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SMases can be classified as Class I or II based on their structural characteristics. Class I exhibits greater dermonecrotic activity than Class II; however, the intracellular mechanisms responsible for this difference remain poorly understood. The differential transcriptomics analysis of human keratinocytes treated with each toxin revealed that Class I primarily activates pathways associated with proteolytic activity and apoptosis. In contrast, Class II uniquely upregulates key genes, including PIM-1, MCL-1, PAI-1, p21, and c-FOS, which support cell survival and inhibit apoptosis. These pathways also facilitate tissue repair and keratinocyte proliferation during wound healing, particularly through signaling mechanisms involving Substance P and VEGF-A. RT-qPCR confirmed these findings, with protein level evaluations indicating the sustained upregulation of VEGF-A exclusively in keratinocytes treated with Class II. We identified Substance P and VEGF-A as potential therapeutic targets for managing cutaneous loxoscelism, providing valuable insights into the cellular mechanisms underlying the distinct toxic effects of the two SMase D isoforms. By elucidating these pathways, this study enhances our understanding of loxoscelism\u2019s pathophysiology and highlights strategies for therapeutic intervention in dermonecrotic injuries caused by spider venom.<\/jats:p>","DOI":"10.3390\/ijms26073012","type":"journal-article","created":{"date-parts":[[2025,3,26]],"date-time":"2025-03-26T04:17:14Z","timestamp":1742962634000},"page":"3012","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":4,"title":["Differential Cellular Responses to Class I and II Sphingomyelinase D: Unraveling the Mechanisms of Loxosceles Venom-Induced Dermonecrosis and Potential Therapeutic Targets"],"prefix":"10.3390","volume":"26","author":[{"given":"Bruna Fernandes","family":"Pinto","sequence":"first","affiliation":[{"name":"Immunochemistry Laboratory, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6670-2564","authenticated-orcid":false,"given":"Priscila Hess","family":"Lopes","sequence":"additional","affiliation":[{"name":"Immunochemistry Laboratory, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"},{"name":"PREVOR, Rue des Chasseurs-Ardennais 3, 4031 Li\u00e8ge, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1216-0418","authenticated-orcid":false,"given":"Carlos Eduardo Madureira","family":"Trufen","sequence":"additional","affiliation":[{"name":"PREVOR, Rue des Chasseurs-Ardennais 3, 4031 Li\u00e8ge, Belgium"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0004-2626-7894","authenticated-orcid":false,"given":"Ana Tung","family":"Ching Ching","sequence":"additional","affiliation":[{"name":"Immunochemistry Laboratory, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"In\u00e1cio de Loyola M.","family":"Junqueira de Azevedo","sequence":"additional","affiliation":[{"name":"Laboratoy of Applied Toxinology, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2410-0562","authenticated-orcid":false,"given":"Milton Yutaka","family":"Nishiyama-Jr","sequence":"additional","affiliation":[{"name":"Laboratoy of Applied Toxinology, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marcelo Medina","family":"de Souza","sequence":"additional","affiliation":[{"name":"Centre of Excellence in New Target Discovery, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8569-8150","authenticated-orcid":false,"given":"Paula C.","family":"Pohl","sequence":"additional","affiliation":[{"name":"Immunochemistry Laboratory, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1896-9074","authenticated-orcid":false,"given":"Denise V.","family":"Tambourgi","sequence":"additional","affiliation":[{"name":"Immunochemistry Laboratory, Butantan Institute, Avenue Vital Brasil, 1500, Butant\u00e3, S\u00e3o Paulo 05503-900, Brazil"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,3,26]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"546","DOI":"10.1016\/S0035-9203(98)90909-9","article-title":"A Clinical and Epidemiological Study of Loxosceles Spider Envenoming in Santa Catarina, Brazil","volume":"92","author":"Sezerino","year":"1998","journal-title":"Trans. R. Soc. Trop. Med. Hyg."