Journal: Liver International

Article Title: Curcumin Targets Crispld2 to Suppress Hepatic Stellate Cell Activation via PI3K / AKT Pathway Inhibition in Hepatic Fibrosis

doi: 10.1111/liv.70696

Figure Lengend Snippet: Curcumin suppresses HSC activation by targeting Crispld2. Animal groups: Control, CCl 4 , CCl 4 + Cur (25 mg/kg), CCl 4 + Cur (50 mg/kg); n = 5. (A) QRT‐PCR measuring the mRNA level of Crispld2. (B) WB determining the protein level of Crispld2. Groupings of LX‐2 cells: Oe‐NC, oe‐Crispld2. (C) QRT‐PCR verifying transfection efficiency. (D) WB confirming transfection efficiency. Groupings of LX‐2 cells with curcumin treatment: Control, TGF‐β + DMSO + oe‐NC, TGF‐β + Cur + oe‐NC, and TGF‐β + Cur + oe‐Crispld2. (E) QRT‐PCR detecting the mRNA levels of Crispld2. (F) WB examining the protein levels of Crispld2. (G) CCK‐8 assay assessing cell viability. (H) Flow cytometry measuring apoptosis. (I) WB detecting the expression of fibrotic proteins α‐SMA, collagen I, fibronectin, and TIMP1. (J, K) ELISA measuring the levels of inflammatory cytokines IL‐6 (J) and TNF‐α (K). * p < 0.05.

Article Snippet: We cultured human HSC line LX‐2 (RRID: CVCL_5792; BNCC337957, BNCC, China) in RPMI‐1640 complete medium (BNCC, China) containing 10% FBS (Beyotime, China) and human embryonic kidney 293 T cells (RRID: CVCL_0063; BNCC353535, BNCC, China) in DMEM‐H complete medium (BNCC, China) containing 10% FBS and 2 mM L‐glutamine (MCE, USA).

Techniques: Activation Assay, Control, Quantitative RT-PCR, Transfection, CCK-8 Assay, Flow Cytometry, Expressing, Enzyme-linked Immunosorbent Assay

Journal: Liver International

Article Title: Curcumin Targets Crispld2 to Suppress Hepatic Stellate Cell Activation via PI3K / AKT Pathway Inhibition in Hepatic Fibrosis

doi: 10.1111/liv.70696

Figure Lengend Snippet: Curcumin treatment suppresses HSC activation by modulating the Crispld2 to mediate the PI3K/AKT axis. (A) KEGG enrichment of Crispld2 in the transitional HSC treatment group. Animal groups: Control, CCl 4 , CCl 4 + Cur (25 mg/kg), CCl 4 + Cur (50 mg/kg); n = 5. (B) WB assessing the protein levels of p‐PI3K, PI3K, p‐AKT, and AKT. Groups of LX‐2 cells treated by TGF‐β: Sh‐NC, sh‐Crispld2. (C) QRT‐PCR quantification of Crispld2 mRNA. (D) WB analysis of Crispld2, p‐PI3K, PI3K, p‐AKT, AKT. Experimental groups of LX‐2 cells with Crispld2 overexpression induced by TGF‐β, following curcumin or LY294002 treatment: Control, Cur + oe‐NC, Cur + oe‐Crispld2, Cur + oe‐Crispld2 + LY294002. ( E) Co‐IP assays confirmed the interaction of CRISPLD2 with PI3K and AKT. (F) CCK‐8 assessing cell viability. (G) Flow cytometry assessing apoptosis. (H) WB for p‐PI3K, PI3K, p‐AKT, AKT and fibrotic proteins (α‐SMA, collagen I, fibronectin, TIMP1). (I, J) ELISA detection of inflammatory cytokines IL‐6 (I) and TNF‐α (J). * p < 0.05.

Article Snippet: We cultured human HSC line LX‐2 (RRID: CVCL_5792; BNCC337957, BNCC, China) in RPMI‐1640 complete medium (BNCC, China) containing 10% FBS (Beyotime, China) and human embryonic kidney 293 T cells (RRID: CVCL_0063; BNCC353535, BNCC, China) in DMEM‐H complete medium (BNCC, China) containing 10% FBS and 2 mM L‐glutamine (MCE, USA).

