metabolism and atherosclerosis. Stuttgart: FK Schattauer
Verlag GmbH; 1982:18-19.
283