Biomedical subjects

G Ogbonna

Publications and source records attributed to G Ogbonna.

5 recordsLinked to original sources

PubMed1995

A competitive chemiluminescent immunoassay for the sensitive measurement of apolipoprotein B100.

OBJECTIVES: We have developed a competitive chemiluminescent immunoassay for the sensitive measurement of apolipoprotein B100 (apoB) and conducted a preliminary evaluation of the method. DESIGN AND METHODS: In the assay, apoB-rabbit immunoglobulin G (IgG) conjugate competes with apoB in the sample for binding to acridinium N-hydroxysuccinimide labelled anti-apoB antibody. Goat anti-rabbit-IgG immobilized to magnetic particles is used to separate the apoB-rabbit IgG conjugate bound to the labelled anti-apoB antibody. Chemiluminescence, measured using a Ciba Corning Magic-Lite chemiluminometer, is inversely proportional to the concentration of apoB in the sample. RESULTS: The assay is completed in 1-2 h. Within-run imprecision (n = 5) determined at 10, 100, and 200 micrograms/L of apoB was found to be 4.8%, 2.4%, and 3.5%, respectively. The between-run imprecision (n = 5) at the same concentrations of apoB was determined to be 14.2%, 9.5%, and 7.7%, respectively. The proposed assay showed reasonable correlation (r = 0.86) with a commercially available immunoturbidimetric method. The lower limit of detection was 2.2 micrograms/L (4.0 pmol/L). CONCLUSIONS: This assay may be useful in tissue culture studies where sensitive measurement of secreted and intracellular concentrations of apoB are required. Our preliminary evaluation of the assay appears to confirm is usefulness.

Acridines↗

PubMed1993

Hormonal regulation of human apolipoprotein E gene expression in HepG2 cells.

1. Hormonal regulation of apolipoprotein E (apoE) gene expression by insulin and thyroid hormone was studied in a human hepatoma cell line, HepG2. 2. Changes at the mRNA level, mRNA translation, in vivo synthesis and secretion were monitored. 3. Both insulin and triiodothyronine were found to have no significant effect on apoE mRNA levels. 4. Insulin treatment caused an inhibition of: (a) the in vitro translation of endogenous apoE mRNA in a HepG2 cell-free system (25%), and (b) the incorporation of radioactivity into newly-synthesized apoE in an in vivo pulse-chase labeling experiment (32%). 5. Interestingly, apoE secretion rate was found to be significantly reduced with insulin (84%) suggesting that a major portion of newly-synthesized apoE may be shunted into a degradative pathway. 6. Using a similar experimental approach, triiodothyronine showed no significant effect on the rate of apoE synthesis or translation (6-15% decrease), however a slight reduction (20%) in secretion rate was shown. 7. Overall, apoE gene expression does not appear to be influenced by triiodothyronine significantly but is modulated by insulin at the translational and post-translational level.

Apolipoproteins E↗

PubMed1992

Thyroid hormone modulates apolipoprotein B gene expression in HepG2 cells.

We have investigated the modulation of apolipoprotein B gene expression in HepG2 cells by thyroid hormone. ApoB secretion rate in serum-free media was found to be significantly increased in the presence of the hormone in long-term cultures (48 h, 37%). This stimulatory effect was dose-dependent. The mechanisms underlying the stimulatory effect of triiodothyronine on apoB production were investigated. Triiodothyronine increased apoB mRNA levels by about 25-36% as determined by slot- and Northern-blot analysis of total RNA. ApoB synthesis rate was also found to be increased both in in vivo pulse-chase experiments (61%) and in in vitro translation studies (54.5%). Despite the 54.5-61% increase in apoB synthesis with triiodothyronine, only a 30% increase in apoB secretion was noted suggesting that part of the increase in the intracellular apoB pool may be lost by degradation. Overall, apoB gene expression appears to be modulated by thyroid hormone at both transcriptional and posttranscriptional levels.

Apolipoproteins B↗

PubMed1992

Apolipoprotein B gene polymorphism and plasma lipids and lipoproteins in a Canadian Caucasian population.

We have investigated the frequency of Hind III DNA polymorphism of the human apolipoprotein B gene in a Canadian Caucasian population with coronary artery disease, as documented by angiography, and a healthy control population. Patients had significantly (p < 0.05) higher levels of cholesterol, triglycerides, LDL-cholesterol, apolipoprotein B, and lower level of apoAI compared to the controls. Restriction fragment-length polymorphism analysis detected nine hybridizable fragments denoted as H1 to H9. The H1, H2, H3, and H7 alleles were polymorphic. The [H4-H9] genotype seems to be the normal genotype within the population studied since it was detected in 69% of the control group. The [H1-H9] genotype was most frequently observed in the patients (frequency = 0.68). We were unable to strongly associate any of the alleles or genotypes detected with the changes in lipids. The additional alleles observed in the patient group may indicate possible mutations at the 3' end of the apolipoprotein B gene locus.

Adult↗

PubMed1990

Rapid purification of human DNA from whole blood for potential application in clinical chemistry laboratories.

A simple, rapid method for isolating human DNA has been developed, which can be routinely used in clinical chemistry laboratories. The entire procedure takes less than 90 min, and as many as 12 blood samples can be handled in one cycle. One milliliter of EDTA-treated blood is lysed and centrifuged to yield a nuclear fraction. The nuclear pellet is treated with sodium dodecyl sulfate/urea and phenol/chloroform to remove contaminating proteins, then the crude DNA extract is purified by use of a Sephadex G-25 spin-column. Typical 260 nm/280 nm absorbance ratio (used to assess purity) and yield for DNA so purified were 1.84 and 24.5 micrograms/mL, respectively. Within- and between-day CVs for recovery of DNA from pooled blood were 8% and 11% respectively. Such DNA preparations were found quite suitable for digestion by a variety of restriction endonucleases and for restriction fragment length polymorphism analysis. We are using this method to isolate DNA from whole blood of myocardial infarction patients for studies on the apolipoprotein B gene.

Apolipoproteins B↗