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Papers by Sarthak Mohanty

Research paper thumbnail of Gli1 Defines a Subset of Fibro‐adipogenic Progenitors that Promote Skeletal Muscle Regeneration With Less Fat Accumulation

Journal of Bone and Mineral Research, 2021

Skeletal muscle has remarkable regenerative ability after injury. Mesenchymal fibro‐adipogenic pr... more Skeletal muscle has remarkable regenerative ability after injury. Mesenchymal fibro‐adipogenic progenitors (FAPs) are necessary, active participants during this repair process, but the molecular signatures of these cells and their functional relevance remain largely unexplored. Here, using a lineage tracing mouse model (Gli1‐CreER Tomato), we demonstrate that Gli1 marks a small subset of muscle‐resident FAPs with elevated Hedgehog (Hh) signaling. Upon notexin muscle injury, these cells preferentially and rapidly expanded within FAPs. Ablation of Gli1+ cells using a DTA mouse model drastically reduced fibroblastic colony‐forming unit (CFU‐F) colonies generated by muscle cells and impaired muscle repair at 28 days. Pharmacologic manipulation revealed that Gli1+ FAPs rely on Hh signaling to increase the size of regenerating myofiber. Sorted Gli1+ FAPs displayed superior clonogenicity and reduced adipogenic differentiation ability in culture compared to sorted Gli1− FAPs. In a glycerol injury model, Gli1+ FAPs were less likely to give rise to muscle adipocytes compared to other FAPs. Further cell ablation and Hh activator/inhibitor treatments demonstrated their dual actions in enhancing myogenesis and reducing adipogenesis after injury. Examining single‐cell RNA‐sequencing dataset of FAPs from normal mice indicated that Gli1+ FAPs with increased Hh signaling provide trophic signals to myogenic cells while restrict their own adipogenic differentiation. Collectively, our findings identified a subpopulation of FAPs that play an essential role in skeletal muscle repair. © 2021 American Society for Bone and Mineral Research (ASBMR).

Research paper thumbnail of Development of a standardized histopathology scoring system for human intervertebral disc degeneration: an Orthopaedic Research Society Spine Section Initiative

JOR SPINE

Background: Rats are a widely accepted preclinical model for evaluating intervertebral disc (IVD)... more Background: Rats are a widely accepted preclinical model for evaluating intervertebral disc (IVD) degeneration and regeneration. IVD morphology is commonly assessed using histology, which forms the foundation for quantifying the state of IVD degeneration. IVD degeneration severity is evaluated using different grading systems that focus on distinct degenerative features. A standard grading system would facilitate more accurate comparison across laboratories and more robust comparisons of different models and interventions. Aims: This study aimed to develop a histology grading system to quantify IVD degeneration for different rat models.

Research paper thumbnail of Limited Scar Resection for Chronic Achilles Tendon Repair: Use of a Rat Model

The American Journal of Sports Medicine

Background: Achilles tendon rupture diagnosis is frequently missed, leading to the development of... more Background: Achilles tendon rupture diagnosis is frequently missed, leading to the development of a chronic rupture that requires surgical intervention to remove scar tissue and return the elongated Achilles tendon to appropriate functional length. The limited scar resection (LSR) intervention strategy may provide an advantage over other techniques, as it is less invasive and nondestructive to other tissues, although there is little evidence comparing outcomes between intervention strategies. Hypothesis: The LSR technique would be a viable treatment option for chronic Achilles tendon ruptures and would perform comparably with a more clinically accepted procedure, the gastrocnemius fascial turndown (GFT), in postintervention functional outcome measures and tendon mechanical and histological properties. Study Design: Controlled laboratory study. Methods: Chronic Achilles tendon ruptures were induced in the right hindlimb of Sprague-Dawley rats by Achilles tendon transection without re...

