Journal: Advanced Science

Article Title: Chrysanthemum Morifolium– Derived Nanovesicles Reduce Lactate‐Associated Histone Lactylation and Modulate Microglia‐Mediated Neuroinflammation in Sepsis‐Associated Encephalopathy

doi: 10.1002/advs.78330

Figure Lengend Snippet: CRELNVs mitigated memory impairment in the hippocampus of SAE mice. (A) Schematic of the experimental procedure used to probe the effect of CRELNVs on memory impairment in the hippocampi of sepsis‐associated encephalopathy (SAE) mice. (B) Center times in open field test, n = 10. (C) Object discrimination times in novel‐object recognition test, n = 10. (D) Spontaneous alternation rates in Y‐Maze, n = 10. (E) Hematoxylin and eosin, Nissl, and TUNEL staining images. (F) Nissl staining quantitative analysis in CA3 region, n = 6. (G) TUNEL staining quantitative analysis in DG region, n = 6. (H) Interleukin‐1β (IL‐1β), (I) tumor necrosis factor‐α (TNF‐α), (J) interleukin‐6 (IL‐6), and (K) interleukin‐10 (IL‐10) levels in the hippocampi of SAE mice, n = 6. # p < 0.05, ## p < 0.01, ### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. lipopolysaccharide (LPS) group.

Article Snippet: IL‐1β (Cat. No. KE10003), IL‐6 (Cat. No. KE10007), TNF‐α (Cat. No. KE10002), and interleukin‐10 (IL‐10; Cat. No. KE1008) Enzyme‐linked immunosorbent assay (ELISA) kits were procured from Proteintech Group (Wuhan, China).

Techniques: TUNEL Assay, Staining, Analysis, Control

Journal: Advanced Science

Article Title: Chrysanthemum Morifolium– Derived Nanovesicles Reduce Lactate‐Associated Histone Lactylation and Modulate Microglia‐Mediated Neuroinflammation in Sepsis‐Associated Encephalopathy

doi: 10.1002/advs.78330

Figure Lengend Snippet: CRELNVs influence microglial homeostasis in the hippocampus of SAE mice. (A) Representative images of immunofluorescence staining for Iba‐1in the hippocampi of SAE mice. (B) Number of Iba‐1 + cells, (C) number of total branch points/cell, and (D) total branch length for panel A, n = 6. (E) IL‐1β, (F) TNF‐α, (G) IL‐6, and (H) IL‐10 mRNA expression in the hippocampi of SAE mice determined using the quantitative polymerase chain reaction (qPCR), n = 3. # p < 0.05, ## p < 0.01, ### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. LPS group).

Article Snippet: IL‐1β (Cat. No. KE10003), IL‐6 (Cat. No. KE10007), TNF‐α (Cat. No. KE10002), and interleukin‐10 (IL‐10; Cat. No. KE1008) Enzyme‐linked immunosorbent assay (ELISA) kits were procured from Proteintech Group (Wuhan, China).

Techniques: Immunofluorescence, Staining, Expressing, Real-time Polymerase Chain Reaction, Control

Journal: Advanced Science

Article Title: Chrysanthemum Morifolium– Derived Nanovesicles Reduce Lactate‐Associated Histone Lactylation and Modulate Microglia‐Mediated Neuroinflammation in Sepsis‐Associated Encephalopathy

doi: 10.1002/advs.78330

Figure Lengend Snippet: The neuro‐protection and Anti‐inflammation activities of CRELNVs in vitro. (A) Protective effect of CRELNVs on SH‐SY5Y cells in an inflammatory environment determined using the CCK8 assay, n = 4. (B) Microplate reader–detected NO contents in BV2 cell supernatants, n = 3. (C) IL‐1β, (D) IL‐6, (E) TNF‐α, and (F) IL‐10 levels in BV2 cell supernatants, n = 3. (G) Representative fluorescence microscopy images acquired to evaluate reactive oxygen species (ROS) content. (H, I) Flow cytometry peaks and ROS contents derived therefrom, n = 3. (J, K) Western blot analysis: NLRP3 levels in BV2 cells, n = 3. # p < 0.05, ## p < 0.01, ### p < 0.001 vs. control group; * p < 0.05, ** p < 0.01, *** p < 0.001 vs. LPS group.

