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mouse anti ppara  (Proteintech)


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    Proteintech mouse anti ppara
    Mouse Anti Ppara, supplied by Proteintech, used in various techniques. Bioz Stars score: 96/100, based on 1294 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/product/mouse+anti+tlr4/TLR4+Antibody/pm41905731-154-42-45
    Average 96 stars, based on 1294 article reviews
    mouse anti ppara - by Bioz Stars, 2026-09
    96/100 stars

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    Related Articles

    Incubation:

    Article Title: Evobrutinib mitigates neuroinflammation after ischemic stroke by targeting M1 microglial polarization via the TLR4/Myd88/NF-κB pathway
    Article Snippet: Protein concentration was determined using a BCA protein assay kit (Beyotime, Shanghai, China), and protein samples were loaded onto 10% SDS-PAGE gels for separation and transferred to PVDF membranes. .. The membranes were blocked with 5% non-fat milk at room temperature for 2 h and then incubated overnight at 4 °C with primary antibodies: Rabbit anti-BTK (1:800, ab208937, abcam), Rabbit anti-BTK phospho Y551 (1:800, ab40770, abcam), Mouse anti-TLR4 (1:800, 66350-1, proteintech), Rabbit anti-Myd88 (1:100, A0980, ABclonal), Rabbit anti-NF-κB (1:800, A2547, ABclonal), Mouse anti-GAPDH (1:1000, KTD101, Abbkine), and Mouse anti-β-actin (1:1000, KTD101, Abbkine). .. The membranes were then incubated with HRP Goat anti-mouse (1:1000, ZB2305, zsbio) and HRP Goat anti-rabbit (1:1000, ZB2301, zsbio) at room temperature for 2 h. ECL chemiluminescence detection was performed using a chemiluminescent imaging system, and protein bands were analyzed using Image J software, with GAPDH and β-actin as internal controls.

    Article Title: Quercetin alleviates neonatal hypoxic-ischaemic brain injury by rebalancing microglial M1/M2 polarization through silent information regulator 1/ high mobility group box-1 signalling.
    Article Snippet: Neonatal hypoxic-ischaemic encephalopathy (HIE) remains one of the major causes of neonatal death and long-term neurological disability.. Due to its complex pathogenesis, there are still many challenges in its treatment.. In our previous studies, we found that quercetin can alleviate neurological dysfunction after hypoxic-ischaemic brain injury (HIBI) in neonatal mice.

    Article Title: Astrocytic AEG-1 drives neuroinflammation and enhances seizure susceptibility.
    Article Snippet: After electrophoresis, the proteins were transferred to nitrocellulose membranes (Amersham, Little Chalfont, UK) using a Bio-Rad mini-protein-III wet transfer unit (Bio-Rad, Hercules, CA, USA) overnight at 4 ◦C. .. The membranes were then incubated with 5 % nonfat milk in TBST (10 mmol/L Tris pH = 7.6, 150 mmol/L NaCl, 0.01 % Tween-20) for 1 h at room temperature, followed by three washes, then were incubated with mouse anti-TLR4 (1:2000; Proteintech, Cat# 66350–1-Ig, RRID:AB_2881730), rabbit anti-AEG-1 (1:2000; Proteintech, Cat# 13860–1-AP, RRID:AB_2266734), rabbit anti-CREB (1:2000; Proteintech, Cat# 12208–1-AP, RRID:AB_2245417), rabbit anti-pCREB (1:2000; Proteintech, Cat# 28792–1-AP, RRID: AB_2918203) and rabbit anti-GAPDH (1:5000; Proteintech, Cat# 10494–1-AP, RRID:AB_2263076) antibodies in TBST overnight at 4 ◦C. .. After several washes with TBST buffer, the membranes were incubated for 1 h with goat-anti rabbit HRP-linked secondary antibody (Boster Biological Technology, Cat# BA1054, RRID:AB_2734136) and goat-anti mouse HRP-linked secondary antibody (Boster Biological Technology, Cat# BA1050, RRID:AB_2904507) diluted 1:5000, followed by four washes.

    Article Title: Evobrutinib mitigates neuroinflammation after ischemic stroke by targeting M1 microglial polarization via the TLR4/Myd88/NF-κB pathway.
    Article Snippet: Protein concentration was determined using a BCA protein assay kit (Beyotime, Shanghai, China), and protein samples were loaded onto 10% SDS-PAGE gels for separation and transferred to PVDF membranes. .. The membranes were blocked with 5% nonfat milk at room temperature for 2 h and then incubated overnight at 4 °C with primary antibodies: Rabbit antiBTK (1:800, ab208937, abcam), Rabbit anti-BTK phospho Y551 (1:800, ab40770, abcam), Mouse anti-TLR4 (1:800, 66350-1, proteintech), Rabbit anti-Myd88 (1:100, A0980, ABclonal), Rabbit anti-NF-κB (1:800, A2547, ABclonal), Mouse anti-GAPDH (1:1000, KTD101, Abbkine), and Mouse anti-β-actin (1:1000, KTD101, Abbkine). .. The membranes were then incubated with HRP Goat antimouse (1:1000, ZB2305, zsbio) and HRP Goat anti-rabbit (1:1000, ZB2301, zsbio) at room temperature for 2 h. ECL chemiluminescence detection was performed using a chemiluminescent imaging system, and protein bands were analyzed using Image J software, with GAPDH and β-actin as internal controls.

    Article Title: Lactobacillus paragasseri LPG-9 reduces placental inflammation in intrahepatic cholestasis of pregnancy by regulating TGR5 in mice.
    Article Snippet: For Co-IP of native proteins, the samples were completely homogenized with a lysis buffer (HY-K0202K-B, MCE, USA), the samples were centrifuged for 10 min at 12,000 rpm and 4 °C , and collected at -80 °C for subsequent testing. .. Next 500 μg of cell lysate was incubated overnight at 4 °C with mouse anti-TLR4 (1:100; 66350, Proteintech, USA) and rabbit antiTGR5 (1:50; A20778, ABclonal, China) antibodies, using IgG (1:1000) as a negative control. .. Subsequently, the complexes were incubated with magnetic beads (HY-K0202K-A, MCE, USA) for 1 h at 25 °C.

