Journal: Developmental dynamics : an official publication of the American Association of Anatomists

Article Title: The endocannabinoid system regulates both ependymoglial and neuronal cell responses to a tail amputation in the axolotl.

doi: 10.1002/dvdy.70035

Figure Lengend Snippet: FIGURE 1 Identification and conservation of the axolotl endocannabinoid receptors. (A) Protein sequence alignment of the putative axolotl CB1 sequence with the rat and zebrafish CB1 sequence. (B) Protein sequence alignment of the putative axolotl CB2 sequence with the rat and zebrafish CB2 sequence. Red asterisks and red boxes indicate amino acids that are conserved between all three species. (C) Western blot using the rat CB1 antibody on axolotl tail tissue demonstrates a single prominent band at 120 kDa (n = 3). (D) Western blot using the rat CB2 antibody on axolotl tail tissue demonstrates two bands at a similar molecular weight of 46 kDa (n = 3). MW = molecular weight (for each band in ladder). (E) Preadsorption control for the CB1 antibody using either CB1 or CB2 antigenic peptides (n = 3). (F) Preadsorption control for the CB2 antibody using CB1 or CB2 antigenic peptides (n = 3).

Article Snippet: Overnight incubations in primary antibodies were then performed at 4 C using 1:1000 CB1 (Alomone Labs) and 1:1000 CB2 (Alomone Labs).

Techniques: Sequencing, Western Blot, Molecular Weight, Control

Journal: Developmental dynamics : an official publication of the American Association of Anatomists

Article Title: The endocannabinoid system regulates both ependymoglial and neuronal cell responses to a tail amputation in the axolotl.

doi: 10.1002/dvdy.70035

Figure Lengend Snippet: FIGURE 2 CB1 and CB2 are upregulated in response to tail amputation. (A) Western blot analysis demonstrates a significant upregulation of CB1 in the first 3 days after tail amputation, compared to uninjured controls (n = 3; F(4,40) = 5.994, p = .0007, one-way ANOVA). (B) No change in CB2 expression is shown in the first 3 days post tail amputation (n = 3; F(4,40) = 2.779, p = .0397, one-way ANOVA). (C) Western blot analysis demonstrates a significant upregulation of CB1 expression at both 7 and 14 days after tail amputation (n = 3; F(2,24) = 15.97, p < .0001, one-way ANOVA). (D) Western blot analysis demonstrates a significant upregulation of CB2 at 7 and 14 days post tail amputation (n = 3; F(2,24) = 10.84, p = .0004, one-way ANOVA). Uninj = uninjured tail tissue. hpa = hours post tail amputation; dpa = days post tail amputation. ns = not significant. *p < .05, **p < .01, ***p < .001, ****p < .0001 compared to uninjured controls. #p < .05.

Article Snippet: Overnight incubations in primary antibodies were then performed at 4 C using 1:1000 CB1 (Alomone Labs) and 1:1000 CB2 (Alomone Labs).

Techniques: Western Blot, Expressing

Journal: Developmental dynamics : an official publication of the American Association of Anatomists

Article Title: The endocannabinoid system regulates both ependymoglial and neuronal cell responses to a tail amputation in the axolotl.

doi: 10.1002/dvdy.70035

Figure Lengend Snippet: FIGURE 4 Inhibiting CB1 and CB2 receptor signaling impairs tail regeneration. (A) Representative images of tail regenerates after a 7-day treatment with the vehicle (control, i), 1 μM AM251 (ii), or 1 μM AM630 (iii). Black dotted line indicates the original plane of amputation. Scale bar: 1 mm. (B, C) Graphs show that the proportional increase in axolotl body length was significantly reduced following either a 7-day treatment with either 1 μM AM251 (n = 8; B) or after a 7-day treatment with 1 μM AM630 (n = 8; C) compared to the vehicle control (unpaired t tests). (D) Graph shows a significant reduction in the proportional increase in axolotl body length (7 days after tail amputation) following only a 1-day pulse treatment with either 1 μM AM251 (n = 10) or 1 μM AM630 (n = 10), compared to vehicle controls (n = 10; F(2,27) = 18.86; p < .0001, one-way ANOVA). (E) Western blot analyses show that treatment with AM251 prevented the upregulation of CB1 that normally occurs in untreated or vehicle-treated control animals at 7-days post tail amputation (n = 3; Constant 7-day bath treatment: F(3,32) = 14.69; p < .0001; 1-day pulse treatment: F(3,32) = 18.60; p < .0001; one-way ANOVAs). Representative blot for 1-day pulse treatment shown. (F) Treatment with AM630 prevented the upregulation of CB2 that normally occurs in untreated or vehicle- treated control animals at 7-days post tail amputation (n = 3; constant treatment: F(3,32) = 24.80; p < .0001; 1-day pulse treatment: F(3,32) = 11.60; p < .0001; one-way ANOVAs). Representative blot for 7-day constant treatment shown. **p < .01, ***p < .001, ****p < .0001 compared to vehicle controls. ###p < .001. ####p < .0001.