Article Snippet: IL‐1β (Cat. No. KE10003), IL‐6 (Cat. No. KE10007), TNF‐α (Cat. No. KE10002), and interleukin‐10 (IL‐10; Cat. No. KE1008) Enzyme‐linked immunosorbent assay (ELISA) kits were procured from Proteintech Group (Wuhan, China).

Techniques: In Vitro, CCK-8 Assay, Fluorescence, Microscopy, Flow Cytometry, Derivative Assay, Western Blot, Analysis, Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: Casticin From Sanse Powder‐ In Vitro Release Medium Inhibits FAK / PI3K / AKT Phosphorylation Reducing Synovial Senescence and Alleviates Inflammation in Knee Osteoarthritis

doi: 10.1111/jcmm.71391

Figure Lengend Snippet: SSP‐IVRM alleviates synovial inflammation and pathological progression in KOA rats. (A) ELISA was used to detect the levels of IL‐1β, IL‐6, CXCL1 and CXCL3 in the serum of rats in each group; (B) Representative images of HE, Masson and Sirius Red staining of knee synovial tissues from each group, along with Krenn's scoring; (C) Representative images of IL‐1β immunofluorescence in knee synovial tissues from each group; (D) Western blot analysis of the protein expression levels of MMP3, MMP9, COL I and TGF‐β in the synovial tissues of rats in each group. (E) qRT‐PCR analysis of MMP3, MMP9, COL1A1 and TGF‐β mRNA expression in synovial tissues. Statistical graphs are presented as Mean ± SD ( n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 versus Ctrl group; # p < 0.05, ## p < 0.01, ### p < 0.001 versus KOA group.

Article Snippet: DMEM high‐glucose medium (Cat# C11995500CP) and fetal bovine serum (Cat# 10099‐141) were purchased from GIBCO (USA); LPS (Cat# L2880) was obtained from Sigma‐Aldrich (USA); Casticin (Cat# B20835 , ≥ 98%) was purchased from Yuanye Biotechnology (China); MIR002 (Cat# HY‐143412), Y15 (Cat# HY‐12444) and VSLRGDTRG acetate (Cat# HY‐P10941A) were obtained from MCE (China); Senescence‐Tracker Senescent Cell Fluorescent Probe (Cat# C0603), Enhanced Mitochondrial Membrane Potential Assay Kit (Cat# C2003S), Reactive Oxygen Species Assay Kit (Cat# S0035S) and Cell Counting Kit‐8 (Cat# C0038) were purchased from Beyotime Biotechnology (China); Rat CXCL1 ELISA Kit (Cat# E‐EL‐R0003), Rat CXCL3 ELISA Kit (Cat# E‐EL‐R0697), Rat IL‐6 ELISA Kit (Cat# E‐EL‐R0015) and Rat IL‐1β ELISA Kit (Cat# E‐EL‐R0012) were obtained from Elabscience (China); TGF‐β Antibody (Cat# 3711), CDC2 Antibody (Cat# 77055), FAK Antibody (Cat# 3285), PI3 Kinase p85 Antibody (Cat# 4292), AKT Antibody (Cat# 9272) and Phospho‐FAK (Tyr397) Antibody (Cat# 3283) were purchased from CST (USA); Phospho‐PI3K p85 (Tyr458)/p55 (Tyr199) Antibody (Cat# AF3242) was obtained from Affinity (China); Phospho‐AKT (Ser473) Antibody (Cat# 80455‐1‐RR), IL‐1β Antibody (Cat# 26048‐1‐AP), IL‐6 Antibody (Cat# 21865‐1‐AP), MMP3 Antibody (Cat# 17873‐1‐AP), MMP9 Antibody (Cat# 10375‐2‐AP), p53 Antibody (Cat# 10442‐1‐AP), p21 Antibody (Cat# 10355‐1‐AP) and GAPDH Antibody (Cat# 60004‐1‐Ig) were purchased from Proteintech (China).

