Mfsd2a-mediated lysolipid transport is important for renal recovery after acute kidney injury.

Acute kidney injury (AKI) is a global public health concern with high mortality and morbidity. In ischemic-reperfusion injury (IRI), a main cause of AKI, the brush border membrane of S3 proximal tubules (PT) is lost to the tubular lumen. How injured tubules reconstitute lost membrane lipids during renal recovery is not known. Here, we identified Mfsd2a, a sodium-dependent lysophosphatidylcholine (LPC) transporter, to be expressed specifically in the basolateral membrane of S3 PT. Using an in vivo activity probe for Mfsd2a, transport activity was found to be specific to the S3 PT. Mice with haploinsufficiency of Mfsd2a exhibited delayed recovery of renal function after acute IRI, with depressed urine osmolality and elevated levels of histological markers of damage, fibrosis, and inflammation, findings corroborated by transcriptomic analysis. Lipidomics revealed a deficiency in docosahexaenoic acid (DHA) containing phospholipids in Mfsd2a haploinsufficiency. Treatment of Mfsd2a haploinsufficient mice with LPC-DHA improved renal function and reduced markers of injury, fibrosis, and inflammation. Additionally, LPC-DHA treatment restored S3 brush border membrane architecture and normalized DHA-containing phospholipid content. These findings indicate that Mfsd2a-mediated transport of LPC-DHA is limiting for renal recovery after AKI and suggest that LPC-DHA could be a promising dietary supplement for improving recovery following AKI.

Deficiency in the omega-3 lysolipid transporter Mfsd2a leads to aberrant oligodendrocyte lineage development and hypomyelination.

Patients with autosomal recessive microcephaly 15 caused by deficiency in the sodium-dependent lysophosphatidylcholine (LPC) transporter major facilitator superfamily domain-containing 2a (Mfsd2a) present with both microcephaly and hypomyelination, suggesting an important role for LPC uptake by oligodendrocytes in the process of myelination. Here we demonstrate that Mfsd2a is specifically expressed in oligodendrocyte precursor cells (OPCs) and is critical for oligodendrocyte development. Single-cell sequencing of the oligodendrocyte lineage revealed that OPCs from OPC-specific Mfsd2a-KO mice (2aOKO mice) underwent precocious differentiation into immature oligodendrocytes and impaired maturation into myelinating oligodendrocytes, correlating with postnatal brain hypomyelination. 2aOKO mice did not exhibit microcephaly, a finding consistent with the notion that microcephaly is the consequence of an absence of LPC uptake at the blood-brain barrier rather than a deficiency in OPCs. Lipidomic analysis showed that OPCs and iOLs from 2aOKO mice had significantly decreased levels of phospholipids containing omega-3 fatty acids, with a corresponding increase in unsaturated fatty acids, the latter being products of de novo synthesis governed by Srebp-1. RNA-Seq indicated activation of the Srebp-1 pathway and defective expression of regulators of oligodendrocyte development. Taken together, these findings indicate that the transport of LPCs by Mfsd2a in OPCs is important for maintaining OPC state to regulate postnatal brain myelination.

Spns1 is a lysophospholipid transporter mediating lysosomal phospholipid salvage.

The lysosome is central to the degradation of proteins, carbohydrates, and lipids and their salvage back to the cytosol for reutilization. Lysosomal transporters for amino acids, sugars, and cholesterol have been identified, and the metabolic fates of these molecules in the cytoplasm have been elucidated. Remarkably, it is not known whether lysosomal salvage exists for glycerophospholipids, the major constituents of cellular membranes. By using a transport assay screen against orphan lysosomal transporters, we identified the major facilitator superfamily protein Spns1 that is ubiquitously expressed in all tissues as a proton-dependent lysophosphatidylcholine (LPC) and lysophosphatidylethanolamine (LPE) transporter, with LPC and LPE being the lysosomal breakdown products of the most abundant eukaryotic phospholipids, phosphatidylcholine and phosphatidylethanolamine, respectively. Spns1 deficiency in cells, zebrafish embryos, and mouse liver resulted in lysosomal accumulation of LPC and LPE species with pathological consequences on lysosomal function. Flux analysis using stable isotope-labeled phospholipid apolipoprotein E nanodiscs targeted to lysosomes showed that LPC was transported out of lysosomes in an Spns1-dependent manner and re-esterified back into the cytoplasmic pools of phosphatidylcholine. Our findings identify a phospholipid salvage pathway from lysosomes to the cytosol that is dependent on Spns1 and critical for maintaining normal lysosomal function.

The lipid transporter Mfsd2a maintains pulmonary surfactant homeostasis.

