Biology subjects

Duran, P.

Publications and source records attributed to Duran, P..

At least 19 recordsLinked to original sources

bioRxiv2026

Endocytosis and Compartmentalized Intracellular Signaling of the Prostaglandin Receptor EP4 Mediate Pain

Prostaglandin E2 (PGE2) is recognized as a major mediator of inflammatory pain. However, the intracellular signaling mechanisms by which PGE2 mediates pain remain unclear. Here, we show that pain-like responses evoked by PGE2 in dorsal root ganglion (DRG) nociceptors are mediated by internalization and compartmentalized signaling of the EP4 receptor. The EP4-selective agonist L-902,688 induced immediate nocifensive behavior and prolonged mechanical allodynia in mice and sensitized isolated DRG nociceptors. Pharmacological inhibition of clathrin- and dynamin-mediated endocytosis (using pitstop2 and dyngo4a) and siRNA knockdown of Dnm1 prevented EP4-induced nociception and sensitization, implicating receptor trafficking in pain signaling. Using genetically-encoded biosensors, we monitored EP4 trafficking and downstream signaling within subcellular compartments of HEK293 cells. PGE2 stimulated dynamin- and {beta}-arrestin-dependent EP4 trafficking from the plasma membrane to early, late and recycling endosomes and Golgi apparatus, and mobilized intracellular EP4 pools from the endoplasmic reticulum. PGE2 induced the assembly of EP4, G proteins and {beta}-arrestin signaling complexes in endosomes and the Golgi apparatus. Inhibition of EP4 endocytosis suppressed intracellular cAMP production and ERK activation. Our findings reveal that EP4-mediated nociceptive signaling originates from intracellular compartments and suggest that modulating EP4 internalization and subcellular signaling dynamics could offer a novel strategy for inflammatory pain management.

cell biology↗

bioRxiv2026

A mouse model of myotonic dystrophy type 1 exhibits pain-like behavior and peripheral nociceptor hyperexcitability

Pain is a prevalent and disabling symptom of myotonic dystrophy type 1 (DM1), yet its underlying mechanisms remain poorly understood. Using HSA LR20b transgenic mice, we found multimodal mechanical and thermal hypersensitivity. Whole-cell electrophysiological recordings demonstrated depolarized resting membrane potentials and increased action potential firing selectively in small- and medium-diameter dorsal root ganglion neurons. These findings identify peripheral nociceptor sensitization as a potential mechanism contributing to pain in DM1.

neuroscience↗

bioRxiv2026

Environmental factors and microbe-microbe interactions drive the structure of the core microbiota of terrestrial microalgae

Plants and other photosynthetic organisms interact with their environment and surrounding microbiota through specialized associations. A global core microbiota has been proposed at high taxonomic levels, such as the order level. However, it remains unclear which environmental factors and how microbe-microbe interactions drive variation of this core microbiota at lower taxonomic resolution. Here, we leveraged the environmental diversity of 141 sites across the southwest of France to characterize algal populations, and their associated bacterial and fungal microbiota. We then performed a meta-analysis, combining these data with published datasets to formally identify the global core microbiota of terrestrial photosynthetic organisms, which comprises seven bacterial and five fungal orders. We next investigated diversity within this core microbiota and the environmental drivers shaping site-specific community composition. While environmental factors have a low impact on the total relative abundance of core orders, the core microbiota at the ASV-level is impacted by climatic factors, edaphic factors, and plant community descriptors. Using interaction network analysis, we finally explored how microbe-microbe interactions contribute to the assembly of stable core communities. Our results show that core ASVs occupy central positions in algal-associated microbial networks and that distinct core orders drive site-specific variation in core microbiota structure. Together, these findings highlight the importance of both environmental context and microbial interactions in shaping the composition and stability of the core microbiota associated with photosynthetic organisms.