Techniques: Enzyme-linked Immunosorbent Assay, Staining, Immunofluorescence, Western Blot, Analysis, Expressing, Quantitative RT-PCR

Journal: Journal of Cellular and Molecular Medicine

Article Title: Casticin From Sanse Powder‐ In Vitro Release Medium Inhibits FAK / PI3K / AKT Phosphorylation Reducing Synovial Senescence and Alleviates Inflammation in Knee Osteoarthritis

doi: 10.1111/jcmm.71391

Figure Lengend Snippet: CAS alleviates synovial inflammation and cellular senescence in KOA rats. (A) Representative images of HE, Masson and Sirius Red staining of synovial tissue from rat knee joints in each group, along with Krenn's scores; (B) Serum levels of IL‐1β, IL‐6, CXCL1 and CXCL3 in each group of rats as measured by ELISA; (C) Representative immunofluorescence images of IL‐1β in synovial tissue from rat knee joints in each group; (D) Protein expression levels of MMP3, MMP9, COL I, TGF‐β, p53, p21, CDC2 and CCNA1 in synovial tissue from each rat group as determined by Western Blot. (E) qRT‐PCR analysis of the mRNA expression levels of MMP3, MMP9, COL1A1, TGF‐β, p53, p21/CDKN1A, CDC2 and CCNA1 in synovial tissues from each rat group. Statistical graphs are presented as Mean ± SD ( n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 versus Ctrl group; # p < 0.05, ## p < 0.01, ### p < 0.001 versus KOA group.

Article Snippet: DMEM high‐glucose medium (Cat# C11995500CP) and fetal bovine serum (Cat# 10099‐141) were purchased from GIBCO (USA); LPS (Cat# L2880) was obtained from Sigma‐Aldrich (USA); Casticin (Cat# B20835 , ≥ 98%) was purchased from Yuanye Biotechnology (China); MIR002 (Cat# HY‐143412), Y15 (Cat# HY‐12444) and VSLRGDTRG acetate (Cat# HY‐P10941A) were obtained from MCE (China); Senescence‐Tracker Senescent Cell Fluorescent Probe (Cat# C0603), Enhanced Mitochondrial Membrane Potential Assay Kit (Cat# C2003S), Reactive Oxygen Species Assay Kit (Cat# S0035S) and Cell Counting Kit‐8 (Cat# C0038) were purchased from Beyotime Biotechnology (China); Rat CXCL1 ELISA Kit (Cat# E‐EL‐R0003), Rat CXCL3 ELISA Kit (Cat# E‐EL‐R0697), Rat IL‐6 ELISA Kit (Cat# E‐EL‐R0015) and Rat IL‐1β ELISA Kit (Cat# E‐EL‐R0012) were obtained from Elabscience (China); TGF‐β Antibody (Cat# 3711), CDC2 Antibody (Cat# 77055), FAK Antibody (Cat# 3285), PI3 Kinase p85 Antibody (Cat# 4292), AKT Antibody (Cat# 9272) and Phospho‐FAK (Tyr397) Antibody (Cat# 3283) were purchased from CST (USA); Phospho‐PI3K p85 (Tyr458)/p55 (Tyr199) Antibody (Cat# AF3242) was obtained from Affinity (China); Phospho‐AKT (Ser473) Antibody (Cat# 80455‐1‐RR), IL‐1β Antibody (Cat# 26048‐1‐AP), IL‐6 Antibody (Cat# 21865‐1‐AP), MMP3 Antibody (Cat# 17873‐1‐AP), MMP9 Antibody (Cat# 10375‐2‐AP), p53 Antibody (Cat# 10442‐1‐AP), p21 Antibody (Cat# 10355‐1‐AP) and GAPDH Antibody (Cat# 60004‐1‐Ig) were purchased from Proteintech (China).

Techniques: Staining, Tissue, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Expressing, Western Blot, Quantitative RT-PCR, Analysis

Journal: Journal of Cellular and Molecular Medicine

Article Title: Casticin From Sanse Powder‐ In Vitro Release Medium Inhibits FAK / PI3K / AKT Phosphorylation Reducing Synovial Senescence and Alleviates Inflammation in Knee Osteoarthritis

doi: 10.1111/jcmm.71391

Figure Lengend Snippet: CAS improves inflammation and cellular senescence in LPS‐induced rat synovial fibroblast‐like cells. (A) CCK‐8 assay to evaluate the cytotoxic effects of CAS on FLS; (B) ELISA to measure the levels of IL‐1β, IL‐6, CXCL1 and CXCL3 in the culture media of rat FLS from each group; (C) Representative immunofluorescence images of IL‐1β and IL‐6 in FLS from each group; (D) Western blot analysis of protein expression levels of MMP3, MMP9, COL I, TGF‐β, p53, p21, CDC2 and CCNA1 in FLS from each group; (E) CM‐H2DCFDA probe to assess ROS levels in FLS from each group; (F) JC‐1 probe to detect mitochondrial membrane potential levels in FLS from each group; (G) Representative images of senescent cells labelled by Senescence‐Tracker probe in FLS from each group. Statistical graphs are presented as Mean ± SD ( n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 versus Ctrl group; # p < 0.05, ## p < 0.01, ### p < 0.001 versus LPS group.

