Journal: Immunity, Inflammation and Disease

Article Title: MAFG Induces the Methylation of CRYAB to Promote the Activation of A1 Astrocyte After Spinal Cord Injury

doi: 10.1002/iid3.70334

Figure Lengend Snippet: The activation of A1 astrocyte was accompanied by MAFG upregulation. (A) In spinal cord tissues of rats, C3‐ and S100A10‐positive cells were checked using immunofluorescence. (B) GSE132242 dataset showed that MAFG was overexpressed in mice after SCI ( n = 4). In spinal cord tissues of rats, (C) RT‐PCR was applied to survey MAFG mRNA expression ( n = 6); (D) MAFG‐positive cells were checked using immunofluorescence; (E) the protein expression of MAFG, C3, and S100A10 was surveyed through Western blot ( n = 6). Data in (B, C, and E) were analyzed using the unpaired t ‑test. * p < 0.05, ** p < 0.01.

Article Snippet: For rat astrocytes, after the indicated treatment, the cells were placed in 4% paraformaldehyde for 15 min. After blocking with 10% goat serum for 1 h, cells were incubated with primary antibodies against MAFG, CRYAB (Abcam); C3, S100A10 (Proteintech, China) at 4°C overnight, maintained with secondary antibodies for 60 min, and treated with 4′,6‐diamidino‐2‐phenylindole (DAPI, Sigma‐Aldrich) for 10 min. After dewaxing, hydration, and antigen retrieval, the spinal cord tissue sections were incubated with primary antibodies against MAFG, C3, and S100A10 at 4°C overnight and treated with secondary antibodies for 1 h. Cells in the tissues were also stained.

Techniques: Activation Assay, Immunofluorescence, Reverse Transcription Polymerase Chain Reaction, Expressing, Western Blot

Journal: Immunity, Inflammation and Disease

Article Title: MAFG Induces the Methylation of CRYAB to Promote the Activation of A1 Astrocyte After Spinal Cord Injury

doi: 10.1002/iid3.70334

Figure Lengend Snippet: Silencing of MAFG inhibited the activation of A1 astrocyte and neuroinflammation. In spinal cord tissues of rats, (A) the mRNA expression of C3 and S100A10 was detected using RT‐PCR ( n = 6); (B) the protein expression of C3 and S100A10 was detected using Western blot ( n = 3); (C) the mRNA expression of TNF‐α, IL‐1β, IL‐6, and IL‐10 was detected using RT‐PCR ( n = 6); (D) the protein expression of TNF‐α, IL‐1β, IL‐6, and IL‐10 was detected using Western blot ( n = 3). All data were analyzed using one‑way ANOVA followed by Tukey's post hoc test. * p < 0.05, ** p < 0.01.

Article Snippet: For rat astrocytes, after the indicated treatment, the cells were placed in 4% paraformaldehyde for 15 min. After blocking with 10% goat serum for 1 h, cells were incubated with primary antibodies against MAFG, CRYAB (Abcam); C3, S100A10 (Proteintech, China) at 4°C overnight, maintained with secondary antibodies for 60 min, and treated with 4′,6‐diamidino‐2‐phenylindole (DAPI, Sigma‐Aldrich) for 10 min. After dewaxing, hydration, and antigen retrieval, the spinal cord tissue sections were incubated with primary antibodies against MAFG, C3, and S100A10 at 4°C overnight and treated with secondary antibodies for 1 h. Cells in the tissues were also stained.

Techniques: Activation Assay, Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: Immunity, Inflammation and Disease

Article Title: MAFG Induces the Methylation of CRYAB to Promote the Activation of A1 Astrocyte After Spinal Cord Injury

doi: 10.1002/iid3.70334

Figure Lengend Snippet: MAFG was highly expressed in the activated A1 astrocyte. (A) In rat astrocytes, MAFG positive cells were checked using immunofluorescence. (B and C) In rat astrocytes, MAFG mRNA expression was detected by RT‐PCR ( n = 3). (D and E) In rat astrocytes, the protein expression of MAFG, Serping1, C3, Sphk1, and S100A10 was detected using Western blot ( n = 3). Data in (B, C, D, and E) were analyzed using the unpaired t ‑test. ** p < 0.01.

Article Snippet: For rat astrocytes, after the indicated treatment, the cells were placed in 4% paraformaldehyde for 15 min. After blocking with 10% goat serum for 1 h, cells were incubated with primary antibodies against MAFG, CRYAB (Abcam); C3, S100A10 (Proteintech, China) at 4°C overnight, maintained with secondary antibodies for 60 min, and treated with 4′,6‐diamidino‐2‐phenylindole (DAPI, Sigma‐Aldrich) for 10 min. After dewaxing, hydration, and antigen retrieval, the spinal cord tissue sections were incubated with primary antibodies against MAFG, C3, and S100A10 at 4°C overnight and treated with secondary antibodies for 1 h. Cells in the tissues were also stained.

