Abstract
A sensitive solution hybridization assay using autologous cRNA probes was developed with the aim to study the simultaneous regulation of hepatic mRNA levels, on a quantitative basis, for the LDL receptor (LDLr), HMG-CoA reductase, and cholesterol 7 alpha-hydroxylase (Cho-7-hx) in C57BL/6J mice. With the purpose to suppress and stimulate transcript levels respectively, animals received established high fat diets, cholesterol-enriched diets, and a diet supplemented with mevinolin and colestipol. One hundred nineteen animals were investigated in six separate experiments. In spite of an eightfold increase in hepatic cholesterol induced by a high fat diet, the LDLr and the HMG-CoA reductase mRNA levels were only reduced to 60-70% and 25-50% of control values, respectively. When the data from all animals were analyzed, a strong positive correlation was obtained between the mRNA levels for the LDLr and HMG-CoA reductase (r = 0.79, P less than 0.001). A significant relation remained when control animals only were analyzed (n = 42, r = 0.59, P less than 0.001). Cho-7-hx mRNA showed a regulatory pattern that differed from that of the LDLr and HMG-CoA reductase; feeding cholesterol at 1.7% and 5% but not at 0.4% elevated the mRNA levels for Cho-7-hx while the LDLr and HMG-CoA reductase mRNA levels were maximally suppressed already at 0.4% of dietary cholesterol. The results show that the mRNA levels for the LDLr and HMG-CoA reductase are regulated in parallel in the liver in vivo during various metabolic perturbations as well as at normal physiologic conditions.(ABSTRACT TRUNCATED AT 250 WORDS)
Highlights
A sensitive solution hybridization assay using autologous Complementary RNA probes (cRNA) probes was developed with the aim to study the simultaneous regulation of hepatic mRNA levels, on a quantitative basis, for the low density lipoproteins (LDL) receptor (LDLr),HMG CoA reductase, and cholesterol 7a-hydroxylase (Cho-7-hx) in C57BL/6J mice
The regulation of HMGCoA reductase is extremely high and rapid due to regulation at the transcriptional, translational, as well as at the protein level (1).The LDLr is believed to be regulated at the transcriptional level; recent reports suggest that the LDL receptor may be regulated at a posttranscriptional level in vitro and in vivo (2,3)
The plasma cholesterol level in various species including humans is determined by the net balance between the input of cholesterol into plasma and the removal rate from plasma, the latter in part dependent on the number of LDL receptors (LDLr)
Summary
A sensitive solution hybridization assay using autologous cRNA probes was developed with the aim to study the simultaneous regulation of hepatic mRNA levels, on a quantitative basis, for the LDL receptor (LDLr),HMG CoA reductase, and cholesterol 7a-hydroxylase (Cho-7-hx) in C57BL/6J mice. Raw data from three separate solution hybridization assays on hepatic RNA from four animals of experiment 2 (left column) and the corresponding standard cuwes (right column) generated with the indicated synthetic mRNA.
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