Journal: Journal of Translational Medicine

Article Title: Psoriatic microRNAs induce NK cell activation via an innate immune crosstalk abrogated by the Toll-like receptor 7/8 antagonist Enpatoran

doi: 10.1186/s12967-026-07909-5

Figure Lengend Snippet: NK cell activation by pso-miR depends on TLR7/8-expressing accessory cells. ( A - D ) PBMCs or highly purified NK cells were stimulated with pso-miR (10 \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\mu $$\end{document} g/ml) or with vehicle alone (NT) for 24 hours. Where indicated, PBMCs were pretreated with Enpatoran (enp, 1 \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\mu $$\end{document} M) for 1 hour. ( A ) After stimulation, PBMCs or highly purified NK cells were cocultured with NK-sensitive K562 cells for 4 hours at the indicated cell ratio. Data are expressed as mean ± SEM ( n = 3) of the percentage of K562 cell lysis. ( B ) PBMCs or highly purified NK cells were stimulated as in A and then cocultured with K562 cells at a 1:1 ratio. CD107a expression was assessed by flow cytometry. Data are expressed as mean ± SEM ( n = 3) of the percentage of CD107a \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$^ + $$\end{document} NK cells. ( C ) Surface expression of CD69 on CD56 + CD3 – NK cells was assessed by flow cytometry. Data are expressed as mean ± SEM ( n = 3) of the percentage of CD69 NK cells. ( D ) for the analysis of intracellular IFN- \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\gamma $$\end{document} , brefeldin a (5 \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\mu $$\end{document} g/ml) was added for the final 5 hours of culture. Left, representative flow cytometry graphs showing intracellular staining of IFN- \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\gamma $$\end{document} in CD56 + CD3 – NK cells in PBMCs (upper panels) or purified NK cells (lower panels). Right, bar graphs from three independent experiments. Data are expressed as mean ± SEM ( n = 3) of the percentage of IFN- \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$${\gamma ^ + }$$\end{document} producing NK cells in PBMCs (upper panel) or purified NK cells (lower panel). (A-D) * p < 0.05 versus “NT” and # p < 0.05 versus “pso-miR” by one-way ANOVA with Tukey’s post-hoc test. ( E ) Highly purified NK cells were labelled with CellTracker Violet (CTV) and added to autologous PBMCs. IFN- \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\gamma $$\end{document} expression in CTV - (untouched) and CTV + (previously purified) NK cells was assessed by flow cytometry after 24 hours of stimulation with pso-miR. One representative experiment out of three is shown. ( F , left panels) real-time PCR showing the expression of TLR7 and TLR8 mRNAs in highly purified NK cells ( n = 5). pDcs ( n = 3) and monocyte-derived DCs (moDC, n = 3) were used as positive controls of TLR7 and TLR8 expression respectively. Data are expressed as mean ± SEM of 2 -ΔCt relative to HPRT. ( F , right panels) indicated cell types, including HEK293 cells transfected to overexpress TLR7 or TLR8 (HEK293–TLR7 and HEK293–TLR8), were lysed and the expression of TLR7, TLR8 and \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\beta $$\end{document} -actin was determined by Western blot. One representative fluorogram out of three is shown

Article Snippet: Equal amounts of extracts were analyzed through SDS-PAGE followed by Western blotting with antibodies against TLR7 (rabbit monoclonal, 5632, Cell Signaling Technology), TLR8 (rabbit monoclonal, 11,886, Cell Signaling Technology), and β-actin (mouse monoclonal, C4, sc-47778, Santa Cruz Biotechnology).

Techniques: Activation Assay, Expressing, Purification, Lysis, Flow Cytometry, Staining, Real-time Polymerase Chain Reaction, Derivative Assay, Transfection, Western Blot

Journal: Journal of Traditional and Complementary Medicine

Article Title: Bushen Huoxue Recipe enhances the immune microenvironment at the maternal-fetal interface via the Hippo signaling pathway in mice with recurrent spontaneous abortion

doi: 10.1016/j.jtcme.2025.02.005

Figure Lengend Snippet: Effect of BSHXR on the expression of inflammatory factors. (A) The mRNA expression of TLR2, TLR4, and TLR7 in decidual tissue (n = 6). (TLR2: Con vs. RSA, p = 0.0036; RSA vs. CsA, p = 0.0443; RSA vs. BSHXR, p = 0.0034; TLR4: Con vs. RSA, p = 0.0097; RSA vs. CsA, p = 0.0260; RSA vs. BSHXR, p = 0.0051; TLR7: Con vs. RSA, p = 0.0104; RSA vs. CsA, p = 0.0315; RSA vs. BSHXR, p = 0.0229). (B) Representative images of immunofluorescence staining for TLR2, TLR4, and TLR7 in decidual tissues. Scale bar: 50 μm. (C) The protein expression of TLR2, TLR4, and TLR7 in decidual tissue. (D) Quantification of the protein levels (n = 3). (TLR2: Con vs. RSA, p = 0.0062; RSA vs. CsA, p = 0.0158; RSA vs. BSHXR, p = 0.0222; TLR4: Con vs. RSA, p = 0.0001; RSA vs. CsA, p = 0.0001; RSA vs. BSHXR, p < 0.0001; TLR7: Con vs. RSA, p = 0.0099; RSA vs. CsA, p = 0.0129; RSA vs. BSHXR, p = 0.0045). The results are expressed as the means ± SEMs. ∗ p < 0.05, ∗∗ p < 0.01 and ∗∗∗ p < 0.001, compared with the control group; # p < 0.05, ## p < 0.01, ### p < 0.001 and, ### p < 0.0001, compared with the RSA group.

Article Snippet: Antibodies against TLR2 (66645-1-Ig), TLR4 (66350-1-Ig), TLR7 (17232-1-AP), and GAPDH (60004-1-Ig) were purchased from Proteintech (Rosemont, IL, USA); antibodies against RORγt (ab207082) and FoxP3 (ab215206) were purchased from Abcam (Cambridge, MA, USA); Alexa Fluor 555 nm (8953) and antibodies against MST (14946), p-MST (49332), LATS (3477S), p-LATS (8654S), YAP (14074S), p-YAP (13008S), TAZ (83669S), and p-TAZ (59971S) were purchased from Cell Signaling Technology (Danvers, MA, USA); and an antibody against Lamin B (A11495) was purchased from ABclonal (Wuhan, China).

Techniques: Expressing, Immunofluorescence, Staining, Control