recombinant human tgf β2  (R&D Systems)

Journal: bioRxiv

Article Title: Macrophage Migration Inhibitory Factor (MIF)-CD74 Signaling Pathway Mediates Trabecular Meshwork Dysfunction in Glaucoma

doi: 10.64898/2026.03.18.712673

Figure Lengend Snippet: Wild-type C57BL/6 mice (n=12/group) were injected intravitreally with either LV-TGF-β2 or LV-null vectors. ( A ) Weekly IOP measurements demonstrate significant and sustained IOP elevation in LV-TGF-β2-injected mice compared with LV-null controls. ( B ) Immunofluorescence staining of anterior segment cross-sections for α-SMA and MIF shows co-localization of MIF with the TM marker α-SMA, indicating increased MIF expression in the TM of LV-TGF-β2 injected mice. ( C-E ) qPCR ( C ) and western blot ( D, E ) analyses of anterior segment tissues show upregulation of MIF and CD74 accompanied by suppression of Blimp-1 in LV-TGF-β2-injected mice. Increased expression of profibrotic markers, including α-SMA, Fn-1, and MYOC, was also evident in LV-TGF-β2-injected mice. Western blot bands were quantified by densitometric analysis using ImageJ and normalized to the housekeeping control β-actin, and protein levels are presented as relative expression (AU) ( E ). Data represent mean ± SD. **p<0.005, ***p<0.0005, and ****p<0.0001. Two-way ANOVA with Tukey’s multiple-comparison test (A), unpaired t-test (C, E).

Article Snippet: Recombinant human TGF-β2 (302-B2-010/CF) protein was purchased from R&D Systems (Minneapolis, MN).

Techniques: Injection, Immunofluorescence, Staining, Marker, Expressing, Western Blot, Control, Comparison

Journal: bioRxiv

Article Title: Macrophage Migration Inhibitory Factor (MIF)-CD74 Signaling Pathway Mediates Trabecular Meshwork Dysfunction in Glaucoma

doi: 10.64898/2026.03.18.712673

Figure Lengend Snippet: Primary HTMCs (n=3) were challenged with TGF-β2 (10ng/mL), rMIF (100ng/mL), or pro-inflammatory cytokine pool (CP; TNF-α, IL-1β, IL-6, and IL-17 100ng/mL) for 24h. (A) qPCR analysis shows significant upregulation of MIF and CD74 transcripts accompanied by reduced Blimp-1 expression in response to glaucomatous stressors. ( B ) Immunofluorescence staining demonstrates increased MIF expression following TGF-β2 or CP treatment. (C) Western blot analysis confirms induction of MIF and CD74 with concurrent suppression of Blimp-1 at the protein level. Data represent mean ± SD. **p<0.005, ***p<0.0005, and ***p<0.0001, One-way ANOVA with Dunnett’s multiple-comparison test.

Article Snippet: Recombinant human TGF-β2 (302-B2-010/CF) protein was purchased from R&D Systems (Minneapolis, MN).

Techniques: Expressing, Immunofluorescence, Staining, Western Blot, Comparison

Journal: bioRxiv

Article Title: Macrophage Migration Inhibitory Factor (MIF)-CD74 Signaling Pathway Mediates Trabecular Meshwork Dysfunction in Glaucoma

doi: 10.64898/2026.03.18.712673

Figure Lengend Snippet: Primary HTMCs (n=3) were challenged with rMIF (100 ng/mL) in the presence or absence of MIF inhibitor 4-IPP (100µM) or the metabolites Agm or Thia (100 ng/mL) for 24h. Cells treated with TGF-β2 (10ng/mL) served as a positive control. (A-B) Western blot analysis confirms activation of the RhoA/ROCK/pMLC signaling pathway in response to MIF treatment, which is significantly attenuated by MIF inhibition with 4-IPP, Agm, or Thia ( A ). Western blot bands were quantified by densitometric analysis using ImageJ and normalized to the housekeeping control β-actin, and protein levels are presented as relative expression (AU) ( B ). (C) Immunofluorescence analysis shows that MIF induces pMLC staining, comparable to that observed with TGF-β2 stimulation, which is significantly attenuated by MIF inhibition with 4-IPP, Agm, or Thia. Data represent mean ± SD. *p<0.05, **p<0.005, ***p<0.0005, and ****p<0.0001, One-way ANOVA with Dunnett’s multiple-comparison test.