Techniques: Activation Assay, Control, Quantitative RT-PCR, Over Expression, Co-Immunoprecipitation Assay, CCK-8 Assay, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Signal Transduction and Targeted Therapy

Article Title: Clinical translational research on stem cell products: prospects and challenges

doi: 10.1038/s41392-026-02582-y

Figure Lengend Snippet: Different stem cell sources. Different types of stem cells are derived from different sources: hematopoietic stem cells (HSCs) are derived from bone marrow, peripheral blood, umbilical cord/placental blood, etc.; pluripotent stem cells are further divided into embryonic stem cells and induced pluripotent stem cells; and embryonic stem cells (ESCs) are derived from the blastocyst cell mass 5–6 days after fertilization, while induced pluripotent stem cells (iPSCs) are reprogrammed from somatic cells (such as skin fibroblasts, peripheral blood mononuclear cells, and kidney epithelial cells shed in urine, etc.). Mesenchymal stem cells (MSCs) can be derived from bone marrow, the umbilical cord, the placenta, fat, menstrual blood, etc., and can also be differentiated from iPSCs. Other sources of adult stem cells are nerves (such as neural stem cells), the gut (intestinal stem cells), and muscle (muscle stem cells)

Article Snippet: Genetically modified HSCs , SKYSONA (bluebird bio, Inc.) , Cerebral adrenoleukodystrophy , America , 2022.

Techniques: Derivative Assay

Journal: Signal Transduction and Targeted Therapy

Article Title: Clinical translational research on stem cell products: prospects and challenges

doi: 10.1038/s41392-026-02582-y

Figure Lengend Snippet: Clinical trials of stem cells. a Cell therapy across various therapy types in different clinical phases. b Clinical trials of HSCs classified by disease type. c Clinical trials classified by different sources of MSCs. d Clinical trials of MSCs classified by disease type. e Clinical trials of iPSCs classified by disease type. f Clinical trials of ESCs classified by disease type. All the data were collected from www.clinicaltrials.gov . HSCs hematopoietic stem cells, MSCs mesenchymal stem cells, iPSCs induced pluripotent stem cells, ESCs embryonic stem cells, Early PhI early PhaseI, PhI PhaseI, PhII PhaseII, PhIII PhaseIII, PhVI PhaseVI

Article Snippet: Genetically modified HSCs , SKYSONA (bluebird bio, Inc.) , Cerebral adrenoleukodystrophy , America , 2022.

Techniques: Clinical Proteomics

Journal: Frontiers in Genetics

Article Title: RUNX1-mediated repression of miR-24 promotes hepatic stellate cell activation and liver fibrosis by targeting the ALK4/Smad3 signaling pathway

doi: 10.3389/fgene.2026.1825784

Figure Lengend Snippet: miR-24 is decreased in fibrotic livers and activated HSCs. (A) Representative images of mouse livers from control and CCl 4 -treated groups. Scale bar = 1 cm. (B) Representative H&E-stained images of mouse livers from control and CCl 4 -treated groups. Scale bar = 50 μm. (C) Representative Sirius red-stained images and quantification of mouse livers from control and CCl 4 -treated groups (n = 4:5). Scale bar = 200 μm. (D) Body weight, spleen weight, and spleen index of mice (n = 7:11). (E) Serum ALT and AST levels in mice (n = 7:11). (F) qRT-PCR analysis of miR-24 levels in liver samples from CCl 4 -induced mice (n = 3:8). (G) qRT-PCR analysis of α-SMA and Col1a1 expressions in HSC-T6s (n = 6 per group). (H) qRT-PCR analysis of α-SMA and Col1a1 expressions in activated primary mouse HSCs (n = 6 per group). (I) qRT-PCR analysis of miR-24 levels in HSC-T6 treated with TGF-β1 (n = 6 per group). (J) qRT-PCR analysis of miR-24 levels in activated primary mouse HSCs (n = 6 per group). (K) miR-24 expression in cirrhotic and healthy liver samples from published miRNA array data ( GSE49012 , n = 12:22). (L) qRT-PCR analysis of miR-24 levels in BRL-3A and HSC-T6 treated with TGF-β1 (n = 5:5:6:6). CCl 4 , carbon tetrachloride; HSC-T6, hepatic stellate cell-T6. Data are expressed as mean ± SD. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: The rat HSC line HSC-T6 was provided by Jiangsu KeyGEN BioTECH Co., Ltd. HEK293T was purchased from the Chinese Academy of Sciences Cell Resource Center.

