{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,5]],"date-time":"2026-10-05T20:19:26Z","timestamp":1791231566900,"version":"4.1.0"},"reference-count":47,"publisher":"MDPI AG","issue":"10","license":[{"start":{"date-parts":[[2025,10,4]],"date-time":"2025-10-04T00:00:00Z","timestamp":1759536000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"NIH NINDS","award":["1R01NS114007-01A1"],"award-info":[{"award-number":["1R01NS114007-01A1"]}]},{"name":"NIH NINDS","award":["R01NS117749-01"],"award-info":[{"award-number":["R01NS117749-01"]}]},{"DOI":"10.13039\/100011781","name":"Shriners Hospitals for Children","doi-asserted-by":"publisher","award":["85115"],"award-info":[{"award-number":["85115"]}],"id":[{"id":"10.13039\/100011781","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100005191","name":"Craig H. Neilsen Foundation Senior Research","doi-asserted-by":"publisher","award":["546798"],"award-info":[{"award-number":["546798"]}],"id":[{"id":"10.13039\/100005191","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100011781","name":"Shriners Viral Core","doi-asserted-by":"publisher","award":["84051-PHI-21"],"award-info":[{"award-number":["84051-PHI-21"]}],"id":[{"id":"10.13039\/100011781","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Bioengineering"],"abstract":"<jats:p>Spinal cord injury (SCI) often results in long-term locomotor impairments, and strategies to enhance functional recovery remain limited. While epidural electrical stimulation (EES) has shown clinical promise, our understanding of the mechanisms by which it improves function remains incomplete. Here, we use genetic tools in an animal model to perform neuromodulation and treadmill rehabilitation in a manner similar to EES, but with the benefit of the genetic tools and animal model allowing for targeted manipulation, precise quantification of the cells and circuits that were manipulated, and the gathering of extensive kinematic data. We used a viral construct that selectively transduces large diameter afferent fibers (LDAFs) with a designer receptor exclusively activated by a designer drug (hM3Dq DREADD; a chemogenetic construct) to increase the excitability of large fibers specifically, in the rat contusion SCI model. As changes in locomotion with afferent stimulation can be subtle, we carried out a detailed characterization of the kinematics of locomotor recovery over time. Adult Long-Evans rats received contusion injuries and direct intraganglionic injections containing AAV2-hSyn-hM3Dq-mCherry, a viral vector that has been shown to preferentially transduce LDAFs, or a control with tracer only (AAV2-hSyn-mCherry). These neurons then had their activity increased by application of the designer drug Clozapine-N-oxide (CNO), inducing tonic excitation during treadmill training in the recovery phase. Kinematic data were collected during treadmill locomotion across a range of speeds over nine weeks post-injury. Data were analyzed using a mixed effects model chosen from amongst several models using information criteria. That model included fixed effects for treatment (DREADDs vs. control injection), time (weeks post injury), and speed, with random intercepts for rat and time point nested within rat. Significant effects of treatment and treatment interactions were found in many parameters, with a sometimes complicated dependence on speed. Generally, DREADDs activation resulted in shorter stance duration, but less reduction in swing duration with speed, yielding lower duty factors. Interestingly, our finding of shorter stance durations with DREADDs activation mimics a past study in the hemi-section injury model, but other changes, including the variability of anterior superior iliac spine (ASIS) height, showed an opposite trend. These may reflect differences in injury severity and laterality (i.e., in the hemi-section injury the contralateral limb is expected to be largely functional). Furthermore, as with that study, withdrawal of DREADDs activation in week seven did not cause significant changes in kinematics, suggesting that activation may have dwindling effects at this later stage. This study highlights the utility of high-resolution kinematics for detecting subtle changes during recovery, and will enable the refinement of neuromechanical models that predict how locomotion changes with afferent neuromodulation, injury, and recovery, suggesting new directions for treatment of SCI.