plant biology↗

bioRxiv2026

Nuclear Speckles Regulate Splicing During Muscle Stem Cell Activation and Aging

Skeletal muscle contains a population of adult stem cells called satellite cells or muscle stem cells (MuSCs) that are responsible for regeneration after injury. MuSCs utilize gene expression programs to maintain quiescence and differentiate after injury and a key regulator of gene expression is splicing, which uniquely changes when transcripts interact with nuclear speckles (NS). NS are membrane-less biomolecular condensates that phase separate proteins, RNAs and chromatin, but how these organelles regulate molecular processes in MuSCs remains unknown. Herein, we build a comprehensive and systems-level understanding of NS influence on alternative splicing, transcriptional regulation and stem cell function before and after injury and in aging. We establish that NS increased in size and number in MuSCs following injury and influence MuSC activation dynamics. We generated a catalog of isoform-resolved splicing events and linked how RNA interactions with NS amplify splicing completion during the injury response. In old age, MuSCs lose NS, yet shifted towards longer, more completely spliced isoforms enriched for RNA binding protein motifs and multivalency. Our studies unveil evidence that RNA interactions with NS shape stem cell state and regenerative responses but are attenuated in old age. HIGHLIGHTSO_LI3D super-resolution imaging of nuclear speckles in muscle stem cells before and after muscle injury shows intricate relationship with activation C_LIO_LIIsoform-resolved profiling of muscle stem cells shows increases in gene expression and splicing during injury response C_LIO_LIMapping RNA interactions with nuclear speckles shows RNAs undergo strongest splicing when proximal to nuclear speckles C_LIO_LIOld aged muscle stem cells lose nuclear speckles and display aberrant splicing, with longer transcripts, more exons, and increased RNA binding protein motifs C_LI

genomics↗

bioRxiv2026

Biotic-response networks are an important organizer of the transcriptome in wild Arabidopsis thaliana populations

Extensive laboratory experimentation has revealed conserved molecular pathways controlling growth and stress responses in plants, yet how these programs operate in natural settings remains poorly understood. We investigated transcriptome organization in wild populations of Arabidopsis thaliana by sampling plants from 60 natural sites in Europe and North America across two seasons. Transcriptomes varied extensively among individuals and showed largely continuous rather than discrete structure across geography and season. Although disease and microbial colonization were common in the wild, wild transcriptomes did not simply recapitulate canonical laboratory stress signatures. Measured microbial infection, environmental, and phenotypic variables explained only a modest fraction of total expression variation, but infection-associated signals accounted for the largest share of the explainable component. Consistent with this, biotic-response networks defined in controlled laboratory experiments were well conserved in wild transcriptomes, whereas control and abiotic-response networks were substantially reorganized. Together, these results suggest that while core transcriptional modules remain recognizable across environments, regulatory relationships among modules differ markedly between laboratory and natural contexts. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=129 SRC="FIGDIR/small/711176v1_ufig1.gif" ALT="Figure 1"> View larger version (28K): org.highwire.dtl.DTLVardef@1c89152org.highwire.dtl.DTLVardef@bcd1acorg.highwire.dtl.DTLVardef@6975a2org.highwire.dtl.DTLVardef@1a9e2fe_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2026

The contribution of endocytic mediators to itch transmission

Chronic itch is a major burden, impacting the quality of life for one in four adults, and is closely associated with increased levels of anxiety, depression, and suicide. Despite its widespread burden, the precise mechanisms driving and sustaining chronic itch remain poorly understood. Non-histaminergic itch transmission in the spinal cord relies on the synaptic vesicle (SV) release of gastrin-releasing peptide (GRP) and the subsequent binding to the Gastrin-Releasing Peptide Receptor (GRPR). While SV exocytosis facilitates neurotransmitter release, such as GRP, SV endocytosis, mediated by key proteins including clathrin, adaptor associated kinase 1 (AAK1), and Dynamin (Dnm) are essential for retrieving and recycling SV from the presynaptic membrane to maintain signaling. Building upon evidence that AAK1- and Dnm-mediated endocytosis are viable targets to reverse pain, we characterized the role of these endocytic mediators in the regulation of synaptic transmission in itch pathways. We localized mRNA encoding AAK1, Dnm1 and Dnm3 within GRP positive neurons of the mouse dorsal root ganglia (DRG). Genetic and pharmacological disruption of AAK1 significantly reduced scratching behavior compared to control groups. This anti-pruritic effect correlated with confirmed Aak1 mRNA knockdown in both the DRG and spinal cord. Similarly, siRNA mediated knockdown of Dnm1 and Dnm3 in the DRGs also reduced scratching behavior. Crucially, these treatments decreased GRP release without altering locomotor activity or anxiety-like behaviors. Together, these findings suggest that the disruption of SV recycling reduces itch related signaling and behavior without affecting normal motor functions, providing a new approach for chronic pruritus.