Techniques: Control, Staining, Quantitative RT-PCR, Expressing

Journal: Frontiers in Genetics

Article Title: RUNX1-mediated repression of miR-24 promotes hepatic stellate cell activation and liver fibrosis by targeting the ALK4/Smad3 signaling pathway

doi: 10.3389/fgene.2026.1825784

Figure Lengend Snippet: miR-24 influences HSC activation. (A) qRT-PCR analysis of miR-24 levels in HSC-T6 transfected with the miR-24 mimic or inhibitor (n = 6 per group). (B) qRT-PCR analysis of α-SMA and Col1a1 expressions in HSC-T6s (n = 6 per group). (C) EdU staining and quantification of EdU-positive HSC-T6s (n = 3 per group). Scale bar = 50 μm. (D) Flow cytometric analysis of the cell cycle distribution in HSC-T6s (n = 6 per group). (E) Transwell migration assay of HSC-T6s (n = 3 per group). Scale bar = 100 μm. HSC-T6, hepatic stellate cell-T6. Data are expressed as the mean ± SD. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: The rat HSC line HSC-T6 was provided by Jiangsu KeyGEN BioTECH Co., Ltd. HEK293T was purchased from the Chinese Academy of Sciences Cell Resource Center.

Techniques: Activation Assay, Quantitative RT-PCR, Transfection, Staining, Transwell Migration Assay

Journal: Frontiers in Genetics

Article Title: RUNX1-mediated repression of miR-24 promotes hepatic stellate cell activation and liver fibrosis by targeting the ALK4/Smad3 signaling pathway

doi: 10.3389/fgene.2026.1825784

Figure Lengend Snippet: miR-24 regulates HSC activation by directly targeting ALK4. (A) Luciferase assay performed by transfection with the miR-24 mimic and interaction site on ALK4 3′UTR (n = 6 per group). (B) qRT-PCR analysis of ALK4 expression in CCl 4 -induced fibrotic livers and TGF-β1-induced HSC-T6s (n = 5 per group). (C) Western blot analysis of ALK4 expression in HSC-T6s (n = 6 per group). (D) qRT-PCR analysis of ALK4 expression in HSC-T6s (n = 6 per group). (E) EdU staining and quantification of EdU-positive HSC-T6s (n = 5 per group). Scale bar = 100 μm. (F) Flow cytometric analysis of the cell cycle distribution in HSC-T6s (n = 5 per group). (G) qRT-PCR analysis of α-SMA expression in HSC-T6s (n = 5:4:5:4). HSC-T6, hepatic stellate cell-T6. Data are expressed as the mean ± SD. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: The rat HSC line HSC-T6 was provided by Jiangsu KeyGEN BioTECH Co., Ltd. HEK293T was purchased from the Chinese Academy of Sciences Cell Resource Center.

Techniques: Activation Assay, Luciferase, Transfection, Quantitative RT-PCR, Expressing, Western Blot, Staining

Journal: Frontiers in Genetics

Article Title: RUNX1-mediated repression of miR-24 promotes hepatic stellate cell activation and liver fibrosis by targeting the ALK4/Smad3 signaling pathway

doi: 10.3389/fgene.2026.1825784

Figure Lengend Snippet: miR-24 suppression promotes HSC activation through Smad3-dependent mechanisms. (A,B) Western blot analysis of p-Smad3, Smad3, and TGF-β1 in HSC-T6s (n = 3 per group). (C) EdU staining and quantification of EdU-positive HSC-T6s (n = 6 per group). Scale bar = 100 μm. (D) Western blot analysis of α-SMA and Col1a1 in HSC-T6s (n = 3 per group). HSC-T6, hepatic stellate cell-T6. Data are expressed as the mean ± SD. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: The rat HSC line HSC-T6 was provided by Jiangsu KeyGEN BioTECH Co., Ltd. HEK293T was purchased from the Chinese Academy of Sciences Cell Resource Center.