Research paper thumbnail of Postoperative Prescription of Low dose Narcotics Yields Equivalent Pain Outcomes Compared to High-dose Narcotics in Opioid-naïve Patients Undergoing Spine Surgery

Research paper thumbnail of An optimized step‐by‐step protocol for isolation of nucleus pulposus, annulus fibrosus, and end plate cells from the mouse intervertebral discs and subsequent preparation of high‐quality intact total RNA

Research paper thumbnail of Author response for "Gli1 defines a subset of fibroadipogenic progenitors that promote skeletal muscle regeneration with less fat accumulation

Research paper thumbnail of Chondrocyte‐like nested cells in the aged intervertebral disc are late‐stage nucleus pulposus cells

Research paper thumbnail of Characterization of Krt19 CreERT allele for targeting the nucleus pulposus cells in the postnatal mouse intervertebral disc

Journal of Cellular Physiology

Intervertebral disc degeneration and associated back pain are relatively common but sparsely unde... more Intervertebral disc degeneration and associated back pain are relatively common but sparsely understood conditions, affecting over 70% of the population during some point of life. Disc degeneration is often associated with a loss of nucleus pulposus (NP) cells. Genetic mouse models offer convenient avenues to understand the cellular and molecular regulation of the disc during its formation, growth, maintenance, and aging. However, due to the lack of inducible driver lines to precisely target NP cells in the postnatal mouse disc, progress in this area of research has been moderate. NP cells are known to express cytokeratin 19 (Krt19), and tamoxifen (Tam)-inducible Krt19 CreERT allele is available. The current study describes the characterization of Krt19 CreERT allele to specifically and efficiently target NP cells in neonatal, skeletally mature, middle-aged, and aged mice using two independent fluorescent reporter lines. The efficiency of recombination at all ages was validated by immunostaining for KRT19. Results show that following Tam induction, Krt19 CreERT specifically drives recombination of NP cells in the spine of neonatal and aged mice, while no recombination was detected in the surrounding tissues. Knee joints from skeletally mature Tam-treated Krt19 CreERT/+ ; R26 tdTOM mouse show the absence of recombination in all tissues and cells of the knee joint. Thus, this study provides evidence for the use of Krt19 CreERT allele for genetic characterization of NP cells at different stages of the mouse life.

Research paper thumbnail of Defects in intervertebral disc and spine during development, degeneration, and pain: New research directions for disc regeneration and therapy

Wiley Interdisciplinary Reviews: Developmental Biology

Research paper thumbnail of Aging of mouse intervertebral disc and association with back pain

Research paper thumbnail of R-spondin-2 is a Wnt agonist that regulates osteoblast activity and bone mass

Bone Research

The R-spondin family of proteins are Wnt agonists, and the complete embryonic disruption of Rspo2... more The R-spondin family of proteins are Wnt agonists, and the complete embryonic disruption of Rspo2 results in skeletal developmental defects that recapitulate the phenotype observed with Lrp5/6 deficiency. Previous work has shown that R-spondin-2 (Rspo2, RSPO2) is both highly expressed in Wnt-stimulated pre-osteoblasts and its overexpression induces osteoblast differentiation in the same cells, supporting its putative role as a positive autocrine regulator of osteoblastogenesis. However, the role of Rspo2 in regulating osteoblastogenesis and bone formation in postnatal bone has not been explored. Here we show that limb-bud progenitor cells from Rspo2 knockout mice undergo reduced mineralization during osteoblastogenesis in vitro and have a corresponding alteration in their osteogenic gene expression profile. We also generated the first Rspo2 conditional knockout (Rspo2 floxed) mouse and disrupted Rspo2 expression in osteoblast-lineage cells by crossing to the Osteocalcin-Cre mouse line (Ocn-Cre + Rspo2 f/f). Ocn-Cre + Rspo2 f/f male and female mice at 1, 3, and 6 months were examined. Ocn-Cre + Rspo2 f/f mice are decreased in overall body size compared to their control littermates and have decreased bone mass. Histomorphometric analysis of 1-month-old mice revealed a similar number of osteoblasts and mineralizing surface per bone surface with a simultaneous decrease in mineral apposition and bone formation rates. Consistent with this observation, serum osteocalcin in 3-month-old Ocn-Cre + Rspo2 f/f was reduced, and bone marrow-mesenchymal stem cells from Ocn-Cre + Rspo2 f/f mice undergo less mineralization in vitro. Finally, gene expression analysis and immunohistochemistry of mature bone shows reduced beta-catenin signaling in Ocn-Cre + Rspo2 f/f. Overall, RSPO2 reduces osteoblastogenesis and mineralization, leading to reduced bone mass.