Article Snippet: DMEM high‐glucose medium (Cat# C11995500CP) and fetal bovine serum (Cat# 10099‐141) were purchased from GIBCO (USA); LPS (Cat# L2880) was obtained from Sigma‐Aldrich (USA); Casticin (Cat# B20835 , ≥ 98%) was purchased from Yuanye Biotechnology (China); MIR002 (Cat# HY‐143412), Y15 (Cat# HY‐12444) and VSLRGDTRG acetate (Cat# HY‐P10941A) were obtained from MCE (China); Senescence‐Tracker Senescent Cell Fluorescent Probe (Cat# C0603), Enhanced Mitochondrial Membrane Potential Assay Kit (Cat# C2003S), Reactive Oxygen Species Assay Kit (Cat# S0035S) and Cell Counting Kit‐8 (Cat# C0038) were purchased from Beyotime Biotechnology (China); Rat CXCL1 ELISA Kit (Cat# E‐EL‐R0003), Rat CXCL3 ELISA Kit (Cat# E‐EL‐R0697), Rat IL‐6 ELISA Kit (Cat# E‐EL‐R0015) and Rat IL‐1β ELISA Kit (Cat# E‐EL‐R0012) were obtained from Elabscience (China); TGF‐β Antibody (Cat# 3711), CDC2 Antibody (Cat# 77055), FAK Antibody (Cat# 3285), PI3 Kinase p85 Antibody (Cat# 4292), AKT Antibody (Cat# 9272) and Phospho‐FAK (Tyr397) Antibody (Cat# 3283) were purchased from CST (USA); Phospho‐PI3K p85 (Tyr458)/p55 (Tyr199) Antibody (Cat# AF3242) was obtained from Affinity (China); Phospho‐AKT (Ser473) Antibody (Cat# 80455‐1‐RR), IL‐1β Antibody (Cat# 26048‐1‐AP), IL‐6 Antibody (Cat# 21865‐1‐AP), MMP3 Antibody (Cat# 17873‐1‐AP), MMP9 Antibody (Cat# 10375‐2‐AP), p53 Antibody (Cat# 10442‐1‐AP), p21 Antibody (Cat# 10355‐1‐AP) and GAPDH Antibody (Cat# 60004‐1‐Ig) were purchased from Proteintech (China).

Techniques: CCK-8 Assay, Enzyme-linked Immunosorbent Assay, Immunofluorescence, Western Blot, Analysis, Expressing, Membrane

Journal: Journal of Cellular and Molecular Medicine

Article Title: Casticin From Sanse Powder‐ In Vitro Release Medium Inhibits FAK / PI3K / AKT Phosphorylation Reducing Synovial Senescence and Alleviates Inflammation in Knee Osteoarthritis

doi: 10.1111/jcmm.71391

Figure Lengend Snippet: CAS reduces the inflammatory state of rat synovial fibroblast‐like cells by lowering cellular senescence levels. (A) Representative images of senescent positive cells in rat FLSs from each group detected using the Senescence‐Tracker probe; (B) Mitochondrial membrane potential levels in rat FLSs from each group measured by the JC‐1 probe; (C) Intracellular ROS levels in rat FLSs from each group assessed with the CM‐H2DCFDA probe; (D) Protein expression levels of MMP3, MMP9, COL I, TGF‐β, p53, p21, CDC2 and CCNA1 in rat FLSs from each group detected by Western blotting (WB); (E) Representative immunofluorescence images of IL‐1β and IL‐6 in rat FLSs from each group; (F) Levels of IL‐1β, IL‐6, CXCL1 and CXCL3 in the culture supernatant of rat FLSs from each group measured by ELISA. Statistical graphs are presented as Mean ± SD ( n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 versus Ctrl group; # p < 0.05, ## p < 0.01, ### p < 0.001 versus LPS group; △ p < 0.05, △△ p < 0.01, △△△ p < 0.001 versus MIR002 group.