neuroscience↗

bioRxiv2026

Nascent extracellular matrix converts biomaterial cues into cell fate decisions

Hydrogels serve as powerful models for investigating cell-extracellular matrix (ECM) interactions. While chemical modifications are routinely used to tune hydrogel properties, it remains unclear whether these modifications mediate cell fate. Previous work has shown that cells deposit newly synthesized (nascent) ECM at the cell-hydrogel interface. Here, we demonstrate that this nascent ECM interface regulates how cells interpret chemical modifications. Using hydrogels with varied chemical modifications, we isolated the effects of chemical modification on nascent ECM and cell fate. Nascent ECM deposition increased as a function of hydrogel modification and with distinct matrisome compositions. While low modification hydrogels promoted cell differentiation, high modifications increased cell proliferation. Perturbing cell-nascent ECM interactions reversed this cell fate. Our findings reveal that nascent ECM regulates cell fate by converting hydrogel cues into signals that control cell fate. This tri-directional interplay among hydrogel chemical modifications, nascent ECM, and cell fate reframes how we interpret cell-hydrogel interactions.

bioengineering↗

bioRxiv2025

Endosomal MrGPRX1 signaling sensitizes TRPV1 to enhance itch

G protein-coupled receptors (GPCRs) and TRPV (transient receptor potential vanilloid) channels are crucial for signal transduction in physiological processes, including neurotransmission, pain, and itch. Downstream effectors of GPCR signaling can directly stimulate TRPV channels or enhance their sensitivity to stimuli, a process known as TRPV sensitization. Traditionally, GPCRs are activated at the cell surface by extracellular agonists, triggering signaling cascades. Recent evidence suggests GPCRs continue to signal from intracellular organelles. The human Mas-related G-protein coupled receptor X1 (MrGPRX1) is a GPCR expressed in primary sensory neurons involved in nociception and pruritus. Recent studies demonstrated how intracellular GPCR signaling regulates neuronal activity. However, there is no evidence characterizing MrGPRX1 trafficking or intracellular signaling. Herein, we characterized MrGPRX1 signaling within the endosomal network and its role in sensitizing TRPV1 channels to enhance itch signaling. Utilizing subcellular targeted biosensors, we demonstrated MrGPRX1 can traffic and signal from endosomes. Immunofluorescence analysis showed that MrGPRX1 internalizes following BAM8-22 stimulation. BRET assays revealed that MrGPRX1 activation induces Gq and {beta}-arrestin-1 recruitment to the plasma membrane and early endosomes. Inhibition of dynamin or clathrin blocked BAM8-22-induced MrGPRX1 endocytosis and decreased nuclear extracellular signal-regulated kinase (ERK) signaling. Calcium signaling confirmed that MrGPRX1-mediated TRPV1 sensitization is mediated by protein kinase C and ERK activation. Our findings reveal a novel role for MrGPRX1 endosomal signaling in TRPV1 sensitization. Understanding the mechanisms of MrGPRX1 signaling offers valuable insights into differentiating between pain and itch pathways, aiding in the development of targeted therapies for chronic pain and persistent itch.

neuroscience↗

bioRxiv2025

Artificial Soil (ArtSoil): recreating soil conditions in synthetic plant growth media