Techniques: Immunofluorescence, Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: Immunity, Inflammation and Disease

Article Title: MAFG Induces the Methylation of CRYAB to Promote the Activation of A1 Astrocyte After Spinal Cord Injury

doi: 10.1002/iid3.70334

Figure Lengend Snippet: Silencing of MAFG inhibited the activation of A1 astrocyte and neuroinflammation in rat astrocytes. (A and B) In rat astrocytes, the positive cells of C3 and S100A10 were checked using immunofluorescence. (C and D) In rat astrocytes, the mRNA expression of Serping1, C3, Sphk1, and S100A10 were checked using RT‐PCR ( n = 3). (E and F) In rat astrocytes, the protein expression of Serping1, C3, Sphk1, and S100A10 were checked using Western blot ( n = 3). (G and H) In supernatant of rat astrocytes, the levels of IL‐1β and IL‐6 were examined by ELISA ( n = 3). Data in (C, D, E, F, G, and H) were analyzed using one‑way ANOVA followed by Tukey's post hoc test. * p < 0.05, ** p < 0.01.

Article Snippet: For rat astrocytes, after the indicated treatment, the cells were placed in 4% paraformaldehyde for 15 min. After blocking with 10% goat serum for 1 h, cells were incubated with primary antibodies against MAFG, CRYAB (Abcam); C3, S100A10 (Proteintech, China) at 4°C overnight, maintained with secondary antibodies for 60 min, and treated with 4′,6‐diamidino‐2‐phenylindole (DAPI, Sigma‐Aldrich) for 10 min. After dewaxing, hydration, and antigen retrieval, the spinal cord tissue sections were incubated with primary antibodies against MAFG, C3, and S100A10 at 4°C overnight and treated with secondary antibodies for 1 h. Cells in the tissues were also stained.

Techniques: Activation Assay, Immunofluorescence, Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Immunity, Inflammation and Disease

Article Title: MAFG Induces the Methylation of CRYAB to Promote the Activation of A1 Astrocyte After Spinal Cord Injury

doi: 10.1002/iid3.70334

Figure Lengend Snippet: Silencing of MAFG inhibited the methylation of CRYAB in vitro which exerted anti‐inflammatory effects. (A) BSP was performed to detect CRYAB methylation (white represented unmethylation, black represented methylation). (B) In rat astrocytes, Western blot was applied to check the changes of key factors in DNA methylation (DNMT1, DNMT3a, and DNMT3b) ( n = 3). (C) In rat astrocytes, the positive cells of CRYAB were checked using immunofluorescence. (D) In rat astrocytes, CRYAB protein expression was checked using Western blot ( n = 3). (E) In rat astrocytes, the mRNA expression of Serping1, C3, Sphk1, and S100A10 was checked by RT‐PCR ( n = 3). (F) In rat astrocytes, the protein expression of Serping1, C3, Sphk1, and S100A10 was checked using Western blot ( n = 3). (G) In supernatant of rat astrocytes, the levels of IL‐1β and IL‐6 were examined by ELISA ( n = 3). Data in (B, D, E, F, and G) were analyzed using one‑way ANOVA followed by Tukey's post hoc test. * p < 0.05, ** p < 0.01.

Article Snippet: For rat astrocytes, after the indicated treatment, the cells were placed in 4% paraformaldehyde for 15 min. After blocking with 10% goat serum for 1 h, cells were incubated with primary antibodies against MAFG, CRYAB (Abcam); C3, S100A10 (Proteintech, China) at 4°C overnight, maintained with secondary antibodies for 60 min, and treated with 4′,6‐diamidino‐2‐phenylindole (DAPI, Sigma‐Aldrich) for 10 min. After dewaxing, hydration, and antigen retrieval, the spinal cord tissue sections were incubated with primary antibodies against MAFG, C3, and S100A10 at 4°C overnight and treated with secondary antibodies for 1 h. Cells in the tissues were also stained.