Pulmonary surfactant is a lipoprotein complex essential for lung function, and insufficiency or altered surfactant composition is associated with major lung diseases, such as acute respiratory distress syndromes, idiopathic pulmonary fibrosis, and chronic obstructive pulmonary disease. Pulmonary surfactant is primarily composed of phosphatidylcholine (PC) in complex with specialized surfactant proteins and secreted by alveolar type 2 (AT2) cells. Surfactant homeostasis on the alveolar surface is balanced by the rates of synthesis and secretion with reuptake and recycling by AT2 cells, with some degradation by pulmonary macrophages and loss up the bronchial tree. However, whether phospholipid (PL) transporters exist in AT2 cells to mediate reuptake of surfactant PL remains to be identified. Here, we demonstrate that major facilitator superfamily domain containing 2a (Mfsd2a), a sodium-dependent lysophosphatidylcholine (LPC) transporter, is expressed at the apical surface of AT2 cells. A mouse model with inducible AT2 cell-specific deficiency of Mfsd2a exhibited AT2 cell hypertrophy with reduced total surfactant PL levels because of reductions in the most abundant surfactants, PC containing dipalmitic acid, and PC species containing the omega-3 fatty acid docosahexaenoic acid. These changes in surfactant levels and composition were mirrored by similar changes in the AT2 cell lipidome. Mechanistically, direct tracheal instillation of fluorescent LPC and PC probes indicated that Mfsd2a mediates the uptake of LPC generated by pulmonary phospholipase activity in the alveolar space. These studies reveal that Mfsd2a-mediated LPC uptake is quantitatively important in maintaining surfactant homeostasis and identify this lipid transporter as a physiological component of surfactant recycling.

The lysolipid transporter Mfsd2a regulates lipogenesis in the developing brain.

Brain development requires a massive increase in brain lipogenesis and accretion of the essential omega-3 fatty acid docosahexaenoic acid (DHA). Brain acquisition of DHA is primarily mediated by the transporter Major Facilitator Superfamily Domain containing 2a (Mfsd2a) expressed in the endothelium of the blood-brain barrier (BBB) and other abundant cell types within the brain. Mfsd2a transports DHA and other polyunsaturated fatty acids (PUFAs) esterified to lysophosphatidylcholine (LPC-DHA). However, the function of Mfsd2a and DHA in brain development is incompletely understood. Here, we demonstrate, using vascular endothelial-specific and inducible vascular endothelial-specific deletion of Mfsd2a in mice, that Mfsd2a is uniquely required postnatally at the BBB for normal brain growth and DHA accretion, with DHA deficiency preceding the onset of microcephaly. In Mfsd2a-deficient mouse models, a lipidomic signature was identified that is indicative of increased de novo lipogenesis of PUFAs. Gene expression profiling analysis of these DHA-deficient brains indicated that sterol regulatory-element binding protein (Srebp)-1 and Srebp-2 pathways were highly elevated. Mechanistically, LPC-DHA treatment of primary neural stem cells down-regulated Srebp processing and activation in a Mfsd2a-dependent fashion, resulting in profound effects on phospholipid membrane saturation. In addition, Srebp regulated the expression of Mfsd2a. These data identify LPC-DHA transported by Mfsd2a as a physiological regulator of membrane phospholipid saturation acting in a feedback loop on Srebp activity during brain development.

Mfsd2a Is a Transporter for the Essential ω-3 Fatty Acid Docosahexaenoic Acid (DHA) in Eye and Is Important for Photoreceptor Cell Development.

Eye photoreceptor membrane discs in outer rod segments are highly enriched in the visual pigment rhodopsin and the ω-3 fatty acid docosahexaenoic acid (DHA). The eye acquires DHA from blood, but transporters for DHA uptake across the blood-retinal barrier or retinal pigment epithelium have not been identified. Mfsd2a is a newly described sodium-dependent lysophosphatidylcholine (LPC) symporter expressed at the blood-brain barrier that transports LPCs containing DHA and other long-chain fatty acids. LPC transport via Mfsd2a has been shown to be necessary for human brain growth. Here we demonstrate that Mfsd2a is highly expressed in retinal pigment epithelium in embryonic eye, before the development of photoreceptors, and is the primary site of Mfsd2a expression in the eye. Eyes from whole body Mfsd2a-deficient (KO) mice, but not endothelium-specific Mfsd2a-deficient mice, were DHA-deficient and had significantly reduced LPC/DHA transport in vivo Fluorescein angiography indicated normal blood-retinal barrier function. Histological and electron microscopic analysis indicated that Mfsd2a KO mice exhibited a specific reduction in outer rod segment length, disorganized outer rod segment discs, and mislocalization of and reduction in rhodopsin early in postnatal development without loss of photoreceptors. Minor photoreceptor cell loss occurred in adult Mfsd2a KO mice, but electroretinography indicated visual function was normal. The developing eyes of Mfsd2a KO mice had activated microglia and up-regulation of lipogenic and cholesterogenic genes, likely adaptations to loss of LPC transport. These findings identify LPC transport via Mfsd2a as an important pathway for DHA uptake in eye and for development of photoreceptor membrane discs.