{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,30]],"date-time":"2026-07-30T14:06:35Z","timestamp":1785420395006,"version":"3.56.0"},"reference-count":36,"publisher":"MDPI AG","issue":"9","license":[{"start":{"date-parts":[[2026,4,25]],"date-time":"2026-04-25T00:00:00Z","timestamp":1777075200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100021037","name":"Taipei City Hospital","doi-asserted-by":"crossref","award":["TPCH-110-69"],"award-info":[{"award-number":["TPCH-110-69"]}],"id":[{"id":"10.13039\/100021037","id-type":"DOI","asserted-by":"crossref"}]},{"award":["TPCH-110-69"],"award-info":[{"award-number":["TPCH-110-69"]}],"id":[{"id":"https:\/\/ror.org\/02gzfb532","id-type":"ROR","asserted-by":"publisher"}]},{"name":"An Nan Hospital, China Medical University","award":["ANHRF111-50"],"award-info":[{"award-number":["ANHRF111-50"]}]},{"name":"National Science and Technology Council","award":["NSTC 112-2221-E-039-002-MY3"],"award-info":[{"award-number":["NSTC 112-2221-E-039-002-MY3"]}]},{"name":"National Science and Technology Council","award":["NSTC 114-2320-B-039-040"],"award-info":[{"award-number":["NSTC 114-2320-B-039-040"]}]},{"award":["NSTC 112-2221-E-039-002-MY3"],"award-info":[{"award-number":["NSTC 112-2221-E-039-002-MY3"]}],"id":[{"id":"https:\/\/ror.org\/00wnb9798","id-type":"ROR","asserted-by":"publisher"}]},{"award":["NSTC 114-2320-B-039-040"],"award-info":[{"award-number":["NSTC 114-2320-B-039-040"]}],"id":[{"id":"https:\/\/ror.org\/00wnb9798","id-type":"ROR","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Peripheral nerve injuries involving critical-sized gaps remain a major clinical challenge. Although autologous nerve grafting is considered the gold standard for peripheral nerve repair, its clinical application is limited by the availability of donor nerve tissue and the risk of donor-site morbidity, including sensory deficits and functional impairment. Therefore, nerve guidance conduits (NGCs) have emerged as a promising alternative when combined with bioactive modulation strategies. In this study, we evaluated bisvinyl sulfonemethyl (BVSM)-crosslinked gelatin conduits integrated with electrical stimulation (ES) at different frequencies (0, 2, 20, and 200 Hz) in a rat sciatic nerve defect model over a 4-week recovery period (n = 10 per group). Structural regeneration was assessed by morphometric analysis, electrophysiology, macrophage infiltration, CGRP immunoreactivity, retrograde Fluorogold tracing, quantitative PCR of growth factors and inflammatory cytokines, and behavioral testing. Among all stimulation paradigms, low-frequency ES at 2 Hz produced the most pronounced regenerative effects. The 2 Hz group demonstrated significantly greater axon number, axonal density, and regenerated nerve area compared with control and high-frequency groups (p &lt; 0.05). Electrophysiological assessments revealed improved nerve conduction velocity, higher MAP amplitudes, and shorter latencies. Enhanced macrophage recruitment and elevated CGRP expression were observed, suggesting coordinated neuroimmune and neurochemical activation. Gene expression analysis indicated upregulation of neurotrophic factors and balanced inflammatory cytokine responses under low-frequency stimulation. In contrast, high-frequency stimulation (200 Hz) failed to enhance overall regeneration and showed reduced axonal metrics, suggesting possible overstimulation-associated suppression. Collectively, these findings demonstrate that BVSM-crosslinked conduits provide a stable and biocompatible regenerative scaffold, and that appropriately tuned low-frequency electrical stimulation (2 Hz) optimally enhances structural, molecular, and functional recovery. The integration of material engineering with bioelectrical modulation represents a promising strategy for next-generation bioelectronic interfaces in peripheral nerve repair.