Article Snippet: DMEM high‐glucose medium (Cat# C11995500CP) and fetal bovine serum (Cat# 10099‐141) were purchased from GIBCO (USA); LPS (Cat# L2880) was obtained from Sigma‐Aldrich (USA); Casticin (Cat# B20835 , ≥ 98%) was purchased from Yuanye Biotechnology (China); MIR002 (Cat# HY‐143412), Y15 (Cat# HY‐12444) and VSLRGDTRG acetate (Cat# HY‐P10941A) were obtained from MCE (China); Senescence‐Tracker Senescent Cell Fluorescent Probe (Cat# C0603), Enhanced Mitochondrial Membrane Potential Assay Kit (Cat# C2003S), Reactive Oxygen Species Assay Kit (Cat# S0035S) and Cell Counting Kit‐8 (Cat# C0038) were purchased from Beyotime Biotechnology (China); Rat CXCL1 ELISA Kit (Cat# E‐EL‐R0003), Rat CXCL3 ELISA Kit (Cat# E‐EL‐R0697), Rat IL‐6 ELISA Kit (Cat# E‐EL‐R0015) and Rat IL‐1β ELISA Kit (Cat# E‐EL‐R0012) were obtained from Elabscience (China); TGF‐β Antibody (Cat# 3711), CDC2 Antibody (Cat# 77055), FAK Antibody (Cat# 3285), PI3 Kinase p85 Antibody (Cat# 4292), AKT Antibody (Cat# 9272) and Phospho‐FAK (Tyr397) Antibody (Cat# 3283) were purchased from CST (USA); Phospho‐PI3K p85 (Tyr458)/p55 (Tyr199) Antibody (Cat# AF3242) was obtained from Affinity (China); Phospho‐AKT (Ser473) Antibody (Cat# 80455‐1‐RR), IL‐1β Antibody (Cat# 26048‐1‐AP), IL‐6 Antibody (Cat# 21865‐1‐AP), MMP3 Antibody (Cat# 17873‐1‐AP), MMP9 Antibody (Cat# 10375‐2‐AP), p53 Antibody (Cat# 10442‐1‐AP), p21 Antibody (Cat# 10355‐1‐AP) and GAPDH Antibody (Cat# 60004‐1‐Ig) were purchased from Proteintech (China).

Techniques: Membrane, Expressing, Western Blot, Immunofluorescence, Enzyme-linked Immunosorbent Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: Casticin From Sanse Powder‐ In Vitro Release Medium Inhibits FAK / PI3K / AKT Phosphorylation Reducing Synovial Senescence and Alleviates Inflammation in Knee Osteoarthritis

doi: 10.1111/jcmm.71391

Figure Lengend Snippet: CAS inhibits FAK/PI3K/AKT phosphorylation process to reduce synovial cell senescence and improve synovial inflammation. (A) Representative immunofluorescence images of p‐FAK and p‐PI3K in FLS from each rat group; (B) Western blot analysis of protein expression levels of p‐FAK, FAK, p‐PI3K, PI3K, p‐AKT and AKT in FLS from each group; (C) Western blot analysis of protein expression levels of MMP9, COL I, TGF‐β, p53, p21, CDC2 and CCNA1 in FLS from each group; (D) Assessment of mitochondrial membrane potential in FLS from each group using the JC‐1 probe; (E) Detection of ROS levels in FLS from each group using the CM‐H2DCFDA probe; (F) Representative images of senescent FLS detected with the Senescence‐Tracker probe in each group; (G) ELISA analysis of IL‐1β, IL‐6, CXCL1 and CXCL3 levels in the culture medium of FLS from each group. Statistical graphs are presented as Mean ± SD ( n = 3). * p < 0.05, ** p < 0.01, *** p < 0.001 versus Ctrl group; # p < 0.05, ## p < 0.01, ### p < 0.001 versus LPS group; △ p < 0.05, △△ p < 0.01, △△△ p < 0.001 versus CAS group.