},{"key":"ref_2","first-page":"e2023568","article-title":"Brown Spiders (Loxosceles) Are Taking Hold in Pernambuco, Brazil: A Case Series, 2018\u20132022","volume":"33","author":"Amorim","year":"2024","journal-title":"Epidemiol. E Servi\u00e7os De Sa\u00fade"},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"463","DOI":"10.4269\/ajtmh.2008.79.463","article-title":"A New Anti-Loxoscelic Serum Produced against Recombinant Sphingomyelinase D: Results of Preclinical Trials","volume":"79","author":"Marcelino","year":"2008","journal-title":"Am. J. Trop. Med. Hyg."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"743","DOI":"10.1016\/j.toxicon.2009.02.013","article-title":"SMase II, a New Sphingomyelinase D from Loxosceles Laeta Venom Gland: Molecular Cloning, Expression, Function and Structural Analysis","volume":"53","author":"Portaro","year":"2009","journal-title":"Toxicon"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"451","DOI":"10.3109\/15563658308990434","article-title":"Loxoscelism and Necrotic Arachnidism","volume":"21","author":"Wasserman","year":"1983","journal-title":"J. Toxicol. Clin. Toxicol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"261","DOI":"10.1097\/00000441-199210000-00008","article-title":"Loxoscelism","volume":"304","author":"Futrell","year":"1992","journal-title":"Am. J. Med. Sci."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"683","DOI":"10.1182\/blood.V95.2.683","article-title":"Loxosceles Intermedia Spider Envenomation Induces Activation of an Endogenous Metalloproteinase, Resulting in Cleavage of Glycophorins from the Erythrocyte Surface and Facilitating Complement-Mediated Lysis","volume":"95","author":"Tambourgi","year":"2000","journal-title":"Blood"},{"key":"ref_8","first-page":"75","article-title":"Necrotic Arachnidism","volume":"38","author":"Mackinnon","year":"1953","journal-title":"An. Fac. Med. Univ. Repub. Montev. Urug."},{"key":"ref_9","first-page":"34","article-title":"A histopathological study on necrotic arachnidism by Loxosceles laeta (author\u2019s transl)","volume":"30","author":"Pizzi","year":"1975","journal-title":"Bol. Chil. Parasitol."},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"4459","DOI":"10.4049\/jimmunol.155.9.4459","article-title":"Incorporation of a 35-Kilodalton Purified Protein from Loxosceles Intermedia Spider Venom Transforms Human Erythrocytes into Activators of Autologous Complement Alternative Pathway","volume":"155","author":"Tambourgi","year":"1995","journal-title":"J. Immunol."},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"366","DOI":"10.1006\/bbrc.1998.9474","article-title":"Sphingomyelinases in the Venom of the Spider Loxosceles Intermedia Are Responsible for Both Dermonecrosis and Complement-Dependent Hemolysis","volume":"251","author":"Tambourgi","year":"1998","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"638","DOI":"10.1016\/S0006-291X(02)02521-4","article-title":"Molecular Cloning and Expression of a Functional Dermonecrotic and Haemolytic Factor from Loxosceles Laeta Venom","volume":"298","author":"Ramos","year":"2002","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"831","DOI":"10.1016\/j.molimm.2004.03.027","article-title":"Molecular Cloning, Expression, Function and Immunoreactivities of Members of a Gene Family of Sphingomyelinases from Loxosceles Venom Glands","volume":"41","author":"Tambourgi","year":"2004","journal-title":"Mol. Immunol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"13658","DOI":"10.1074\/jbc.M412437200","article-title":"Structural Basis for Metal Ion Coordination and the Catalytic Mechanism of Sphingomyelinases D","volume":"280","author":"Murakami","year":"2005","journal-title":"J. Biol. Chem."