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Articles |
1
Department of Cardiovascular Biochemistry, St. Bartholomew’s and The Royal London School of Medicine and Dentistry, Charterhouse Square, London EC1 M 6BQ, United Kingdom.
2
Section of Metabolism, Endocrinology, and Nutrition,
111E Carl T. Hayden VA Medical Center, 650 East Indian School Road,
Phoenix, AZ 85012-1892.
a Author for correspondence. Fax 602-200-6004; e-mail eliot.brinton{at}med.va.gov
Background: Very small apolipoprotein (apo) A-I-containing lipoprotein (Sm LpA-I) particles with pre-ß electrophoretic mobility may play key roles as "nascent" and/or "senescent" HDL; however, methods for their isolation are difficult and often semiquantitative.
Methods: We developed a preparative method for separating Sm LpA-I particles from human plasma by high-performance size-exclusion chromatography (HP-SEC), using two gel permeation columns (Superdex 200 and Superdex 75) in series and measuring apo A-I content in column fractions in 30 subjects with HDL-cholesterol (HDL-C) concentrations of 0.4–3.83 mmol/L.
Results: Three major sizes of apo A-I-containing particles were detected: an ~15-nm diameter (~700 kDa) species; a 7.5–12 nm (100–450 kDa) species; and a 5.8–6.3 nm species (40–60 kDa, Sm LpA-I particles), containing 0.2–3%, 80–96%, and 2–15% of plasma total apo A-I, respectively. Two subjects with severe HDL deficiency had increased relative apo A-I content in Sm LpA-I: 25% and 37%, respectively. The percentage of apo A-I in Sm LpA-I correlated positively with fasting plasma triglyceride concentrations (r = 0.581; P <0.0005) and inversely with total apo A-I (r = -0.551; P <0.0013) and HDL-C concentrations (r = -0.532; P <0.0017), although the latter two relationships were largely attributable to extremely hypoalphalipoproteinemic subjects. The percentage of apo A-I in Sm LpA-I correlated with that in pre-ß-migrating species by crossed immunoelectrophoresis (r = 0.98; P <0.0001; n = 24) and with that in the d >1.21 kg/L fraction by ultracentrifugation (r = 0.86; P <0.001; n = 20). Sm LpA-I particles, on average, appear to contain two apo A-I and four phospholipid molecules but little or no apo A-II, triglyceride, or cholesterol.
Conclusions: We present a new HP-SEC method for size separation of native HDL particles from plasma, including Sm Lp A-I, which may play important roles in the metabolism of HDL and in its contribution(s) to protection against atherosclerosis. This method provides a basis for further studies of the structure and function of Sm Lp A-I.
The following articles in journals at HighWire Press have cited this article:
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