<\/jats:p>","DOI":"10.3390\/ijms27093820","type":"journal-article","created":{"date-parts":[[2026,4,28]],"date-time":"2026-04-28T11:33:37Z","timestamp":1777376017000},"page":"3820","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Low-Frequency Electrical Stimulation Optimizes Neurotrophic and Neuroimmune Signaling in Bisvinyl Sulfonemethyl-Based Nerve Guidance Conduits"],"prefix":"10.3390","volume":"27","author":[{"given":"Ching-Feng","family":"Su","sequence":"first","affiliation":[{"name":"Graduate Institute of Acupuncture Science, Graduate Institute of Integrated Medicine, School of Chinese Medicine, Department of Biomedical Engineering, China Medical University, Taichung 404, Taiwan"},{"name":"Division of Family Medicine, Pingtung Christian Hospital, Pingtung 900, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Chung-Chia","family":"Chen","sequence":"additional","affiliation":[{"name":"Linsen Chinese Medicine and Kunming Branch, Taipei City Hospital, Taipei 103, Taiwan"},{"name":"School of Chinese Medicine for Post Baccalaureate, I-Shou University, Kaohsiung City 840, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wei-Cheng","family":"Hsu","sequence":"additional","affiliation":[{"name":"Graduate Institute of Acupuncture Science, Graduate Institute of Integrated Medicine, School of Chinese Medicine, Department of Biomedical Engineering, China Medical University, Taichung 404, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ming-Hsuan","family":"Lu","sequence":"additional","affiliation":[{"name":"Graduate Institute of Acupuncture Science, Graduate Institute of Integrated Medicine, School of Chinese Medicine, Department of Biomedical Engineering, China Medical University, Taichung 404, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Joanna Pi-Jung","family":"Lee","sequence":"additional","affiliation":[{"name":"Division of Plastic Surgery, Department of Surgery, An-Nan Hospital, China Medical University, Tainan 709, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8756-5113","authenticated-orcid":false,"given":"Yung-Hsiang","family":"Chen","sequence":"additional","affiliation":[{"name":"Graduate Institute of Acupuncture Science, Graduate Institute of Integrated Medicine, School of Chinese Medicine, Department of Biomedical Engineering, China Medical University, Taichung 404, Taiwan"},{"name":"Department of Psychology, College of Medical and Health Science, Department of Bioinformatics and Medical Engineering, Asia University, Taichung 413, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yueh-Sheng","family":"Chen","sequence":"additional","affiliation":[{"name":"Graduate Institute of Acupuncture Science, Graduate Institute of Integrated Medicine, School of Chinese Medicine, Department of Biomedical Engineering, China Medical University, Taichung 404, Taiwan"},{"name":"Department of Psychology, College of Medical and Health Science, Department of Bioinformatics and Medical Engineering, Asia University, Taichung 413, Taiwan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,25]]},"reference":[{"key":"ref_1","first-page":"586","article-title":"Synergistic effects of electrical and chemical cues with biodegradable scaffolds for large peripheral nerve defect regeneration","volume":"49","author":"Bordett","year":"2025","journal-title":"Bioact. Mater."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"3498","DOI":"10.1021\/acsabm.4c02000","article-title":"Development of Electrospun Nerve Guidance Conduits by a Milk-Derived Protein with Biodegradable Polymers for Peripheral Nerve Regeneration","volume":"8","author":"Kim","year":"2025","journal-title":"ACS Appl. Bio Mater."},{"key":"ref_3","doi-asserted-by":"crossref","unstructured":"Zhang, S., Sun, X., Yang, X., Fan, Y., Liang, Y., Li, J., and Ling, J. (2025). Research progress on composite nerve guidance conduits with immune-regulatory functions. Front. Immunol., 16.","DOI":"10.3389\/fimmu.2025.1622508"},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"e25633","DOI":"10.4161\/biom.25633","article-title":"Mechanical and biocompatible characterization of a cross-linked collagen-hyaluronic acid wound dressing","volume":"3","author":"Kirk","year":"2013","journal-title":"Biomatter"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"541","DOI":"10.1177\/0885328218804338","article-title":"Fabrication and characterization of 3D microtubular collagen scaffolds for peripheral nerve repair","volume":"33","author":"Li","year":"2018","journal-title":"J. Biomater. Appl."},{"key":"ref_6","doi-asserted-by":"crossref","unstructured":"Sapula, P., Bialik-Was, K., and Malarz, K. (2023). Are Natural Compounds a Promising Alternative to Synthetic Cross-Linking Agents in the Preparation of Hydrogels?. Pharmaceutics, 15.","DOI":"10.3390\/pharmaceutics15010253"},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"32","DOI":"10.1007\/s10856-021-06570-2","article-title":"A comprehensive review on methods for promotion of mechanical features and biodegradation rate in amniotic membrane scaffolds","volume":"33","author":"Sarvari","year":"2022","journal-title":"J. Mater. Sci. Mater. Med."