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"323","DOI":"10.1016\/j.bbrc.2006.01.123","article-title":"Structural Insights into the Catalytic Mechanism of Sphingomyelinases D and Evolutionary Relationship to Glycerophosphodiester Phosphodiesterases","volume":"342","author":"Murakami","year":"2006","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"421","DOI":"10.1016\/j.toxicon.2004.08.022","article-title":"Variations in Loxosceles Spider Venom Composition and Toxicity Contribute to the Severity of Envenomation","volume":"45","author":"Piazza","year":"2005","journal-title":"Toxicon"},{"key":"ref_17","doi-asserted-by":"crossref","unstructured":"Pedroso, A., Matioli, S.R., Murakami, M.T., Pidde-Queiroz, G., and Tambourgi, D.V. (2015). Adaptive Evolution in the Toxicity of a Spider\u2019s Venom Enzymes. BMC Evol. Biol., 15.","DOI":"10.1186\/s12862-015-0561-4"},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"725","DOI":"10.1111\/j.0022-202X.2005.23654.x","article-title":"Loxosceles Sphingomyelinase Induces Complement-Dependent Dermonecrosis, Neutrophil Infiltration, and Endogenous Gelatinase Expression","volume":"124","author":"Tambourgi","year":"2005","journal-title":"J. Investig. Dermatol."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"61","DOI":"10.1038\/sj.jid.5700049","article-title":"Role of Matrix Metalloproteinases in HaCaT Keratinocytes Apoptosis Induced by Loxosceles Venom Sphingomyelinase D","volume":"126","author":"Tambourgi","year":"2006","journal-title":"J. Investig. Dermatol."},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"1410","DOI":"10.1038\/sj.jid.5700688","article-title":"Tetracycline Protects against Dermonecrosis Induced by Loxosceles Spider Venom","volume":"127","author":"Okamoto","year":"2007","journal-title":"J. Investig. Dermatol."},{"key":"ref_21","doi-asserted-by":"crossref","unstructured":"Corr\u00eaa, M.A., Okamoto, C.K., Gon\u00e7alves-de-Andrade, R.M., van den Berg, C.W., and Tambourgi, D.V. (2016). Sphingomyelinase D from Loxosceles Laeta Venom Induces the Expression of MMP7 in Human Keratinocytes: Contribution to Dermonecrosis. PLoS ONE, 11.","DOI":"10.1371\/journal.pone.0153090"},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"3285","DOI":"10.1007\/s00204-023-03602-4","article-title":"Role of ErbB and IL-1 Signaling Pathways in the Dermonecrotic Lesion Induced by Loxosceles Sphingomyelinases D","volume":"97","author":"Pinto","year":"2023","journal-title":"Arch. Toxicol."},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1809","DOI":"10.1016\/j.jid.2019.02.004","article-title":"SOX2 Epidermal Overexpression Promotes Cutaneous Wound Healing via Activation of EGFR\/MEK\/ERK Signaling Mediated by EGFR Ligands","volume":"139","author":"Uchiyama","year":"2019","journal-title":"J. Investig. Dermatol."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"937","DOI":"10.1038\/labinvest.3780304","article-title":"Sox18 Is Transiently Expressed during Angiogenesis in Granulation Tissue of Skin Wounds with an Identical Expression Pattern to Flk-1 mRNA","volume":"81","author":"Darby","year":"2001","journal-title":"Lab. Investig."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"225","DOI":"10.1089\/wound.2014.0581","article-title":"Metalloproteinases and Wound Healing","volume":"4","author":"Caley","year":"2015","journal-title":"Adv. Wound Care"},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"1290","DOI":"10.1046\/j.1523-1747.2002.19626.x","article-title":"Substance P Enhances the Production of Interferon-Induced Protein of 10 kDa by Human Keratinocytes in Synergy with Interferon-Gamma","volume":"119","author":"Kanda","year":"2002","journal-title":"J. Investig. Dermatol."},{"key":"ref_27","doi-asserted-by":"crossref","unstructured":"Redkiewicz, P. (2022). The Regenerative Potential of Substance P. Int. J. Mol. Sci., 23.","DOI":"10.3390\/ijms23020750"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"786","DOI":"10.1097\/WNR.0b013e3283572206","article-title":"Substance P Induces M2-Type Macrophages after Spinal Cord Injury","volume":"23","author":"Jiang","year":"2012","journal-title":"Neuroreport"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"179","DOI":"10.1016\/j.bbrc.2014.09.090","article-title":"Substance P Ameliorates Collagen II-Induced Arthritis in Mice via Suppression of the Inflammatory Response","volume":"453","author":"Hong","year":"2014","journal-title":"Biochem. Biophys. Res. Commun."