Techniques: Methylation, In Vitro, Western Blot, DNA Methylation Assay, Immunofluorescence, Expressing, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

Journal: Immunity, Inflammation and Disease

Article Title: MAFG Induces the Methylation of CRYAB to Promote the Activation of A1 Astrocyte After Spinal Cord Injury

doi: 10.1002/iid3.70334

Figure Lengend Snippet: Silencing of MAFG inhibited the activation of A1 astrocyte and neuroinflammation via CRYAB methylation. (A and B) In rat astrocytes, the mRNA expression of Serping1, C3, Sphk1, and S100A10 was checked by RT‐PCR ( n = 3). (C and D) In rat astrocytes, the protein expression of Serping1, C3, Sphk1, and S100A10 was checked using Western blot ( n = 3). (E and F) In supernatant of rat astrocytes, the levels of IL‐1β and IL‐6 were examined by ELISA ( n = 3). All data were analyzed using one‑way ANOVA followed by Tukey's post hoc test. * p < 0.05, ** p < 0.01.

Article Snippet: For rat astrocytes, after the indicated treatment, the cells were placed in 4% paraformaldehyde for 15 min. After blocking with 10% goat serum for 1 h, cells were incubated with primary antibodies against MAFG, CRYAB (Abcam); C3, S100A10 (Proteintech, China) at 4°C overnight, maintained with secondary antibodies for 60 min, and treated with 4′,6‐diamidino‐2‐phenylindole (DAPI, Sigma‐Aldrich) for 10 min. After dewaxing, hydration, and antigen retrieval, the spinal cord tissue sections were incubated with primary antibodies against MAFG, C3, and S100A10 at 4°C overnight and treated with secondary antibodies for 1 h. Cells in the tissues were also stained.

Techniques: Activation Assay, Methylation, Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Fluids and Barriers of the CNS

Article Title: Age‑dependent and post‑intraventricular hemorrhage remodeling of the ependymal glycocalyx in mice

doi: 10.1186/s12987-025-00725-x

Figure Lengend Snippet: Visualization and glycan profiling of the ependymal Gcx in young adult mice. ( a ) Scanning electron microscopy (SEM) images of ventricular ependymal cells. Gcx is not visible under conventional glutaraldehyde fixation (left) but clearly visualized surrounding the cell surface and cilia with lanthanum staining (right). ( b ) Alcian blue + hematoxylin-eosin (HE) staining. Glycans covering the apical surface of the ependyma are stained blue with Alcian blue. White scale bar: 20 μm. ( c ) Low-vacuum SEM image of a serial section corresponding to ( b ). A three-dimensional view of the Gcx layer covering the base of the cilia and surrounding the ciliary shafts at the apical surface. Enlarged view on the right. ( d ) Double immunofluorescence staining of lectin (PNA, red) and S100β (green) in frozen sections. A distinct glycan layer is observed at the apical surface of the ependyma, comparable to that seen in electron microscopy. White scale bar: 10 μm. ( e ) Fluorescence intensity plot along the measurement line (yellow) in ( d ), from the ventricular lumen (left) toward the brain parenchyma (right); red indicates lectin, green indicates S100β. The plot confirms localization of the Gcx at the ependymal cell surface. ( f ) Bar graph showing the mean fluorescence intensity of the ependymal Gcx for each of the 21 lectins examined ( n = 3 mice; total of 30 cells). ( g ) Merged images of lectin (red) and S100β (green) staining. Representative examples are shown: strong positive (RCA-I, STL), moderate positive (WGA, PHA-E), and negative (UEA-I, S-WGA). White scale bar: 10 μm

Article Snippet: Subsequently, biotinylated lectins (1:200 dilution) and an ependymal cell marker, the S100β antibody (mouse monoclonal, 1:500 dilution; sc-393919, Santa Cruz Biotechnology), were applied and incubated overnight at 4 °C.

Techniques: Glycoproteomics, Electron Microscopy, Staining, Double Immunofluorescence Staining, Fluorescence

Journal: Fluids and Barriers of the CNS

Article Title: Age‑dependent and post‑intraventricular hemorrhage remodeling of the ependymal glycocalyx in mice