    Staining:

    Article Title: Platelet-rich plasma improves cyclophosphamide-induced interstitial cystitis in rat models through the toll-like receptor 4/nuclear factor-kappa B signalling pathway
    Article Snippet: .. The following reagents and instruments were obtained: CYP (B0150, Yuchun Bio, Shanghai, China); haematoxylin and eosin Toluidine blue stain; Masson’s trichrome stain; PureLink ribonucleic acid (RNA) (Ambion, Austin, Texas, USA); Ezol Reagent (B002-v001, GenePharma, Shanghai, China); GoScript RT system (A5001, Promega, Madison, WI, USA); GoTaq qPCR Master Mix (A6001, Promega, Madison, WI, USA); qRT-PCR Plus System (Stratagene, La Jolla, CA, USA); Mouse anti-TLR4 (66350-1-Ig; Proteintech, CHI, USA); Rabbit anti-ZO-1 (AF5145; Affinity, OU, USA); Rabbit anti-E-cadherin (ab40772; Abcam, Cambridge, UK); Rabbit anti-NF-kB p65 (10745-1-AP; Proteintech, CHI, USA); primary and secondary antibodies and washing solutions (ZO-1, E-Cadherin, TLR4, NF-kB p65, ABcam, Cambridge, UK); DMEM/F-12 K medium (31800–105, Gibco, USA); cell counting kit-8 (CCK-8) (GW770, Dojindo, Tokyo, Japan); a microplate reader (Multiskan SkyHigh, Thermo Fisher Scientific); Cell-Light TM EdU Apollo567 In vitro Kit (RiboBio, Guangzhou, China); a flow cytometer (BD Biosciences); CycleTESTTM PLUS DNA Reagent Kit (BD Biosciences); pentobarbital sodium (Jiangsu Hengfengqiang Biotechnology Co., Ltd.); an F3 catheter (150301J, Shanghai Shangyi Kangge Medical Equipment Co., Ltd.); a 15-channel physiological recorder (MP150-WSW, BIOPAC, USA); a 5% CO 2 incubator (XD-101, SANYO, Japan); and a – 80 °C refrigerator. ..

    Real-time Polymerase Chain Reaction:

    Article Title: Platelet-rich plasma improves cyclophosphamide-induced interstitial cystitis in rat models through the toll-like receptor 4/nuclear factor-kappa B signalling pathway
    Article Snippet: .. The following reagents and instruments were obtained: CYP (B0150, Yuchun Bio, Shanghai, China); haematoxylin and eosin Toluidine blue stain; Masson’s trichrome stain; PureLink ribonucleic acid (RNA) (Ambion, Austin, Texas, USA); Ezol Reagent (B002-v001, GenePharma, Shanghai, China); GoScript RT system (A5001, Promega, Madison, WI, USA); GoTaq qPCR Master Mix (A6001, Promega, Madison, WI, USA); qRT-PCR Plus System (Stratagene, La Jolla, CA, USA); Mouse anti-TLR4 (66350-1-Ig; Proteintech, CHI, USA); Rabbit anti-ZO-1 (AF5145; Affinity, OU, USA); Rabbit anti-E-cadherin (ab40772; Abcam, Cambridge, UK); Rabbit anti-NF-kB p65 (10745-1-AP; Proteintech, CHI, USA); primary and secondary antibodies and washing solutions (ZO-1, E-Cadherin, TLR4, NF-kB p65, ABcam, Cambridge, UK); DMEM/F-12 K medium (31800–105, Gibco, USA); cell counting kit-8 (CCK-8) (GW770, Dojindo, Tokyo, Japan); a microplate reader (Multiskan SkyHigh, Thermo Fisher Scientific); Cell-Light TM EdU Apollo567 In vitro Kit (RiboBio, Guangzhou, China); a flow cytometer (BD Biosciences); CycleTESTTM PLUS DNA Reagent Kit (BD Biosciences); pentobarbital sodium (Jiangsu Hengfengqiang Biotechnology Co., Ltd.); an F3 catheter (150301J, Shanghai Shangyi Kangge Medical Equipment Co., Ltd.); a 15-channel physiological recorder (MP150-WSW, BIOPAC, USA); a 5% CO 2 incubator (XD-101, SANYO, Japan); and a – 80 °C refrigerator. ..

    Quantitative RT-PCR:

    Article Title: Platelet-rich plasma improves cyclophosphamide-induced interstitial cystitis in rat models through the toll-like receptor 4/nuclear factor-kappa B signalling pathway
    Article Snippet: .. The following reagents and instruments were obtained: CYP (B0150, Yuchun Bio, Shanghai, China); haematoxylin and eosin Toluidine blue stain; Masson’s trichrome stain; PureLink ribonucleic acid (RNA) (Ambion, Austin, Texas, USA); Ezol Reagent (B002-v001, GenePharma, Shanghai, China); GoScript RT system (A5001, Promega, Madison, WI, USA); GoTaq qPCR Master Mix (A6001, Promega, Madison, WI, USA); qRT-PCR Plus System (Stratagene, La Jolla, CA, USA); Mouse anti-TLR4 (66350-1-Ig; Proteintech, CHI, USA); Rabbit anti-ZO-1 (AF5145; Affinity, OU, USA); Rabbit anti-E-cadherin (ab40772; Abcam, Cambridge, UK); Rabbit anti-NF-kB p65 (10745-1-AP; Proteintech, CHI, USA); primary and secondary antibodies and washing solutions (ZO-1, E-Cadherin, TLR4, NF-kB p65, ABcam, Cambridge, UK); DMEM/F-12 K medium (31800–105, Gibco, USA); cell counting kit-8 (CCK-8) (GW770, Dojindo, Tokyo, Japan); a microplate reader (Multiskan SkyHigh, Thermo Fisher Scientific); Cell-Light TM EdU Apollo567 In vitro Kit (RiboBio, Guangzhou, China); a flow cytometer (BD Biosciences); CycleTESTTM PLUS DNA Reagent Kit (BD Biosciences); pentobarbital sodium (Jiangsu Hengfengqiang Biotechnology Co., Ltd.); an F3 catheter (150301J, Shanghai Shangyi Kangge Medical Equipment Co., Ltd.); a 15-channel physiological recorder (MP150-WSW, BIOPAC, USA); a 5% CO 2 incubator (XD-101, SANYO, Japan); and a – 80 °C refrigerator. ..