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"1376","DOI":"10.1111\/j.1462-5822.2012.01802.x","article-title":"The Vascular Endothelial Growth Factor (VEGF)-E Encoded by Orf Virus Regulates Keratinocyte Proliferation and Migration and Promotes Epidermal Regeneration","volume":"14","author":"Wise","year":"2012","journal-title":"Cell Microbiol."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1023\/A:1006995023982","article-title":"Loxosceles Deserta Spider Venom Induces the Expression of Vascular Endothelial Growth Factor (VEGF) in Keratinocytes","volume":"24","author":"Desai","year":"2000","journal-title":"Inflammation"},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"1351","DOI":"10.1038\/jid.2008.363","article-title":"Mcl-1 Functions as Major Epidermal Survival Protein Required for Proper Keratinocyte Differentiation","volume":"129","author":"Sitailo","year":"2009","journal-title":"J. Investig. Dermatol."},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"495","DOI":"10.1006\/excr.1998.4359","article-title":"VEGF Prevents Apoptosis of Human Microvascular Endothelial Cells via Opposing Effects on MAPK\/ERK and SAPK\/JNK Signaling","volume":"247","author":"Gupta","year":"1999","journal-title":"Exp. Cell Res."},{"key":"ref_34","doi-asserted-by":"crossref","unstructured":"Macdonald, A., Campbell, D.G., Toth, R., McLauchlan, H., Hastie, C.J., and Arthur, J.S.C. (2006). Pim Kinases Phosphorylate Multiple Sites on Bad and Promote 14-3-3 Binding and Dissociation from Bcl-XL. BMC Cell Biol., 7.","DOI":"10.1186\/1471-2121-7-1"},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"1640","DOI":"10.1039\/c0pp00244e","article-title":"UVB-Induced p21 Degradation Promotes Apoptosis of Human Keratinocytes","volume":"9","author":"Lei","year":"2010","journal-title":"Photochem. Photobiol. Sci."},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"86","DOI":"10.12659\/MSMBR.893608","article-title":"The Role of p21 in Apoptosis, Proliferation, Cell Cycle Arrest, and Antioxidant Activity in UVB-Irradiated Human HaCaT Keratinocytes","volume":"21","author":"Chen","year":"2015","journal-title":"Med. Sci. Monit. Basic. Res."},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"321","DOI":"10.1089\/wound.2014.0611","article-title":"Targeted Inhibition of PAI-1 Activity Impairs Epithelial Migration and Wound Closure Following Cutaneous Injury","volume":"4","author":"Simone","year":"2015","journal-title":"Adv. Wound Care"},{"key":"ref_38","doi-asserted-by":"crossref","first-page":"791","DOI":"10.1111\/j.0022-202X.2004.22319.x","article-title":"Regulation of MAPK Activation, AP-1 Transcription Factor Expression and Keratinocyte Differentiation in Wounded Fetal Skin","volume":"122","author":"Gangnuss","year":"2004","journal-title":"J. Investig. Dermatol."},{"key":"ref_39","doi-asserted-by":"crossref","first-page":"23","DOI":"10.1038\/nrc2986","article-title":"For Better or for Worse: The Role of Pim Oncogenes in Tumorigenesis","volume":"11","author":"Nawijn","year":"2011","journal-title":"Nat. Rev. Cancer"},{"key":"ref_40","doi-asserted-by":"crossref","first-page":"L240","DOI":"10.1152\/ajplung.00156.2013","article-title":"Pim1 Kinase Protects Airway Epithelial Cells from Cigarette Smoke-Induced Damage and Airway Inflammation","volume":"307","author":"Heijink","year":"2014","journal-title":"Am. J. Physiol. Lung Cell Mol. Physiol."},{"key":"ref_41","doi-asserted-by":"crossref","first-page":"4536","DOI":"10.1182\/blood-2003-11-3827","article-title":"Identification of Flk-1 Target Genes in Vasculogenesis: Pim-1 Is Required for Endothelial and Mural Cell Differentiation in Vitro","volume":"103","author":"Zippo","year":"2004","journal-title":"Blood"},{"key":"ref_42","doi-asserted-by":"crossref","first-page":"2051","DOI":"10.3892\/mmr.2014.2139","article-title":"Use of Regulators and Inhibitors of Pim-1, a Serine\/threonine Kinase, for Tumour Therapy (review)","volume":"9","author":"Liang","year":"2014","journal-title":"Mol. Med. Rep."