Journal: Frontiers in Immunology

Article Title: Cannabinoid receptor-2 attenuates neuroinflammation by promoting autophagy-mediated degradation of the NLRP3 inflammasome post spinal cord injury

doi: 10.3389/fimmu.2022.993168

Figure Lengend Snippet: Expression patterns of CB2R in cord tissues post-SCI. (A) Representative protein band of CB2R on day 3 and weeks 1, 2, 4, and 6 post-SCI. (B) Quantitative analysis showing that CB2R expression was significantly increased on day 3 and weeks 1 and 2 post-SCI. (C) IF staining showing that CB2R (red) in cords of mice within 6W post-SCI. (D) IF staining of CB2R (red) with GFAP, Neun, or IBA-1 (green) in cords of sham mice. (E) IF staining of CB2R (red) with GFAP, Neun, or IBA-1 (green) in SCI mice on day 7 post-SCI. Scale bar = 100 μm. * p < 0.05.

Article Snippet: At 80% confluence, the microglia and HEK-293T cells were pretreated for 24 h with the CB2R activator JWH-133 (10 nM; B7941; APExBIO Technology LLC, Houston, TX, USA) or antagonist AM630 (40 nM; A3168; APExBIO Technology LLC) dissolved in 0.1% dimethyl sulfoxide (KGT5131; Nanjing KeyGen Biotech.

Techniques: Expressing, Staining

Journal: Frontiers in Immunology

Article Title: Cannabinoid receptor-2 attenuates neuroinflammation by promoting autophagy-mediated degradation of the NLRP3 inflammasome post spinal cord injury

doi: 10.3389/fimmu.2022.993168

Figure Lengend Snippet: CB2R regulates inflammation-mediated polarization of microglia. (A) IF staining of iNOS (red) and ARG-1 (green) in BV2 microglia after LPS insult for 24 h with CB2R activation by JWH-133 (J) or inactivation by AM630 (A) . Scale bar = 50 μm. (B) Quantitative analysis indicating that CB2R significantly altered expression of the M1 marker iNOS during inflammation. (C) Quantitative analysis indicating that CB2R significantly altered expression of the M2 marker ARG-1 after LPS stimulation. (D) Flow cytometry results showing the relative proportion of M1 microglia (red) after LPS insult with and without CB2R activation or inactivation. (E) Quantitative analysis of M1 microglia. (F) Flow cytometry results showing the relative proportion of M2 microglia (red) after LPS insult with and without CB2R activation or inactivation. (G) Quantitative analysis of M2 microglia. ELISA results of IL-1β (H) , IL-6 (I) , and IL-10 (J) in the control, LPS, LPS+J (JWH-133), and LPS+A (AM630) groups. * p < 0.05. n.s means no significance.

Article Snippet: At 80% confluence, the microglia and HEK-293T cells were pretreated for 24 h with the CB2R activator JWH-133 (10 nM; B7941; APExBIO Technology LLC, Houston, TX, USA) or antagonist AM630 (40 nM; A3168; APExBIO Technology LLC) dissolved in 0.1% dimethyl sulfoxide (KGT5131; Nanjing KeyGen Biotech.

Techniques: Staining, Activation Assay, Expressing, Marker, Flow Cytometry, Enzyme-linked Immunosorbent Assay

Journal: Frontiers in Immunology

Article Title: Cannabinoid receptor-2 attenuates neuroinflammation by promoting autophagy-mediated degradation of the NLRP3 inflammasome post spinal cord injury

doi: 10.3389/fimmu.2022.993168

Figure Lengend Snippet: CB2R promoted ubiquitination and autophagy-induced degradation of NLRP3 via the AMPK/ULK signaling pathway. (A) IF staining of CB2R (red) and NLRP3 (green) after LPS stimuli for 24 h with CB2R activation by JWH-133 (J) or inactivation by AM630 (A) . Scale bar = 50 μm. (B) Quantitative analysis of CB2R. (C) Quantitative analysis of NLRP3. (D) Representative protein bands of p-AMPK, AMPK, p-ULK1, ULK1, LC3A, and LC3B in each group after 24 h of LPS insult. (E) Quantitative analysis of the p-AMPK/AMPK ratio. (F) Quantitative analysis of the p-ULK1/ULK1 ratio. (G) Quantitative analysis of the LC3B/LC3A ratio. (H) Representative protein bands of p-p65, p65, NLRP3, and p62 after 24 h of LPS stimulation. (I) Quantitative analysis of the p-p65/p65 ratio. (J) Quantitative analysis of NLRP3. (K) Quantitative analysis of p62. (L) IF staining of CB2R (red) and NLRP3 (green) after LPS stimulation for 24 h with CB2R activation by JWH-133 or CB2R inactivation by AM630. Scale bar = 50 μm. (M) Representative protein bands of NLRP3 and p62 in BV2 microglia after IP assay using an Ab against NLRP3. (N) Representative protein bands of ubiquitin linkage-specific K48 bound to NLRP3. * p < 0.05.