<\/jats:p>","DOI":"10.3390\/bioengineering12101080","type":"journal-article","created":{"date-parts":[[2025,10,6]],"date-time":"2025-10-06T08:10:51Z","timestamp":1759738251000},"page":"1080","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Detailed Kinematic Analysis Reveals Subtleties of Recovery from Contusion Injury in the Rat Model with DREADDs Afferent Neuromodulation"],"prefix":"10.3390","volume":"12","author":[{"given":"Gavin Thomas","family":"Koma","sequence":"first","affiliation":[{"name":"Department of Bioengineering, Temple University, Philadelphia, PA 19122, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kathleen M.","family":"Keefe","sequence":"additional","affiliation":[{"name":"Department of Biomedical Education and Data Science, Lewis Katz School of Medicine, Temple University, Philadelphia, PA 19140, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"George","family":"Moukarzel","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc., North Wales, PA 19446, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hannah","family":"Sobotka-Briner","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc., North Wales, PA 19446, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bradley C.","family":"Rauscher","sequence":"additional","affiliation":[{"name":"Department of Biomedical Engineering, Boston University, Boston, MA 02215, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Julia","family":"Capaldi","sequence":"additional","affiliation":[{"name":"Merck & Co., Inc., North Wales, PA 19446, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jie","family":"Chen","sequence":"additional","affiliation":[{"name":"Department of Neural Sciences, Lewis Katz School of Medicine, Temple University, Philadelphia, PA 19140, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"suffix":"3rd","given":"Thomas J.","family":"Campion","sequence":"additional","affiliation":[{"name":"Department of Pathology and Laboratory Medicine, Children\u2019s Hospital of Philadelphia, Philadelphia, PA 19104, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jacquelynn","family":"Rajavong","sequence":"additional","affiliation":[{"name":"Department of Neural Sciences, Lewis Katz School of Medicine, Temple University, Philadelphia, PA 19140, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Kaitlyn","family":"Rauscher","sequence":"additional","affiliation":[{"name":"Department of Bioengineering, Temple University, Philadelphia, PA 19122, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Benjamin D.","family":"Robertson","sequence":"additional","affiliation":[{"name":"XCMR Inc., Narberth, PA 19072, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2614-1624","authenticated-orcid":false,"given":"George M.","family":"Smith","sequence":"additional","affiliation":[{"name":"Department of Neural Sciences, Lewis Katz School of Medicine, Temple University, Philadelphia, PA 19140, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrew J.","family":"Spence","sequence":"additional","affiliation":[{"name":"Department of Bioengineering, Temple University, Philadelphia, PA 19122, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,10,4]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"91","DOI":"10.1016\/j.expneurol.2011.02.009","article-title":"Rehabilitative training and plasticity following spinal cord injury","volume":"235","author":"Fouad","year":"2012","journal-title":"Exp. Neurol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"1841","DOI":"10.1089\/neu.2016.4577","article-title":"Rehabilitation Strategies after Spinal Cord Injury: Inquiry into the Mechanisms of Success and Failure","volume":"34","author":"Murray","year":"2017","journal-title":"J. Neurotrauma"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"188","DOI":"10.1038\/sc.1996.37","article-title":"Effect of surgery on motor recovery following traumatic spinal cord injury","volume":"34","author":"Waters","year":"1996","journal-title":"Spinal Cord"},{"key":"ref_4","doi-asserted-by":"crossref","unstructured":"Bilchak, J.N., Caron, G., and C\u00f4t\u00e9, M.-P. (2021). Exercise-Induced Plasticity in Signaling Pathways Involved in Motor Recovery after Spinal Cord Injury. Int. J. Mol. Sci., 22.","DOI":"10.3390\/ijms22094858"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"260","DOI":"10.1038\/s41591-021-01663-5","article-title":"Activity-dependent spinal cord neuromodulation rapidly restores trunk and leg motor functions after complete paralysis","volume":"28","author":"Rowald","year":"2022","journal-title":"Nat. Med."