Journal: International Journal of Molecular Sciences

Article Title: Preventive Effect of Epigallocatechin-3-Gallate on Postoperative Cognitive Dysfunction in Aged Rats via Modulation of Microglial Differentiation: An Experimental Animal Study

doi: 10.3390/ijms262311326

Figure Lengend Snippet: Differentiation for the phenotype expression of microglia, M1 phenotype as a pro-inflammatory producer ( A ) and M2 phenotype as an anti-inflammatory producer ( B ). Colors: DAPI (blue, nuclei); microglial expression (yellow). White arrows indicate the target cells. Tissue collection: brains collected 24 h after anesthesia; anesthesia: isoflurane 1.5 vol% for 4 h (endotracheal intubation, controlled ventilation). * p < 0.05 vs. Control group. Abbreviations: DAPI, 4′,6-diamidino-2-phenylindole staining; Iba1, ionized calcium-binding adapter molecule 1 staining; CD, cluster of differentiation; Merged, merged image of DAPI staining, Iba1 staining, and CD16 (for M1 phenotype) or CD206 staining (for M2 phenotype).

Article Snippet: Cluster of differentiation 16 (CD16) antibody (Bioss Antibodies, Woburn, MA, USA, cat bs-6028R) for the M1 phenotype and CD206 antibody (Bioss Antibodies, cat bsm-60761R) for the M2 phenotype were used for the detection of differentiation.

Techniques: Expressing, Control, Staining, Binding Assay

Journal: Journal of Inflammation Research

Article Title: The Role of Bone Morphogenetic Protein 4 in Microglial Polarization in the Process of Neuropathic Pain

doi: 10.2147/JIR.S356531

Figure Lengend Snippet: Primers for Each Gene for PCR Analysis

Article Snippet: All sections were rinsed in 0.2%Triton X-100 mixed with 5% donkey serum for 40 min at room temperature and then incubated overnight with primary antibodies at 4°C at the following dilutions: mouse anti-CD11b (SM262PS, 1:50, OriGene, MD, US), rabbit anti-CD16 (1:50, Affinity) and rabbit anti-ARG1 (1:100, Affinity).

Techniques:

Journal: Journal of Inflammation Research

Article Title: The Role of Bone Morphogenetic Protein 4 in Microglial Polarization in the Process of Neuropathic Pain

doi: 10.2147/JIR.S356531

Figure Lengend Snippet: Exogenous BMP4-induced allodynia, microglial activation and polarization. ( A ) PWT in rats after intrathecal BMP4 application showed a significant effect on time: F (4.143, 66.29) = 16.09, P<0.01, left; F (3.960, 63.35) = 12.67, P<0.01, right, intrathecal application: F (1, 16) = 161.3, P<0.01, left; F (1, 16) = 160.5, P<0.01, right and interaction: F (7, 112) = 11.76, P<0.01, left; F (7, 112) = 5.653, P<0.01, right. Compared with the Sham group, rats in the BMP4 group developed a significant decrease in bilateral PWT for the whole first week, P<0.01, n=9 at each time point for both groups. ( B ) Representative Western blotting showed a sustained increase of CD11b (P<0.01) and CD16 (P<0.01) expressions for the whole 1st week in the BMP4 group compared with the Sham group. Meanwhile, ARG-1 levels increased at day 1 (P<0.01) and day 4 (P<0.01), then fell below the levels of the Sham group (P<0.01). n=3 for each column. ( C and D ) Double-immunofluorescence further detected that compared with the Sham group (P7), CD11b expression was elevated in the dorsal horn of spinal cord after BMP4 treatment. Moreover, expressions of CD16 and ARG-1 showed similar pattern with Western blotting results, which were both mainly accumulated with CD11b + cells. n=3 for each column. Two-way ANOVA, followed by a Bonferroni test ( A ) and one-way ANOVA, followed by Sidak’s multiple comparisons test ( B – D ) were performed to analyze the statistical differences. *Represented P<0.05 and **Represented P<0.01 compared with the Sham group.

Article Snippet: All sections were rinsed in 0.2%Triton X-100 mixed with 5% donkey serum for 40 min at room temperature and then incubated overnight with primary antibodies at 4°C at the following dilutions: mouse anti-CD11b (SM262PS, 1:50, OriGene, MD, US), rabbit anti-CD16 (1:50, Affinity) and rabbit anti-ARG1 (1:100, Affinity).