Article Snippet: DMEM high‐glucose medium (Cat# C11995500CP) and fetal bovine serum (Cat# 10099‐141) were purchased from GIBCO (USA); LPS (Cat# L2880) was obtained from Sigma‐Aldrich (USA); Casticin (Cat# B20835 , ≥ 98%) was purchased from Yuanye Biotechnology (China); MIR002 (Cat# HY‐143412), Y15 (Cat# HY‐12444) and VSLRGDTRG acetate (Cat# HY‐P10941A) were obtained from MCE (China); Senescence‐Tracker Senescent Cell Fluorescent Probe (Cat# C0603), Enhanced Mitochondrial Membrane Potential Assay Kit (Cat# C2003S), Reactive Oxygen Species Assay Kit (Cat# S0035S) and Cell Counting Kit‐8 (Cat# C0038) were purchased from Beyotime Biotechnology (China); Rat CXCL1 ELISA Kit (Cat# E‐EL‐R0003), Rat CXCL3 ELISA Kit (Cat# E‐EL‐R0697), Rat IL‐6 ELISA Kit (Cat# E‐EL‐R0015) and Rat IL‐1β ELISA Kit (Cat# E‐EL‐R0012) were obtained from Elabscience (China); TGF‐β Antibody (Cat# 3711), CDC2 Antibody (Cat# 77055), FAK Antibody (Cat# 3285), PI3 Kinase p85 Antibody (Cat# 4292), AKT Antibody (Cat# 9272) and Phospho‐FAK (Tyr397) Antibody (Cat# 3283) were purchased from CST (USA); Phospho‐PI3K p85 (Tyr458)/p55 (Tyr199) Antibody (Cat# AF3242) was obtained from Affinity (China); Phospho‐AKT (Ser473) Antibody (Cat# 80455‐1‐RR), IL‐1β Antibody (Cat# 26048‐1‐AP), IL‐6 Antibody (Cat# 21865‐1‐AP), MMP3 Antibody (Cat# 17873‐1‐AP), MMP9 Antibody (Cat# 10375‐2‐AP), p53 Antibody (Cat# 10442‐1‐AP), p21 Antibody (Cat# 10355‐1‐AP) and GAPDH Antibody (Cat# 60004‐1‐Ig) were purchased from Proteintech (China).

Techniques: Phospho-proteomics, Immunofluorescence, Western Blot, Analysis, Expressing, Membrane, Enzyme-linked Immunosorbent Assay

Journal: Biology

Article Title: Temporal Evolution and Dissociation of Neuroendocrine Stress Inflammatory Responses Neurological Dysfunction and Multiorgan Injury Following Experimental Polytrauma in Mice

doi: 10.3390/biology15191788

Figure Lengend Snippet: Counter-regulatory cytokine responses in serum and injured brain tissue following polytrauma. ( A ) Serum IL-4. ( B ) IL-4 in injured brain tissue. ( C ) Serum IL-10. ( D ) IL-10 in injured brain tissue. ( E ) Serum TGF-β. ( F ) TGF-β in injured brain tissue. ELISA: n = 6 per group at each time point. Brain-tissue values were normalized to total protein. Data are mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001; ns—not significant.

Article Snippet: Concentrations of IL-1β, IL-6, TNF-α, IL-4, IL-10 and TGF-β in peripheral serum and injured brain tissue homogenates were determined using enzyme-linked immunosorbent assay kits (CSB-E08054m, CSB-E04639m, CSB-E04741m, CSB-E04634m-IS, CSB-E04594m-IS, CSB-E04726m-IS, CUSABIO, https://www.cusabio.com/ ) according to the manufacturer’s instructions.

Techniques: Tissue, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Molecular Biosciences

Article Title: Integrated transcriptome and single-cell RNA sequencing identifies transient receptor potential channel-related biomarkers in ulcerative colitis

doi: 10.3389/fmolb.2026.1902447

Figure Lengend Snippet: Primer sequences for RT-qPCR experiment.

Article Snippet: For Western blot analysis, equal amounts of protein (50 μg) were denatured in SDS loading buffer, separated by SDS-PAGE (80 V for 25 min, then 120 V for 50 min), and transferred to PVDF membranes (Millipore, MA, United States) at 200 mA for 2 h. Membranes were blocked with blocking buffer (NCM Bio, P30500 ) for 10 min at room temperature and subsequently incubated overnight at 4 °C with primary antibodies against pro-IL-1β (Proteintech, 16806-1-AP, 1:1,000), IL-1β (Immunoway, YM8498, 1:1,000), and Pik3r3 (Proteintech, 27035-1-AP, 1:1,000).

Techniques:

Journal: Frontiers in Molecular Biosciences

Article Title: Integrated transcriptome and single-cell RNA sequencing identifies transient receptor potential channel-related biomarkers in ulcerative colitis

doi: 10.3389/fmolb.2026.1902447

Figure Lengend Snippet: Identification of 22 candidate genes associated with TRP channel in UC. (A) Volcano plot of DEGs between UC and HC groups. Red and blue dots represent significantly upregulated and downregulated genes respectively. The top 10 up- and downregulated genes are labeled; (B) Venn diagram showing the intersection of 3,247 DEGs and 147 TRP channel-related genes, yielding 22 candidate genes; (C) PPI network of the 22 candidate genes; (D) GO and KEGG pathway enrichment analysis of the 22 candidate genes. The top 5 enriched pathways were presented.