Article Snippet: Overnight incubations in primary antibodies were then performed at 4 C using 1:1000 CB1 (Alomone Labs) and 1:1000 CB2 (Alomone Labs).

Techniques: Control, Western Blot

Journal: Chinese Medicine

Article Title: Electroacupuncture exerts antipruritic and anti-inflammatory effects on atopic dermatitis by activating CB2 receptor

doi: 10.1186/s13020-025-01102-4

Figure Lengend Snippet: EA further promotes the expression of CB2R and modulates endocannabinoid metabolism in AD skin lesions. A Comparative gene expression fold changes (FC) are illustrated on a color scale, with blue indicates underexpression (FC < 2) and yellow indicates overexpression (FC > 2). The comparisons are shown for AD patients’ lesional skin compared to healthy human skin (left panel), and within AD patients between lesional skin (LS) and non-lesional skin (NLS) (right panel). B Expression levels of CNR2 mRNA were determined using RT-qPCR, normalization to GAPDH. C Representative WB diagram for CB2R. D Quantitative analysis of CB2R expression, normalized to α-tubulin. E Immunofluorescence images of skin lesions demonstrate the presence of CB2R (red), while nuclei of the cells were stained using DAPI (blue). Scale bars = 100 μm. F Summary data illustrate the quantity of CB2R + cells located in the dermis of skin lesions. G-H Expression levels of endocannabinoid metabolism-related enzymes, including DAGLβ, NAPE-PLD, MAGL and FAAH mRNA, were assessed via RT-qPCR, normalized to GAPDH. All data are shown as mean ± S.E.M (n = 6 per group). Analysis was performed using one-way ANOVA complemented by Tukey's post hoc test for multiple comparisons, * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Adult male C57BL/6 J mice were obtained from Beijing Vital River Laboratory Animal Technology and male CB2 receptor knockout (CB2R −/− ) mice on C57BL/6 background aged 6–8 weeks from Jackson Laboratories (Strain #:005786).

Techniques: Expressing, Gene Expression, Over Expression, Quantitative RT-PCR, Immunofluorescence, Staining

Journal: Chinese Medicine

Article Title: Electroacupuncture exerts antipruritic and anti-inflammatory effects on atopic dermatitis by activating CB2 receptor

doi: 10.1186/s13020-025-01102-4

Figure Lengend Snippet: CB2R knockout mitigates the therapeutic efficacy of EA on AD-like lesions and chronic pruritus. A The protocol regarding the establishment of the AD model, the EA treatment and the detection time of chronic pruritus behavior in WT and CB2R −/− mice. B Time-course of SCORAD scores(n = 8). C Representative images of skin lesions from each group of mice on day8. D Representative H&E staining images of each group. Scale bar = 100 μm. E Quantitative measurement of epidermal thickness based on the H&E images (n = 8). F Total number of scratches performed by mice in each group within one hour on day 8 (n = 12). All data are shown as mean ± S.E.M. Analysis was performed using two-way ANOVA complemented by Sidak's test for multiple comparisons, * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Adult male C57BL/6 J mice were obtained from Beijing Vital River Laboratory Animal Technology and male CB2 receptor knockout (CB2R −/− ) mice on C57BL/6 background aged 6–8 weeks from Jackson Laboratories (Strain #:005786).