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Eisdorfer, J.T., Smit, R.D., Keefe, K.M., Lemay, M.A., Smith, G.M., and Spence, A.J. (2020). Epidural Electrical Stimulation: A Review of Plasticity Mechanisms That Are Hypothesized to Underlie Enhanced Recovery from Spinal Cord Injury with Stimulation. Front. Mol. Neurosci., 13.","DOI":"10.3389\/fnmol.2020.00163"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"32","DOI":"10.1310\/sci1301-32","article-title":"A systematic review of the efficacy of gait rehabilitation strategies for spinal cord injury","volume":"13","author":"Lam","year":"2007","journal-title":"Top. Spinal Cord Inj. Rehabil."},{"key":"ref_8","first-page":"99","article-title":"Functional neuromuscular stimulation for mobility in people with spinal cord injuries. The Parastep I System","volume":"19","author":"Chaplin","year":"1996","journal-title":"J. Spinal Cord Med."},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1089\/neu.1995.12.1","article-title":"A sensitive and reliable locomotor rating scale for open field testing in rats","volume":"12","author":"Basso","year":"1995","journal-title":"J. Neurotrauma"},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Ageeva, T., Sabirov, D., Sufianov, A., Davletshin, E., Plotnikova, E., Shigapova, R., Sufianova, G., Timofeeva, A., Chelyshev, Y., and Rizvanov, A. (2024). The Impact of Treadmill Training on Tissue Integrity, Axon Growth, and Astrocyte Modulation. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms25073772"},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Cai, J., Wang, Y., Zhai, C., Jiang, K., Wang, Z., Fang, L., Li, X., Zhu, C., Liu, W., and Wang, T. (2025). Body weight-supported treadmill training reduces glial scar overgrowth in SCI rats by decreasing the reactivity of astrocytes during the subacute phase. BMC Neurosci., 26.","DOI":"10.1186\/s12868-025-00947-7"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"1938","DOI":"10.1016\/S0140-6736(11)60547-3","article-title":"Effect of epidural stimulation of the lumbosacral spinal cord on voluntary movement, standing, and assisted stepping after motor complete paraplegia: A case study","volume":"377","author":"Harkema","year":"2011","journal-title":"Lancet"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"65","DOI":"10.1038\/s41586-018-0649-2","article-title":"Targeted neurotechnology restores walking in humans with spinal cord injury","volume":"563","author":"Wagner","year":"2018","journal-title":"Nature"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"540","DOI":"10.1038\/s41586-022-05385-7","article-title":"The neurons that restore walking after paralysis","volume":"611","author":"Kathe","year":"2022","journal-title":"Nature"},{"key":"ref_15","doi-asserted-by":"crossref","unstructured":"Dorrian, R.M., Berryman, C.F., Lauto, A., and Leonard, A.V. (2023). Electrical stimulation for the treatment of spinal cord injuries: A review of the cellular and molecular mechanisms that drive functional improvements. Front. Cell. Neurosci., 17.","DOI":"10.3389\/fncel.2023.1095259"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"2367","DOI":"10.4103\/1673-5374.313017","article-title":"Epidural electrical stimulation for spinal cord injury","volume":"16","author":"Choi","year":"2021","journal-title":"Neural Regen. Res."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"eadp9607","DOI":"10.1126\/scitranslmed.adp9607","article-title":"High-frequency epidural electrical stimulation reduces spasticity and facilitates walking recovery in patients with spinal cord injury","volume":"17","author":"Romeni","year":"2025","journal-title":"Sci. Transl. Med."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1333","DOI":"10.1038\/nn.2401","article-title":"Transformation of nonfunctional spinal circuits into functional states after the loss of brain input","volume":"12","author":"Courtine","year":"2009","journal-title":"Nat. Neurosci."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"576","DOI":"10.1038\/s41593-018-0093-5","article-title":"Cortico-reticulo-spinal circuit reorganization enables functional recovery after severe spinal cord contusion","volume":"21","author":"Asboth","year":"2018","journal-title":"Nat. Neurosci."