Controlled plant growth in laboratories can be achieved by cultivating plants in sterile or axenic conditions on pre-defined synthetic growth media typically supplemented with sugar. In nature, plants do not receive exogenous sugar supply, form symbiosis with microbes, and plant growth is influenced by soil edaphic factors. Thus, physiological and multi-omic analyses from plants grown on synthetic media will differ from those of soil-grown plants due to the influence of sucrose, and the lack of influence from microbiota and soil edaphic factors on plant growth. The rapid advancements in spatial-omics call for accurate characterization of plants grown in conditions similar to soil. To address the issue, we developed Artificial Soil (ArtSoil), a growth medium containing essential nutrients for plant growth, and aqueous soil extract (ASE) to maintain soil microbiomes and edaphic factors, simultaneously eliminating the need for sugar supplementation in the medium. We compared Arabidopsis thaliana grown on conventional media to ArtSoil under various growth conditions, and showed that complex soil microbiota in ArtSoil promote plant growth without physiological side effects induced by sucrose. We demonstrate an application for ArtSoil in single-cell transcriptomics and report microbiota-induced cell-type specificity in immune and nitrogen signaling. We tested ArtSoil with six types of ASEs to present the potential of ArtSoil in decoupling the effects of nutrients from microbiota in plant growth. We conclude that ArtSoil recapitulates the soil environment compared to conventional media, hence, enabling physiologically relevant plant growth. Significance statementLaboratory plant growth commonly relies on sugar-supplemented media and sterile growth conditions, which introduces physiological artifacts and fails to capture the natural influences of soil microbiota and edaphic factors, leading to analyses that diverge from native conditions. We developed Artificial Soil (ArtSoil), a medium containing aqueous soil extract that supports microbiota and sugar-independent growth, and demonstrate its utility in recapitulating native soil properties and enabling physiologically relevant plant development in vitro.

plant biology↗

bioRxiv2024

Mouse models of non-dystrophic and dystrophic myotonia exhibit nociplastic pain-like behaviors

Pain is a common and disabling feature of myotonic disorders, yet its biological basis remains poorly understood and no targeted analgesic therapies currently exist. Here, we demonstrate that skeletal muscle hyperexcitability is sufficient to initiate a persistent pain state independent of inflammation, nerve injury, or overt tissue damage. Using complementary pharmacological and genetic models of myotonia resulting from loss of the voltage-gated skeletal muscle chloride channel ClC-1 function, we show that transient and chronic myotonia produce robust mechanical, thermal, and cold hypersensitivity, as well as spontaneous pain-like behavior. Notably, pain-like behaviors induced by transient myotonia persist long after overt motor symptoms have resolved, suggesting that a transient episode of muscle hyperexcitability is sufficient to trigger prolonged alterations in nociceptive processing. Physiological recordings revealed altered excitability of dorsal root ganglion and superficial dorsal horn neurons and enhanced sensory-evoked activity in the parabrachial nucleus, indicating altered nociceptive processing across multiple levels of the pain neuraxis. Transient myotonia increased total sodium current density in sensory neurons, with a shift toward a greater tetrodotoxin-resistant current fraction. Pharmacological inhibition with the NaV1.8-directed analgesic Suzetrigine markedly attenuated pain-like behaviors in both models of myotonia. Together, these findings establish a link between myotonia and persistent alterations in nociceptive processing and identify NaV1.8-directed analgesia as a promising therapeutic strategy for myotonia-associated pain.

neuroscience↗

bioRxiv2024

Uncoupling the CRMP2-CaV2.2 interaction reduces pain-like behavior in a preclinical osteoarthritis model

Osteoarthritis (OA) represents a significant pain challenge globally, as current treatments are limited and come with substantial and adverse side effects. Voltage-gated calcium channels have proved to be pharmacologically effective targets, with multiple FDA-approved CaV2.2 modulators available for the treatment of pain. Although effective, drugs targeting CaV2.2 are complicated by the same obstacles facing other pain therapeutics-invasive routes of administration, narrow therapeutic windows, side effects, and addiction potential. We have identified a key regulator of CaV2.2 channels, collapsing response mediator protein 2 (CRMP2), that allows us to indirectly regulate CaV2.2 expression and function. We developed a peptidomimetic modulator of CRMP2, CBD3063, that effectively reverses neuropathic and inflammatory pain without negative side effects by reducing membrane expression of CaV2.2. Using a rodent model of OA, we demonstrate the intraperitoneal administration of CBD3063 alleviates both evoked and non-evoked behavioral hallmarks of OA pain. Further, we reveal that CBD3063 reduces OA-induced increased neural activity in the parabrachial nucleus, a key supraspinal site modulating the pain experience. Together, these studies suggest CBD3063 is an effective analgesic for OA pain.