Research paper thumbnail of P78. Socioeconomic and community disparities in presenting physical functionality and mental distress characteristics of patients undergoing spine surgery

Research paper thumbnail of Gli1 Defines a Subset of Fibro‐adipogenic Progenitors that Promote Skeletal Muscle Regeneration With Less Fat Accumulation

Journal of Bone and Mineral Research, 2021

Skeletal muscle has remarkable regenerative ability after injury. Mesenchymal fibro‐adipogenic pr... more Skeletal muscle has remarkable regenerative ability after injury. Mesenchymal fibro‐adipogenic progenitors (FAPs) are necessary, active participants during this repair process, but the molecular signatures of these cells and their functional relevance remain largely unexplored. Here, using a lineage tracing mouse model (Gli1‐CreER Tomato), we demonstrate that Gli1 marks a small subset of muscle‐resident FAPs with elevated Hedgehog (Hh) signaling. Upon notexin muscle injury, these cells preferentially and rapidly expanded within FAPs. Ablation of Gli1+ cells using a DTA mouse model drastically reduced fibroblastic colony‐forming unit (CFU‐F) colonies generated by muscle cells and impaired muscle repair at 28 days. Pharmacologic manipulation revealed that Gli1+ FAPs rely on Hh signaling to increase the size of regenerating myofiber. Sorted Gli1+ FAPs displayed superior clonogenicity and reduced adipogenic differentiation ability in culture compared to sorted Gli1− FAPs. In a glycerol injury model, Gli1+ FAPs were less likely to give rise to muscle adipocytes compared to other FAPs. Further cell ablation and Hh activator/inhibitor treatments demonstrated their dual actions in enhancing myogenesis and reducing adipogenesis after injury. Examining single‐cell RNA‐sequencing dataset of FAPs from normal mice indicated that Gli1+ FAPs with increased Hh signaling provide trophic signals to myogenic cells while restrict their own adipogenic differentiation. Collectively, our findings identified a subpopulation of FAPs that play an essential role in skeletal muscle repair. © 2021 American Society for Bone and Mineral Research (ASBMR).

Research paper thumbnail of Development of a standardized histopathology scoring system for human intervertebral disc degeneration: an Orthopaedic Research Society Spine Section Initiative

JOR SPINE

Background: Rats are a widely accepted preclinical model for evaluating intervertebral disc (IVD)... more Background: Rats are a widely accepted preclinical model for evaluating intervertebral disc (IVD) degeneration and regeneration. IVD morphology is commonly assessed using histology, which forms the foundation for quantifying the state of IVD degeneration. IVD degeneration severity is evaluated using different grading systems that focus on distinct degenerative features. A standard grading system would facilitate more accurate comparison across laboratories and more robust comparisons of different models and interventions. Aims: This study aimed to develop a histology grading system to quantify IVD degeneration for different rat models.

Research paper thumbnail of Limited Scar Resection for Chronic Achilles Tendon Repair: Use of a Rat Model

The American Journal of Sports Medicine

Background: Achilles tendon rupture diagnosis is frequently missed, leading to the development of... more Background: Achilles tendon rupture diagnosis is frequently missed, leading to the development of a chronic rupture that requires surgical intervention to remove scar tissue and return the elongated Achilles tendon to appropriate functional length. The limited scar resection (LSR) intervention strategy may provide an advantage over other techniques, as it is less invasive and nondestructive to other tissues, although there is little evidence comparing outcomes between intervention strategies. Hypothesis: The LSR technique would be a viable treatment option for chronic Achilles tendon ruptures and would perform comparably with a more clinically accepted procedure, the gastrocnemius fascial turndown (GFT), in postintervention functional outcome measures and tendon mechanical and histological properties. Study Design: Controlled laboratory study. Methods: Chronic Achilles tendon ruptures were induced in the right hindlimb of Sprague-Dawley rats by Achilles tendon transection without re...