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"683","DOI":"10.1016\/j.neuron.2016.01.040","article-title":"DREADDs for Neuroscientists","volume":"89","author":"Roth","year":"2016","journal-title":"Neuron"},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Eisdorfer, J.T., Sobotka-Briner, H., Schramfield, S., Moukarzel, G., Chen, J., Campion, T.J., Smit, R., Rauscher, B.C., Lemay, M.A., and Smith, G.M. (2022). Chemogenetic modulation of sensory afferents induces locomotor changes and plasticity after spinal cord injury. Front. Mol. Neurosci., 15.","DOI":"10.3389\/fnmol.2022.872634"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"464","DOI":"10.1016\/j.mcn.2012.03.002","article-title":"AAV8(gfp) preferentially targets large diameter dorsal root ganglion neurones after both intra-dorsal root ganglion and intrathecal injection","volume":"49","author":"Jacques","year":"2012","journal-title":"Mol. Cell. Neurosci."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Eisdorfer, J.T., Phelan, M.A., Keefe, K.M., Rollins, M.M., Campion, T.J., Rauscher, K.M., Sobotka-Briner, H., Senior, M., Gordon, G., and Smith, G.M. (2021). Addition of angled rungs to the horizontal ladder walking task for more sensitive probing of sensorimotor changes. PLoS ONE, 16.","DOI":"10.1371\/journal.pone.0246298"},{"key":"ref_24","doi-asserted-by":"crossref","unstructured":"Jendryka, M., Palchaudhuri, M., Ursu, D., van der Veen, B., Liss, B., K\u00e4tzel, D., Nissen, W., and Pekcec, A. (2019). Pharmacokinetic and pharmacodynamic actions of clozapine-N-oxide, clozapine, and compound 21 in DREADD-based chemogenetics in mice. Sci. Rep., 9.","DOI":"10.1038\/s41598-019-41088-2"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"254","DOI":"10.1302\/0301-620X.90B2.19808","article-title":"The characteristics of dorsal-root ganglia and sensory innervation of the hip in rats","volume":"90-B","author":"Nakajima","year":"2008","journal-title":"J. Bone Jt. Surg. Br."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"9831","DOI":"10.1128\/JVI.00878-06","article-title":"The 37\/67-Kilodalton Laminin Receptor Is a Receptor for Adeno-Associated Virus Serotypes 8, 2, 3, and 9","volume":"80","author":"Akache","year":"2006","journal-title":"J. Virol."},{"key":"ref_27","first-page":"50111","article-title":"A Contusion Model of Severe Spinal Cord Injury in Rats","volume":"17","author":"Krishna","year":"2013","journal-title":"J. Vis. Exp. JoVE"},{"key":"ref_28","doi-asserted-by":"crossref","unstructured":"Maghsoudi, O.H., Tabrizi, A.V., Robertson, B., and Spence, A. (November, January 29). Superpixels based marker tracking vs. hue thresholding in rodent biomechanics application. Proceedings of the 2017 51st Asilomar Conference on Signals, Systems, and Computers, Pacific Grove, CA, USA.","DOI":"10.1109\/ACSSC.2017.8335168"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"1","DOI":"10.2307\/1005513","article-title":"Anatomy of the Rat","volume":"27","author":"Greene","year":"1935","journal-title":"Trans. Am. Philos. Soc."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"559","DOI":"10.1007\/s00586-004-0834-x","article-title":"Characteristics of sensory DRG neurons innervating the lumbar facet joints in rats","volume":"14","author":"Ishikawa","year":"2005","journal-title":"Eur. Spine J."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"31","DOI":"10.1186\/1744-8069-6-31","article-title":"Differential adeno-associated virus mediated gene transfer to sensory neurons following intrathecal delivery by direct lumbar puncture","volume":"6","author":"Vulchanova","year":"2010","journal-title":"Mol. Pain"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"596","DOI":"10.1038\/s41386-024-01993-1","article-title":"Role of serotonin neurons in the dorsal raphe nucleus in heroin self-administration and punishment","volume":"50","author":"Li","year":"2025","journal-title":"Neuropsychopharmacology"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"13","DOI":"10.1016\/j.celrep.2019.08.083","article-title":"Dissecting the Roles of GABA and Neuropeptides from Rat Central Amygdala CRF Neurons in Anxiety and Fear Learning","volume":"29","author":"Pomrenze","year":"2019","journal-title":"Cell Rep."