neuroscience↗

bioRxiv2024

Cellular mechanotransduction of human osteoblasts in microgravity

Astronauts experience significant and rapid bone loss as a result of an extended stay in space, making the International Space Station (ISS) the perfect laboratory for studying osteoporosis due to the accelerated nature of bone loss on the ISS. This prompts the question, how does the lack of load due to zero-gravity propagate to bone-forming cells, human fetal osteoblasts (hFOBs), altering their maturation to mineralization? Here, we aim to study the mechanotransduction mechanisms by which bone loss occurs in microgravity. Two automated experiments, 4 microfluidic chips capable of measuring single-cell mechanics of hFOBs via aspiration and cell spheroids incubated in pressure-controlled chambers, were each integrated into a CubeLab deployed to the ISS National Laboratory. For the first experiment, we report protrusion measurements of aspirated cells after exposure to microgravity at the ISS and compare these results to ground control conducted inside the CubeLab. Our analysis revealed slightly elongated protrusions for space samples compared to ground samples indicating softening of hFOB cells in microgravity. In the second experiment, we encapsulated osteoblast spheroids in collagen gel and incubated the samples in pressure-controlled chambers. We found that microgravity significantly reduced filamentous actin levels in the hFOB spheroids. When subjected to pressure, the spheroids exhibited increased pSMAD1/5/9 expression, regardless of the microgravity condition. Moreover, microgravity reduced YAP expression, while pressure increased YAP levels, thus restoring YAP expression for spheroids in microgravity. Our study provides insights into the influence of microgravity on the mechanical properties of bone cells and the impact of compressive pressure on cell behavior and signaling in space.

cell biology↗

bioRxiv2024

Quantification of local matrix deposition during muscle stem cell activation using engineered hydrogels

Adult stem cells occupy a niche that contributes to their function, but how stem cells remodel their microenvironment remains an open-ended question. Herein, biomaterials-based systems and metabolic labeling were utilized to evaluate how skeletal muscle stem cells deposit extracellular matrix. Muscle stem cells and committed myoblasts were observed to generate less nascent matrix than muscle resident fibro-adipogenic progenitors. When cultured on substrates that matched the stiffness of physiological uninjured and injured muscles, the increased nascent matrix deposition was associated with stem cell activation. Reducing the ability to deposit nascent matrix in muscle stem cells attenuated function and mimicked impairments observed from muscle stem cells isolated from old aged muscles, which could be rescued with therapeutic supplementation of insulin-like growth factors. These results highlight how nascent matrix production is critical for maintaining healthy stem cell function.

bioengineering↗

bioRxiv2023

Intranasal CRMP2-Ubc9 Inhibitor Regulates NaV1.7 to Alleviate Trigeminal Neuropathic Pain