Techniques: Activation Assay, Western Blot, Staining

Journal: Frontiers in Genetics

Article Title: RUNX1-mediated repression of miR-24 promotes hepatic stellate cell activation and liver fibrosis by targeting the ALK4/Smad3 signaling pathway

doi: 10.3389/fgene.2026.1825784

Figure Lengend Snippet: RUNX1 regulates miR-24 generation during HSC activation. (A) Promoter region analysis of the miR-24 host gene. (B) Western blot analysis of RUNX1 in HSC-T6s (n = 3 per group). (C) qRT-PCR analysis of RUNX1 expression in HSC-T6s (n = 6 per group). (D) qRT-PCR analysis of miR-24 expression in HSC-T6s (n = 6 per group). (E) Interaction between RUNX1 and the miR-24 promoter motif (PDB ID: 1HJC). (F) ChIP assay to determine RUNX1 enrichment at the miR-24 promoter in HSC-T6s (n = 3 per group). (G) EdU staining and quantification of EdU-positive HSC-T6s (n = 6 per group). Scale bar = 100 μm. (H) qRT-PCR analysis of α-SMA expression in HSC-T6s (n = 6 per group). (I) EdU staining and quantification of EdU-positive HSC-T6s (n = 5 per group). Scale bar = 100 μm. (J) qRT-PCR analysis of α-SMA and Col1a1 expressions in HSC-T6 cells (n = 5:5:3:3). HSC-T6, hepatic stellate cell-T6. Data are expressed as mean ± SD. *, p < 0.05; **, p < 0.01; ***, p < 0.001.

Article Snippet: The rat HSC line HSC-T6 was provided by Jiangsu KeyGEN BioTECH Co., Ltd. HEK293T was purchased from the Chinese Academy of Sciences Cell Resource Center.

Techniques: Activation Assay, Western Blot, Quantitative RT-PCR, Expressing, Staining

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet: (a) Representative images of 5-HT 1B and 5-HT 1D expression in cultured primary mouse Schwann cells (mSCs, scale bar: 10 μm, n=6) and ( b) in mouse sciatic nerve tissues (scale bar: 20μm, n=6). SOX10 and S100 are markers of SCs. (c) Representative RNAscope images of Htr1b and Htr1d mRNA and S100 protein expression in mouse sciatic nerve (scale bar: 5μm, n=6). (d) RT-qPCR for Htr1b and Htr1d mRNA in primary mSCs and mouse sciatic nerve tissues. (e) Dose dependent periorbital mechanical allodynia and cumulative data of facial grimacing (mouse grimace scale, MGS) in C57BL/6J mice treated with CGRP (0.1 mg/kg, i.p.) and pretreated with sumatriptan (Suma, 0.2-0.6 mg/kg) (f) and pretreated with GR125743 (0.5 mg/kg, i.p.) or vehicle (Veh). (g) Representative images and colocalization data (Rcoloc) of human SCs (hSCs) showing internalization of 5-HT 1B -EGFP or 5-HT 1D -EGFP in Ras-related protein (Rab5a)-RFP endosomes after sumatriptan exposure in presence of GR125743 or Veh (scale bar: 2μm n=6). (h) Schematic representation of 5-HT 1B/D trafficking to early endosomes using the NanoLuc Binary Technology (NanoBiT) assay. (i) Typical traces and cumulative data of 5-HT 1B and 5-HT 1D internalization after exposure to sumatriptan (10 µM) in presence of GR125743 (10 µM). (replicates: Veh=8, Suma=8, GR125743=8) Data are mean ± s.e.m. (n=8 mice/group). Student’s t-test, 1-way and 2-way ANOVA. ***p<0.001, ****p<0.0001 vs Veh/Veh; # p<0.05, ## p<0.01, #### p<0.0001 vs CGRP/Veh or CGRP/Veh/Veh.