Article Snippet: Recombinant human TGF-β2 (302-B2-010/CF) protein was purchased from R&D Systems (Minneapolis, MN).

Techniques: Positive Control, Western Blot, Activation Assay, Inhibition, Control, Expressing, Immunofluorescence, Staining, Comparison

Journal: bioRxiv

Article Title: Macrophage Migration Inhibitory Factor (MIF)-CD74 Signaling Pathway Mediates Trabecular Meshwork Dysfunction in Glaucoma

doi: 10.64898/2026.03.18.712673

Figure Lengend Snippet: OHT was induced in WT C57BL/6 mice (n=12) by intravitreal injection of LV-TGF-β2. LV-null-injected mcie served as controls. Following OHT induction, extracellular vesicle-loaded agmatine (EV-Agm; 0.1 µg/eye) was applied topically once daily. (A) Weekly IOP measurements over 9 weeks show that EV-Agm treatment significantly reduces IOP compared with untreated LV-TGF-β2-injected mice. (B) qPCR analysis of anterior segment tissue demonstrates that EV-Agm significantly reduces expression of Mif, Cd74, pro-inflammatory cytokines ( Il-1 β , Il-6 ), and cytoskeletal/ECM markers (α -Sma, Myoc, Fn-1 ), while restoring Blimp-1 expression compared with untreated LV-TGF-β2-injected mice. *p<0.05, **p<0.005, ###,***p<0.0005, and ####,****p<0.0001. Two-way ANOVA with Tukey’s multiple comparison (A), one-way ANOVA with Tukey’s multiple comparison (B). In panel A, * denotes LV-null vs. LV-TGF-β2, and # denotes LV-TGF-β2 vs. LV-TGF-β2+EV-Agm comparisons.

Article Snippet: Recombinant human TGF-β2 (302-B2-010/CF) protein was purchased from R&D Systems (Minneapolis, MN).

Techniques: Injection, Expressing, Comparison

Journal: Journal of Translational Medicine

Article Title: Micheliolide suppresses epithelial-mesenchymal transition of lens epithelial cells via downregulating matrix metalloproteinase 8 to ameliorate posterior capsular opacification

doi: 10.1186/s12967-026-07890-z

Figure Lengend Snippet: Regulation of TGF-β2-induced LECs proliferation, migration, and EMT by MCL in vitro A , Viability test of LECs at gradient concentrations of MCL, n = 8. B , qRT-PCR analysis of α-SMA, Col I, and FN in LECs pretreated with or without 5 μmol/L MCL and stimulated with 10 ng/mL TGF-β2 for 48 hours, n = 3. C , Western blot and quantitative analysis of α-SMA, Col I, and FN in LECs pretreated with or without 5 μmol/L MCL and stimulated with 10 ng/mL TGF-β2 for 48 hours, n = 5. D , Representative images of LECs that migrated to the lower chamber obtained by Transwell assay with TGF-β2 (10 ng/mL) stimulation with or without pretreatment of MCL (5 μmol/L), n = 5, scale bar = 100 μm. E , Representative images of the scratch wound assay in LECs at 0, 24, and 48 hours. Cells were pretreated with or without 5 μmol/L MCL and then stimulated with 10 ng/mL TGF-β2, n = 3, scale bar = 500 μm. F , Quantification of cells that migrated to the lower chamber. G , The wound healing rate was quantified at 0, 24, and 48 hours after the scratch

Article Snippet: To examine the impact of MCL (Cat. HY-N0847, MedChemExpress, New Jersey, USA) on TGF-β2 (Cat. HZ-1092, Proteintech, Chicago, USA)-induced EMT in vitro, cells were pre-treated with 5 μmol/L MCL and starved overnight at 50%-60% confluency.