Article Snippet: For Western blot analysis, equal amounts of protein (50 μg) were denatured in SDS loading buffer, separated by SDS-PAGE (80 V for 25 min, then 120 V for 50 min), and transferred to PVDF membranes (Millipore, MA, United States) at 200 mA for 2 h. Membranes were blocked with blocking buffer (NCM Bio, P30500 ) for 10 min at room temperature and subsequently incubated overnight at 4 °C with primary antibodies against pro-IL-1β (Proteintech, 16806-1-AP, 1:1,000), IL-1β (Immunoway, YM8498, 1:1,000), and Pik3r3 (Proteintech, 27035-1-AP, 1:1,000).

Techniques: Labeling, Analysis

Journal: Frontiers in Molecular Biosciences

Article Title: Integrated transcriptome and single-cell RNA sequencing identifies transient receptor potential channel-related biomarkers in ulcerative colitis

doi: 10.3389/fmolb.2026.1902447

Figure Lengend Snippet: Identification of TRP channel-related feature genes in UC. (A) LASSO regression analysis with 5-fold cross-validation. The vertical dashed line indicates the optimal penalty parameter (λ). (B) Feature selection performed by the SVM-RFE algorithm; (C) Venn diagram showing the overlap between LASSO- and SVM-RFE-selected genes, resulting in five shared feature genes; (D) ROC analysis of IL-1β; (E) ROC analysis of PIK3R3; (F) ROC analysis of ADCY3; (G) ROC analysis of ADCY7; (H) ROC analysis of TRPV1.

Article Snippet: For Western blot analysis, equal amounts of protein (50 μg) were denatured in SDS loading buffer, separated by SDS-PAGE (80 V for 25 min, then 120 V for 50 min), and transferred to PVDF membranes (Millipore, MA, United States) at 200 mA for 2 h. Membranes were blocked with blocking buffer (NCM Bio, P30500 ) for 10 min at room temperature and subsequently incubated overnight at 4 °C with primary antibodies against pro-IL-1β (Proteintech, 16806-1-AP, 1:1,000), IL-1β (Immunoway, YM8498, 1:1,000), and Pik3r3 (Proteintech, 27035-1-AP, 1:1,000).

Techniques: Analysis, Biomarker Discovery, Selection

Journal: Frontiers in Molecular Biosciences

Article Title: Integrated transcriptome and single-cell RNA sequencing identifies transient receptor potential channel-related biomarkers in ulcerative colitis

doi: 10.3389/fmolb.2026.1902447

Figure Lengend Snippet: Nomogram construction and validation based on the expression of the 5 feature genes. (A) Violin plot of ADCY3, ADCY7, IL-1β, PIK3R3 and TRPV1 expression in the training cohort ( GSE87466 ); (B) Violin plot of ADCY3, ADCY7, IL-1β, PIK3R3 and TRPV1 expression in the validation cohort ( GSE75214 ); (C) ROC analysis of IL-1β, PIK3R3, ADCY3, ADCY7 and TRPV1 in the validation cohort ( GSE75214 ); (D) Nomogram based on the expression of IL-1β, PIK3R3, ADCY3, ADCY7 and TRPV1; (E) Calibration curves for the predicted probability; (F) DCA of nomogram for UC; (G) ROC curve analysis of nomogram for UC probability.

Article Snippet: For Western blot analysis, equal amounts of protein (50 μg) were denatured in SDS loading buffer, separated by SDS-PAGE (80 V for 25 min, then 120 V for 50 min), and transferred to PVDF membranes (Millipore, MA, United States) at 200 mA for 2 h. Membranes were blocked with blocking buffer (NCM Bio, P30500 ) for 10 min at room temperature and subsequently incubated overnight at 4 °C with primary antibodies against pro-IL-1β (Proteintech, 16806-1-AP, 1:1,000), IL-1β (Immunoway, YM8498, 1:1,000), and Pik3r3 (Proteintech, 27035-1-AP, 1:1,000).

