Journal: Clinics

Article Title: Pregabalin as a neuroprotective alternative to methylprednisolone in peripheral nerve regeneration: an experimental study

doi: 10.1016/j.clinsp.2025.100838

Figure Lengend Snippet: Evaluation of the groups’ ELISA results. (a, b, c, d and e) There is no statistically significant difference between groups in columns symbolized by the same letter. There is a statistically significant difference between groups in columns symbolized by different letters. CNTF, Ciliary Neurotrophic Factor; NGF, Nerve Growth Factor; TGF-β1, Transforming Growth Factor β; MBP, Myelin Basic Protein.

Article Snippet: Serum NGF, Ciliary Neurotrophic Factor (CNTF), Myelin Basic Protein (MBP), and Transforming Growth Factor Beta (TGF-β) values were determined using the ELISA method with a Rat NGF ELISA kit (BTLAB, cat n° E0539Ra, China), a Rat CNTF ELISA kit (BTLAB, cat n° E0358Ra, China), a Rat MBP ELISA kit (BTLAB, cat n° E0576Ra, China), and a Rat TGF-β ELISA kit (BTLAB, cat n° E0778Ra, China) in accordance with the manufacturer’s recommendations.

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Stem Cells Translational Medicine

Article Title: TGFBR3 dependent mechanism of TGFB2 in smooth muscle cell differentiation and implications for TGFB2-related aortic aneurysm

doi: 10.1093/stcltm/szae101

Figure Lengend Snippet: TGFB1 and TGFB2 have similar effects on hiPSC-derived SMC differentiation. (A) Relative expression of SMC markers in CPC-SMCs after differentiation with either 2 ng/mL active TGFB1, 2 ng/mL active TGFB2, or control (vehicle/no ligand). The average expression in the TGFB1-treated samples was set to 1. n = 6 biological replicates. (B) Relative expression of smooth muscle cell (SMC) markers in cardiovascular progenitor cells (CPC)-SMCs after differentiation with either 2 ng/mL active TGFB2 or 2 ng/mL full-length TGFB2 (FL-TGFB2). The average expression in the active TGFB2-treated samples was set to 1. n = 6 biological replicates. (C) Top: western blots of CPC-SMC protein extracts after differentiation with either active TGFB1 or TGFB2. Bottom: quantification of western blot data showing relative protein SMC marker levels in CPC-SMCs. n = 6 biological replicates. (D) Top: western blots of canonical and non-canonical TGFβ signaling mediators in CPC-SMCs after 1-hour stimulation with either active TGFB1, TGFB2, or vehicle. Bottom: quantification of western blot data for pSMAD2, pSMAD3, pERK, and pAKT (T308) after the stimulation. The average protein levels in the TGFB1 treated samples were set to 1. n = 6 biological replicates. 1-way analysis of variance (ANOVA) with multiple comparisons test.

Article Snippet: Active human TGFB2 protein levels were assessed in tissue ring lysates using a TGFB2 enzyme-linked immunosorbent assay (ELISA) kit (Catalog No. DB250, R&D Systems).

Techniques: Derivative Assay, Expressing, Control, Western Blot, Marker

Journal: Stem Cells Translational Medicine

Article Title: TGFBR3 dependent mechanism of TGFB2 in smooth muscle cell differentiation and implications for TGFB2-related aortic aneurysm

doi: 10.1093/stcltm/szae101

Figure Lengend Snippet: TGFBR3 facilitates TGFB2 signaling transduction during SMC differentiation. (A and B), Relative expression of SMC markers after TGFBR3-siRNA treatment. CPC-SMCs were transfected with 40nM TGFBR3-siRNA or control siRNA for 4 days during differentiation with either active TGFB1 or TGFB2. n = 6 biological replicates. ns: not significant; unpaired t-test or Mann-Whitney U test. (C) pSMAD3 and pSMAD2 western blots of CPC-SMC protein extracts stimulated with either active TGFB1, TGFB2 or vehicle for 1 hour. CPC-SMCs were transfected with 40 nM target siRNA (TGFBR1-siRNA or TGFBR3-siRNA) or control siRNA for 2 days. (D) Quantification of pSMAD3 western blot data. Each quantification was normalized to its vehicle control. The average levels in control siRNA-treated samples were set to 1. n = 5 biological replicates. ns: not significant; Kruskal-Wallis test with multiple comparisons test.

Article Snippet: Active human TGFB2 protein levels were assessed in tissue ring lysates using a TGFB2 enzyme-linked immunosorbent assay (ELISA) kit (Catalog No. DB250, R&D Systems).

Techniques: Transduction, Expressing, Transfection, Control, MANN-WHITNEY, Western Blot

Journal: Stem Cells Translational Medicine

Article Title: TGFBR3 dependent mechanism of TGFB2 in smooth muscle cell differentiation and implications for TGFB2-related aortic aneurysm

doi: 10.1093/stcltm/szae101

Figure Lengend Snippet: TGFB2 and TGFBR3 expression show enrichment in the human aortic root. (A) Relative expression of TGFB2 in aortic media of root, ascending, and arch samples. TGFB2 expression level in each aortic root sample was set to 1. (B) Relative expression of TGFBR3 in aortic media of root, ascending, and arch samples. TGFBR3 expression level in each aortic root sample was set to 1. (C) TGFB2 and TGFBR3 western blots of tunica media protein extracts from different thoracic aorta regions. (D) Quantification of western blot data showing TGFB2 and TGFBR3 enrichment in the aortic root media. n = 4 biological replicates. Kruskal-Wallis test with multiple comparisons test.

Article Snippet: Active human TGFB2 protein levels were assessed in tissue ring lysates using a TGFB2 enzyme-linked immunosorbent assay (ELISA) kit (Catalog No. DB250, R&D Systems).

Techniques: Expressing, Western Blot

Journal: Stem Cells Translational Medicine

Article Title: TGFBR3 dependent mechanism of TGFB2 in smooth muscle cell differentiation and implications for TGFB2-related aortic aneurysm

doi: 10.1093/stcltm/szae101

Figure Lengend Snippet: TGFB2 haploinsufficiency causes SMC differentiation defects. (A) The Sanger sequencing results for TGFB2 +/+ and TGFB2 KO/+ showing the position of 7 base-pair deletion resulting in a premature stop codon in TGFB2 gene. (B) Left: relative TGFB2 expression in TGFB2 +/+ and TGFB2 KO/+ SMCs. n = 6 biological replicates. unpaired t-test. Right: relative TGFB2 protein levels in TGFB2 +/+ and TGFB2 KO/+ SMCs. n = 3 biological replicates; unpaired t-test. (C) Diagram showing the co-culture assay design with TGFB2 +/+ and TGFB2 KO/+ CPC-SMCs. (D) Relative SMC marker levels in the following conditions: TGFB2 +/+ CPC-SMCs co-cultured with TGFB2 +/+ CPC-SMCs (black columns); TGFB2 KO/+ CPC-SMCs co-cultured with TGFB2 KO/+ CPC-SMCs (red columns) and TGFB2 KO/+ CPC-SMCs co-cultured with TGFB2 +/+ CPC-SMCs (gray columns). n = 6 biological replicates. ns: not significant; 1-way ANOVA with multiple comparisons test. (E) Relative SMC marker levels in TGFB2 KO/+ CPC-SMCs supplemented with either active TGFB2 or vehicle for 7 days during CPC-SMC differentiation. The average expression in vehicle samples was set to 1. n = 6 biological replicates. ns: not significant; unpaired t-test or Mann-Whitney U test. (F) Relative SMC marker levels in TGFB2 +/+ CPC-SMCs treated with 1ug/mL TGFB2 neutralization antibody. The average expression in the control samples was set to 1. n = 6 biological replicates. ns: not significant; unpaired t-test.

Article Snippet: Active human TGFB2 protein levels were assessed in tissue ring lysates using a TGFB2 enzyme-linked immunosorbent assay (ELISA) kit (Catalog No. DB250, R&D Systems).

Techniques: Sequencing, Expressing, Co-culture Assay, Marker, Cell Culture, MANN-WHITNEY, Neutralization, Control

Journal: Stem Cells Translational Medicine

Article Title: TGFBR3 dependent mechanism of TGFB2 in smooth muscle cell differentiation and implications for TGFB2-related aortic aneurysm

doi: 10.1093/stcltm/szae101

Figure Lengend Snippet: TGFBR3 KO/KO causes reduced TGFB2 sensitivity in TGFB2 KO/+ SMCs. (A) The Sanger sequencing results showing 2 independent TGFBR3 KO/KO hiPSC clones with premature TGFBR3 stop codons generated by CRISPR/Cas9 gene editing. (B) Relative TGFBR3 expression in different conditions. n = 6 biological replicates; 1-way ANOVA with multiple comparisons test. (C) Relative SMC marker expression in TGFB2 KO/+ , 2 TGFB2 KO/+ TGFBR3 KO/KO clones and isogenic control ( TGFB2 +/+ TGFBR3 +/+ ) CPC-SMCs differentiated with TGFB2. n = 6 biological replicates; ns: not significant; 1-way ANOVA with multiple comparisons test. (D-E) MYH11 staining intensity and cell density in TGFB2 KO/+ , TGFB2 KO/+ TGFBR3 KO/KO , and isogenic control ( TGFB2 +/+ TGFBR3 +/+ ) tissue rings generated using TGFB2. The average values in TGFB2 +/+ TGFBR3 +/+ rings were set to 1. n = 6 biological replicates; ns: not significant; 1-way ANOVA with multiple comparisons test.

Article Snippet: Active human TGFB2 protein levels were assessed in tissue ring lysates using a TGFB2 enzyme-linked immunosorbent assay (ELISA) kit (Catalog No. DB250, R&D Systems).

Techniques: Sequencing, Clone Assay, Generated, CRISPR, Expressing, Marker, Control, Staining

Journal: Stem Cells Translational Medicine

Article Title: TGFBR3 dependent mechanism of TGFB2 in smooth muscle cell differentiation and implications for TGFB2-related aortic aneurysm

doi: 10.1093/stcltm/szae101

Figure Lengend Snippet: TGFB2 G276R variant disrupts the mechanical properties of vascular tissue rings. (A) The Sanger sequencing results showing the position of c.826G > A variant and its CRISPR/Cas9-based genetic correction (black rectangle). (B) Representative stress-strain curves and maximum tensile stress values from Patient G276R/+ , Patient +/+ tissue rings as well as Patient G276R/+ rings treated with TGFB2. n = 6 biological replicates; 1-way ANOVA with multiple comparisons test. (C-D) MYH11 staining intensity and cell density in Patient G276R/+ , Patient +/+ tissue rings as well as Patient G276R/+ rings supplemented with TGFB2. The average values in Patient G276R/+ samples were set to 1. n = 6 biological replicates; ns: not significant; 1-way ANOVA with multiple comparisons test. (E) The Sanger sequencing results for TGFB2 +/+ and TGFB2 G276R/+ hiPSCs. (F) Representative stress-strain curves and maximum tensile stress values from TGFB2 +/+ , TGFB2 G276R/+ as well as TGFB2 G276R/+ tissue rings treated with TGFB2. n = 6 biological replicates; 1-way ANOVA with multiple comparisons test. (G-H) MYH11 staining intensity and cell density in TGFB2 +/+ , TGFB2 G276R/+ as well as TGFB2 G276R/+ tissue rings supplemented with TGFB2. The average values in Patient G276R/+ samples were set to 1. n = 6 biological replicates; ns: not significant; 1-way ANOVA with multiple comparisons test.

Article Snippet: Active human TGFB2 protein levels were assessed in tissue ring lysates using a TGFB2 enzyme-linked immunosorbent assay (ELISA) kit (Catalog No. DB250, R&D Systems).

Techniques: Variant Assay, Sequencing, CRISPR, Staining

Journal: Journal of Biomedical Science

Article Title: Periostin promotes ovarian cancer metastasis by enhancing M2 macrophages and cancer-associated fibroblasts via integrin-mediated NF-κB and TGF-β2 signaling

