Rhian Stavely

Multisystemic smooth muscle dysfunction syndrome (MSMDS) is a rare disorder caused by ACTA2 mutations, including the R179H variant, which alters actin filament stability and dynamics and smooth muscle contractility. Cardiovascular complications dominate its clinical presentation, but gastrointestinal (GI) dysfunction significantly affects quality of life. To investigate the structural, functional, and cellular basis of gut dysmotility in MSMDS, we reviewed clinical data from 24 patients with MSMDS and studied the ACTA2 R179H mouse model. Patients exhibited severe gut dysmotility, with 75% requiring medication for chronic constipation. ACTA2 mutant mice displayed cecal and colonic dilatation, reduced intestinal length, and disrupted colonic migrating motor complexes. Delayed whole-gut transit and impaired contractile responses to electrical and pharmacological stimulation were observed. Transcriptomic analysis revealed significant actin cytoskeleton-related gene changes in smooth muscle cells, and immune profiling identified increased lymphocytic infiltration. Despite functional abnormalities, there were no obvious changes in the enteric nervous system. These findings establish ACTA2 mice as a robust model for studying GI pathology in MSMDS, elucidating the role of smooth muscle dysfunction in gut dysmotility. This model provides a foundation for developing targeted therapies aimed at restoring intestinal motility by directly addressing actin cytoskeletal disruptions in smooth muscle cells.

Peripheral nerve injuries are associated with significant morbidity, particularly when primary surgical repair is delayed or impossible due to extensive nerve gaps. While advances in biomedical engineering have led to commercially available nerve guidance conduits tailored for such injuries, rates of sensory and motor recovery remain suboptimal following neurotmesis with gap defects beyond 3 cm. Cell therapy represents a promising treatment strategy to heal the injured peripheral nervous system, thought to promote tissue regeneration and enhance endogenous mechanisms of nerve repair to restore functionality. In this study, we explore the potential utility and efficacy of subcutaneous adipose tissue-derived neural stem cells in a nerve transection injury model. Plp1-EGFP mice, which express GFP in Schwann cells, underwent surgical excision of a 5 mm segment of the left sciatic nerve. Nerves were then immediately repaired using silicone nerve guidance conduits with a residual 5 mm defect between nerve stumps. Conduits were loaded with cell culture media alone or with subcutaneous adipose tissue-derived neural stem cells harvested from Wnt1-tdTomato neural crest reporter mice, the latter enabling cell tracing post-transplantation. Subcutaneous adipose tissue-derived neural stem cells persisted through postoperative day 56 and contributed structurally to the reformed sciatic nerve. Integration between Wnt1-tdTomato neural stem cells and endogenous Plp1-EGFP Schwann cells occurred at the distal and proximal transection margins. Furthermore, neural stem cells predominantly differentiated into Schwann-like cells following transplantation, aiding in myelination of the reformed nerve, but not undesirable cell types such as neurons. Gait testing indicated that adipose-derived neural stem cells significantly improved hindlimb motor recovery compared to conduit repair alone by postoperative day 56. Using cell tracer models, we confirm that adipose-derived neural stem cells can be therapeutically delivered to injured peripheral nerves, integrate with recipient axons and Schwann cells, and differentiate into myelinating Schwann-like cells to enhance motor recovery. These findings indicate that subcutaneous adipose tissue-derived neural stem cells could fill a critical gap in the treatment of peripheral nerve injuries, representing a readily available, autologous source of regenerative cells to optimize functional recovery after injury.

Anti-Müllerian hormone is a factor secreted by granulosa cells of growing follicles that regulates many aspects of ovarian function, including primordial follicle activation and early follicle development through inhibitory feedback. Treatment with exogenous anti-Müllerian hormone at supraphysiological levels can inhibit follicular development and prevent ovulation in mice, rats, and cats. Single-cell transcriptomic analysis of ovaries from mice treated postnatally with anti-Müllerian hormone identified a distinct quiescence signature in granulosa cells. Herein, we further characterized the anti-Müllerian hormone -induced quiescent cell state and drew parallels to developmental senescence by describing a reversible “quiescence-associated secretory phenotype.” This state was characterized by the induction of several hallmarks of senescence, including inhibition of proliferation (reduced KI67), upregulation of markers of senescence (Cdkn1a, Fn1, Cebpb, Timp3), and chemokines and their receptors (Cxcl14, Cxcl12, Ccl12, Ccl21, Cxcr2). However, quiescence-associated secretory phenotype did not recapitulate other senescence hallmarks, such as activation of beta-galactosidase activity or permanent exit from the cell cycle. Similar to classical senescence, we found that anti-Müllerian hormone induced quiescence-associated secretory phenotype, in part, through the upregulation of cyclin-dependent kinase inhibitors (Cdkn1a, Cdkn1b, and Cdkn1c) in granulosa cells and oocytes of mouse and human ovaries. Finally, we showed that the quiescence-associated secretory phenotype state could be reversed within a month of discontinuing anti-Müllerian hormone treatment. This recovery was marked by the return of growing primary, secondary, and antral follicles. In conclusion, these data suggest that anti-Müllerian hormone can pause follicle development by inducing quiescence-associated secretory phenotype, a reversible senescent-like state, which may benefit fertility preservation.

