GLUT1 Is Redundant in Hypoxic and Glycolytic Nucleus Pulposus Cells of the Intervertebral Disc

Glycolysis is central to homeostasis of nucleus pulposus (NP) cells in the avascular intervertebral disc. Since the glucose transporter, GLUT1, is a highly enriched phenotypic marker of NP cells, we hypothesized that it is vital for the development and postnatal maintenance of the disc. Surprisingly, primary NP cells treated with 2 well-characterized GLUT1 inhibitors maintained normal rates of glycolysis and ATP production, indicating intrinsic compensatory mechanisms. We showed in vitro that NP cells mitigated GLUT1 loss by rewiring glucose import through GLUT3. Of note, we demonstrated that substrates, such as glutamine and palmitate, did not compensate for glucose restriction resulting from dual inhibition of GLUT1/3, and inhibition compromised long-term cell viability. To investigate the redundancy of GLUT1 function in NP, we generated 2 NP-specific knockout mice: Krt19CreERT Glut1fl/fl and Foxa2Cre Glut1fl/fl. There were no apparent defects in postnatal disc health or development and maturation in mutant mice. Microarray analysis verified that GLUT1 loss did not cause transcriptomic alterations in the NP, supporting that cells are refractory to GLUT1 loss. These observations provide the first evidence to our knowledge of functional redundancy in GLUT transporters in the physiologically hypoxic intervertebral disc and underscore the importance of glucose as the indispensable substrate for NP cells

The Role of HIF-2α in the Intervertebral Disc

Low back pain, and associated intervertebral disc degeneration, is the leading cause of disability and has a substantial economic burden. Identifying links between disc degeneration and low back pain is essential. However, understanding basic intervertebral disc biology is critical to this endeavor. The intervertebral disc is a unique, avascular, and thus physiologically hypoxic organ comprising three major compartments. The innermost nucleus pulposus (NP) is encompassed by the annulus fibrosus and capped superiorly and inferiorly by cartilaginous end plates. Cells within this hypoxic niche have adapted to survive through the unique regulation of genes controlled by hypoxia inducible factors (HIFs). Our prior work has found that hypoxia inducible factor 1α (HIF-1α) expression is required for the development and maintenance of the NP and overall disc structure. Although the function of HIF-1α in the disc has been extensively explored, neither the role of HIF-2α (a homologue of HIF-1α) nor its transcriptome is thoroughly investigated in the disc context. Additionally, glucose transporter 1 (GLUT1/Slc2a1), a target of HIF-1α and a phenotypic marker of NP cells, is the main glucose transporter critical to the function of cartilage and bone. Even though GLUT1 is highly enriched in the NP, we do not know the importance of GLUT1 in the adult intervertebral disc. Accordingly, this work had two main aims: (1) to examine and elucidate the role of HIF-2α in NP cells, and (2) to evaluate the importance and the role of GLUT1 in the adult disc. Using an NP-specific HIF-2α conditional knockout mouse model, we show that HIF-2α is not necessary for NP compartment development, and rather than compromising NP compartment, like loss of HIF-1α, loss of HIF-2α provides transient protection against age-related disc generation. These are novel observations that support the notion that HIF-2α may be independent of the functions of HIF-1α. Furthermore, using an NP-specific overexpression HIF-2α mouse model, we show a very mild but significant increase in the NP grades of degeneration in 24-month-old HIF-2α overexpressing mice. Microarray analysis from the NP of HIF-2α overexpressing mice suggests HIF-2α to be important for regulating genes and associated with CompBio themes linked to cilia. Both conditional knockout and overexpression NP-specific HIF-2α mouse models explored here provide strong evidence for the differences in roles of HIF-2α vs. HIF-1α in the disc. Furthermore, in contrast to many cell types, we show using an adult NP-specific conditional knockout mouse model of GLUT1 that loss of GLUT1 does not impact overall disc morphology nor alter NP transcriptomic profile. We observed that the maintenance of mRNA and protein levels of alternative glucose transporters (i.e., GLUT3) in the NP of GLUT1 knockout mice shows that these alternative glucose transporters can provide NP with sufficient glucose concentrations. Overall, the studies presented here add insights into the biological role of HIF-2α, confirms the redundancy of glucose transporters, and reaffirm the importance of glucose utilization in NP cells. These results contribute to a new understanding of the specific role of HIF-2α and GLUT1 in NP cells, which help advance our understanding of the unique biological functions of the intervertebral disc

Long-term treatment with senolytic drugs Dasatinib and Quercetin ameliorates age-dependent intervertebral disc degeneration in mice

Intervertebral disc degeneration is a leading cause of chronic back pain and disability. Here the authors show that long term treatment with senolytic compounds Dasatinib and Quercetin reduces disc senescence burden and ameliorates age-dependent degeneration in mice