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"17489","DOI":"10.1038\/s41598-017-17792-2","article-title":"Biodegradable Bisvinyl Sulfonemethyl-crosslinked Gelatin Conduit Promotes Regeneration after Peripheral Nerve Injury in Adult Rats","volume":"7","author":"Ko","year":"2017","journal-title":"Sci. Rep."},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Viezuina, D.M., Musa, I., Aldea, M., Matache, I.M., Rotaru Zavaleanu, A.D., Gresita, A., Veronica, S., and Mitran, S.I. (2025). Gelatin-Based Hydrogels for Peripheral Nerve Regeneration: A Multifunctional Vehicle for Cellular, Molecular, and Pharmacological Therapy. Gels, 11.","DOI":"10.3390\/gels11070490"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1584","DOI":"10.1007\/s10439-019-02259-4","article-title":"DC Electric Fields Induce Perpendicular Alignment and Enhanced Migration in Schwann Cell Cultures","volume":"47","author":"Bunn","year":"2019","journal-title":"Ann. Biomed. Eng."},{"key":"ref_11","doi-asserted-by":"crossref","unstructured":"Gordon, T. (2024). Brief Electrical Stimulation Promotes Recovery after Surgical Repair of Injured Peripheral Nerves. Int. J. Mol. Sci., 25.","DOI":"10.3390\/ijms25010665"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"113397","DOI":"10.1016\/j.expneurol.2020.113397","article-title":"Electrical stimulation to enhance peripheral nerve regeneration: Update in molecular investigations and clinical translation","volume":"332","author":"Zuo","year":"2020","journal-title":"Exp. Neurol."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"101755","DOI":"10.1016\/j.mtbio.2025.101755","article-title":"Conductive MeCbl\/PEDOT:PSS\/HA hydrogels with electrical stimulation for enhanced peripheral nerve regeneration","volume":"32","author":"Liu","year":"2025","journal-title":"Mater. Today Bio"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"3553","DOI":"10.1113\/JP270892","article-title":"The use of brief post-surgical low frequency electrical stimulation to enhance nerve regeneration in clinical practice","volume":"594","author":"Chan","year":"2016","journal-title":"J. Physiol."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"46460","DOI":"10.1021\/acsami.3c08070","article-title":"Machine Learning Integrated Workflow for Predicting Schwann Cell Viability on Conductive MXene Biointerfaces","volume":"15","author":"Chung","year":"2023","journal-title":"ACS Appl. Mater. Interfaces"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"758","DOI":"10.1002\/glia.24309","article-title":"Electrical stimulation accelerates Wallerian degeneration and promotes nerve regeneration after sciatic nerve injury","volume":"71","author":"Li","year":"2023","journal-title":"Glia"},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"8516","DOI":"10.1021\/la0605384","article-title":"Gelatin hydrogels cross-linked with bisvinyl sulfonemethyl. 2. The physical and chemical networks","volume":"22","author":"Djabourov","year":"2006","journal-title":"Langmuir"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"1248","DOI":"10.1016\/j.neurom.2021.12.009","article-title":"Transcutaneous Electrical Nerve Stimulation in Nerve Regeneration: A Systematic Review of In Vivo Animal Model Studies","volume":"25","author":"Alarcon","year":"2022","journal-title":"Neuromodulation"},{"key":"ref_19","first-page":"572949","article-title":"The parameters of transcutaneous electrical nerve stimulation are critical to its regenerative effects when applied just after a sciatic crush lesion in mice","volume":"2014","author":"Martinez","year":"2014","journal-title":"Biomed. Res. Int."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"102525","DOI":"10.1016\/j.hansur.2025.102525","article-title":"Electrical stimulation in peripheral nerve injuries: A review of current literature","volume":"44","author":"Johnson","year":"2025","journal-title":"Hand Surg. Rehabil."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"347","DOI":"10.1016\/j.expneurol.2007.01.040","article-title":"Electrical stimulation promotes sensory neuron regeneration and growth-associated gene expression","volume":"205","author":"Geremia","year":"2007","journal-title":"Exp. Neurol."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"379","DOI":"10.1023\/B:CEMN.0000022770.66463.f7","article-title":"Electrical stimulation accelerates and enhances expression of regeneration-associated genes in regenerating rat femoral motoneurons","volume":"24","author":"Tam","year":"2004","journal-title":"Cell Mol. Neurobiol."