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"1030","DOI":"10.1523\/JNEUROSCI.2143-18.2018","article-title":"A Corticotropin Releasing Factor Network in the Extended Amygdala for Anxiety","volume":"39","author":"Pomrenze","year":"2019","journal-title":"J. Neurosci."},{"key":"ref_35","doi-asserted-by":"crossref","unstructured":"Hedrick, T.L. (2008). Software techniques for two- and three-dimensional kinematic measurements of biological and biomimetic systems. Bioinspir. Biomim., 3.","DOI":"10.1088\/1748-3182\/3\/3\/034001"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1281","DOI":"10.1038\/s41593-018-0209-y","article-title":"DeepLabCut: Markerless pose estimation of user-defined body parts with deep learning","volume":"21","author":"Mathis","year":"2018","journal-title":"Nat Neurosci."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"2152","DOI":"10.1038\/s41596-019-0176-0","article-title":"Using DeepLabCut for 3D markerless pose estimation across species and behaviors","volume":"14","author":"Nath","year":"2019","journal-title":"Nat Protoc."},{"key":"ref_38","doi-asserted-by":"crossref","unstructured":"He, K., Zhang, X., Ren, S., and Sun, J. (2015). Deep Residual Learning for Image Recognition. arXiv.","DOI":"10.1109\/CVPR.2016.90"},{"key":"ref_39","doi-asserted-by":"crossref","unstructured":"Insafutdinov, E., Pishchulin, L., Andres, B., Andriluka, M., and Schiele, B. (2016). DeeperCut: A Deeper, Stronger, and Faster Multi-Person Pose Estimation Model. arXiv.","DOI":"10.1007\/978-3-319-46466-4_3"},{"key":"ref_40","doi-asserted-by":"crossref","unstructured":"Biewener, A., and Patek, S. (2018). Animal Locomotion, Oxford University Press. [2nd ed.].","DOI":"10.1093\/oso\/9780198743156.001.0001"},{"key":"ref_41","unstructured":"Pinheiro, J., Bates, D., and R-Core (2025, July 16). nlme Linear and Nonlinear Mixed Effects Models. R Package Version 3.1-153.\u2014References\u2014Scientific Research Publishing. Available online: https:\/\/www.scirp.org\/reference\/referencespapers?referenceid=3610088."},{"key":"ref_42","doi-asserted-by":"crossref","unstructured":"Prat-Ortega, G., Ensel, S., Donadio, S., Borda, L., Boos, A., Yadav, P., Verma, N., Ho, J., Frazier-Kim, S., and Fields, D.P. (2024). Targeted Stimulation of the Sensory Afferents Improves Motoneuron Function in Humans with Spinal Muscular Atrophy. medRxiv.","DOI":"10.1101\/2024.02.14.24302709"},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"814","DOI":"10.1016\/j.neuron.2016.01.009","article-title":"Mechanisms Underlying the Neuromodulation of Spinal Circuits for Correcting Gait and Balance Deficits after Spinal Cord Injury","volume":"89","author":"Moraud","year":"2016","journal-title":"Neuron"},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"251","DOI":"10.1016\/j.asd.2004.06.003","article-title":"Mechanical aspects of legged locomotion control","volume":"33","author":"Koditschek","year":"2004","journal-title":"Arthropod Struct. Dev."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"641","DOI":"10.1113\/jphysiol.2006.118711","article-title":"Modelling spinal circuitry involved in locomotor pattern generation: Insights from the effects of afferent stimulation","volume":"577","author":"Rybak","year":"2006","journal-title":"J. Physiol."},{"key":"ref_46","first-page":"e20220615","article-title":"Coordinated neurostimulation promotes circuit rewiring and unlocks recovery after spinal cord injury","volume":"220","author":"Burattini","year":"2022","journal-title":"J. Exp Med."},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"114496","DOI":"10.1016\/j.expneurol.2023.114496","article-title":"Spinal control of locomotion before and after spinal cord injury","volume":"368","author":"Danner","year":"2023","journal-title":"Exp. Neurol."}],"container-title":["Bioengineering"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2306-5354\/12\/10\/1080\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,11,19]],"date-time":"2025-11-19T09:28:31Z","timestamp":1763544511000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2306-5354\/12\/10\/1080"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,10,4]]},"references-count":47,"journal-issue":{"issue":"10","published-online":{"date-parts":[[2025,10]]}},"alternative-id":["bioengineering12101080"],"URL":"https:\/\/doi.org\/10.3390\/bioengineering12101080","relation":{},"ISSN":["2306-5354"],"issn-type":[{"value":"2306-5354","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,10,4]]}}}