Dysregulation of voltage-gated sodium NaV1.7 channels in sensory neurons contributes to chronic pain conditions, including trigeminal neuropathic pain. We previously reported that chronic pain results in part from increased SUMOylation of collapsin response mediator protein 2 (CRMP2), leading to an increased CRMP2/NaV1.7 interaction and increased functional activity of NaV1.7. Targeting this feed-forward regulation, we developed compound 194, which inhibits CRMP2 SUMOylation mediated by the SUMO-conjugating enzyme Ubc9. We further demonstrated that 194 effectively reduces the functional activity of NaV1.7 channels in dorsal root ganglia neurons and alleviated inflammatory and neuropathic pain. Here, we employed a comprehensive array of investigative approaches, encompassing biochemical, pharmacological, genetic, electrophysiological, and behavioral analyses, to assess the functional implications of NaV1.7 regulation by CRMP2 in trigeminal ganglia (TG) neurons. We confirmed the expression of Scn9a, Dpysl2, and UBE2I within TG neurons. Furthermore, we found an interaction between CRMP2 and NaV1.7, with CRMP2 being SUMOylated in these sensory ganglia. Disrupting CRMP2 SUMOylation with compound 194 uncoupled the CRMP2/NaV1.7 interaction, impeded NaV1.7 diffusion on the plasma membrane, and subsequently diminished NaV1.7 activity. Compound 194 also led to a reduction in TG neuron excitability. Finally, when intranasally administered to rats with chronic constriction injury of the infraorbital nerve (CCI-ION), 194 significantly decreased nociceptive behaviors. Collectively, our findings underscore the critical role of CRMP2 in regulating NaV1.7 within TG neurons, emphasizing the importance of this indirect modulation in trigeminal neuropathic pain.

neuroscience↗

bioRxiv2022

Identification and targeting of a unique NaV1.7 domain driving chronic pain

Despite identification of several small molecules directly targeting the voltage-gated sodium channel NaV1.7, none has been clinically successful. We reported that preventing addition of a small ubiquitin-like modifier (SUMO) on the NaV1.7-interacting cytosolic collapsin response mediator protein 2 (CRMP2) blocked NaV1.7 functions and was antinociceptive in rodents. Here, we discovered a 15 amino acid CRMP2 regulatory sequence (CRS) unique to NaV1.7 that is essential for this regulatory coupling. CRMP2 preferentially bound to the NaV1.7 CRS over other isoforms. Substitution of the NaV1.7 CRS with the homologous domains from the other eight voltage-gated sodium channel isoforms decreased tetrodotoxin-sensitive NaV1.7 currents in rodent sensory neurons. A cell-penetrant version of NaV1.7-CRS reduced NaV1.7 currents and trafficking, decreased presynaptic NaV1.7 localization, reduced spinal neurotransmitter release, and reversed mechanical allodynia in a rat spared nerve injury model of neuropathic pain. Interfering with NaV1.7-CRMP2 coupling did not produce motor impairment and spared thermal, inflammatory, and post-surgical nociception. As proof-of-concept for NaV1.7-targeted gene therapy, we found that NaV1.7-CRS packaged into an adeno-associated virus recapitulated the effects on NaV1.7 function in both rodent and rhesus macaque sensory neurons and both reversed and prevented the development of mechanical allodynia in a neuropathic pain model in male and female rodents. One Sentence SummaryA novel regulatory domain on the voltage gated sodium channel NaV1.7 that can be targeted to produce analgesia.

neuroscience↗

bioRxiv2022

Repeated birth injuries lead to pelvic floor muscle dysfunction and impairment in regeneration

ObjectivesChildbirth is a key risk factor for pelvic floor muscle (PFM) injury and dysfunction, and subsequent pelvic floor disorders (PFDs). Multiparity further exacerbates these risks. Using the pre-clinical rat model of simulated birth injury (SBI), we previously identified that an SBI leads to PFM atrophy and fibrosis. We hypothesized that multiple SBIs further overwhelm PFM regenerative capacity, leading to functionally relevant pathological alterations long-term. Study DesignRats underwent SBI and were allowed to recover for 8 weeks to undergo another SBI. Animals were sacrificed at acute, subacute, and long-term time points post-second injury (N=3-6/time point), and pubocaudalis (PCa) was harvested to assess ex vivo muscle function, histomorphological properties and gene expression. ResultsAcutely following the 1st SBI, PCa force was decreased relative to controls. At 4 weeks, PCa force was recovered and remained unchanged at 8 weeks. Similarly, lower PCa force was observed immediately after repeated SBI. In contrast to functional recovery after 1st SBI, PCa force remained lower at 4 weeks post-2nd SBI and continued to be decreased even after 12 weeks after repeated injury. Fiber size was smaller at the long-term time points after 2nd SBI compared to controls and single SBI groups. As opposed to the resolution of centralized nuclei at 8 weeks post-1st SBI, regenerating myofibers persisted even at 12 weeks post-2nd SBI. In contrast to the peak of collagen content at 4 weeks post-1st SBI, this parameter raised progressively over 12 weeks after repeated SBIs. Prolonged inflammatory response, impairment in muscle anabolism, and sustained expression of ECM remodeling genes were observed after repeated SBIs. ConclusionsRepeated birth injuries delay PFM regeneration and impair function in the pre-clinical rat model.