    CCK-8 Assay:

    Article Title: Platelet-rich plasma improves cyclophosphamide-induced interstitial cystitis in rat models through the toll-like receptor 4/nuclear factor-kappa B signalling pathway
    Article Snippet: .. The following reagents and instruments were obtained: CYP (B0150, Yuchun Bio, Shanghai, China); haematoxylin and eosin Toluidine blue stain; Masson’s trichrome stain; PureLink ribonucleic acid (RNA) (Ambion, Austin, Texas, USA); Ezol Reagent (B002-v001, GenePharma, Shanghai, China); GoScript RT system (A5001, Promega, Madison, WI, USA); GoTaq qPCR Master Mix (A6001, Promega, Madison, WI, USA); qRT-PCR Plus System (Stratagene, La Jolla, CA, USA); Mouse anti-TLR4 (66350-1-Ig; Proteintech, CHI, USA); Rabbit anti-ZO-1 (AF5145; Affinity, OU, USA); Rabbit anti-E-cadherin (ab40772; Abcam, Cambridge, UK); Rabbit anti-NF-kB p65 (10745-1-AP; Proteintech, CHI, USA); primary and secondary antibodies and washing solutions (ZO-1, E-Cadherin, TLR4, NF-kB p65, ABcam, Cambridge, UK); DMEM/F-12 K medium (31800–105, Gibco, USA); cell counting kit-8 (CCK-8) (GW770, Dojindo, Tokyo, Japan); a microplate reader (Multiskan SkyHigh, Thermo Fisher Scientific); Cell-Light TM EdU Apollo567 In vitro Kit (RiboBio, Guangzhou, China); a flow cytometer (BD Biosciences); CycleTESTTM PLUS DNA Reagent Kit (BD Biosciences); pentobarbital sodium (Jiangsu Hengfengqiang Biotechnology Co., Ltd.); an F3 catheter (150301J, Shanghai Shangyi Kangge Medical Equipment Co., Ltd.); a 15-channel physiological recorder (MP150-WSW, BIOPAC, USA); a 5% CO 2 incubator (XD-101, SANYO, Japan); and a – 80 °C refrigerator. ..

    In Vitro:

    Article Title: Platelet-rich plasma improves cyclophosphamide-induced interstitial cystitis in rat models through the toll-like receptor 4/nuclear factor-kappa B signalling pathway
    Article Snippet: .. The following reagents and instruments were obtained: CYP (B0150, Yuchun Bio, Shanghai, China); haematoxylin and eosin Toluidine blue stain; Masson’s trichrome stain; PureLink ribonucleic acid (RNA) (Ambion, Austin, Texas, USA); Ezol Reagent (B002-v001, GenePharma, Shanghai, China); GoScript RT system (A5001, Promega, Madison, WI, USA); GoTaq qPCR Master Mix (A6001, Promega, Madison, WI, USA); qRT-PCR Plus System (Stratagene, La Jolla, CA, USA); Mouse anti-TLR4 (66350-1-Ig; Proteintech, CHI, USA); Rabbit anti-ZO-1 (AF5145; Affinity, OU, USA); Rabbit anti-E-cadherin (ab40772; Abcam, Cambridge, UK); Rabbit anti-NF-kB p65 (10745-1-AP; Proteintech, CHI, USA); primary and secondary antibodies and washing solutions (ZO-1, E-Cadherin, TLR4, NF-kB p65, ABcam, Cambridge, UK); DMEM/F-12 K medium (31800–105, Gibco, USA); cell counting kit-8 (CCK-8) (GW770, Dojindo, Tokyo, Japan); a microplate reader (Multiskan SkyHigh, Thermo Fisher Scientific); Cell-Light TM EdU Apollo567 In vitro Kit (RiboBio, Guangzhou, China); a flow cytometer (BD Biosciences); CycleTESTTM PLUS DNA Reagent Kit (BD Biosciences); pentobarbital sodium (Jiangsu Hengfengqiang Biotechnology Co., Ltd.); an F3 catheter (150301J, Shanghai Shangyi Kangge Medical Equipment Co., Ltd.); a 15-channel physiological recorder (MP150-WSW, BIOPAC, USA); a 5% CO 2 incubator (XD-101, SANYO, Japan); and a – 80 °C refrigerator. ..

    Flow Cytometry:

    Article Title: Platelet-rich plasma improves cyclophosphamide-induced interstitial cystitis in rat models through the toll-like receptor 4/nuclear factor-kappa B signalling pathway
    Article Snippet: .. The following reagents and instruments were obtained: CYP (B0150, Yuchun Bio, Shanghai, China); haematoxylin and eosin Toluidine blue stain; Masson’s trichrome stain; PureLink ribonucleic acid (RNA) (Ambion, Austin, Texas, USA); Ezol Reagent (B002-v001, GenePharma, Shanghai, China); GoScript RT system (A5001, Promega, Madison, WI, USA); GoTaq qPCR Master Mix (A6001, Promega, Madison, WI, USA); qRT-PCR Plus System (Stratagene, La Jolla, CA, USA); Mouse anti-TLR4 (66350-1-Ig; Proteintech, CHI, USA); Rabbit anti-ZO-1 (AF5145; Affinity, OU, USA); Rabbit anti-E-cadherin (ab40772; Abcam, Cambridge, UK); Rabbit anti-NF-kB p65 (10745-1-AP; Proteintech, CHI, USA); primary and secondary antibodies and washing solutions (ZO-1, E-Cadherin, TLR4, NF-kB p65, ABcam, Cambridge, UK); DMEM/F-12 K medium (31800–105, Gibco, USA); cell counting kit-8 (CCK-8) (GW770, Dojindo, Tokyo, Japan); a microplate reader (Multiskan SkyHigh, Thermo Fisher Scientific); Cell-Light TM EdU Apollo567 In vitro Kit (RiboBio, Guangzhou, China); a flow cytometer (BD Biosciences); CycleTESTTM PLUS DNA Reagent Kit (BD Biosciences); pentobarbital sodium (Jiangsu Hengfengqiang Biotechnology Co., Ltd.); an F3 catheter (150301J, Shanghai Shangyi Kangge Medical Equipment Co., Ltd.); a 15-channel physiological recorder (MP150-WSW, BIOPAC, USA); a 5% CO 2 incubator (XD-101, SANYO, Japan); and a – 80 °C refrigerator. ..

    other:

    Article Title: Harnessing dual-channel probiotics to synergistically correct intestinal and vaginal dysbiosis after antibiotic disruption
    Article Snippet: Mouse anti-TLR4 , Proteintech , 66350-1-Ig , 1:1000.