},{"key":"ref_23","doi-asserted-by":"crossref","unstructured":"Liao, C.F., Hsu, S.T., Chen, C.C., Yao, C.H., Lin, J.H., Chen, Y.H., and Chen, Y.S. (2020). Effects of Electrical Stimulation on Peripheral Nerve Regeneration in a Silicone Rubber Conduit in Taxol-Treated Rats. Materials, 13.","DOI":"10.3390\/ma13051063"},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"e37880","DOI":"10.1002\/jbm.a.37880","article-title":"Liquid Metal-Based Conductive Nerve Guidance Conduit Combined With Electrical Stimulation Boosts Peripheral Nerve Repair","volume":"113","author":"Zhu","year":"2025","journal-title":"J. Biomed. Mater. Res. A"},{"key":"ref_25","first-page":"168","article-title":"Thirty minutes of low intensity electrical stimulation promotes nerve regeneration after sciatic nerve crush injury in a rat model","volume":"110","author":"Alrashdan","year":"2010","journal-title":"Acta Neurol. Belg."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"337","DOI":"10.1016\/j.hcl.2024.03.001","article-title":"Physiology of Nerve Regeneration: Key Factors Affecting Clinical Outcomes","volume":"40","author":"Gordon","year":"2024","journal-title":"Hand Clin."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"490","DOI":"10.1177\/1545968315604399","article-title":"Electrical Stimulation to Promote Peripheral Nerve Regeneration","volume":"30","author":"Willand","year":"2016","journal-title":"Neurorehabil. Neural Repair"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"1257","DOI":"10.1109\/TBME.2015.2492998","article-title":"Stimulation Frequency Alters the Dorsal Root Ganglion Neurite Growth and Directionality In Vitro","volume":"63","author":"Kumar","year":"2016","journal-title":"IEEE Trans. Biomed. Eng."},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"367","DOI":"10.1177\/1545968307313507","article-title":"Effects of electrical stimulation at different frequencies on regeneration of transected peripheral nerve","volume":"22","author":"Lu","year":"2008","journal-title":"Neurorehabil. Neural Repair"},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2602","DOI":"10.1523\/JNEUROSCI.20-07-02602.2000","article-title":"Brief electrical stimulation promotes the speed and accuracy of motor axonal regeneration","volume":"20","author":"Neumann","year":"2000","journal-title":"J. Neurosci."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"35","DOI":"10.1186\/s12984-015-0022-8","article-title":"The impact of the stimulation frequency on closed-loop control with electrotactile feedback","volume":"12","author":"Paredes","year":"2015","journal-title":"J. Neuroeng. Rehabil."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"605","DOI":"10.1007\/s00401-015-1482-4","article-title":"Role of macrophages in Wallerian degeneration and axonal regeneration after peripheral nerve injury","volume":"130","author":"Chen","year":"2015","journal-title":"Acta Neuropathol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"e16281","DOI":"10.1111\/jnc.16281","article-title":"Neuronal TRPV1-CGRP axis regulates peripheral nerve regeneration through ERK\/HIF-1 signaling pathway","volume":"169","author":"Che","year":"2025","journal-title":"J. Neurochem."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"1344","DOI":"10.1038\/s41598-024-52029-z","article-title":"Calcitonin gene-related peptide regulates periodontal tissue regeneration","volume":"14","author":"Miki","year":"2024","journal-title":"Sci. Rep."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"115","DOI":"10.1016\/S0165-0270(01)00532-5","article-title":"On the use of fast blue, fluoro-gold and diamidino yellow for retrograde tracing after peripheral nerve injury: Uptake, fading, dye interactions, and toxicity","volume":"115","author":"Navarro","year":"2002","journal-title":"J. Neurosci. Methods"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"279","DOI":"10.1002\/jor.20986","article-title":"Low affinity NGF receptor (p75 neurotrophin receptor) inhibitory antibody reduces pain behavior and CGRP expression in DRG in the mouse sciatic nerve crush model","volume":"28","author":"Fukui","year":"2010","journal-title":"J. Orthop. Res."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/9\/3820\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,4,29]],"date-time":"2026-04-29T04:23:11Z","timestamp":1777436591000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/27\/9\/3820"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,4,25]]},"references-count":36,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2026,5]]}},"alternative-id":["ijms27093820"],"URL":"https:\/\/doi.org\/10.3390\/ijms27093820","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,4,25]]}}}