physiology↗

bioRxiv2021

The role of muscle stem cells and fibro-adipogenic progenitors in female pelvic floor muscle regeneration following birth injury

Pelvic floor muscle (PFM) injury during childbirth is a key risk factor for subsequent pelvic floor disorders that affect millions of women worldwide. Muscle stem cells (MuSCs) play a central role in the regeneration of injured skeletal muscles, where they activate, proliferate, and differentiate to assure myogenesis needed for muscle recovery. For robust regenerative function, MuSCs require the support of fibro-adipogenic progenitors (FAPs) and immune cells. To elucidate the role of MuSCs, FAPs, and immune infiltrate in female PFM regeneration, we used radiation to perturb the system and followed PFM recovery in a simulated birth injury (SBI) rat model. Non-irradiated and irradiated rats were euthanized at 3,7, 10, and 28 days after SBI; PFMs were harvested and prepared for immunohistochemistry. Cross sectional area (CSA) of all PFM myofibers 28 days after injury in irradiated animals was significantly lower relative to non-irradiated injured controls, indicating impairment of PFM recovery. Following SBI in non-irradiated animals, the number of MuSCs and FAPs expanded significantly at 7 and 3 days after injury, respectively; this expansion did not occur in irradiated animals at the same time points. CSA of embryonic myosin heavy chain (eMyHC, marker of newly regenerated myofibers) positive fibers was also significantly smaller following SBI in irradiated muscles compared to PFMs from non-irradiated injured controls at 7 days. Our results demonstrate that loss of function and decreased expansion of MuSCs and FAPs associated with irradiation results in impaired PFM recovery, signifying essential roles for MuSCs and FAPs in the regenerative process of female PFMs after birth injury. These findings can inform the identification of novel preventative and therapeutic targets and the development of new treatments for PFM dysfunction and associated pelvic floor disorders.

cell biology↗

bioRxiv2021

Pro-regenerative Extracellular Matrix Hydrogel Prevents and Mitigates Pathological Alterations of Pelvic Muscles Following Birth Injury

Pelvic floor disorders, which include pelvic organ prolapse, and urinary and fecal incontinence, affect millions of women globally and represent a major public health concern. Pelvic floor muscle (PFM) dysfunction has been identified as one of the leading risk factors for the development of these morbid conditions. Even though childbirth, specifically vaginal delivery, has been long recognized as the most important potentially modifiable risk factor for PFM injury, the precise mechanisms of PFM dysfunction following childbirth remain elusive. In this study we demonstrate that PFMs undergo atrophy and severe fibrosis in parous women with symptomatic pelvic organ prolapse compared to age-matched nulliparous cadaveric donors without history of pelvic floor disorders. These pathological alterations are recapitulated in the pre-clinical rat model of simulated birth injury. The transcriptional signature of PFMs post-injury demonstrates a sustained inflammatory response, impairment in muscle anabolism, and persistent expression of extracellular matrix (ECM) remodeling genes. Next, we evaluated the administration of acellular injectable skeletal muscle extracellular matrix hydrogel for the prevention and mitigation of these pathological alterations. Treatment of PFMs with the biomaterial either at the time of birth injury or 4 weeks post-injury reduced muscle atrophy and mitigated fibrotic degeneration. By evaluating gene expression, we demonstrate that these changes are mainly driven by the hydrogel-induced modulation of the immune response and intramuscular fibrosis, as well as enhancement of the endogenous myogenesis. This work furthers our understanding of PFM birth injury and demonstrates proof-of-concept for a new pragmatic pro-regenerative biomaterial approach for treating injured PFMs.

bioengineering↗