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: Expressing, Cell Culture, RNAscope, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet: ( a ) Representative images of 5-HT 1B e 5-HT 1D expression in human Schwann cells (hSCs, scale bar: 10 μm, n=6) and ( b) in human sciatic nerve tissues (scale bar: 20 μm, n=6). SOX10 and S100 are markers of SCs. (c) Representative RNAscope images of HTR1B and HTR1D mRNA and S100 protein expression in human sciatic nerve (scale bar: 20 μm, n=6). (d) RT-qPCR for HTR1B and HTR1D mRNA in commercial and primary (isolated from lingual, sublingual and inferior alveolar nerves biopsies) hSCs and in human nerve tissues. (e) Typical traces and cumulative data of the effects of sumatriptan (Suma, 30 μM) on CGRP (3 μM)-evoked Rab5a cAMP formation in the presence of GR125743 (1 μM) or vehicle (Veh) in primary isolated hSCs (cells number: Veh/Veh/Veh=29, CGRP/Veh/Veh =26, CGRP/Suma/Veh =26, CGRP/Suma/GR125743 =29) and in (f) primary mouse SCs (mSCs) (cells number: Veh/Veh/Veh=40, CGRP/Veh/Veh =50, CGRP/Suma/Veh =37, CGRP/Suma/GR125743 =41). (g) Representative images colocalization data (Rcoloc) of 5-HT 1B and 5-HT 1D silencing in SCs in Plp-Htr1b-Htr1d and Control mice. Data are mean ± s.e.m. Student’s t-test and 1-way ANOVA, Bonferroni correction.

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: Expressing, RNAscope, Quantitative RT-PCR, Isolation, Control

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet: (a) Typical traces, concentration response curve (CRC) and cumulative data showing the effects of graded concentrations of CGRP (3 nM-30 µM) or vehicle (Veh) on cAMP formation at early endosomes in human Schwann cells (hSCs) (cells number: 3 nM =38, 100 nM=54, 1 µM=70, 30 µM=57; EC 50 CI=506-787 nM and EC 80 CI=2-3.1 µM). Ras-related protein (Rab5a) is an early endosome marker. (b) Typical traces, CRC and cumulative data showing the effects of graded concentrations of sumatriptan (Suma, 100 nM-300 µM) or Veh on CGRP (3 µM)-evoked Rab5a associated cAMP formation in hSCs (cells number: 100 nM=75, 1 µM=55, 3 µM=108, 10 µM=53, 300 µM=109; IC 50 CI=3.2-20 µM; IC 80 CI=13-82 µM) and (c) in the presence of GR125743 (1 μM) or Veh (cells number: Veh/Veh/Veh=20, CGRP/Veh/Veh =44, CGRP/Suma/Veh =32, CGRP/Suma/GR125743 =39). (d) Schematic representation of Cre-dependent dicistronic adeno AAV strategy to selectively silence 5-HT 1B/D receptors in SCs in Plp Cre mice. (e) Time dependent periorbital mechanical allodynia and facial grimacing (mouse grimace scale, MGS) induced by CGRP (0.1 mg/kg, i.p.) in Control, Plp-Htr1b-Htr1d and (f) Plp-scrambled mice pretreated with Suma (0.6 mg/kg, i.p.) or Veh. Data are mean ± s.e.m. (n=8 mice/group). 1-way and 2-way ANOVA, Bonferroni correction, ****p<0.0001 vs Control/Veh/Veh or Plp-Scrambled /Veh/Veh.