    Article Title: Hippocampal GPR35 is involved in the depression-like behaviors induced by inflammation and mediates the antidepressant effects of fluoxetine in mice.
    Article Snippet: Background: Neuroinflammation plays a pivotal role in the pathogenesis of depression.. G protein-coupled receptor 35 (GPR35) is expressed in the brain and plays a role in regulating inflammatory processes.. However, its specific role in depression remains unclear.

    Negative Control:

    Article Title: Lactobacillus paragasseri LPG-9 reduces placental inflammation in intrahepatic cholestasis of pregnancy by regulating TGR5 in mice.
    Article Snippet: For Co-IP of native proteins, the samples were completely homogenized with a lysis buffer (HY-K0202K-B, MCE, USA), the samples were centrifuged for 10 min at 12,000 rpm and 4 °C , and collected at -80 °C for subsequent testing. .. Next 500 μg of cell lysate was incubated overnight at 4 °C with mouse anti-TLR4 (1:100; 66350, Proteintech, USA) and rabbit antiTGR5 (1:50; A20778, ABclonal, China) antibodies, using IgG (1:1000) as a negative control. .. Subsequently, the complexes were incubated with magnetic beads (HY-K0202K-A, MCE, USA) for 1 h at 25 °C.



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    Nr-CWS affects MARCO expression through <t>TLR4.</t> The expression of TLR4 significantly increased after Nr-CWS treatment [ (D-G) , scale bars = 100 μm], while the expression TLR2 was unchanged (A-C) . Subsequent inhibition of TLR4 expression resulted in decreased levels of MARCO [ (H-J) , scale bars = 100 μm]. n = 3, * P < 0.05, ** P < 0.01, *** P < 0.001.
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    Image Search Results


    Nr-CWS affects MARCO expression through TLR4. The expression of TLR4 significantly increased after Nr-CWS treatment [ (D-G) , scale bars = 100 μm], while the expression TLR2 was unchanged (A-C) . Subsequent inhibition of TLR4 expression resulted in decreased levels of MARCO [ (H-J) , scale bars = 100 μm]. n = 3, * P < 0.05, ** P < 0.01, *** P < 0.001.

    Journal: Frontiers in Immunology

    Article Title: Nocardia rubra cell wall skeleton-induced MARCO expression: implications for improved phagocytosis and cytokine secretion in tumor-associated macrophages

    doi: 10.3389/fimmu.2026.1611476

    Figure Lengend Snippet: Nr-CWS affects MARCO expression through TLR4. The expression of TLR4 significantly increased after Nr-CWS treatment [ (D-G) , scale bars = 100 μm], while the expression TLR2 was unchanged (A-C) . Subsequent inhibition of TLR4 expression resulted in decreased levels of MARCO [ (H-J) , scale bars = 100 μm]. n = 3, * P < 0.05, ** P < 0.01, *** P < 0.001.

    Article Snippet: The following commercial antibodies (vendor, catalog number and dilution) utilized for western blot and immunohistochemical staining were employed in accordance with the manufacturers’ guidelines: mouse anti-CD68 antibody (Abcam, ab201340, 1:200), rabbit anti-MARCO antibody (Abcam, ab231046, 1:1000 for western blot and 1:100 for immunohistochemistry), rabbit anti-CD163 antibody (Abways Technology, CY6845, 1:500), mouse anti-CD86 antibody (Proteintech, 68674-2-Ig, 1:5000), mouse anti-TLR4 antibody (Santa Cruz Biotechnology, sc-293072, 1:1000 for western blot and 1:200 for immunohistochemistry), rabbit anti-TLR2 antibody(Abways Technology, CY5102, 1:2000), mouse anti-GAPDH antibody (Proteintech, 6004-1-Ig, 1:50000), HRP-goat anti-mouse recombinant secondary antibody (H+L)(Proteintech, RGAM001, 1:5000), goat anti-mouse IgG (H+L) Alexa Fluor 594 (Abways Technology, AB0152, 1:300), HRP-goat anti-rabbit recombinant secondary antibody (H+L) (Proteintech, RGAR001, 1:5000), goat anti-rabbit IgG (H+L) secondary antibody DyLightTM 594 (Report Biotech, S7002, 1:500), goat anti-mouse IgG (H+L) secondary antibody DyLightTM 488 (Report Biotech, S6001, 1:500), goat anti-mouse IgG (H+L) secondary antibody DyLightTM 594 (Abways Technology, AB0152, 1:500).

    Techniques: Expressing, Inhibition

    EA-induced analgesia is associated with MD2, TLR4, and p-p65/p65 in the ST36 acupoint and MD2 and TLR4 interactions. (A) The experimental protocol timeline is illustrated, with the duration of each intervention clearly indicated in the figure. (B) The location of the ST36 acupoint in mice (up) and the swelling of paw comparison before (right) and after (left) CFA injection. (C) PTWTs were detected on the D-3 to D-1, D1pre, D1post, D3, D5, and D7 after interventions (n = 8), ** P < 0.01 vs. the CON, ※※ P < 0.01 vs. the SHAM. (D) The expression levels of PKC protein in the spinal cord were determined by WB (n = 3), ** P < 0.01 vs. the CON, ** P < 0.01 vs. the SHAM. (E, F) The expression levels of CX3CL1 (E) , IL-1β (F) in the spinal cord were determined by Elisa (n = 6), ** P < 0.01 vs. the CON, ** P < 0.01 vs. the SHAM. (G) Data sourced from https://cn.string-db.org/ . (H) Co-IP assay with TLR4 antibody (or IgG) in the ST36 acupoint, followed by WB analysis of MD2 and TLR4. (I–K) The expression levels of MD2 (I) , TLR4 (J) , and p-p65/p65 (K) were determined using WB analysis in the ST36 acupoint on D7 (n = 3), * P < 0.05 vs. the CON, ** P < 0.01 vs. the CON, ※※ P < 0.01 vs. the SHAM.