Involvement of the muscularis propria accompanies a poorer prognosis in IBD. This study characterizes muscularis macrophage subpopulations during colitis, highlighting the loss of anti-inflammatory LYVE-1⁺ macrophages and inflammatory plasticity in resident CX3CR1⁺ macrophages, providing insights into prognostic and therapeutic targets.

The goal of this study was to determine if transplantation of enteric neural stem cells (ENSCs) can rescue the enteric nervous system, restore gut motility, reduce colonic inflammation, and improve survival in the Ednrb-KO mouse model of Hirschsprung disease (HSCR). ENSCs were isolated from mouse intestine, expanded to form neurospheres, and microinjected into the colons of recipient Ednrb-KO mice. Transplanted ENSCs were identified in recipient colons as cell clusters in “neo-ganglia.” Immunohistochemical evaluation demonstrated extensive cell migration away from the sites of cell delivery and across the muscle layers. Electrical field stimulation and optogenetics showed significantly enhanced contractile activity of aganglionic colonic smooth muscle following ENSC transplantation and confirmed functional neuromuscular integration of the transplanted ENSC-derived neurons. ENSC injection also partially restored the colonic migrating motor complex. Histological examination revealed a significant reduction in inflammation in ENSC-transplanted aganglionic recipient colon compared with that of sham-operated mice. Interestingly, mice that received cell transplant also had prolonged survival compared with controls. This study demonstrates that ENSC transplantation can improve outcomes in HSCR by restoring gut motility and reducing the severity of Hirschsprung-associated enterocolitis, the leading cause of death in human HSCR.

Oxidative stress is increasingly recognized as a central player in a range of gastrointestinal (GI) disorders, as well as complications stemming from therapeutic interventions. This article presents an overview of the mechanisms of oxidative stress in GI conditions and highlights a link between oxidative insult and disruption to the enteric nervous system (ENS), which controls GI functions. The dysfunction of the ENS is characteristic of a spectrum of disorders, including neurointestinal diseases and conditions such as inflammatory bowel disease (IBD), diabetic gastroparesis, and chemotherapy-induced GI side effects. Neurons in the ENS, while essential for normal gut function, appear particularly vulnerable to oxidative damage. Mechanistically, oxidative stress in enteric neurons can result from intrinsic nitrosative injury, mitochondrial dysfunction, or inflammation-related pathways. Although antioxidant-based therapies have shown limited efficacy, recognizing the multifaceted role of oxidative stress in GI diseases offers a promising avenue for future interventions. This comprehensive review summarizes the literature to date implicating oxidative stress as a critical player in the pathophysiology of GI disorders, with a focus on its role in ENS injury and dysfunction, and highlights opportunities for the development of targeted therapeutics for these diseases.

The enteric nervous system (ENS) is principally derived from vagal neural crest cells that migrate caudally along the entire length of the gastrointestinal tract, giving rise to neurons and glial cells in two ganglionated plexuses. Incomplete migration of enteric neural crest-derived cells (ENCDC) leads to Hirschsprung disease, a congenital disorder characterized by the absence of enteric ganglia along variable lengths of the colorectum. Our previous work strongly supported the essential role of the avian ceca, present at the junction of the midgut and hindgut, in hindgut ENS development, since ablation of the cecal buds led to incomplete ENCDC colonization of the hindgut. In situ hybridization shows bone morphogenetic protein-4 (BMP4) is highly expressed in the cecal mesenchyme, leading us to hypothesize that cecal BMP4 is required for hindgut ENS development. To test this, we modulated BMP4 activity using embryonic intestinal organ culture techniques and retroviral infection. We show that overexpression or inhibition of BMP4 in the ceca disrupts hindgut ENS development, with GDNF playing an important regulatory role. Our results suggest that these two important signaling pathways are required for normal ENCDC migration and enteric ganglion formation in the developing hindgut ENS.