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: Concentration Assay, Marker, Control

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet: (a) Time dependent periorbital mechanical allodynia (PMA) and facial grimacing (mouse grimace scale, MGS) in male and female C57BL/6J (B6) mice during/after repeated administration of sumatriptan (Suma, 0.6 mg/kg once daily over 7 consecutive days) or vehicle (Veh). (b) PMA and facial grimacing induced by Suma or Veh in B6 mice pretreated with GR125743 (0.5 mg/kg, i.p.) or Veh. (c) PMA and facial grimacing induced by Suma or Veh in Plp-5-Htr1b-Htr1d, Control or Plp-Scrambled mice. (d) Differentially methylated regions (DMRs), differentially methylated genes (DMGs) and the cumulative size of DMRs in human Schwann cells (hSCs) stimulated with Suma (10 µM) for 6h, 24h, and 48h. (e) Density plots of methylation changes distribution (Δβ) for all DMRs identified at 6h, 24h, and 48h. (f) Butterfly plot of the DMGs distribution across different genomic features Consistent with the global trend described in panel d (p=1.057×10 −11 ). (g) Multidimensional scaling plot representing relationships between different time points after hSCs stimulation with Suma. Each point represents a single sample. (h) Heatmap of differentially expressed genes (DEGs) in hSCs across the indicated time points. Each column represents an individual sample. Conditions are ordered from left to right as time 0, 6h, 24h, and 48h. List of genes is available in (i) Volcano plots showing the log2-fold change (lfc) and –log10(padj) for all DEGs between hSCs at time 0 vs. 6h, 24h and 48h of Suma stimulation. The top 200 differentially expressed genes are provided in the , while the complete list of genes is available in the Data file S1. (j) RT-qPCR for TGFB3 , IL-15 and IL-32 mRNA in hSCs after Suma stimulation (48 h). ( k) PMA and facial grimacing induced by Suma or Veh in B6 mice pretreated with mAb-TGF-β3, mAb-IL-15 (all, 200 μg/mice) or IgG. (l) RT-qPCR for Tgfb3 and Il-15 mRNA in mouse trigeminal nerve homogenates. (m) ELISA assay for TGF-β3 and IL-15 in trigeminal nerve tissue homogenates from B6 mice after Suma or Veh and pretreated with mAb-TGF-β3, mAb-IL-15 or IgG. (n) PMA and facial grimacing after Suma or Veh administration in B6 mice pretreated with LY2109761 (50 mg/kg, i.g.) or Veh. Black arrows indicate Suma administration. Data are mean ± s.e.m. (n=8 mice/group). Student’s t-test, 1-way and 2-way ANOVA, Bonferroni correction. ****p<0.0001 vs Veh♂, Veh♀, Veh/Veh, Control/Veh, Veh/IgG; ### p<0.001, #### p<0.0001 vs Suma/Veh, Control/Suma or Suma/IgG. Statistical significance was assessed (d) using the Cochran-Armitage test for trend. DMRs p value < 2.2 x 10 -16 , DMGs p value = 1.057 10 -11 , and cumulative DMR size p value < 2.2 x 10 -16 .

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: Control, Methylation, Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet: (a) RT-qPCR for BETAGLYCAN mRNA and (b) methylation changes distribution (Δβ) for CpG sites in the BETAGLYCAN intron 2 in human Schwann cells (hSCs) stimulated with sumatriptan (10 μM) (Suma). (c) Enrichment analysis on TGF-β pathways in hSCs treated for 24h and 48h with Suma. Numbers indicated the ratio of genes differentially expressed. (d) Schematic representation illustrating the sequence of events induced by chronic stimulation: Suma activates SCs 5-HT 1B/D receptors, (2) promoting BETAGLYCAN intronic hypermethylation, (3) BETAGLYCAN over-expression, (4) BETAGLYCAN enhanced non-canonical TGF-βR intracellular signaling and a TGFΒ3 upregulation establishing a feed-forward mechanism. (e) RT-qPCR for TGFB3 and BETAGLYCAN mRNA and (f) ELISA assay for TGF-β3 in hSCs stimulated with Suma or vehicle (Veh) and pretreated with TGF-βRs inhibitor LY2109761 (10 μM) or Veh. (g) RT-qPCR for BETAGLYCAN mRNA in hSC stimulated with Suma or Veh and pretreated with 5-Aza-2′-deossicitidina (5-AZA-CdR, 1 μM) or Veh. (h) Periorbital mechanical allodynia and facial grimacing (mouse grimace scale, MGS) in C57BL/6J mice after repeated (once daily over 7 consecutives days) administration of suma (0.6 mg/kg), 5-AZA-CdR (4 mg/kg, i.p.) or Veh. Red arrows indicate Suma and 5-AZA-CdR administration. hSCs are stimulated with Suma in all the experiments for 48h. Data are mean ± s.e.m. (n=8 mice/group). Student’s t-test, 1-way and 2-way ANOVA, Bonferroni correction. ****p<0.0001 vs Veh/Veh, ## p<0.01, #### p<0.0001 vs Suma/Veh.