    Journal: Frontiers in Immunology

    Article Title: Electroacupuncture alleviates pain by activating the MD2/TLR4/NF-κB pathway in the ST36 acupoint

    doi: 10.3389/fimmu.2025.1626755

    Figure Lengend Snippet: EA-induced analgesia is associated with MD2, TLR4, and p-p65/p65 in the ST36 acupoint and MD2 and TLR4 interactions. (A) The experimental protocol timeline is illustrated, with the duration of each intervention clearly indicated in the figure. (B) The location of the ST36 acupoint in mice (up) and the swelling of paw comparison before (right) and after (left) CFA injection. (C) PTWTs were detected on the D-3 to D-1, D1pre, D1post, D3, D5, and D7 after interventions (n = 8), ** P < 0.01 vs. the CON, ※※ P < 0.01 vs. the SHAM. (D) The expression levels of PKC protein in the spinal cord were determined by WB (n = 3), ** P < 0.01 vs. the CON, ** P < 0.01 vs. the SHAM. (E, F) The expression levels of CX3CL1 (E) , IL-1β (F) in the spinal cord were determined by Elisa (n = 6), ** P < 0.01 vs. the CON, ** P < 0.01 vs. the SHAM. (G) Data sourced from https://cn.string-db.org/ . (H) Co-IP assay with TLR4 antibody (or IgG) in the ST36 acupoint, followed by WB analysis of MD2 and TLR4. (I–K) The expression levels of MD2 (I) , TLR4 (J) , and p-p65/p65 (K) were determined using WB analysis in the ST36 acupoint on D7 (n = 3), * P < 0.05 vs. the CON, ** P < 0.01 vs. the CON, ※※ P < 0.01 vs. the SHAM.

    Article Snippet: Tissue lysate (1 mg protein) was incubated with either 2 μg of mouse monoclonal anti-TLR4 antibody or 2 μg of mouse IgG isotype control antibody (5415, Cell Signaling Technology, USA) for 1 hour at room temperature.

    Techniques: Comparison, Injection, Expressing, Enzyme-linked Immunosorbent Assay, Co-Immunoprecipitation Assay

    The effects of modulating MD2 in the ST36 acupoint on EA-induced analgesia and on the protein levels of MD2, TLR4, and p-p65/p65. (A) Lentivirus injection, behavioral testing, and acupoint stimulation experimental schedule. (B) Detection of lentivirus transduction in the ST36 acupoint by IF assay after MD2 knockdown lentivirus injection (n=3). (C) IF assay detection of lentivirus transduction in ST36 acupoint after overexpression lentivirus injection (n=3). (D) The transfection efficiency of the control lentivirus (FR-376) in ST36 acupoint (n=3). (E) The transfection efficiency of the control lentivirus (FV-115) in ST36 acupoint (n=3). (F) The expression of MD2 after injection of three different target knockdown lentiviruses were detected by WB (n=4). (G) The expression of MD2 after injection of overexpression was detected by WB (n=3). (H) The PTWTs were detected on D-3 to D-1, D1pre, D1post to D7 (every other day) (n = 6), ** P < 0.01 vs. the AIA+LVcon, # P < 0.05 vs. the AIA+LVcon+EA, ## P < 0.01 vs. the AIA+LVcon+EA. (I–K) The expression level of MD2 (I) , TLR4 (J) , and p-p65/p65 (K) (n = 3), ** P < 0.01 vs. the AIA+LVcon, # P < 0.05 vs. the AIA+LVcon+EA, ## P < 0.01 vs. the AIA+LVcon+EA. (L) PTWTs were detected on D-3 to D-1, D1pre, D1post to D7 (every other day) (n = 6), * P < 0.05 vs. the AIA+OEcon, ** P < 0.01 vs. the AIA+OEcon. (M–O) The expression level of MD2 (M) , TLR4 (N) , and p-p65/p65 (O) (n = 3), ** P < 0.01 vs. the AIA+OEcon.

    Journal: Frontiers in Immunology

    Article Title: Electroacupuncture alleviates pain by activating the MD2/TLR4/NF-κB pathway in the ST36 acupoint

    doi: 10.3389/fimmu.2025.1626755

    Figure Lengend Snippet: The effects of modulating MD2 in the ST36 acupoint on EA-induced analgesia and on the protein levels of MD2, TLR4, and p-p65/p65. (A) Lentivirus injection, behavioral testing, and acupoint stimulation experimental schedule. (B) Detection of lentivirus transduction in the ST36 acupoint by IF assay after MD2 knockdown lentivirus injection (n=3). (C) IF assay detection of lentivirus transduction in ST36 acupoint after overexpression lentivirus injection (n=3). (D) The transfection efficiency of the control lentivirus (FR-376) in ST36 acupoint (n=3). (E) The transfection efficiency of the control lentivirus (FV-115) in ST36 acupoint (n=3). (F) The expression of MD2 after injection of three different target knockdown lentiviruses were detected by WB (n=4). (G) The expression of MD2 after injection of overexpression was detected by WB (n=3). (H) The PTWTs were detected on D-3 to D-1, D1pre, D1post to D7 (every other day) (n = 6), ** P < 0.01 vs. the AIA+LVcon, # P < 0.05 vs. the AIA+LVcon+EA, ## P < 0.01 vs. the AIA+LVcon+EA. (I–K) The expression level of MD2 (I) , TLR4 (J) , and p-p65/p65 (K) (n = 3), ** P < 0.01 vs. the AIA+LVcon, # P < 0.05 vs. the AIA+LVcon+EA, ## P < 0.01 vs. the AIA+LVcon+EA. (L) PTWTs were detected on D-3 to D-1, D1pre, D1post to D7 (every other day) (n = 6), * P < 0.05 vs. the AIA+OEcon, ** P < 0.01 vs. the AIA+OEcon. (M–O) The expression level of MD2 (M) , TLR4 (N) , and p-p65/p65 (O) (n = 3), ** P < 0.01 vs. the AIA+OEcon.