Techniques: Activation Assay, Western Blot, Immunofluorescence, Expressing

Journal: Journal of Inflammation Research

Article Title: The Role of Bone Morphogenetic Protein 4 in Microglial Polarization in the Process of Neuropathic Pain

doi: 10.2147/JIR.S356531

Figure Lengend Snippet: Exogenous BMP4 induced a distinct pattern of M1/M2 gene expressions. RT-PCR analysis showed that M1-gene markers, including CD16, TNF-α and MHC-II, significantly increased throughout the whole 1st week, while M2-gene markers including ARG-1, CD-204 and IL-4 increased at an early stage (P1 or P4), then all fell to the Sham level at day 7. n=3 for each time point for both groups. One-way ANOVA, followed by Sidak’s multiple comparisons test were performed to analyze the statistical differences. *Represented P<0.05 and **Represented P<0.01 compared with the Sham group.

Article Snippet: All sections were rinsed in 0.2%Triton X-100 mixed with 5% donkey serum for 40 min at room temperature and then incubated overnight with primary antibodies at 4°C at the following dilutions: mouse anti-CD11b (SM262PS, 1:50, OriGene, MD, US), rabbit anti-CD16 (1:50, Affinity) and rabbit anti-ARG1 (1:100, Affinity).

Techniques: Reverse Transcription Polymerase Chain Reaction

Journal: Journal of Inflammation Research

Article Title: The Role of Bone Morphogenetic Protein 4 in Microglial Polarization in the Process of Neuropathic Pain

doi: 10.2147/JIR.S356531

Figure Lengend Snippet: Noggin relieved allodynia, attenuated microglial activation, and changed the polarized pattern after SNL. ( A ) PWT in rats receiving intrathecal Noggin application after SNL showed a significant effect on time: F (3.463, 55.41) = 162.3, P<0.01, intrathecal application: F (1, 16) = 44.85, P<0.01 and interaction: F (7, 112) = 3.540, P<0.01. Compared with the SNL group, Noggin application provided marked pain relief at day 3, day 4, day 5 and day 7 after SNL. n=9 at each time point for both groups. ( B and C ) Representative Western blotting showed SNL induced CD11b and CD16 upregulation at the first week, while ARG-1 expression remained unchanged compared with the Sham group; Moreover, Noggin effectively decreased the CD11b expressions at P1 and P4 and the CD16 expressions consecutively for the whole week. Simultaneously, the ARG-1 levels significantly elevated from P1 to P7 after Noggin treatment. n=3 for each column. Two-way ANOVA, followed by a Bonferroni test (A) and one-way ANOVA, followed by Sidak’s multiple comparisons test (B and C) were performed to analyze the statistical differences. **Represented P<0.01 compared with the Sham group, # Represented P<0.05 and ## Represented P<0.01 compared with the SNL group.

Article Snippet: All sections were rinsed in 0.2%Triton X-100 mixed with 5% donkey serum for 40 min at room temperature and then incubated overnight with primary antibodies at 4°C at the following dilutions: mouse anti-CD11b (SM262PS, 1:50, OriGene, MD, US), rabbit anti-CD16 (1:50, Affinity) and rabbit anti-ARG1 (1:100, Affinity).

Techniques: Activation Assay, Western Blot, Expressing

Journal: Journal of Inflammation Research

Article Title: The Role of Bone Morphogenetic Protein 4 in Microglial Polarization in the Process of Neuropathic Pain

doi: 10.2147/JIR.S356531

Figure Lengend Snippet: Noggin decreased M1-gene levels and induced a late-stage elevation of M2-gene levels after SNL. The mRNA levels of M1 and M2 subtype were determined by the RT-PCR method. The mRNA relative expressions from the Sham group were set as reference (equal to 1.0). As for the M1 genes, a significant increase in gene expressions were detected in CD16 from P1 to P7 and TNF-α from P4 to P7 in the SNL group compared with the Sham group, except for the MHC-II, which stayed unchanged. Then, in the SNL+NOG group, Noggin application markedly decreased CD16 and TNF-α levels for the 1st week. As for the M2 genes, SNL induced statistical elevation of ARG-1 from P4 and P7 and CD204 from P1 to P7, while IL-4 stayed comparable with the Sham group. Then, in the SNL+NOG group, Noggin treatment prominently increased ARG-1 at P7, CD204 at P4 and P7 and IL-4 at P7. n=3 for each time point. One-way ANOVA, followed by Sidak’s multiple comparisons test were performed to analyze the statistical differences. *Represented P<0.05 and **Represented P<0.01 compared with the Sham group; # Represented P<0.05 and ## Represented P<0.01 compared with the SNL group.

Article Snippet: All sections were rinsed in 0.2%Triton X-100 mixed with 5% donkey serum for 40 min at room temperature and then incubated overnight with primary antibodies at 4°C at the following dilutions: mouse anti-CD11b (SM262PS, 1:50, OriGene, MD, US), rabbit anti-CD16 (1:50, Affinity) and rabbit anti-ARG1 (1:100, Affinity).

Techniques: Reverse Transcription Polymerase Chain Reaction