doi: 10.1186/s12987-025-00725-x

Figure Lengend Snippet: Comparison of ependymal Gcx between young adult and aged mice. ( a ) Heatmap showing the mean fluorescence intensity per pixel of the ependymal glycocalyx (Gcx) for 21 lectins in young adult and aged mice ( n = 3 mice per group; 10 cells per mouse). ( b ) Representative PNA-based fluorescence images of the ependymal Gcx in young adult (top) and aged (bottom) mice. White scale bar, 5 μm. ( c ) Quantification of Gcx coverage along the periventricular circumference of the lateral ventricle ( n = 3 mice per group; two-sided t-test; mean ± S.E.M.). ( d ) Transmission electron microscopy (TEM) images of ependymal cells. In young adults (left), the Gcx is visible on the apical surface (black arrowheads), microvilli (black arrows), and cilia (small black arrow). In aged mice (right), loss of the Gcx is evident on the apical surface (white arrowheads) and microvilli (white arrows). White scale bar, 500 nm. ( e ) Schematic illustration of the ependymal surface Gcx based on lectin staining and TEM. ( f ) Double fluorescence staining for LEL (red) and S100β (green) (left) with corresponding fluorescence intensity profiles (right; red = LEL, green = S100β; dots indicate inflection points). White scale bar, 5 μm. ( g ) Correlation between inflection point distance and manually measured Gcx thickness using PNA staining ( n = 20 measurement sites; Pearson correlation). ( h ) Comparison of Gcx thickness between young adult and aged mice determined by inflection point distances for LEL, PNA, and RCA-I ( n = 3 mice per group; 16 sites per mouse; two-sided t-test; mean ± S.E.M.). ( i ) Representative double fluorescence staining for PNA (red) and S100β (green) showing enhanced cytoplasmic PNA positivity in aged ependymal cells. White scale bar, 5 μm. ( j ) Cytoplasmic PNA-positive rate along the ventricular wall ( n = 3 per group; two-sided t-test; mean ± S.E.M.). ( k ) Heatmap of mean fluorescence intensity in the ependymal cytoplasm for 21 lectins ( n = 3 per group; 10 cells per mouse)

Article Snippet: Subsequently, biotinylated lectins (1:200 dilution) and an ependymal cell marker, the S100β antibody (mouse monoclonal, 1:500 dilution; sc-393919, Santa Cruz Biotechnology), were applied and incubated overnight at 4 °C.

Techniques: Comparison, Fluorescence, Transmission Assay, Electron Microscopy, Staining

Journal: BMC Cancer

Article Title: Increased cytoplasmic and nuclear S100A6 expression is associated with improved prognosis in ovarian cancer

doi: 10.1186/s12885-026-15631-0

Figure Lengend Snippet: ( A ) Western blot of S100A6 in OVCA433 ovarian cancer cells, with an expected single band of 10 kDa. Full-length blots with an additional cell line (SKOV-3) are presented in Supplementary Fig. 2. Representative images of IHC staining for low ( B-D ) and high ( E-G ) cytoplasmic and nuclear expression of S100A6. 10X images with a 20X index box. Scale bar 100 μm

Article Snippet: Specificity of the S100A6 antibody (10245-1-AP, Proteintech) was initially determined via Western Blotting, to assess suitability for subsequent IHC based studies.

Techniques: Western Blot, Immunohistochemistry, Expressing

Journal: BMC Cancer

Article Title: Increased cytoplasmic and nuclear S100A6 expression is associated with improved prognosis in ovarian cancer

doi: 10.1186/s12885-026-15631-0

Figure Lengend Snippet: Kaplan-Meier analysis of ovarian cancer overall survival with low (black) and high (red) S100A6 expression in the cytoplasm ( A ) and nucleus ( B ). Significance was determined using the log-rank test

Article Snippet: Specificity of the S100A6 antibody (10245-1-AP, Proteintech) was initially determined via Western Blotting, to assess suitability for subsequent IHC based studies.

Techniques: Expressing

Journal: BMC Cancer

Article Title: Increased cytoplasmic and nuclear S100A6 expression is associated with improved prognosis in ovarian cancer

doi: 10.1186/s12885-026-15631-0

Figure Lengend Snippet: Kaplan-Meier analysis of ovarian cancer overall survival with low (black) and high (red) S100A6 expression in the cytoplasm ( A , C , E ) and nucleus ( B , D , F ) in high-grade serous carcinoma (HGSC) ( A - B ), mucinous carcinoma ( C - D ), and endometrioid carcinoma ( E - F ). Significance was determined using the log-rank test

Article Snippet: Specificity of the S100A6 antibody (10245-1-AP, Proteintech) was initially determined via Western Blotting, to assess suitability for subsequent IHC based studies.

Techniques: Expressing

Journal: BMC Cancer

Article Title: Increased cytoplasmic and nuclear S100A6 expression is associated with improved prognosis in ovarian cancer

doi: 10.1186/s12885-026-15631-0

Figure Lengend Snippet: Kaplan-Meier analysis of overall survival ( A ) and progression free survival ( B ) with low (black) and high (red) mRNA expression of S100A6 in ovarian cancer. Significance was determined using the log-rank test

Article Snippet: Specificity of the S100A6 antibody (10245-1-AP, Proteintech) was initially determined via Western Blotting, to assess suitability for subsequent IHC based studies.

Techniques: Expressing