    Article Snippet: Tissue lysate (1 mg protein) was incubated with either 2 μg of mouse monoclonal anti-TLR4 antibody or 2 μg of mouse IgG isotype control antibody (5415, Cell Signaling Technology, USA) for 1 hour at room temperature.

    Techniques: Injection, Transduction, Knockdown, Over Expression, Transfection, Control, Expressing

    EA treatment can increase the co-expression of MD2/TLR4, TLR4-Vimentin, TLR4-Tryptase, as well as TLR4-F4/80 at the ST36 acupoint. (A) Double-labeling immunofluorescence of TLR4 (green), MD2 (red) and DAPI (blue), scale bar = 20 µm. (B) Double-labeling immunofluorescence of TLR4 (green), fibroblasts (red) and DAPI (blue), scale bar = 20 µm. (C) Double-labeling immunofluorescence of TLR4 (green) and Tryptase (red), scale bar = 20µm. (D) Double-labeling immunofluorescence of TLR4 (green) and F4/80 (red), DAPI (blue), scale bar = 20µm. (E) Immunofluorescence quantitative analysis of MD2/TLR4 (n = 3), * P < 0.05 vs. the CON, ## P < 0.01 vs. the AIA. (F) Immunofluorescence quantitative analysis of TLR4-Vimentin (n = 3), ** P < 0.01 vs. the CON, ## P < 0.01 vs. the AIA. (G) Immunofluorescence quantitative analysis of TLR4-Tryptase (n = 3), # P < 0.05 vs. the AIA. (H) Immunofluorescence quantitative analysis of TLR4-F4/80 (n = 3).

    Journal: Frontiers in Immunology

    Article Title: Electroacupuncture alleviates pain by activating the MD2/TLR4/NF-κB pathway in the ST36 acupoint

    doi: 10.3389/fimmu.2025.1626755

    Figure Lengend Snippet: EA treatment can increase the co-expression of MD2/TLR4, TLR4-Vimentin, TLR4-Tryptase, as well as TLR4-F4/80 at the ST36 acupoint. (A) Double-labeling immunofluorescence of TLR4 (green), MD2 (red) and DAPI (blue), scale bar = 20 µm. (B) Double-labeling immunofluorescence of TLR4 (green), fibroblasts (red) and DAPI (blue), scale bar = 20 µm. (C) Double-labeling immunofluorescence of TLR4 (green) and Tryptase (red), scale bar = 20µm. (D) Double-labeling immunofluorescence of TLR4 (green) and F4/80 (red), DAPI (blue), scale bar = 20µm. (E) Immunofluorescence quantitative analysis of MD2/TLR4 (n = 3), * P < 0.05 vs. the CON, ## P < 0.01 vs. the AIA. (F) Immunofluorescence quantitative analysis of TLR4-Vimentin (n = 3), ** P < 0.01 vs. the CON, ## P < 0.01 vs. the AIA. (G) Immunofluorescence quantitative analysis of TLR4-Tryptase (n = 3), # P < 0.05 vs. the AIA. (H) Immunofluorescence quantitative analysis of TLR4-F4/80 (n = 3).

    Article Snippet: Tissue lysate (1 mg protein) was incubated with either 2 μg of mouse monoclonal anti-TLR4 antibody or 2 μg of mouse IgG isotype control antibody (5415, Cell Signaling Technology, USA) for 1 hour at room temperature.

    Techniques: Expressing, Labeling, Immunofluorescence

    The pattern of the effect of EA treatment on MD2, TLR4 in ST36 acupoint. (A) Behavioral testing, acupoint stimulation and molecular detection experimental schedule. (B–D) PTWTs were detected on D1post (B) (n=8), D3 (C) (n=6), and D5 (D) (n=6), ** P < 0.01 vs. the CON, ※ P < 0.05 vs. the SHAM, ※※ P < 0.01 vs. the SHAM. (E–H) The expression level of MD2 in the ST36 acupoint was determined by WB on D1post (E) , D3 (F) and D5 (G) (n = 3); the TLR4 was determined on the D5 (H) (n = 3), * P < 0.05 vs. the CON, ** P < 0.01 vs. the CON, # P < 0.05 vs. the AIA. ※ P < 0.05 vs. the SHAM.

    Journal: Frontiers in Immunology

    Article Title: Electroacupuncture alleviates pain by activating the MD2/TLR4/NF-κB pathway in the ST36 acupoint

    doi: 10.3389/fimmu.2025.1626755

    Figure Lengend Snippet: The pattern of the effect of EA treatment on MD2, TLR4 in ST36 acupoint. (A) Behavioral testing, acupoint stimulation and molecular detection experimental schedule. (B–D) PTWTs were detected on D1post (B) (n=8), D3 (C) (n=6), and D5 (D) (n=6), ** P < 0.01 vs. the CON, ※ P < 0.05 vs. the SHAM, ※※ P < 0.01 vs. the SHAM. (E–H) The expression level of MD2 in the ST36 acupoint was determined by WB on D1post (E) , D3 (F) and D5 (G) (n = 3); the TLR4 was determined on the D5 (H) (n = 3), * P < 0.05 vs. the CON, ** P < 0.01 vs. the CON, # P < 0.05 vs. the AIA. ※ P < 0.05 vs. the SHAM.

    Article Snippet: Tissue lysate (1 mg protein) was incubated with either 2 μg of mouse monoclonal anti-TLR4 antibody or 2 μg of mouse IgG isotype control antibody (5415, Cell Signaling Technology, USA) for 1 hour at room temperature.