},{"key":"ref_43","doi-asserted-by":"crossref","first-page":"699","DOI":"10.1111\/1523-1747.ep12324482","article-title":"Differentiation-Associated Expression of the Proto-Oncogene Pim-1 in Cultured Human Keratinocytes","volume":"105","author":"Stewart","year":"1995","journal-title":"J. Investig. Dermatol."},{"key":"ref_44","doi-asserted-by":"crossref","first-page":"C905","DOI":"10.1152\/ajpcell.00200.2003","article-title":"Increased Vascular Endothelial Growth Factor May Account for Elevated Level of Plasminogen Activator Inhibitor-1 via Activating ERK1\/2 in Keloid Fibroblasts","volume":"286","author":"Wu","year":"2004","journal-title":"Am. J. Physiol. Cell Physiol."},{"key":"ref_45","doi-asserted-by":"crossref","first-page":"2886","DOI":"10.1182\/blood-2004-05-1760","article-title":"VEGF Induces Mcl-1 up-Regulation and Protects Multiple Myeloma Cells against Apoptosis","volume":"104","author":"Podar","year":"2004","journal-title":"Blood"},{"key":"ref_46","doi-asserted-by":"crossref","first-page":"4448","DOI":"10.1073\/pnas.1000803107","article-title":"IL-33 Augments Substance P-Induced VEGF Secretion from Human Mast Cells and Is Increased in Psoriatic Skin","volume":"107","author":"Theoharides","year":"2010","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_47","doi-asserted-by":"crossref","first-page":"400","DOI":"10.1038\/nrc2657","article-title":"p21 in Cancer: Intricate Networks and Multiple Activities","volume":"9","author":"Abbas","year":"2009","journal-title":"Nat. Rev. Cancer"},{"key":"ref_48","doi-asserted-by":"crossref","first-page":"207","DOI":"10.1111\/j.0022-202X.2004.22702.x","article-title":"AKT Status Controls Susceptibility of Malignant Keratinocytes to the Early-Activated and UVB-Induced Apoptotic Pathway","volume":"123","author":"Decraene","year":"2004","journal-title":"J. Investig. Dermatol."},{"key":"ref_49","doi-asserted-by":"crossref","first-page":"265","DOI":"10.1016\/S0021-9258(19)52451-6","article-title":"Protein Measurement with the Folin Phenol Reagent","volume":"193","author":"Lowry","year":"1951","journal-title":"J. Biol. Chem."},{"key":"ref_50","doi-asserted-by":"crossref","first-page":"680","DOI":"10.1038\/227680a0","article-title":"Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4","volume":"227","author":"Laemmli","year":"1970","journal-title":"Nature"},{"key":"ref_51","doi-asserted-by":"crossref","first-page":"1","DOI":"10.2174\/1875036201307010001","article-title":"Comparison of Sequencing Utility Programs","volume":"7","author":"Aronesty","year":"2013","journal-title":"Open Bioinform. J."},{"key":"ref_52","doi-asserted-by":"crossref","unstructured":"Langmead, B., Trapnell, C., Pop, M., and Salzberg, S.L. (2009). Ultrafast and Memory-Efficient Alignment of Short DNA Sequences to the Human Genome. Genome Biol., 10.","DOI":"10.1186\/gb-2009-10-3-r25"},{"key":"ref_53","doi-asserted-by":"crossref","first-page":"15","DOI":"10.1093\/bioinformatics\/bts635","article-title":"STAR: Ultrafast Universal RNA-Seq Aligner","volume":"29","author":"Dobin","year":"2013","journal-title":"Bioinformatics"},{"key":"ref_54","doi-asserted-by":"crossref","first-page":"923","DOI":"10.1093\/bioinformatics\/btt656","article-title":"featureCounts: An Efficient General Purpose Program for Assigning Sequence Reads to Genomic Features","volume":"30","author":"Liao","year":"2014","journal-title":"Bioinformatics"},{"key":"ref_55","doi-asserted-by":"crossref","first-page":"896","DOI":"10.1038\/nbt.2931","article-title":"Normalization of RNA-Seq Data Using Factor Analysis of Control Genes or Samples","volume":"32","author":"Risso","year":"2014","journal-title":"Nat. Biotechnol."