Article Snippet: At 80% confluence, the microglia and HEK-293T cells were pretreated for 24 h with the CB2R activator JWH-133 (10 nM; B7941; APExBIO Technology LLC, Houston, TX, USA) or antagonist AM630 (40 nM; A3168; APExBIO Technology LLC) dissolved in 0.1% dimethyl sulfoxide (KGT5131; Nanjing KeyGen Biotech.

Techniques: Staining, Activation Assay

Journal: Frontiers in Immunology

Article Title: Cannabinoid receptor-2 attenuates neuroinflammation by promoting autophagy-mediated degradation of the NLRP3 inflammasome post spinal cord injury

doi: 10.3389/fimmu.2022.993168

Figure Lengend Snippet: CB2R alleviated inflammation by promoting NLRP3-autophagosome formation in injured cords post-SCI. On day 3 post-SCI with CB2R activation by JWH-133 or inactivation by AM630, (A) IF staining of iNOS (red) and IBA-1 (green); (B) IF staining of ARG-1 (red) and IBA-1 (green). Scale bar = 100 μm. (C) Quantitative analysis of the number of iNOS + microglia. (D) Quantitative analysis of the number of ARG-1 + microglia. (E) IF staining of NLRP3 (red) and LC3B (green) on day 3 post-SCI. Scale bar = 100 μm. * p < 0.05.

Article Snippet: At 80% confluence, the microglia and HEK-293T cells were pretreated for 24 h with the CB2R activator JWH-133 (10 nM; B7941; APExBIO Technology LLC, Houston, TX, USA) or antagonist AM630 (40 nM; A3168; APExBIO Technology LLC) dissolved in 0.1% dimethyl sulfoxide (KGT5131; Nanjing KeyGen Biotech.

Techniques: Activation Assay, Staining

Journal: Frontiers in Immunology

Article Title: Cannabinoid receptor-2 attenuates neuroinflammation by promoting autophagy-mediated degradation of the NLRP3 inflammasome post spinal cord injury

doi: 10.3389/fimmu.2022.993168

Figure Lengend Snippet: CB2R ameliorated secondary neuronal apoptosis and demyelination post-SCI. (A) LFB staining of spinal cords on day 7 post-SCI. Scale bar = 200 μm. (B) Quantitative analysis of LFB-labeled area. (C) IF staining of NF200 or myelin basic protein (green) on day 7 post-SCI. Scale bar = 500 μm. (D) Quantitative analysis of NF200. (E) Quantitative analysis of myelin basic protein. (F) TUNEL staining of spinal cords on day 7 post-SCI. Scale bar = 100 μm. (G) Quantitative analysis of TUNEL-positive cells. (H) IF staining of Neun (green) and Annexin V (red) on day 7 post-SCI. Scale bar = 100 μm. (I) Quantitative analysis of Annexin V-positive neurons. (J) Nissl staining of spinal cords on day 7 post-SCI. Scale bar = 200 μm. (K) Quantitative analysis of the number of neurons. * p < 0.05. n.s means no significance.

Article Snippet: At 80% confluence, the microglia and HEK-293T cells were pretreated for 24 h with the CB2R activator JWH-133 (10 nM; B7941; APExBIO Technology LLC, Houston, TX, USA) or antagonist AM630 (40 nM; A3168; APExBIO Technology LLC) dissolved in 0.1% dimethyl sulfoxide (KGT5131; Nanjing KeyGen Biotech.

Techniques: Staining, Labeling, TUNEL Assay

Journal: Frontiers in Immunology

Article Title: Cannabinoid receptor-2 attenuates neuroinflammation by promoting autophagy-mediated degradation of the NLRP3 inflammasome post spinal cord injury

doi: 10.3389/fimmu.2022.993168

Figure Lengend Snippet: CB2R mitigated expansion of neuronal damage and formation of glial scarring post-SCI. (A) IF staining of GFAP (green) and IBA-1 (red) on day 7 post-SCI. Scale bar = 500 μm. Quantitative analysis of the area of injured foci inside the white dotted line (B) , microglia-occupied area, (C) and astrocyte-occupied area (D) on day 7 post-SCI. (E) IF staining of GFAP (green) and IBA-1 (red) on day 28 post-SCI. Scale bar = 500 μm. Quantitative analysis of the area of injured foci inside the white dotted line (F) , microglia-occupied area, (G) and astrocyte-occupied area (H) on day 28 post-SCI. * p < 0.05.

