Combination of lipid metabolism alterations and their sensitivity to inflammatory cytokines in human lipin-1-deficient myoblasts

Abstract

Lipin-1 deficiency is associated with massive rhabdomyolysis episodes in humans, precipitated by febrile illnesses. Despite well-known roles of lipin-1 in lipid biosynthesis and transcriptional regulation, the pathogenic mechanisms leading to rhabdomyolysis remain unknown. Here we show that primary myoblasts from lipin-1-deficient patients exhibit a dramatic decrease in LPIN1 expression and phosphatidic acid phosphatase 1 activity, and a significant accumulation of lipid droplets (LD). The expression levels of LPIN1-target genes [peroxisome proliferator-activated receptors delta and alpha (PPARδ, PPARα), peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC-1α), acyl-coenzyme A dehydrogenase, very long (ACADVL), carnitine palmitoyltransferase IB and 2 (CPT1B and CPT2)] were not affected while lipin-2 protein level, a closely related member of the family, was increased. Microarray analysis of patients’ myotubes identified 19 down-regulated and 51 up-regulated genes, indicating pleiotropic effects of lipin-1 deficiency. Special attention was paid to the up-regulated ACACB (acetyl-CoA carboxylase beta), a key enzyme in the fatty acid synthesis/oxidation balance. We demonstrated that overexpression of ACACB was associated with free fatty acid accumulation in patients’ myoblasts whereas malonyl-carnitine (as a measure of malonyl-CoA) and CPT1 activity were in the normal range in basal conditions accordingly to the normal daily activity reported by the patients. Remarkably ACACB invalidation in patients’ myoblasts decreased LD number and size while LPIN1 invalidation in controls induced LD accumulation. Further, pro-inflammatory treatments tumor necrosis factor alpha + Interleukin-1beta(TNF1α + IL-1β) designed to mimic febrile illness, resulted in increased malonyl-carnitine levels, reduced CPT1 activity and enhanced LD accumulation, a phenomenon reversed by dexamethasone and TNFα or IL-1β inhibitors. Our data suggest that the pathogenic mechanism of rhabdomyolysis in lipin-1-deficient patients combines the predisposing constitutive impairment of lipid metabolism and its exacerbation by pro-inflammatory cytokines.

Rhabdomyolysis-Associated Mutations in Human LPIN1 Lead to Loss of Phosphatidic Acid Phosphohydrolase Activity

“…Previous work showed that PAP activity was markedly reduced in isolated myocytes from three patients with lipin 1 mutations, including the E766-S838_del mutation studied herein (Michot et al 2013). However, those cells exhibited a complete loss of lipin 1 mRNA expression, and a distinction regarding impaired transcriptional regulatory function or PAP activity could not be made.…”

Altered Lipid Synthesis by Lack of Yeast Pah1 Phosphatidate Phosphatase Reduces Chronological Life Span

“…These phenotypic changes in lipid metabolism are directly or indirectly coupled to other phenotypes, such as an irregular expansion of the nuclear/ER membrane, marked vacuole fragmentation, and increased fatty acid-induced lipotoxicity (11,29,34,35,38). Similarly, in mammalian cells, loss of lipin PAP activity results in metabolic disorders that include lipodystrophy, insulin resistance, peripheral neuropathy, rhabdomyolysis, and inflammation (30,(93)(94)(95)(96)(97)(98)(99)(100)(101).…”

Cross-talk Phosphorylations by Protein Kinase C and Pho85p-Pho80p Protein Kinase Regulate Pah1p Phosphatidate Phosphatase Abundance in Saccharomyces cerevisiae

“…1A) (10,14). The losses of or defects in mammalian lipin PAP enzymes underlie metabolic disorders that include lipodystrophy and insulin resistance, peripheral neuropathy, myoglobinuria, and inflammation (7,(15)(16)(17)(18)(19)(20)(21)(22)(23)(24)(25)(26). Together, these observations highlight the importance of PAP in lipid metabolism and cell physiology and the need to understand the genetic and biochemical regulations of the enzyme.…”