Techniques: Biomarker Discovery, Expressing, Analysis

Journal: Frontiers in Molecular Biosciences

Article Title: Integrated transcriptome and single-cell RNA sequencing identifies transient receptor potential channel-related biomarkers in ulcerative colitis

doi: 10.3389/fmolb.2026.1902447

Figure Lengend Snippet: Functional pathway enrichment analysis of 5 TRP channel-related feature genes. (A) GSEA of IL-1β; (B) GSEA of PIK3R3; (C) GSEA of ADCY3; (D) GSEA of ADCY7; (E) GSEA of TRPV1; (F) Hotmap of correlation between 5 TRP channel-related feature genes and 50 shared pathways.

Article Snippet: For Western blot analysis, equal amounts of protein (50 μg) were denatured in SDS loading buffer, separated by SDS-PAGE (80 V for 25 min, then 120 V for 50 min), and transferred to PVDF membranes (Millipore, MA, United States) at 200 mA for 2 h. Membranes were blocked with blocking buffer (NCM Bio, P30500 ) for 10 min at room temperature and subsequently incubated overnight at 4 °C with primary antibodies against pro-IL-1β (Proteintech, 16806-1-AP, 1:1,000), IL-1β (Immunoway, YM8498, 1:1,000), and Pik3r3 (Proteintech, 27035-1-AP, 1:1,000).

Techniques: Functional Assay, Analysis

Journal: Frontiers in Molecular Biosciences

Article Title: Integrated transcriptome and single-cell RNA sequencing identifies transient receptor potential channel-related biomarkers in ulcerative colitis

doi: 10.3389/fmolb.2026.1902447

Figure Lengend Snippet: Pseudotime trajectory analysis and the expression dynamics of the five TRP channel-related feature genes in B and Plasma cells. (A) The top 50 PCs selection of B and Plasma cells; (B) 9 cell clusters were identified by tSNE reduced dimensionality in B and Plasma cells; (C) Pseudotime trajectory analysis from timing differences in cell differentiation, different cell clusters, developmental stages and different samples (HC and UC); (D) The expression dynamics of the five TRP channel-related feature genes along the pseudotime trajectory of B and plasma cells; (E) IL-1β and PIK3R3 expression in 5 developmental stages.

Article Snippet: For Western blot analysis, equal amounts of protein (50 μg) were denatured in SDS loading buffer, separated by SDS-PAGE (80 V for 25 min, then 120 V for 50 min), and transferred to PVDF membranes (Millipore, MA, United States) at 200 mA for 2 h. Membranes were blocked with blocking buffer (NCM Bio, P30500 ) for 10 min at room temperature and subsequently incubated overnight at 4 °C with primary antibodies against pro-IL-1β (Proteintech, 16806-1-AP, 1:1,000), IL-1β (Immunoway, YM8498, 1:1,000), and Pik3r3 (Proteintech, 27035-1-AP, 1:1,000).

Techniques: Analysis, Expressing, Clinical Proteomics, Selection, Cell Differentiation

Journal: Frontiers in Molecular Biosciences

Article Title: Integrated transcriptome and single-cell RNA sequencing identifies transient receptor potential channel-related biomarkers in ulcerative colitis

doi: 10.3389/fmolb.2026.1902447

Figure Lengend Snippet: Validation of IL-1β and PIK3R3 expression in UC mouse model. (A) ssGSEA of IL-1β expression and immune-infiltrating cells; (B) ssGSEA of and PIK3R3 expression and immune-infiltrating cells; (C) mIHC images of the colonic tissues of mice between the UC and control groups; (D) Protein expression levels of pro-IL-1β, IL-1β, and Pik3r3 in the UC and control groups ( ** P < 0.01); (E) Relative mRNA expression levels of IL-1β, and Pik3r3 in the UC and control groups ( ** P < 0.01).

Article Snippet: For Western blot analysis, equal amounts of protein (50 μg) were denatured in SDS loading buffer, separated by SDS-PAGE (80 V for 25 min, then 120 V for 50 min), and transferred to PVDF membranes (Millipore, MA, United States) at 200 mA for 2 h. Membranes were blocked with blocking buffer (NCM Bio, P30500 ) for 10 min at room temperature and subsequently incubated overnight at 4 °C with primary antibodies against pro-IL-1β (Proteintech, 16806-1-AP, 1:1,000), IL-1β (Immunoway, YM8498, 1:1,000), and Pik3r3 (Proteintech, 27035-1-AP, 1:1,000).

Techniques: Biomarker Discovery, Expressing, Control