Biology subjects

Risbud, M. V.

Publications and source records attributed to Risbud, M. V..

5 recordsLinked to original sources

bioRxiv2024

OPA1 protects intervertebral disc and knee joint health in aged mice by maintaining the structure and metabolic functions of mitochondria

Due to their glycolytic nature and limited vascularity, nucleus pulposus (NP) cells of the intervertebral disc and articular chondrocytes were long thought to have minimal reliance on mitochondrial function. Recent studies have challenged this long-held view and highlighted the increasingly important role of mitochondria in the physiology of these tissues. We investigated the role of mitochondrial fusion protein OPA1 in maintaining the spine and knee joint health in aging mice. OPA1 knockdown in NP cells altered mitochondrial size and cristae shape and increased the oxygen consumption rate without affecting ATP synthesis. OPA1 governed the morphology of multiple organelles, and its loss resulted in the dysregulation of NP cell autophagy. Metabolic profiling and 13C-flux analyses revealed TCA cycle anaplerosis and altered metabolism in OPA1-deficient NP cells. Noteworthy, Opa1AcanCreERT2 mice showed age- dependent disc, and cartilage degeneration and vertebral osteopenia. Our findings suggest that OPA1 regulation of mitochondrial dynamics and multi-organelle interactions is critical in preserving metabolic homeostasis of disc and cartilage. TeaserOPA1 is necessary for the maintenance of intervertebral disc and knee joint health in aging mice

cell biology↗

bioRxiv2023

Increased HIF-2α Activity in the Nucleus Pulposus Causes Intervertebral Disc Degeneration in the Aging Mouse Spine

Hypoxia-inducible factors (HIFs) are essential to the homeostasis of hypoxic tissues. Although HIF-2, is expressed in nucleus pulposus (NP) cells, consequences of elevated HIF-2 activity on disc health remains unknown. We expressed HIF-2 with proline to alanine substitutions (P405A;P531A) in the Oxygen-dependent degradation domain (HIF-2dPA) in the NP tissue using an inducible, nucleus pulposus-specific K19CreERT allele to study HIF-2 function in the adult intervertebral disc. Expression of HIF-2 in NP impacted disc morphology, as evident from small but significantly higher scores of degeneration in NP of 24-month-old K19CreERT; HIF-2dPA (K19-dPA) mice. Noteworthy, comparisons of grades within each genotype between 14 months and 24 months indicated that HIF-2 overexpression contributed to more pronounced changes than aging alone. The annulus fibrosus (AF) compartment in the 14-month-old K19-dPA mice exhibited lower collagen turnover and Fourier transform-infrared (FTIR) spectroscopic imaging analyses showed changes in the biochemical composition of the 14-and 24-month-old K19-dPA mice. Moreover, there were changes in aggrecan, chondroitin sulfate, and COMP abundance without alterations in NP phenotypic marker CA3, suggesting the overexpression of HIF-2 had some impact on matrix composition but not the cell phenotype. Mechanistically, the global transcriptomic analysis showed enrichment of differentially expressed genes in themes closely related to NP cell function such as cilia, SLIT/ROBO pathway, and HIF/Hypoxia signaling at both 14- and 24-months. Together, these findings underscore the role of HIF-2 in the pathogenesis of disc degeneration in the aged spine.

cell biology↗

bioRxiv2023

SDC4 deletion perturbs intervertebral disc matrix homeostasis and promotes early osteopenia in the aging spine

Syndecan 4 (SDC4), a cell surface heparan sulfate proteoglycan, is known to regulate matrix catabolism by nucleus pulposus cells in an inflammatory milieu. However, the role of SDC4 in the aging spine has never been explored. Here we analyzed the spinal phenotype of SDC4 global knockout (KO) mice as a function of age. Micro-computed tomography showed that SDC4 deletion severely reduced vertebral trabecular and cortical bone mass, and biomechanical properties of vertebrae were significantly altered in SDC4 KO mice. These changes in vertebral bone were due to elevated osteoclastic activity in KO mice. The histological assessment also showed subtle phenotypic changes in the intervertebral discs. Imaging-Fourier transform-infrared (FTIR) analyses showed a reduced relative ratio of mature collagen crosslink in young adult NP and AF compartments of KO compared to wildtype (WT) mice. Additionally, relative chondroitin sulfated glycosaminoglycan (GAG) levels increased in the NP compartment of the KO mice. Transcriptomic analysis of NP tissue using CompBio, an AI-based tool showed biological themes associated with prominent dysregulation of heparan sulfate GAG degradation, mitochondria metabolism, autophagy, and endoplasmic reticulum to Golgi protein processing. Overall, this study highlights the important role of SDC4 in fine-tuning vertebral bone homeostasis and extracellular matrix homeostasis in the intervertebral disc.

cell biology↗

bioRxiv2022

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 importer, GLUT1, is a highly enriched phenotypic marker of NP cells, we hypothesized that it is vital for the development and post-natal maintenance of the disc. Surprisingly, primary NP cells treated with two well-characterized GLUT1 inhibitors maintained normal rates of glycolysis and ATP production, indicating intrinsic compensatory mechanisms. We show in vitro that NP cells mitigate GLUT1 loss by rewiring glucose import through GLUT3. Noteworthy, we demonstrate that substrates, such as glutamine and palmitate, do not compensate for glucose restriction resulting from dual inhibition of GLUT1/3 and inhibition compromises long-term cell viability. To investigate the redundancy of GLUT1 function in NP, we generated two NP-specific knockout mice: Krt19CreERT; Glut1f/f and Foxa2Cre; Glut1f/f. Noteworthy, there were no apparent defects in post-natal disc health or development and maturation in mutant mice. Microarray analysis confirmed 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 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.

cell biology↗

bioRxiv2022

The cGAS-STING pathway affects vertebral bone but does not promote intervertebral disc cell senescence or degeneration

The DNA-sensing cGAS-STING pathway promotes the senescence-associated secretory phenotype (SASP) and mediates type-I interferon inflammatory responses to foreign viral and bacterial DNA as well as self-DNA. Studies of the intervertebral disc in humans and mice demonstrate associations between aging, increased cell senescence, and disc degeneration. Herein we assessed the role of STING in SASP promotion in STING gain- (N153S) and loss-of-function mouse models. N153S mice evidenced elevated circulating levels of proinflammatory markers including IL-1{beta}, IL-6, and TNF- and exhibited a mild trabecular and cortical bone phenotype in caudal vertebrae. Interestingly, despite systemic inflammation, the structural integrity of the disc and knee articular joint remained intact, and cells did not show a loss of their phenotype or elevated SASP. Transcriptomic analysis of N153S tissues demonstrated an upregulated immune response by disc cells, which did not closely resemble inflammatory changes in human tissues. Interestingly, STING-/- mice also showed a mild vertebral bone phenotype, but the absence of STING did not improve the age-associated disc phenotype or reduce the abundance of SASP markers. Overall, the analyses of N153S and STING-/- mice that the cGAS-STING pathway is not a major contributor to SASP induction and consequent disc aging and degeneration but may play a minor role in the maintenance of trabecular bone in the vertebrae. This work contributes to a growing body of work demonstrating that systemic inflammation is not a key driver of disc degeneration.

cell biology↗