Techniques: Knock-Out, Drug discovery, Staining

Journal: Chinese Medicine

Article Title: Electroacupuncture exerts antipruritic and anti-inflammatory effects on atopic dermatitis by activating CB2 receptor

doi: 10.1186/s13020-025-01102-4

Figure Lengend Snippet: EA influences mast cell and CD4 + T cell proliferation in AD through CB2R. A Representative toluidine blue–stained images of skin lesions. Scale bars (overview) = 100 μm and scale bars (magnified) = 50 μm. B-D Statistical results of the total number ( B ), the number of degranulation ( C ) and the degranulation rate of mast cells ( D ) (n = 8). E Immunofluorescence images of skin lesions show the CD4 + T cells (green). Nuclei of the cells were stained using DAPI (blue). Scale bars (overview) = 100 μm and scale bars (magnified) = 50 μm F Summarized data present the count of CD4 + T cells within skin sections (n = 6). All data are shown as mean ± S.E.M. Analysis was performed using two-way ANOVA complemented by Sidak's test for multiple comparisons, * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Adult male C57BL/6 J mice were obtained from Beijing Vital River Laboratory Animal Technology and male CB2 receptor knockout (CB2R −/− ) mice on C57BL/6 background aged 6–8 weeks from Jackson Laboratories (Strain #:005786).

Techniques: Staining, Immunofluorescence

Journal: Chinese Medicine

Article Title: Electroacupuncture exerts antipruritic and anti-inflammatory effects on atopic dermatitis by activating CB2 receptor

doi: 10.1186/s13020-025-01102-4

Figure Lengend Snippet: EA modulates cytokine release and receptor activation via CB2R and its downstream ERK pathway in AD mice. A Schematic diagram illustrating the procedures for RT-qPCR in each group. B–F Expression levels of IL4, IL13, IL31, IL4R and IL31R mRNA as determined by RT-qPCR, normalized to GAPDH. G Schematic representation of the role of CB2R activation in modulating inflammatory responses by targeting the ERK phosphorylation. H Representative WB results for p-ERK and ERK. I Quantitative assessment of p-ERK levels, with expression levels normalized to ERK. All data are shown as mean ± S.E.M (n = 6 per group). Analysis was performed using Two-way ANOVA complemented by Sidak's test for multiple comparisons, * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Adult male C57BL/6 J mice were obtained from Beijing Vital River Laboratory Animal Technology and male CB2 receptor knockout (CB2R −/− ) mice on C57BL/6 background aged 6–8 weeks from Jackson Laboratories (Strain #:005786).

Techniques: Activation Assay, Quantitative RT-PCR, Expressing, Phospho-proteomics

Journal: Chinese Medicine

Article Title: Electroacupuncture exerts antipruritic and anti-inflammatory effects on atopic dermatitis by activating CB2 receptor

doi: 10.1186/s13020-025-01102-4

Figure Lengend Snippet: Schematic diagram of this study's findings on the impact of EA on AD Mice. This schematic diagram presents the pathophysiological alterations in AD and the therapeutic interventions of EA treatment. Initially, upon the induction of AD, there is an escalation in the number of mast cells and CD4 + T cells within the skin lesions, along with upregulation of cytokines IL4, IL13, and IL31, and their receptors IL4R and IL31R in the cervical DRG. These changes are linked to the exacerbation of chronic itching and skin inflammation. Despite an upregulation of the CB2R, the endogenous levels are not adequate to effectively inhibit ERK phosphorylation, a key driver of inflammatory processes (left side). In contrast, EA treatment, applied via specific acupoints corresponding to the affected dermatome, initiates a cascade of anti-inflammatory effects. EA stimulates axon reflexes that enhance the synthesis of endocannabinoids and reduce their degradation in the lesional skin tissue. This results in elevated levels of the primary endocannabinoids such as AEA and 2-AG, which in turn upregulate the expression of CB2R. Stimulation of CB2R by endocannabinoids leads to a significant reduction in ERK hyperphosphorylation, an essential process in curbing inflammatory response. As a result, EA treatment diminishes the number of mast cells and CD4 + T cells in lesional skin, reduces the expression of pro-inflammatory cytokines IL4, IL13, and IL31, and decreases the expression of IL4R and IL31R in the cervical DRG. Collectively, these effects contribute to the alleviation of itching and the inhibition of inflammation development (right side)

Article Snippet: Adult male C57BL/6 J mice were obtained from Beijing Vital River Laboratory Animal Technology and male CB2 receptor knockout (CB2R −/− ) mice on C57BL/6 background aged 6–8 weeks from Jackson Laboratories (Strain #:005786).

Techniques: Phospho-proteomics, Expressing, Inhibition