Journal: Histochemistry and Cell Biology

Article Title: Immunohistochemical analysis to detect a molecular signature in intervertebral disc degeneration

doi: 10.1007/s00418-025-02434-w

Figure Lengend Snippet: Detection of FOXO3a ( a ), SOD2 ( b ), HIF1α ( c ), GLUT1 ( d ), and Bry ( e ) expression by immunohistochemistry. Immunohistochemical analysis was performed on IVD tissues with different Pfirrmann grades of degeneration. Protein levels were quantified by densitometric analysis of immunostaining using ImageJ software and expressed as the percentage of positive cells per area (five sections per sample; Pfirrmann I–II group, n = 6; Pfirrmann III group, n = 40 and Pfirrmann IV–V group, n = 9). Results are reported as a whisker box plot representing the min to max (the line indicates median). * p < 0.01 (Pfirrmann III group vs. Pfirrmann I–II group); ^ p < 0.01 (Pfirrmann IV–V group vs. Pfirrmann I–II group); § p < 0.01 (Pfirrmann III group vs. Pfirrmann IV–V group). Scale bars 20 μm

Article Snippet: For immunohistochemical evaluation, sections were incubated overnight (4 °C) with a primary antibody against FOXO3a (#ab70315, rabbit anti-human, 1:100 dilution; Abcam, Cambridge, UK), SOD2 (#sc-133134, mouse anti-human, 1:100 dilution; Santa Cruz biotech., Dallas, USA), HIF1α (clone H1alpha67, mouse anti-human, 1:200 dilution; Novusbio, Centennial, USA), GLUT1 (#TA301678, mouse anti-human, 1:200 dilution; Origene, Rockville, USA), and BRY (#ab209665, rabbit anti-human, 1:500 dilution; Abcam), followed by treatment with Vectastain ABC solution (#MP-7500; Vectorlabs, Burlingame, USA) for 30 min. To achieve specificity, the working concentrations of primary antibodies were selected by following the dilution guidelines and technical specifications provided by the antibody’s manufacturer.

Techniques: Expressing, Immunohistochemistry, Immunohistochemical staining, Immunostaining, Software, Whisker Assay

Journal: Histochemistry and Cell Biology

Article Title: Immunohistochemical analysis to detect a molecular signature in intervertebral disc degeneration

doi: 10.1007/s00418-025-02434-w

Figure Lengend Snippet: Analysis of FOXO3a, SOD2, HIF1α, GLUT1, and Bry expression in IVD tissues with Pfirrmann grade III. Protein levels were assessed by immunohistochemistry and quantified by densitometric analysis of immunostaining using ImageJ software. Results were expressed as the percentage of positive cells per area, based on five representative sections per sample ( n = 40, Pfirrmann grade III). Data are presented as box-and-whisker plots, displaying min to max (the line indicates the median). Comparisons were made on the basis of the following parameters: a sex (female, male), age (18–40, 41–60, > 60 years), smoking status (non-smoker [NS], former smoker [FS], current smoker [CS]), and body mass index (BMI) (normal weight [NW], overweight [OW], obese [OB]); b anatomical site of surgery (L2–L3, L3–L4, L4–L5, L5–S1), duration of symptoms before surgery (< 6 months, ≥ 6 months), and area of infiltration (moderate, abundant)

Article Snippet: For immunohistochemical evaluation, sections were incubated overnight (4 °C) with a primary antibody against FOXO3a (#ab70315, rabbit anti-human, 1:100 dilution; Abcam, Cambridge, UK), SOD2 (#sc-133134, mouse anti-human, 1:100 dilution; Santa Cruz biotech., Dallas, USA), HIF1α (clone H1alpha67, mouse anti-human, 1:200 dilution; Novusbio, Centennial, USA), GLUT1 (#TA301678, mouse anti-human, 1:200 dilution; Origene, Rockville, USA), and BRY (#ab209665, rabbit anti-human, 1:500 dilution; Abcam), followed by treatment with Vectastain ABC solution (#MP-7500; Vectorlabs, Burlingame, USA) for 30 min. To achieve specificity, the working concentrations of primary antibodies were selected by following the dilution guidelines and technical specifications provided by the antibody’s manufacturer.

Techniques: Expressing, Immunohistochemistry, Immunostaining, Software, Whisker Assay

Journal: Histochemistry and Cell Biology

Article Title: Immunohistochemical analysis to detect a molecular signature in intervertebral disc degeneration

doi: 10.1007/s00418-025-02434-w

Figure Lengend Snippet: Correlation between FOXO3a, SOD2, HIF1α, GLUT1, and Bry expression levels and the onset of chronic pain or relapse. Results were expressed as percentage of positive cells per area, based on five representative sections per sample ( n = 24, Pfirrmann grade III). Data are presented as box-and-whisker plots, displaying min to max (the line indicates the median). Patients without chronic pain after surgery (green), with joint inflammatory symptoms after several months (blue), and with short-term relapses (red)

Article Snippet: For immunohistochemical evaluation, sections were incubated overnight (4 °C) with a primary antibody against FOXO3a (#ab70315, rabbit anti-human, 1:100 dilution; Abcam, Cambridge, UK), SOD2 (#sc-133134, mouse anti-human, 1:100 dilution; Santa Cruz biotech., Dallas, USA), HIF1α (clone H1alpha67, mouse anti-human, 1:200 dilution; Novusbio, Centennial, USA), GLUT1 (#TA301678, mouse anti-human, 1:200 dilution; Origene, Rockville, USA), and BRY (#ab209665, rabbit anti-human, 1:500 dilution; Abcam), followed by treatment with Vectastain ABC solution (#MP-7500; Vectorlabs, Burlingame, USA) for 30 min. To achieve specificity, the working concentrations of primary antibodies were selected by following the dilution guidelines and technical specifications provided by the antibody’s manufacturer.

Techniques: Expressing, Whisker Assay