الملخص الإنجليزي
Background: Dyslipidemia is a risk factor for CVDs. Bile acids are produced by
hepatocytes in conjugated forms and secreted into the intestine in which they are
converted to secondary unconjugated BA by the gut microbiota. Due to their ability to
modulate gene expressions, BAs involve in the regulation of numerous metabolic
processes like; lipid and cholesterol homeostasis. BA production represents the most
efficient cholesterol elimination route. Serum level of 7-alpha-hydroxy-4-cholesten-3-
one (C4) is widely used as a marker for CYPA71, a key enzyme in the BA synthesis
pathway. This enzyme is regulated by BAs themselves and by the fibroblast growth
factor-19 (FGF19). HPLC coupled with tandem mass spectrometry (LC-MS/MS) is
currently the technique of choice for accurate BA detection and quantification in a
variety of biological matrices. Hence, we present a validated LC-MS/MS method for the
combined quantitative analysis of the 15 major human bile acids. In addition, we sought
to explore the underlying causes of reduced/altered bile acid levels in patients with
premature acute coronary syndrome (PACS)/myocardial infarction (MI), and whether
variations in circulating BAs are predictive of CAD among Omanis.
Methods: LC-MS/MS method for the combined quantification of human BAs and C4
was established and validated based on the recommendations of the FDA. The methods’
linearity, limits of detection and quantification, method precision and accuracy, and
carryover, in addition to matrix effect and recovery were evaluated. Using this method,
the BA profile and C4 levels were analyzed in serum samples of PACS Patients (n=91)
and healthy controls (n=146). Serum FGF19 level was quantified using a commercially
available ELISA kit.
Results: Validation of the method showed high linear responses, good sensitivity,
accuracy, and precision with limited matrix effects, fair analyte recoveries and no
carryover. In addition, results obtained from the developed method were compared to a
sensitive colorimetric assay which showed a good correlation. PACS patients had
significantly lower unconjugated BAs (p=0.026), and elevated taurine-conjugated BAs
in serum (p=0.021). On the other hand, equal median C4, and similar FGF19 levels (p =
0.83) were observed. Of the BA groups, conjugated BAs were predictive of CAD after
controlling for age and gender with odds ratio (OR) (95%CI) of [1.401 (1.04- 1.89), p
<0.05]
Conclusion: A straightforward, simple, and sensitive LC-MS/MS method for the
simultaneous determination of the 15 human bile acids and their precursor C4 in serum
was developed, validated, and applied for the profiling of serum BAs in PACS patients.
Patients had an altered BA composition. An in-depth investigation into the causative
factors of the observed alteration might provide novel prognostic and therapeutic
approaches.