    Techniques: Expressing

    Biglycan (BGN) promotes proliferation and migration of esophageal squamous cell carcinoma (ESCC) cells through its receptor TLR4. ( A ) Expression of TLR4 in ESCC cells was confirmed by Western Blotting. ( B ) Double immunofluorescence staining for BGN (green) and TLR4 (red) was performed in ESCC cells treated with recombinant human BGN (rhBGN; 100 ng/mL). Nuclei were counterstained with DAPI (blue). ( C , D ) MTS assays ( C ) and Transwell migration assays ( D ) were conducted to evaluate changes in the proliferation ( C ) and migration ( D ) of TE-9, TE-10, and TE-15 cells following treatment with rhBGN (100 ng/mL) in the presence of a TLR4-neutralizing antibody (1 μg/mL) or control immunoglobulin G (IgG; 1 μg/mL). Migrated cells were counted in five representative microscopic fields after 48 h, and representative images are shown beneath the graphs ( D ). The data are presented as the mean ± standard error of the mean (SEM) from three independent experiments ( C , D ). ** p < 0.01, *** p < 0.001. Scale bars: 20 μm ( B ) and 100 μm ( D ).

    Journal: International Journal of Molecular Sciences

    Article Title: BGN Secreted by Cancer-Associated Fibroblasts Promotes Esophageal Squamous Cell Carcinoma Progression via Activation of TLR4-Mediated Erk and NF-κB Signaling Pathways

    doi: 10.3390/ijms262412024

    Figure Lengend Snippet: Biglycan (BGN) promotes proliferation and migration of esophageal squamous cell carcinoma (ESCC) cells through its receptor TLR4. ( A ) Expression of TLR4 in ESCC cells was confirmed by Western Blotting. ( B ) Double immunofluorescence staining for BGN (green) and TLR4 (red) was performed in ESCC cells treated with recombinant human BGN (rhBGN; 100 ng/mL). Nuclei were counterstained with DAPI (blue). ( C , D ) MTS assays ( C ) and Transwell migration assays ( D ) were conducted to evaluate changes in the proliferation ( C ) and migration ( D ) of TE-9, TE-10, and TE-15 cells following treatment with rhBGN (100 ng/mL) in the presence of a TLR4-neutralizing antibody (1 μg/mL) or control immunoglobulin G (IgG; 1 μg/mL). Migrated cells were counted in five representative microscopic fields after 48 h, and representative images are shown beneath the graphs ( D ). The data are presented as the mean ± standard error of the mean (SEM) from three independent experiments ( C , D ). ** p < 0.01, *** p < 0.001. Scale bars: 20 μm ( B ) and 100 μm ( D ).

    Article Snippet: The primary antibodies used as follows: rabbit BGN (#16409-1-AP, R&D Systems), rabbit phosphorylated Erk1/2 (pErk; #9101, Cell signaling Technology; CST, Danvers, MA, USA), rabbit total Erk1/2 (tErk; #9102, CST), rabbit phosphorylated NF-κB p65 (pNF-κB; #3033, CST), rabbit total NF-κB p65 (tNF-κB; #8242, CST), mouse TLR4 (#sc-293072, Santa Cruz Biotechmology, Dallas, TX, USA), sheep FAP (#AF3715, R&D Systems), rabbit αSMA (#ab5694, Abcam, Cambridge, UK), rabbit IL-6 (#ab6672, Abcam), mouse CD163 (#NCL-L-CD163; Leica Biosystems, Wetzlar, Germany), mouse CD206 (#sc-376108, Santa Cruz), and rabbit β-actin (#4970, CST) antibodies.

    Techniques: Migration, Expressing, Western Blot, Double Immunofluorescence Staining, Recombinant, Control

    Biglycan (BGN) promotes proliferation, migration, and activation of macrophages and fibroblasts. ( A ) Cell–cell communication network analysis of BGN -expressing cancer-associated fibroblasts (CAFs) in esophageal squamous cell carcinoma (ESCC) tissues. Fibroblasts in single-cell RNA sequencing (scRNAseq) datasets from ESCC tissues were stratified into CAF_ BGN _High (scaled expression > 3) and CAF_ BGN _Low (≤3) groups based on BGN expression levels. Cell–cell communication inferred by CellChat revealed that CAF_ BGN _High cells exhibited strong outgoing signaling toward epithelial and myeloid cells, as well as prominent autocrine signaling within the CAF_ BGN _High population. ( B , C ) MTS assay ( B ) and Transwell migration assay ( C ) showing increased proliferation and migration of mesenchymal stem cells (MSCs) treated with recombinant human BGN (rhBGN; 100 ng/mL), respectively. ( D ) Western Blot analysis showing that fibroblast activation protein (FAP) and α-smooth muscle actin (αSMA) (CAF markers) were upregulated in MSCs treated with rhBGN for 24 h, whereas interleukin-6 (IL6) expression remained unchanged. TLR4 expression was detected; however, phosphorylation of NF-κB and Erk was not increased by rhBGN. ( E , F ) MTS assay ( E ) and Transwell migration assay ( F ) showing that rhBGN-induced proliferation and migration of MSCs were not suppressed by a TLR4-neutralizing antibody (1 μg/mL). ( G , H ) MTS assay ( G ) and Transwell migration assay ( H ) showing increased proliferation and migration of macrophages treated with rhBGN (100 ng/mL), respectively. ( I ) Western Blot analysis showing that rhBGN treatment upregulated the expression of CD163 and CD206 (M2 macrophage markers) and increased NF-κB phosphorylation, while TLR4 expression was also detected, but phosphorylated Erk was not changed. ( J , K ) MTS assay ( J ) and Transwell migration assay ( K ) showing that rhBGN-induced proliferation and migration of macrophages were attenuated by treatment with a TLR4-neutralizing antibody (1 μg/mL), respectively. ( L , M ) MTS assay ( L ) and Transwell migration assay ( M ) showing that treatment with the NF-κB pathway inhibitor Bay 11-7082 (1 μM) attenuated rhBGN-induced proliferation and migration to a similar extent, respectively. Data are presented as the mean ± standard error of the mean (SEM) from three independent experiments ( B , C , E – H , J – M ). * p < 0.05, ** p < 0.01, *** p < 0.001. N.S., not significant. Scale bars: 100 μm ( C , F , H , K , M ).