Article Snippet: At 80% confluence, the microglia and HEK-293T cells were pretreated for 24 h with the CB2R activator JWH-133 (10 nM; B7941; APExBIO Technology LLC, Houston, TX, USA) or antagonist AM630 (40 nM; A3168; APExBIO Technology LLC) dissolved in 0.1% dimethyl sulfoxide (KGT5131; Nanjing KeyGen Biotech.

Techniques: Staining

Journal: Frontiers in Immunology

Article Title: Cannabinoid receptor-2 attenuates neuroinflammation by promoting autophagy-mediated degradation of the NLRP3 inflammasome post spinal cord injury

doi: 10.3389/fimmu.2022.993168

Figure Lengend Snippet: CB2R attenuated histological and functional damage post-SCI. (A) Representative images of H&E staining of spinal cords on day 3 post-SCI (injured cords in black frames). Scale bar (i) = 600 μm (ii) = 100 μm. (B) Quantitative analysis of defected area by H&E staining. (C) H&E staining of spinal cords on day 3 post-SCI (injured cords in black frames). Scale bar: (i) = 600 μm, (ii) = 100 μm. (D) Quantitative analysis of defected area by H&E staining. (E) Mouse footprint assay on day 28 post-SCI. Quantitative analysis of stride length (F) and width (G) of the hindlimbs of mice post-SCI. (H) Representative images of a mouse swimming on day 28 post-SCI. Statistical analysis of the Louisville Swim Scale score (I) and Basso Mouse Scale (J) over a 28-day period. * p < 0.05. n.s means no significance.

Article Snippet: At 80% confluence, the microglia and HEK-293T cells were pretreated for 24 h with the CB2R activator JWH-133 (10 nM; B7941; APExBIO Technology LLC, Houston, TX, USA) or antagonist AM630 (40 nM; A3168; APExBIO Technology LLC) dissolved in 0.1% dimethyl sulfoxide (KGT5131; Nanjing KeyGen Biotech.

Techniques: Functional Assay, Staining

Journal: bioRxiv

Article Title: Cannabinoid CB2R regulates T cell gut-homing in preclinical Crohn’s model

doi: 10.1101/2025.09.05.674422

Figure Lengend Snippet: (A) Integrin β7 promoter activity assessed using dual luciferase reporter assay after 24 h cannabinoid treatment in the presence of 1μM retinoic acid. (B) Integrin β7 mRNA expression levels assessed using RT-PCR following 24 h cannabinoid treatment. (C) Flow cytometry analysis of integrin expression levels at a surface protein level by means of mean fluorescence intensity (MFI) following 48 h CB 2 R activation in Jurkat T cells. ( D ) Representative histogram overlay of the average MFI values for each treatment group, displaying alterations in surface α4β7 expression on T cells following CB 2 R activation. ( E ) Flow cytometry analysis used to make comparisons between total and surface integrin α4β7 protein expression levels in Jurkat T cells after 48 h CB 2 R activation. ( F ) RT-PCR studies investigating integrin β7 mRNA expression levels in CRISPRi CNR2 -/- Jurkat T cells following 24 h cannabinoid treatment. Results represent mean±SEM for n≥3 independent experiments.* p<0.05, ** p<0.01, *** p<0.001.

Article Snippet: Jurkat T cells plated at 2.5x10 6 cells/ml were treated for 48hr with Retinoic Acid (RA; 1μM; Merck; Cat No. R2625), with(out) CB 2 R agonist JWH-133 (1μM; Tocris BioScience) or CB 2 R inverse agonist GP-1a (10μM; Tocris BioScience) or vehicle as appropriate.

Techniques: Activity Assay, Luciferase, Reporter Assay, Expressing, Reverse Transcription Polymerase Chain Reaction, Flow Cytometry, Fluorescence, Activation Assay

Journal: BioMed Research International

Article Title: CB2 Receptor Activation Ameliorates the Proinflammatory Activity in Acute Lung Injury Induced by Paraquat

doi: 10.1155/2014/971750

Figure Lengend Snippet: Effects of JWH133 on lung W/D ratio and histopathological changes in lung tissues of PQ-induced acute lung injury rats. JWH133 was intraperitoneally administered 1 h before intraperitoneal administration of PQ. The lung W/D ratio (a, b) and lung histological evaluation (c, d, HE staining) were determined after PQ administration for 72 h. L-JWH133: JWH133 at the low dose of 5 mg/kg and H-JWH133: JWH133 at the high dose of 20 mg/kg. The values presented are the mean ± SD. * P < 0.05 versus PQ group; # P < 0.05 versus L-JWH133 group.