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: Quantitative RT-PCR, Methylation, Sequencing, Over Expression, Enzyme-linked Immunosorbent Assay

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet: (a ) UMAP plot of the nociceptive lineage cells highlighted by the pain state identified by aggregation of Leiden clusters. (b) Schematic representation of dorsal root ganglia (DRG) neurons and mouse Schwann cells (mSCs) in co-culture device. (c) RT-qPCR for Atf3, Atf4, Onecut2, Fols2, Sox11, Klf6 and Arid5a in DRG neurons alone or co-cultured with mSCs and stimulated with sumatriptan (Suma) or vehicle (Veh) and pretreated with GR125743 (1 μM), LY2109761 (10 μM), mAb-TGF-β3 (5 μg/ml), IgG (5 μg/ml) or Veh. Time dependent periorbital mechanical allodynia (PMA) and facial grimacing (mouse grimace scale, MGS) in Control, and (d) Adv-Tgfbr1-Tgfbr2 and (e) Adv-Scrambled during repeated administration of Suma (0.6 mg/kg once daily over 7 consecutive days) or vehicle (Veh). (f) RT-qPCR for Tgfb3 and Betaglycan in Control, Adv-Tgfbr1-Tgfbr2 and Adv-Scrambled mice treated with Suma or Veh. Black arrows indicate Suma administration Data are mean ± s.e.m. (n=8 mice/group). Student’s t-test, 1-way and 2-way ANOVA, Bonferroni correction. ***p<0.001, ****p<0.0001 vs Veh/Veh; ### p<0.001, #### p<0.0001 vs Control/Suma

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: Co-Culture Assay, Quantitative RT-PCR, Cell Culture, Control

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet: Time dependent periorbital mechanical allodynia (PMA) and facial grimacing (mouse grimace scale, MGS) in C57BL/6J (B6) mice during/after repeated (once daily over 7 consecutives days) administration of naratriptan (0.2 mg/kg i.p.), eletriptan (1 mg/kg, i.p.) or vehicle (Veh). (c) RT-qPCR for BETAGLYCAN and TGFB3 in human Schwann cells (hSCs) stimulated with naratriptan, eletriptan or Veh. (d) ELISA assay for TGF-β3 in trigeminal nerve homogenates from naratriptan, eletriptan and Veh treated B6 mice. PMA and facial grimacing induced by repeated naratriptan, eletriptan or Veh in Control and (e) Plp-Htr1b-Htr1d, (f) Plp-Tgfb3, (g) Plp-Betaglycan , (h) Adv-Tgfbr1-Tgfbr2 mice. Black arrows indicate drugs administration. Data are mean ± s.e.m. (n=8 mice/group). Student’s t test, 1-way and 2-way ANOVA, Bonferroni correction. ***p<0.001, ****p<0.0001 vs Veh/Veh; # p<0.05, ### p<0.001, #### p<0.0001 vs Control/naratriptan/eletriptan.

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: Quantitative RT-PCR, Enzyme-linked Immunosorbent Assay, Control

Journal: bioRxiv

Article Title: Triptans reprogram Schwann cells to drive medication-overuse headache via β-glycan/TGF-β3 signaling

doi: 10.64898/2026.04.24.720547

Figure Lengend Snippet:

Article Snippet: Commercial human primary Schwann cells (hSCs) (#P10351, Innoprot) were grown and maintained in Schwann cell medium (#P60123, Innoprot) at 37 °C in 5% CO 2 and 95% O 2 .

Techniques: