jam a sirnas  (Santa Cruz Biotechnology)

Journal: Journal of Translational Autoimmunity

Article Title: A novel epigenetic regulation of JAM-A by EZH2-DNMT3A cascade contributes to T cell adhesion via the activation of Rap1a in lupus patients

doi: 10.1016/j.jtauto.2026.100362

Figure Lengend Snippet: Increased expression of JAM-A in SLE patient PBMCs and peripheral T cells. (A – D) FCM analysis of JAM-A + PBMCs and JAM-A + T cells. (A) JAM-A was stained with anti-human JAM-A-PE antibodies in the PBMCs of healthy individuals (red), new SLE patients (blue), SLE treated (green) and isotype controls (black). (B) Quantification of JAM-A + PBMCs from (A). (C) Double staining with both anti-human JAM-A-PE and anti-human CD3-FITC antibodies. (D) Quantification of JAM-A + T cells from (C), and the error bars indicate the SEM. Healthy, (healthy volunteers, n = 24); new SLE (SLE patients without clinical treatments, n = 9); SLE treated (SLE patients with clinical treatments, n = 15). (E – F) The correlations of EZH2 mRNA/β-actin and the JAM-A mRNA/β-actin in PBMCs ( E ) and T cells ( F ) from SLE patients. ∗ P < 0.05, ∗∗∗ P < 0.001.

Article Snippet: Cells were stimulated overnight with anti-CD3 and anti-CD28 antibodies and transfected with 100 nM EZH2 siRNAs (siG09121182926-1-5/si-h-EZH2_001 and siG09121182954-1-5/si-h-EZH2_002, RiboBio, China), JAM-A siRNAs (SC-43139, Santa Cruz Biotechnology, USA), and negative control siRNA (siN0000001-1-5 for EZH2, SC-37007 for JAM-A) in 6-well plates.

Techniques: Expressing, Staining, Double Staining

Journal: Journal of Translational Autoimmunity

Article Title: A novel epigenetic regulation of JAM-A by EZH2-DNMT3A cascade contributes to T cell adhesion via the activation of Rap1a in lupus patients

doi: 10.1016/j.jtauto.2026.100362

Figure Lengend Snippet: The crosstalk between EZH2/H3K27me3 and DNMT3A/JAM-A. (A, D) The protein expression in Jurkat cells or T cells treated with 5 μM GSK126 (A) or EZH2-siRNA (D), GAPDH or H3 protein as loading control. (B, E) The methylation frequency in JAM-A gene promoter of T cells under EZH2 inhibitor 5 μM GSK126 (B) or EZH2-siRNA treated (E). (C, F) Immunoprecipitated JAM-A with anti-DNMT3A or DNMT3A with anti-H3K27me3 ChIP-grade antibodies in T cells under 5 μM GSK126 (C) or EZH2-siRNA treated (F). The values were normalized to the chromatin levels with 1% input samples. (G, H) The negative correlations of EZH2 mRNA/β-actin and DNMT3A mRNA/β-actin (G) , DNMT3A mRNA/β-actin and JAM-A mRNA/β-actin (H) in T cells from SLE patients. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001, and error bars indicate the SEM.

Article Snippet: Cells were stimulated overnight with anti-CD3 and anti-CD28 antibodies and transfected with 100 nM EZH2 siRNAs (siG09121182926-1-5/si-h-EZH2_001 and siG09121182954-1-5/si-h-EZH2_002, RiboBio, China), JAM-A siRNAs (SC-43139, Santa Cruz Biotechnology, USA), and negative control siRNA (siN0000001-1-5 for EZH2, SC-37007 for JAM-A) in 6-well plates.

Techniques: Expressing, Control, Methylation, Immunoprecipitation

Journal: Journal of Translational Autoimmunity

Article Title: A novel epigenetic regulation of JAM-A by EZH2-DNMT3A cascade contributes to T cell adhesion via the activation of Rap1a in lupus patients

doi: 10.1016/j.jtauto.2026.100362

Figure Lengend Snippet: Inhibition of EZH2 and JAM-A suppresses β1 integrin-mediated T cells and Jurkat cells adhering to ECM. ( A ) The adhesion capacity in T cells from Healthy and SLE patients in vitro . ( B-C ) The adhesion capacity of T cells isolated from healthy or Jurkat cells blocked by 5 μM GSK126 (B) or EZH2-siRNA (C) in vitro . ( D ) Compared the adhesion capacity of T cells from SLE patients, or Jurkat cells with/out neutralizing anti-JAM-A, 1 μg/ml J10.4. (E) Protein expression levels in Jurkat cells overexpressing or knockdown of JAM-A. (F) The adhesion capacity of Jurkat cells with JAM-A regulation. ∗P < 0.05, ∗∗P < 0.01, ∗∗∗ P < 0.001, and error bars indicate the SEM.

Article Snippet: Cells were stimulated overnight with anti-CD3 and anti-CD28 antibodies and transfected with 100 nM EZH2 siRNAs (siG09121182926-1-5/si-h-EZH2_001 and siG09121182954-1-5/si-h-EZH2_002, RiboBio, China), JAM-A siRNAs (SC-43139, Santa Cruz Biotechnology, USA), and negative control siRNA (siN0000001-1-5 for EZH2, SC-37007 for JAM-A) in 6-well plates.

Techniques: Inhibition, In Vitro, Isolation, Expressing, Knockdown

Journal: Journal of Translational Autoimmunity

Article Title: A novel epigenetic regulation of JAM-A by EZH2-DNMT3A cascade contributes to T cell adhesion via the activation of Rap1a in lupus patients

doi: 10.1016/j.jtauto.2026.100362

Figure Lengend Snippet: Inhibition of EZH2 with GSK126 ameliorates lupus-like disease phenotypes in MRL/ lpr mice. (A) Eight-week-old female MRL/ lpr mice were adaptively fed for 2 weeks (wks, n = 15) in the SPF animal room. Mice were randomly divided into a vehicle group (n = 8) and a GSK126-treated group (n = 7) and were treated with an intraperitoneal (IP) injection of β-SEB (20%) or GSK126 (50 mg/kg) every other day for one month. (B) Survival curves of the vehicle and treated group. (C) Anti-dsDNA antibody levels in the plasma. (D) The mRNA expression of IL-10 and TGF-β in splenomegaly. (E) Photomicrographic representation of renal damage (arrow). (F) The percentage plot of glomerulus damage. (G) The Representative histological images of immunohistochemical staining of JAM-A and β1-integrin antibody. (H) The quantification of positive staining areas was quantified by Saiviewer software. (I) The protein expression of EZH2, DNMT3A, JAM-A, Rap1a, β1 integrin and H3K27me3 in splenocytes detected by western blotting. (J) A total of 7 mice per group were analyzed for densitometry by image J software from (I). The symbols represent individual mice, and error bars indicate the SEM. ∗P < 0.05, ∗∗∗P < 0.001, ∗∗∗P < 0.001.

Article Snippet: Cells were stimulated overnight with anti-CD3 and anti-CD28 antibodies and transfected with 100 nM EZH2 siRNAs (siG09121182926-1-5/si-h-EZH2_001 and siG09121182954-1-5/si-h-EZH2_002, RiboBio, China), JAM-A siRNAs (SC-43139, Santa Cruz Biotechnology, USA), and negative control siRNA (siN0000001-1-5 for EZH2, SC-37007 for JAM-A) in 6-well plates.

Techniques: Inhibition, Injection, Clinical Proteomics, Expressing, Immunohistochemical staining, Staining, Software, Western Blot

Journal: Journal of Translational Autoimmunity

Article Title: A novel epigenetic regulation of JAM-A by EZH2-DNMT3A cascade contributes to T cell adhesion via the activation of Rap1a in lupus patients

doi: 10.1016/j.jtauto.2026.100362

Figure Lengend Snippet: Proposed schematic model of the EZH2/miR-26a-5p signaling axis involved in the development of SLE. Increased EZH2 expression is mediated by miR-26a-5p, which in turn is epigenetically repressed by EZH2-mediated H3K27me3 trimethylation within the miR-26a-5p promoter, thus forming a vicious cycle. Crosstalk between H3K27me3 and CpG island methylation mediated by DNMT3A within the JAM-A promoter, resulted in increased expression of JAM-A. Increased expression of JAM-A up-regulated the expression of Rap1a, a regulator of β1-integrin, which is functionally relevant to T cell adhesion. Combined with the increased transcriptional level of Rap1a mediated by miR-26a-5p, ultimately, Rap1a led to increased expression of β1-integrin, which is involved in increasing T cell adhesion.

Article Snippet: Cells were stimulated overnight with anti-CD3 and anti-CD28 antibodies and transfected with 100 nM EZH2 siRNAs (siG09121182926-1-5/si-h-EZH2_001 and siG09121182954-1-5/si-h-EZH2_002, RiboBio, China), JAM-A siRNAs (SC-43139, Santa Cruz Biotechnology, USA), and negative control siRNA (siN0000001-1-5 for EZH2, SC-37007 for JAM-A) in 6-well plates.

Techniques: Expressing, Methylation

Journal: International journal of molecular medicine

Article Title: Jam3 promotes migration and suppresses apoptosis of renal carcinoma cell lines.

doi: 10.3892/ijmm.2018.3854

Figure Lengend Snippet: Figure 1. mRNA and protein expression of Jam3 in HK‑2, Caki‑1 and 786‑0 cells. (A) Western blot analysis was performed to detect the protein level of Jam3, which showed that the protein expression of Jam3 in Caki‑1 and 786‑0 cells was increased compared with that in HK‑2 cells (*P<0.05). (B) Reverse transcription‑polymerase chain reaction analysis was performed to determine the mRNA expression levels of Jam3. Caki‑1 and 786‑0 cells were compared with HK‑2 cells. β‑actin was used as a loading control (*P<0.05). Jam3, junctional adhesion molecule 3; GAPDH, glyceraldehyde 3‑phosphate dehydrogenase.

Article Snippet: Jam3 small interfering (si)RNA (sc-43872) and negative siRNAs (sc-37007) were obtained from Santa Cruz Biotechnology, Inc.

Techniques: Expressing, Western Blot, Reverse Transcription Polymerase Chain Reaction, Control

Journal: International journal of molecular medicine

Article Title: Jam3 promotes migration and suppresses apoptosis of renal carcinoma cell lines.

doi: 10.3892/ijmm.2018.3854

Figure Lengend Snippet: Figure 2. Increased apoptosis of Caki‑1 and 786‑0 cells transfected with Jam3 siRNA. (A) Cellular Jam3 protein was collected from Caki‑1 and 786‑0 cells transfected with non‑specific siRNA or Jam3 siRNA, and the protein levels of Jam3 in each group were assessed by western blot analysis. GAPDH was used as a loading control (*P<0.05). (B) Effects of Jam3 knockdown on the apoptosis of Caki‑1 and 786‑0 cells were measured using flow cytometry (*P<0.05). Jam3, junctional adhesion molecule 3; GAPDH, glyceraldehyde 3‑phosphate dehydrogenase; siRNA, small interfering RNA; PI, propidium iodide.

Article Snippet: Jam3 small interfering (si)RNA (sc-43872) and negative siRNAs (sc-37007) were obtained from Santa Cruz Biotechnology, Inc.

Techniques: Transfection, Western Blot, Control, Knockdown, Flow Cytometry, Small Interfering RNA

Journal: International journal of molecular medicine

Article Title: Jam3 promotes migration and suppresses apoptosis of renal carcinoma cell lines.

doi: 10.3892/ijmm.2018.3854

Figure Lengend Snippet: Figure 3. Jam3 mediates the migration of Caki‑1 and 786‑0 cells. (A) Wound sites/interval between the two wound sites of Caki‑1 cells) were detected and images were captured. Images show the repair of the wound in the two groups (*P<0.05). (B) Wound sites/interval between the two wound sites of 786‑0 cells were measured and images were captured. Images show the repair of the wound in the two groups (*P<0.05). (C) Migration of Caki‑1 and 786‑0 cells was measured using a Transwell chamber (*P<0.05). Original magnification of all images, x200. Jam3, junctional adhesion molecule 3; siRNA, small interfering RNA.

Article Snippet: Jam3 small interfering (si)RNA (sc-43872) and negative siRNAs (sc-37007) were obtained from Santa Cruz Biotechnology, Inc.

Techniques: Migration, Small Interfering RNA

Journal: International journal of molecular medicine

Article Title: Jam3 promotes migration and suppresses apoptosis of renal carcinoma cell lines.

doi: 10.3892/ijmm.2018.3854

Figure Lengend Snippet: Figure 4. Differential expression of E‑cadherin, N‑cadherin, integrin β1 and MMP‑2 in Caki‑1 and 786‑0 cells transfected with non‑specific siRNA and Jam3 siRNA. (A) Cellular protein was collected from Caki‑1 and 786‑0 cells transfected with non‑specific siRNA or Jam3 siRNA. Western blot analysis was performed to assess the levels of E‑cadherin, N‑cadherin, integrin β1, MMP‑2, Bax and Bcl‑2. GAPDH was used as a loading control. (B) Average grey values represented as a histogram (*P<0.05). Jam3, junctional adhesion molecule 3; MMP‑2, matrix metalloproteinase 2; Bcl‑2, B‑cell lymphoma 2; Bax, Bcl‑2‑associated X protein; GAPDH, glyceraldehyde 3‑phosphate dehydrogenase; siRNA, small interfering RNA.

Article Snippet: Jam3 small interfering (si)RNA (sc-43872) and negative siRNAs (sc-37007) were obtained from Santa Cruz Biotechnology, Inc.

Techniques: Quantitative Proteomics, Transfection, Western Blot, Control, Small Interfering RNA

Journal: Cell Death & Disease

Article Title: JAM-A knockdown accelerates the proliferation and migration of human keratinocytes, and improves wound healing in rats via FAK/Erk signaling

doi: 10.1038/s41419-018-0941-y

Figure Lengend Snippet: a Time points selected for the in vitro study. Generally, siRNA duplexes for scramble (Scr) or JAM-A were transfected into keratinocyte cultures on day 0. After 24-h reaction, the mixture was replaced with fresh medium, cells were then cultured for another 24 h followed by different assays to examine the protein expression, cell proliferation, and migration at appointed times. b Western blot showing an ~70% knockdown of JAM-A expression in keratinocytes 2 days after RNAi treatment. Densitometric analysis of JAM-A immunoblotting data normalized against actin with scramble RNAi arbitrarily set at 1. Each bar represents mean ± SD from four experiments. ** P < 0.01. c The proliferation of keratinocytes after JAM-A silencing was evaluated by MTT assay at 0, 24, 48 h. Bars represent the mean ± SD, n = 4. d The migration of keratinocytes after JAM-A silencing was evaluated by cell scratch assay. Notably, keratinocytes were pre-treated with mitomycin C for 1 h before scratch assay. Images at 0, 24, and 48 h post-scratching were captured to display the process of gap closure. Scale bars = 100 µm. e Densitometric analysis of the data from scratch assay normalized against the gap length at 0 h in scramble RNAi group, which was designated as 1. Bars are means ± SD, n = 4. * P < 0.05; ** P < 0.01

Article Snippet: To silence JAM-A, a mixture of 100 nM JAM-A siRNA duplexes (50 nM sequence #1: 5′-GGAUAGUGAUGCCUACGAAdTdT-3′ + 50 nM sequence #2: 5′-dTdTCCUAUCACUACGGAUGCUU-3′, RiboBio Co. Ltd., Guangzhou, China) vs. 100 nM nontargeting scramble siRNA duplexes was used.

Techniques: In Vitro, Transfection, Cell Culture, Expressing, Migration, Western Blot, Knockdown, MTT Assay, Wound Healing Assay

Journal: Cell Death & Disease

Article Title: JAM-A knockdown accelerates the proliferation and migration of human keratinocytes, and improves wound healing in rats via FAK/Erk signaling

doi: 10.1038/s41419-018-0941-y

Figure Lengend Snippet: a Time points selected for the in vivo study. Generally, on day 0 the dorsal skin of SD rats received 1 × 1 cm 2 full-thickness excision followed by immediate RNAi transfection at the wound edge, the RNAi transfection was repeated one more time on day 2. b Skin tissues at the wound edge from day-5 rats were collected, followed by western blot analysis. Densitometric analysis of JAM-A immunoblotting data normalized against actin with scramble RNAi arbitrarily set at 1. Each bar represents mean ± SD from four experiments. ** P < 0.01. c In wound healing assay, DMSO, PF-562271, or PD98059 was co-administrated with siRNA on day 0 and 2. Images showing the healing process of rat dorsal skin wounds were captured and recorded on day 1, 5, and 8. Scale bars = 1 cm. d Densitometric analysis of the in vivo wound healing data normalized against the wound area on day 1 in (Scr RNAi + Veh) group which was designated as 1. Each bar represents the mean ± SD, n = 4. ** P < 0.01

Article Snippet: To silence JAM-A, a mixture of 100 nM JAM-A siRNA duplexes (50 nM sequence #1: 5′-GGAUAGUGAUGCCUACGAAdTdT-3′ + 50 nM sequence #2: 5′-dTdTCCUAUCACUACGGAUGCUU-3′, RiboBio Co. Ltd., Guangzhou, China) vs. 100 nM nontargeting scramble siRNA duplexes was used.

Techniques: In Vivo, Transfection, Western Blot, Wound Healing Assay