Dataset Information

Sortilin facilitates VLDL-B100 secretion by insulin sensitive McArdle RH7777 cells.

ABSTRACT: Studies examining the relationship between cellular sortilin and VLDL-B100 secretion demonstrate inconsistent results. Current studies explore the possibility that discrepancies may be related to insulin sensitivity. McArdle RH7777 cells (McA cells) cultured under serum enriched conditions lose sensitivity to insulin. Following incubation in serum-free DMEM containing 1% BSA, McA cells become insulin responsive and demonstrate reduced apo B secretion. Current studies indicate that insulin sensitive McA cells express lower cellular sortilin that corresponds with reduction in VLDL-B100 secretion without changes in mRNA of either sortilin or apo B. When sortilin expression is further reduced by siRNA knockdown (KD), there are additional decreases in VLDL-B100 secretion. A crystal structure of human sortilin (hsortilin) identifies two binding sites on the luminal domain for the N- and C-termini of neurotensin (NT). A small organic compound (cpd984) was identified that has strong theoretical binding to the N-terminal site. Both cpd984 and NT bind hsortilin by surface plasmon resonance. In incubations with insulin sensitive McA cells, cpd984 was shown to enhance VLDL-B100 secretion at each level of sortilin KD suggesting cpd984 acted through sortilin in mediating its effect. Current results support a role for sortilin to facilitate VLDL-B100 secretion which is limited to insulin sensitive McA cells. Inconsistent reports of the relationship between VLDL-B100 secretion and sortilin in previous studies may relate to differing functions of sortilin in VLDL-B100 secretion depending upon insulin sensitivity.

SUBMITTER: Sparks RP

PROVIDER: S-EPMC5002383 | biostudies-literature | 2016 Sep

REPOSITORIES: biostudies-literature

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Sortilin facilitates VLDL-B100 secretion by insulin sensitive McArdle RH7777 cells.

Sparks Robert P RP Guida Wayne C WC Sowden Mark P MP Jenkins Jermaine L JL Starr Matthew L ML Fratti Rutilio A RA Sparks Charles E CE Sparks Janet D JD

Biochemical and biophysical research communications 20160802 2

Studies examining the relationship between cellular sortilin and VLDL-B100 secretion demonstrate inconsistent results. Current studies explore the possibility that discrepancies may be related to insulin sensitivity. McArdle RH7777 cells (McA cells) cultured under serum enriched conditions lose sensitivity to insulin. Following incubation in serum-free DMEM containing 1% BSA, McA cells become insulin responsive and demonstrate reduced apo B secretion. Current studies indicate that insulin sensit ...[more]

PMID: 27495870

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Project description:RATIONALE:Genome-wide association studies identified single-nucleotide polymorphisms near the SORT1 locus strongly associated with decreased plasma LDL-C (low-density lipoprotein cholesterol) levels and protection from atherosclerotic cardiovascular disease and myocardial infarction. The minor allele of the causal SORT1 single-nucleotide polymorphism locus creates a putative C/EBP? (CCAAT/enhancer-binding protein ?)-binding site in the SORT1 promoter, thereby increasing in homozygotes sortilin expression by 12-fold in liver, which is rich in this transcription factor. Our previous studies in mice have showed reductions in plasma LDL-C and its principal protein component, apoB (apolipoprotein B) with increased SORT1 expression, and in vitro studies suggested that sortilin promoted the presecretory lysosomal degradation of apoB associated with the LDL precursor, VLDL (very-low-density lipoprotein). OBJECTIVE:To determine directly that SORT1 overexpression results in apoB degradation and to identify the mechanisms by which this reduces apoB and VLDL secretion by the liver, thereby contributing to understanding the clinical phenotype of lower LDL-C levels. METHODS AND RESULTS:Pulse-chase studies directly established that SORT1 overexpression results in apoB degradation. As noted above, previous work implicated a role for lysosomes in this degradation. Through in vitro and in vivo studies, we now demonstrate that the sortilin-mediated route of apoB to lysosomes is unconventional and intersects with autophagy. Increased expression of sortilin diverts more apoB away from secretion, with both proteins trafficking to the endosomal compartment in vesicles that fuse with autophagosomes to form amphisomes. The amphisomes then merge with lysosomes. Furthermore, we show that sortilin itself is a regulator of autophagy and that its activity is scaled to the level of apoB synthesis. CONCLUSIONS:These results strongly suggest that an unconventional lysosomal targeting process dependent on autophagy degrades apoB that was diverted from the secretory pathway by sortilin and provides a mechanism contributing to the reduced LDL-C found in individuals with SORT1 overexpression.

Project description:ContextThe exact mechanisms responsible for increased plasma triglyceride (TG) concentration in obese people are unclear, and it is not known whether excess energy intake per se is involved in the pathophysiology of this abnormality.ObjectiveThe purpose of our study was to examine how excess energy intake from a balanced diet for 1 day affects very-low-density lipoprotein (VLDL)-TG kinetics and its putative regulators hepatic insulin sensitivity and plasma free fatty acid availability.Subjects and designWe used stable isotope-labeled tracer methods to evaluate glucose and lipid kinetics in 8 overweight and obese men (age, 38 ± 3 years; body mass index, 33.7 ± 1.7 kg/m(2); means ± SEM) on 2 occasions (randomized crossover design): once, the day after they consumed a balanced diet that provided an amount of energy that matched their energy expenditure, and another time, the day after they consumed a balanced diet that provided 30% excess calories. Eight healthy, lean men (34 ± 1 years; 22.5 ± 0.6 kg/m(2)) were studied under isocaloric conditions only to provide a reference for normal lipid kinetics.ResultsVLDL-TG and VLDL-apolipoprotein B-100 (apoB-100) concentrations and secretion rates were significantly greater (P < .01) in overweight/obese compared with lean men. Hypercaloric, compared with isocaloric, feeding in overweight/obese men increased glucose rate of appearance in plasma (904 ± 21 vs 873 ± 26 μmol/min), the hepatic insulin resistance index (10.9 ± 2.2 vs 8.3 ± 1.8), and VLDL-apoB-100 concentration and secretion rate (1.91 ± 0.24 vs. 1.53 ± 0.13 nmol/min), whereas VLDL-apoB-100 plasma clearance rate, VLDL-TG secretion and plasma clearance rates, and free fatty acid rate of appearance in plasma were not affected by overfeeding.ConclusionOne day of moderate overfeeding (30% excess energy intake) stimulates hepatic glucose and VLDL-apo B-100 secretion rates but has no effect on hepatic and adipose tissue fatty acid metabolism in overweight/obese men.

Dataset Information

Sortilin facilitates VLDL-B100 secretion by insulin sensitive McArdle RH7777 cells.

Publications

Sortilin facilitates VLDL-B100 secretion by insulin sensitive McArdle RH7777 cells.

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