Research paper thumbnail of Postoperative Prescription of Low dose Narcotics Yields Equivalent Pain Outcomes Compared to High-dose Narcotics in Opioid-naïve Patients Undergoing Spine Surgery

Research paper thumbnail of An optimized step‐by‐step protocol for isolation of nucleus pulposus, annulus fibrosus, and end plate cells from the mouse intervertebral discs and subsequent preparation of high‐quality intact total RNA

Research paper thumbnail of Author response for "Gli1 defines a subset of fibroadipogenic progenitors that promote skeletal muscle regeneration with less fat accumulation

Research paper thumbnail of Chondrocyte‐like nested cells in the aged intervertebral disc are late‐stage nucleus pulposus cells

Research paper thumbnail of Characterization of Krt19 CreERT allele for targeting the nucleus pulposus cells in the postnatal mouse intervertebral disc

Journal of Cellular Physiology

Intervertebral disc degeneration and associated back pain are relatively common but sparsely unde... more Intervertebral disc degeneration and associated back pain are relatively common but sparsely understood conditions, affecting over 70% of the population during some point of life. Disc degeneration is often associated with a loss of nucleus pulposus (NP) cells. Genetic mouse models offer convenient avenues to understand the cellular and molecular regulation of the disc during its formation, growth, maintenance, and aging. However, due to the lack of inducible driver lines to precisely target NP cells in the postnatal mouse disc, progress in this area of research has been moderate. NP cells are known to express cytokeratin 19 (Krt19), and tamoxifen (Tam)-inducible Krt19 CreERT allele is available. The current study describes the characterization of Krt19 CreERT allele to specifically and efficiently target NP cells in neonatal, skeletally mature, middle-aged, and aged mice using two independent fluorescent reporter lines. The efficiency of recombination at all ages was validated by immunostaining for KRT19. Results show that following Tam induction, Krt19 CreERT specifically drives recombination of NP cells in the spine of neonatal and aged mice, while no recombination was detected in the surrounding tissues. Knee joints from skeletally mature Tam-treated Krt19 CreERT/+ ; R26 tdTOM mouse show the absence of recombination in all tissues and cells of the knee joint. Thus, this study provides evidence for the use of Krt19 CreERT allele for genetic characterization of NP cells at different stages of the mouse life.

Research paper thumbnail of Defects in intervertebral disc and spine during development, degeneration, and pain: New research directions for disc regeneration and therapy

Wiley Interdisciplinary Reviews: Developmental Biology

Research paper thumbnail of Aging of mouse intervertebral disc and association with back pain

Research paper thumbnail of R-spondin-2 is a Wnt agonist that regulates osteoblast activity and bone mass

Bone Research

The R-spondin family of proteins are Wnt agonists, and the complete embryonic disruption of Rspo2... more The R-spondin family of proteins are Wnt agonists, and the complete embryonic disruption of Rspo2 results in skeletal developmental defects that recapitulate the phenotype observed with Lrp5/6 deficiency. Previous work has shown that R-spondin-2 (Rspo2, RSPO2) is both highly expressed in Wnt-stimulated pre-osteoblasts and its overexpression induces osteoblast differentiation in the same cells, supporting its putative role as a positive autocrine regulator of osteoblastogenesis. However, the role of Rspo2 in regulating osteoblastogenesis and bone formation in postnatal bone has not been explored. Here we show that limb-bud progenitor cells from Rspo2 knockout mice undergo reduced mineralization during osteoblastogenesis in vitro and have a corresponding alteration in their osteogenic gene expression profile. We also generated the first Rspo2 conditional knockout (Rspo2 floxed) mouse and disrupted Rspo2 expression in osteoblast-lineage cells by crossing to the Osteocalcin-Cre mouse line (Ocn-Cre + Rspo2 f/f). Ocn-Cre + Rspo2 f/f male and female mice at 1, 3, and 6 months were examined. Ocn-Cre + Rspo2 f/f mice are decreased in overall body size compared to their control littermates and have decreased bone mass. Histomorphometric analysis of 1-month-old mice revealed a similar number of osteoblasts and mineralizing surface per bone surface with a simultaneous decrease in mineral apposition and bone formation rates. Consistent with this observation, serum osteocalcin in 3-month-old Ocn-Cre + Rspo2 f/f was reduced, and bone marrow-mesenchymal stem cells from Ocn-Cre + Rspo2 f/f mice undergo less mineralization in vitro. Finally, gene expression analysis and immunohistochemistry of mature bone shows reduced beta-catenin signaling in Ocn-Cre + Rspo2 f/f. Overall, RSPO2 reduces osteoblastogenesis and mineralization, leading to reduced bone mass.

Research paper thumbnail of P78. Socioeconomic and community disparities in presenting physical functionality and mental distress characteristics of patients undergoing spine surgery