Article Snippet: CB2R-agonist JWH133 was obtained from Enzo Life Sciences Ltd. (UK).

Techniques: Staining

Journal: BioMed Research International

Article Title: CB2 Receptor Activation Ameliorates the Proinflammatory Activity in Acute Lung Injury Induced by Paraquat

doi: 10.1155/2014/971750

Figure Lengend Snippet: Effects of JWH133 on the PaO 2 in the arterial blood, MPO activity, and cytokine (IL-1 β and TNF- α ) levels in the BALF of PQ-induced acute lung injury rats. JWH133 was intraperitoneally administered 1 h before intraperitoneal administration of PQ. The arterial blood and BALF were collected after PQ administration for 4, 8, 24, and 72 h to analyze PaO 2 level (a), MPO activity (b), and levels of TNF- α and IL-1 β (c, d). L-JWH133: JWH133 at the low dose of 5 mg/kg and H-JWH133: JWH133 at the high dose of 20 mg/kg. The values presented are the mean ± SD. * P < 0.05 versus PQ group; # P < 0.05 versus L-JWH133 group.

Article Snippet: CB2R-agonist JWH133 was obtained from Enzo Life Sciences Ltd. (UK).

Techniques: Activity Assay

Journal: BioMed Research International

Article Title: CB2 Receptor Activation Ameliorates the Proinflammatory Activity in Acute Lung Injury Induced by Paraquat

doi: 10.1155/2014/971750

Figure Lengend Snippet: Effects of JWH133 on CB2 receptor in lung tissues of paraquat- (PQ-) induced acute lung injury rats. JWH133 was intraperitoneally administered 1 h before intraperitoneal administration of PQ. Lung tissue was collected after PQ administration for 72 h to determine the expression of CB2 receptor by immunohistochemistry (a) and Western blotting (b). L-JWH133: JWH133 at the low dose of 5 mg/kg and H-JWH133: JWH133 at the high dose of 20 mg/kg. The values presented are the mean ± SD. * P < 0.05 versus PQ group; # P < 0.05 versus L-JWH133 group.

Article Snippet: CB2R-agonist JWH133 was obtained from Enzo Life Sciences Ltd. (UK).

Techniques: Expressing, Immunohistochemistry, Western Blot

Journal: BioMed Research International

Article Title: CB2 Receptor Activation Ameliorates the Proinflammatory Activity in Acute Lung Injury Induced by Paraquat

doi: 10.1155/2014/971750

Figure Lengend Snippet: Effects of JWH133 on p38 MAPK, ERK1/2, and JNK1/2 signal transduction in lung tissues of PQ-induced acute lung injury rats. JWH133 was intraperitoneally administered 1 h before intraperitoneal administration of PQ. Lung tissue was collected after PQ administration for 72 h to determine the expression of p38 MAPK, ERK1/2, and JNK1/2 by Western blotting. L-JWH133: JWH133 at the low dose of 5 mg/kg and H-JWH133: JWH133 at the high dose of 20 mg/kg. The values presented are the mean ± SD. * P < 0.05 versus PQ group; # P < 0.05 versus L-JWH133 group.

Article Snippet: CB2R-agonist JWH133 was obtained from Enzo Life Sciences Ltd. (UK).

Techniques: Transduction, Expressing, Western Blot

Journal: BioMed Research International

Article Title: CB2 Receptor Activation Ameliorates the Proinflammatory Activity in Acute Lung Injury Induced by Paraquat

doi: 10.1155/2014/971750

Figure Lengend Snippet: Effects of JWH133 on NF- κ B signal transduction in lung tissues of PQ-induced acute lung injury rats. JWH133 was intraperitoneally administered 1 h before intraperitoneal administration of PQ. Lung tissue was collected after PQ administration for 72 h to determine the expression of NF- κ B p65, I κ B- α , and pI κ B- α by Western blotting. L-JWH133: JWH133 at the low dose of 5 mg/kg and H-JWH133: JWH133 at the high dose of 20 mg/kg. The values presented are the mean ± SD. * P < 0.05 versus PQ group; # P < 0.05 versus L-JWH133 group.

Article Snippet: CB2R-agonist JWH133 was obtained from Enzo Life Sciences Ltd. (UK).

Techniques: Transduction, Expressing, Western Blot