Abstract
In cultured human fibroblasts, SNAT transporters (System A) account for the accumulation of non-essential neutral amino acids, are adaptively up-regulated upon amino acid deprivation and play a major role in cell volume recovery upon hypertonic stress. No information is instead available on the expression and activity of SNAT transporters in human bone marrow mesenchymal stromal cells (MSC), although they are increasingly investigated for their staminal and immunomodulatory properties and used for several therapeutic applications. The uptake of glutamine and proline, two substrates of SNAT1 and SNAT2 transporters, was measured in primary human MSC and an MSC line. The amino acid analogue MeAIB, a specific substrate of these carriers, has been used to selectively inhibit SNAT-dependent transport of glutamine and, through its sodium-dependent transport, as an indicator of SNAT1/2 activity. SNAT1/2 expression and localization were assessed with RT-PCR and confocal microscopy, respectively. Cell volume was assessed from urea distribution space. In all these experiments, primary human fibroblasts were used as the positive control for SNAT expression and activity. Compared with fibroblasts, MSC have a lower SNAT1 expression and hardly detectable membrane localization of both SNAT1 and SNAT2. Moreover, they exhibit no sodium-dependent MeAIB uptake or MeAIB-inhibitable glutamine transport, and exhibit a lower ability to accumulate glutamine and proline than fibroblasts. MSC exhibited an only marginal increase in MeAIB transport upon amino acid starvation and did not recover cell volume after hypertonic stress. In conclusion, the activity of SNAT transporters is low in human MSC. MSC adaptation to amino acid shortage is expected to rely on intracellular synthesis, given the absence of an effective up-regulation of the SNAT transporters.
Highlights
Bone marrow mesenchymal stem cells (MSC) are self-renewable, multipotent cells capable of multi-lineage differentiation into adipocytes, chondrocytes or osteoblasts [1], and, widely used for different applications in regenerative medicine [2,3]
We shown that, compared with human fibroblasts, when cultured in standard growth medium, human bone marrow-derived MSC are endowed with lower activity of System A transporters, which is associated with a reduced Gln cell content and impaired ability to recover cell volume upon hypertonic stress
Sci. 2020, 21, 1899 was lower in either primary MSC or in hTERT-MSC than in IMR-90 fibroblasts (Figure 1b). This result is consistent with the absence of System A activity and suggest that in MSC Gln uptake mainly occurs through the activity of an ASCT2 exchanger transporter
Summary
Bone marrow mesenchymal stem cells (MSC) are self-renewable, multipotent cells capable of multi-lineage differentiation into adipocytes, chondrocytes or osteoblasts [1], and, widely used for different applications in regenerative medicine [2,3]. MSC secrete a variety of soluble mediators that influence various types of immune cells and have distinctive immunomodulatory properties that widen the range of their potential therapeutic applications [4,5]. Metabolism is considered an important determinant of MSC properties, their metabolic features are yet to be completely defined [6]. Metabolic features may be of particular relevance for the nutritional support that MSC can provide to other cells of the microenvironment. In the case of Acute Lymphoblastic Leukemia (ALL), MSC produce and secrete asparagine (Asn) to support the growth of leukemic blasts, which are auxotroph for Asn [7,8]. The nutritional support of ALL blasts by MSC is only possible if Gln availability is adequate
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