Techniques: Migration, In Vitro, Quantitative RT-PCR, Western Blot, Transwell Assay, Scratch Wound Assay Assay

Journal: Journal of Translational Medicine

Article Title: Micheliolide suppresses epithelial-mesenchymal transition of lens epithelial cells via downregulating matrix metalloproteinase 8 to ameliorate posterior capsular opacification

doi: 10.1186/s12967-026-07890-z

Figure Lengend Snippet: MCL modulates the expression of EMT markers in TGF-β2-induced LECs. A , Immunofluorescence staining of Col I, FN, and α-SMA (green) shows that MCL inhibited the expression of TGF-β2-induced EMT marker proteins in LECs. Nuclei were counterstained with DAPI (blue), n = 4, scale bar = 100um. B , Quantification of immunofluorescence intensity

Article Snippet: To examine the impact of MCL (Cat. HY-N0847, MedChemExpress, New Jersey, USA) on TGF-β2 (Cat. HZ-1092, Proteintech, Chicago, USA)-induced EMT in vitro, cells were pre-treated with 5 μmol/L MCL and starved overnight at 50%-60% confluency.

Techniques: Expressing, Immunofluorescence, Staining, Marker

Journal: Journal of Translational Medicine

Article Title: Micheliolide suppresses epithelial-mesenchymal transition of lens epithelial cells via downregulating matrix metalloproteinase 8 to ameliorate posterior capsular opacification

doi: 10.1186/s12967-026-07890-z

Figure Lengend Snippet: Identification of Micheliolide, a potential MMP8 regulator. A , qRT-PCR analysis of MMP8 expression in capsular tissues 24 hours after ECLE surgery, combined with anterior chamber injection of saline or MCL, n = 3, N = 18. B , Western blot analysis and quantification of MMP8 protein levels in capsular tissues 24 hours after ECLE surgery with anterior chamber injection of saline or MCL, n = 4, N = 20. C , qRT-PCR analysis of MMP8 expression in LECs pretreated with or without 5 μmol/L MCL and stimulated with 10 ng/mL TGF-β2 for 48 hours, n = 6. D , Western blot and corresponding quantification of MMP8 protein expression in LECs pretreated with or without 5 μmol/L MCL followed by stimulation with 10 ng/mL TGF-β2 for 48 hours, n = 4. E , Molecular docking simulation of Micheliolide and MCL, with the interacting MMP8 amino acids represented by blue sticks. F , CETSA was used to assess the thermal instability of the interaction between MMP8 and MCL over the temperature range of 58 °C to 78 °C, n = 4

Article Snippet: To examine the impact of MCL (Cat. HY-N0847, MedChemExpress, New Jersey, USA) on TGF-β2 (Cat. HZ-1092, Proteintech, Chicago, USA)-induced EMT in vitro, cells were pre-treated with 5 μmol/L MCL and starved overnight at 50%-60% confluency.

Techniques: Quantitative RT-PCR, Expressing, Injection, Saline, Western Blot

Journal: Journal of Translational Medicine

Article Title: Micheliolide suppresses epithelial-mesenchymal transition of lens epithelial cells via downregulating matrix metalloproteinase 8 to ameliorate posterior capsular opacification

doi: 10.1186/s12967-026-07890-z

Figure Lengend Snippet: MCL inhibits EMT by regulating MMP8 expression. A , Viability of LECs treated with a gradient concentration of M8I, n = 8. B , Western blot analysis of EMT-related markers in LECs induced by TGF-β2 and treated with MCL alone or in combination with M8I, n = 4. C , Quantitative analysis of the Western blot results shown in ( B ), normalized to GAPDH. D , Coimmunofluorescence staining for αSMA (red) and MMP8 (green) in LECs induced by TGFβ2 and treated with MCL alone or in combination with M8I, n = 3, scale bar = 100um. E , Quantification of immunofluorescence intensity

Article Snippet: To examine the impact of MCL (Cat. HY-N0847, MedChemExpress, New Jersey, USA) on TGF-β2 (Cat. HZ-1092, Proteintech, Chicago, USA)-induced EMT in vitro, cells were pre-treated with 5 μmol/L MCL and starved overnight at 50%-60% confluency.

Techniques: Expressing, Concentration Assay, Western Blot, Staining, Immunofluorescence