doi: 10.1186/s12929-022-00888-x

Figure Lengend Snippet: POSTN increases expression of TGF-β2 capable of inducing CAF markers and is correlated with abundant CAFs in tumor microenvironment. A The qRT-PCR analysis of α-SMA and FAP mRNAs from conditioned medium treated hADSCs. The conditioned media were collected from SKOV3/ctrl or SKOV3/POSTN cells cultured for 2 days. B MTT proliferation assay of SKOV3 cells incubated with conditioned medium from unprimed, SKOV3/ctrl cells or SKOV3/POSTN cells primed hADSC cells. C , D RNA and protein levels of TGF-β1 and TGF-β2 in POSTN-overexpressing SKOV3 cells were determined qRT-PCR and ELISA, respectively. E Expression levels of α-SMA and FAP transcripts in TGF-β2 treated normal human adipose derived stromal cells (hADSCs) were determined by qRT-PCR. F Representative immunohistochemical analysis for the expression of α-SMA in POSTN-overexpressing versus control SKOV3-derived primary and metastatic tumors in the orthotopic metastasis mouse model (left panel). Quantification results of the immunohistochemical analysis of anti-α-SMA (right panel). Six female NOD/SCID mice were randomly assigned to each group and were followed up for 3 weeks in orthotopic metastasis mouse model. G Western blot analysis for measuring the POSTN level in SKOV-I6 cells infected with lentiviral plasmid encoding shPOSTN or a scrambled control. H The diagram illustrates the co-culture system used for cancer cell-induced CAF markers activation. I Representative immunofluorescence images showe the α-SMA levels of hADSC in non-co-cultured (left), co-cultured with SKOV-I6/SC (middle) or co-cultured with SKOV-I6/shPOSTN#1 cells (right). Scale bar = 50 μm. J Western blot analysis for detecting TGFB2 and POSTN in POSTN overexpressing SKOV3 cells infected with lentiviral vector shTGFB2 or a scrambled control. K The diagram illustrates the co-culture system used for cancer cell-induced CAF markers activation. L Representative immunofluorescence images showing the α-SMA levels of hADSC in co-cultured with SKOV3/ctrl (left), SKOV3/POSTN-scrambled control (middle) or SKOV3/POSTN-shTGFB2#2 cells (right). * p < 0.05; ** p < 0.01; *** p < 0.001

Article Snippet: The expression level of secreted TGF-β1 or TGF-β2 was detected by using human TGF-beta1 ELISA kit (#ELH-TGFb1) or human TGF-beta2 ELISA kit (#ELH-TGFb2-1) (RayBiotech, Peachtree Corners, GA, USA) according to user manual.

Techniques: Expressing, Quantitative RT-PCR, Cell Culture, Proliferation Assay, Incubation, Enzyme-linked Immunosorbent Assay, Derivative Assay, Immunohistochemical staining, Western Blot, Infection, Plasmid Preparation, Co-Culture Assay, Activation Assay, Immunofluorescence

Journal: Journal of Translational Medicine

Article Title: Role of epidermal growth factor receptor inhibitor-induced interferon pathway signaling in the head and neck squamous cell carcinoma therapeutic response

doi: 10.1186/s12967-021-02706-8

Figure Lengend Snippet: Variation in CXCL10 and IL6 induction by the pan-ERBB inhibitor, AZD8931, and MEK inhibitor, trametinib, in human HNSCC cell lines. a UMSCC8 and UMSCC25 cells were treated for 1 to 6 days with DMSO, AZD8931 (100 nM) or trametinib (10 nM) and conditioned media was analyzed by ELISA for CXCL10 and IL6. Levels were normalized to total cellular protein and presented as pg/µg protein. The data points represent single determinations at five distinct time points per treatment. b The indicated HNSCC cell lines were treated with DMSO or AZD8931 (100 nM) for 4 days and conditioned media was submitted to ELISA for CXCL10 and IL6. The color bar indicates the expression levels of CXCL10 and IL6 after normalization to cellular protein

Article Snippet: Chemokine levels were measured using the Invitrogen ELISA Kit (Quantikine mouse/human CXCL10/IP-10, IL6, TGFB2 ELISA kits; R&D Systems, Minneapolis, MN) following manufacturer’s instructions.

Techniques: Enzyme-linked Immunosorbent Assay, Expressing

Journal: Journal of Translational Medicine

Article Title: Role of epidermal growth factor receptor inhibitor-induced interferon pathway signaling in the head and neck squamous cell carcinoma therapeutic response

doi: 10.1186/s12967-021-02706-8

Figure Lengend Snippet: Induction of chemokine and cytokine expression by EGFR/ERBB and MEK inhibitor treatment in B4B8 cells. a B4B8 cells were treated with DMSO, AZD8931 (100 nM) or trametinib (10 nM) for 1–9 days. Conditioned media was collected and submitted to ELISA for CXCL10, IL6 and TGFB2. The data are the means and SEM three independent experiments (CXCL10) or single determinations at 6 distinct time points (IL6 and TGFB2) and presented as pg per µg of cellular protein. b B4B8 cells were treated for the indicated time with DMSO or AZD8931 (100 nM) and conditioned media was submitted to Luminex analysis for the indicated secreted analytes. The data points represent single determinations at four distinct time points per treatment

Article Snippet: Chemokine levels were measured using the Invitrogen ELISA Kit (Quantikine mouse/human CXCL10/IP-10, IL6, TGFB2 ELISA kits; R&D Systems, Minneapolis, MN) following manufacturer’s instructions.

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Luminex