},{"key":"ref_56","doi-asserted-by":"crossref","first-page":"139","DOI":"10.1093\/bioinformatics\/btp616","article-title":"edgeR: A Bioconductor Package for Differential Expression Analysis of Digital Gene Expression Data","volume":"26","author":"Robinson","year":"2010","journal-title":"Bioinformatics"},{"key":"ref_57","doi-asserted-by":"crossref","unstructured":"Robinson, M.D., and Oshlack, A. (2010). A Scaling Normalization Method for Differential Expression Analysis of RNA-Seq Data. Genome Biol., 11.","DOI":"10.1186\/gb-2010-11-3-r25"},{"key":"ref_58","doi-asserted-by":"crossref","unstructured":"Chen, Y., Lun, A.T.L., and Smyth, G.K. (2014). Differential Expression Analysis of Complex RNA-Seq Experiments Using edgeR. Statistical Analysis of Next Generation Sequencing Data, Springer International Publishing.","DOI":"10.1007\/978-3-319-07212-8_3"},{"key":"ref_59","doi-asserted-by":"crossref","first-page":"946","DOI":"10.1214\/16-AOAS920","article-title":"Robust Hyperparameter Estimation Protects Against Hypervariable Genes And Improves Power To Detect Differential Expression","volume":"10","author":"Phipson","year":"2016","journal-title":"Ann. Appl. Stat."},{"key":"ref_60","doi-asserted-by":"crossref","first-page":"4288","DOI":"10.1093\/nar\/gks042","article-title":"Differential Expression Analysis of Multifactor RNA-Seq Experiments with Respect to Biological Variation","volume":"40","author":"McCarthy","year":"2012","journal-title":"Nucleic Acids Res."},{"key":"ref_61","doi-asserted-by":"crossref","first-page":"289","DOI":"10.1111\/j.2517-6161.1995.tb02031.x","article-title":"Controlling the False Discovery Rate: A Practical and Powerful Approach to Multiple Testing","volume":"57","author":"Benjamini","year":"1995","journal-title":"J. R. Stat. Soc."},{"key":"ref_62","doi-asserted-by":"crossref","first-page":"15545","DOI":"10.1073\/pnas.0506580102","article-title":"Gene Set Enrichment Analysis: A Knowledge-Based Approach for Interpreting Genome-Wide Expression Profiles","volume":"102","author":"Subramanian","year":"2005","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_63","doi-asserted-by":"crossref","first-page":"D258","DOI":"10.1093\/nar\/gkh036","article-title":"The Gene Ontology (GO) Database and Informatics Resource","volume":"32","author":"Harris","year":"2004","journal-title":"Nucleic Acids Res."},{"key":"ref_64","doi-asserted-by":"crossref","first-page":"D605","DOI":"10.1093\/nar\/gkaa1074","article-title":"The STRING Database in 2021: Customizable Protein-Protein Networks, and Functional Characterization of User-Uploaded Gene\/measurement Sets","volume":"49","author":"Szklarczyk","year":"2021","journal-title":"Nucleic Acids Res"},{"key":"ref_65","doi-asserted-by":"crossref","first-page":"2498","DOI":"10.1101\/gr.1239303","article-title":"Cytoscape: A Software Environment for Integrated Models of Biomolecular Interaction Networks","volume":"13","author":"Shannon","year":"2003","journal-title":"Genome Res."},{"key":"ref_66","doi-asserted-by":"crossref","first-page":"W90","DOI":"10.1093\/nar\/gkw377","article-title":"Enrichr: A Comprehensive Gene Set Enrichment Analysis Web Server 2016 Update","volume":"44","author":"Kuleshov","year":"2016","journal-title":"Nucleic Acids Res."},{"key":"ref_67","doi-asserted-by":"crossref","first-page":"e45","DOI":"10.1093\/nar\/29.9.e45","article-title":"A New Mathematical Model for Relative Quantification in Real-Time RT-PCR","volume":"29","author":"Pfaffl","year":"2001","journal-title":"Nucleic Acids Res."},{"key":"ref_68","doi-asserted-by":"crossref","unstructured":"Shapiro, S.S., and Wilk, M.B. (1965). An Analysis of Variance Test for Normality (complete Samples). Biometrika, 52.","DOI":"10.2307\/2333709"},{"key":"ref_69","doi-asserted-by":"crossref","first-page":"W566","DOI":"10.1093\/nar\/gkv468","article-title":"ClustVis: A Web Tool for Visualizing Clustering of Multivariate Data Using Principal Component Analysis and Heatmap","volume":"43","author":"Metsalu","year":"2015","journal-title":"Nucleic Acids Res."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/7\/3012\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,9]],"date-time":"2025-10-09T17:00:19Z","timestamp":1760029219000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/26\/7\/3012"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2025,3,26]]},"references-count":69,"journal-issue":{"issue":"7","published-online":{"date-parts":[[2025,4]]}},"alternative-id":["ijms26073012"],"URL":"https:\/\/doi.org\/10.3390\/ijms26073012","relation":{},"ISSN":["1422-0067"],"issn-type":[{"value":"1422-0067","type":"electronic"}],"subject":[],"published":{"date-parts":[[2025,3,26]]}}}