    Journal: International Journal of Molecular Sciences

    Article Title: BGN Secreted by Cancer-Associated Fibroblasts Promotes Esophageal Squamous Cell Carcinoma Progression via Activation of TLR4-Mediated Erk and NF-κB Signaling Pathways

    doi: 10.3390/ijms262412024

    Figure Lengend Snippet: Biglycan (BGN) promotes proliferation, migration, and activation of macrophages and fibroblasts. ( A ) Cell–cell communication network analysis of BGN -expressing cancer-associated fibroblasts (CAFs) in esophageal squamous cell carcinoma (ESCC) tissues. Fibroblasts in single-cell RNA sequencing (scRNAseq) datasets from ESCC tissues were stratified into CAF_ BGN _High (scaled expression > 3) and CAF_ BGN _Low (≤3) groups based on BGN expression levels. Cell–cell communication inferred by CellChat revealed that CAF_ BGN _High cells exhibited strong outgoing signaling toward epithelial and myeloid cells, as well as prominent autocrine signaling within the CAF_ BGN _High population. ( B , C ) MTS assay ( B ) and Transwell migration assay ( C ) showing increased proliferation and migration of mesenchymal stem cells (MSCs) treated with recombinant human BGN (rhBGN; 100 ng/mL), respectively. ( D ) Western Blot analysis showing that fibroblast activation protein (FAP) and α-smooth muscle actin (αSMA) (CAF markers) were upregulated in MSCs treated with rhBGN for 24 h, whereas interleukin-6 (IL6) expression remained unchanged. TLR4 expression was detected; however, phosphorylation of NF-κB and Erk was not increased by rhBGN. ( E , F ) MTS assay ( E ) and Transwell migration assay ( F ) showing that rhBGN-induced proliferation and migration of MSCs were not suppressed by a TLR4-neutralizing antibody (1 μg/mL). ( G , H ) MTS assay ( G ) and Transwell migration assay ( H ) showing increased proliferation and migration of macrophages treated with rhBGN (100 ng/mL), respectively. ( I ) Western Blot analysis showing that rhBGN treatment upregulated the expression of CD163 and CD206 (M2 macrophage markers) and increased NF-κB phosphorylation, while TLR4 expression was also detected, but phosphorylated Erk was not changed. ( J , K ) MTS assay ( J ) and Transwell migration assay ( K ) showing that rhBGN-induced proliferation and migration of macrophages were attenuated by treatment with a TLR4-neutralizing antibody (1 μg/mL), respectively. ( L , M ) MTS assay ( L ) and Transwell migration assay ( M ) showing that treatment with the NF-κB pathway inhibitor Bay 11-7082 (1 μM) attenuated rhBGN-induced proliferation and migration to a similar extent, respectively. Data are presented as the mean ± standard error of the mean (SEM) from three independent experiments ( B , C , E – H , J – M ). * p < 0.05, ** p < 0.01, *** p < 0.001. N.S., not significant. Scale bars: 100 μm ( C , F , H , K , M ).

    Article Snippet: The primary antibodies used as follows: rabbit BGN (#16409-1-AP, R&D Systems), rabbit phosphorylated Erk1/2 (pErk; #9101, Cell signaling Technology; CST, Danvers, MA, USA), rabbit total Erk1/2 (tErk; #9102, CST), rabbit phosphorylated NF-κB p65 (pNF-κB; #3033, CST), rabbit total NF-κB p65 (tNF-κB; #8242, CST), mouse TLR4 (#sc-293072, Santa Cruz Biotechmology, Dallas, TX, USA), sheep FAP (#AF3715, R&D Systems), rabbit αSMA (#ab5694, Abcam, Cambridge, UK), rabbit IL-6 (#ab6672, Abcam), mouse CD163 (#NCL-L-CD163; Leica Biosystems, Wetzlar, Germany), mouse CD206 (#sc-376108, Santa Cruz), and rabbit β-actin (#4970, CST) antibodies.

    Techniques: Migration, Activation Assay, Expressing, RNA Sequencing, MTS Assay, Transwell Migration Assay, Recombinant, Western Blot, Phospho-proteomics

    A schematic illustration of the role of BGN in the tumor–stromal interactions among ESCC cells, CAFs, and macrophages. BGN secreted from MSCs that have undergone CAF transition upon direct contact with ESCC cells promotes ESCC cell proliferation and migration through the TLR4–Erk/NF-κB signaling pathways. In addition, BGN enhances the proliferation and migration of MSCs and induces their differentiation into CAFs. Furthermore, BGN promotes the proliferation and migration of macrophages and drives their polarization toward the M2 phenotype through the TLR4–NF-κB signaling pathways.

    Journal: International Journal of Molecular Sciences

    Article Title: BGN Secreted by Cancer-Associated Fibroblasts Promotes Esophageal Squamous Cell Carcinoma Progression via Activation of TLR4-Mediated Erk and NF-κB Signaling Pathways

    doi: 10.3390/ijms262412024

    Figure Lengend Snippet: A schematic illustration of the role of BGN in the tumor–stromal interactions among ESCC cells, CAFs, and macrophages. BGN secreted from MSCs that have undergone CAF transition upon direct contact with ESCC cells promotes ESCC cell proliferation and migration through the TLR4–Erk/NF-κB signaling pathways. In addition, BGN enhances the proliferation and migration of MSCs and induces their differentiation into CAFs. Furthermore, BGN promotes the proliferation and migration of macrophages and drives their polarization toward the M2 phenotype through the TLR4–NF-κB signaling pathways.

    Article Snippet: The primary antibodies used as follows: rabbit BGN (#16409-1-AP, R&D Systems), rabbit phosphorylated Erk1/2 (pErk; #9101, Cell signaling Technology; CST, Danvers, MA, USA), rabbit total Erk1/2 (tErk; #9102, CST), rabbit phosphorylated NF-κB p65 (pNF-κB; #3033, CST), rabbit total NF-κB p65 (tNF-κB; #8242, CST), mouse TLR4 (#sc-293072, Santa Cruz Biotechmology, Dallas, TX, USA), sheep FAP (#AF3715, R&D Systems), rabbit αSMA (#ab5694, Abcam, Cambridge, UK), rabbit IL-6 (#ab6672, Abcam), mouse CD163 (#NCL-L-CD163; Leica Biosystems, Wetzlar, Germany), mouse CD206 (#sc-376108, Santa Cruz), and rabbit β-actin (#4970, CST) antibodies.

    Techniques: Migration, Protein-Protein interactions