Secretion of apolipoprotein A-I by calf liver parenchymal cells

Toru Miyamoto From the Laboratory of Biochemistry, National Institute of Animal Health, 3-1-1 Kannondai, Tsukuba, Ibaraki 305, Japan.

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Norio Katoh From the Laboratory of Biochemistry, National Institute of Animal Health, 3-1-1 Kannondai, Tsukuba, Ibaraki 305, Japan.

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Abstract

Objective

To determine: whether apolipoprotein A-I (apoA-I) is secreted by calf liver parenchymal cells; the isoprotein pattern and association with lipids of secreted apoA-I; and effects of steroid hormones on apoA-I secretion.

Sample Population

6 male Holstein calves (1 to 2 weeks old) as a cell culture source.

Procedure

apoA-I in culture medium was detected by immunoblot analysis, and its concentration was measured by use of an ELISA. The isoprotein pattern was analyzed by use of two-dimensional polyacrylamide gel electrophoresis. Associations of apoA-I with cholesterol and phospholipids were examined by ultracentrifugal analysis of the culture medium.

Results

Concentration of apoA-I in culture medium increased linearly up to 24 hours. The protein synthesis inhibitors, actinomycin D and cycloheximide, suppressed apoA-I accumulation in the medium in dose-dependent manner. Molecular mass of this protein in culture medium was 28 kd, and was indistinguishable from apoA-I of plasma. Four isoproteins with different isoelectric points (1 = 5.75; 2 = 5.67; 3 = 5.58; and 4 = 5.46) were detected. Of these, isoprotein 2 was the major species. By comparison, isoprotein 4 was the predominant species in plasma, and isoprotein 5 (isoelectric point = 5.38) was newly detected instead of isoprotein 1. Approximately 33% of apoA-I in culture medium was found in lipid-rich fractions, whereas the rest was found in nonlipoprotein fractions. Dexamethasone (10−8 to 10−6M) significantly (P < 0.001) increased apoA-I concentration in the medium, and the stimulatory effect was significantly (P < 0.001) suppressed by the simultaneous addition of 10−5M progesterone. Progesterone itself (10−6 to 10−5M) had little effect on apoA-I secretion; estradiol (10−14 to 10−6M) also had no significant effect.

Conclusions

apoA-I is synthesized by calf liver parenchymal cells, and the secreted protein is modified during circulation. Moreover, apoA-I synthesis and secretion by the cells appear not to be largely influenced by hormones, except for dexamethasone. (Am J Vet Res 1997;58:811–815)

Abstract

Objective

To determine: whether apolipoprotein A-I (apoA-I) is secreted by calf liver parenchymal cells; the isoprotein pattern and association with lipids of secreted apoA-I; and effects of steroid hormones on apoA-I secretion.

Sample Population

6 male Holstein calves (1 to 2 weeks old) as a cell culture source.

Procedure

apoA-I in culture medium was detected by immunoblot analysis, and its concentration was measured by use of an ELISA. The isoprotein pattern was analyzed by use of two-dimensional polyacrylamide gel electrophoresis. Associations of apoA-I with cholesterol and phospholipids were examined by ultracentrifugal analysis of the culture medium.

Results

Concentration of apoA-I in culture medium increased linearly up to 24 hours. The protein synthesis inhibitors, actinomycin D and cycloheximide, suppressed apoA-I accumulation in the medium in dose-dependent manner. Molecular mass of this protein in culture medium was 28 kd, and was indistinguishable from apoA-I of plasma. Four isoproteins with different isoelectric points (1 = 5.75; 2 = 5.67; 3 = 5.58; and 4 = 5.46) were detected. Of these, isoprotein 2 was the major species. By comparison, isoprotein 4 was the predominant species in plasma, and isoprotein 5 (isoelectric point = 5.38) was newly detected instead of isoprotein 1. Approximately 33% of apoA-I in culture medium was found in lipid-rich fractions, whereas the rest was found in nonlipoprotein fractions. Dexamethasone (10−8 to 10−6M) significantly (P < 0.001) increased apoA-I concentration in the medium, and the stimulatory effect was significantly (P < 0.001) suppressed by the simultaneous addition of 10−5M progesterone. Progesterone itself (10−6 to 10−5M) had little effect on apoA-I secretion; estradiol (10−14 to 10−6M) also had no significant effect.

Conclusions

apoA-I is synthesized by calf liver parenchymal cells, and the secreted protein is modified during circulation. Moreover, apoA-I synthesis and secretion by the cells appear not to be largely influenced by hormones, except for dexamethasone. (Am J Vet Res 1997;58:811–815)

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