Journal: Cellular & Molecular Biology Letters

Article Title: Dichotomous SMAD2/3 regulation and selective antihypertrophic activity of heparin during in vitro chondrogenesis of mesenchymal stromal cells

doi: 10.1186/s11658-026-00899-8

Figure Lengend Snippet: MSC in vitro chondrogenesis in heparin–PEG hydrogels versus pellet controls. MSCs were cultured as pellets (pellet control [ctrl]; 5 × 10 5 cells) or in heparin–PEG hydrogels (H–PEG hydrogel; 1.2 × 10 6 cells) containing 22.4 mg/mL crosslinked heparin and 120 ng TGF-β1 for 4 weeks in TGF-β1-containing (10 ng/mL) standard chondrogenic medium in vitro. A , B Microsections of day-28 samples were assessed either via immunohistochemistry to detect type II collagen or aggrecan, and type X collagen, or by enzymatic activity staining to visualize alkaline phosphatase, as indicated (scale bar: overview = 200 µm, magnification = 50 µm; positive and negative staining controls are shown in Supplementary Fig. S2). One representative result out of n = 5 experiments with independent MSC donor populations is shown. C Day-28 gene expression levels of chondrocyte and D hypertrophy markers, with CPSF6 and HPRT used as reference genes. The hypertrophy markers in day-28 samples are expressed as ratio to COL2A1 . n = 4 experiments using independent MSC donor populations. Box plots were built as described in the statistics section. * p \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\le $$\end{document} ≤ 0.05, Mann–Whitney U test

Article Snippet: Human type II collagens, type X collagens, or aggrecans were visualized after incubation with a mouse anti-type II collagen antibody (1:1000, same as for western blotting), a mouse anti-type X collagen antibody (1:1, as above), and a mouse anti-aggrecan antibody (1:25, clone HAG7D4 (7D4), no. SM1353, Acris/OriGene, Rockville, MD, USA), respectively, and an ALP-coupled goat anti-mouse immunoglobulin G secondary antibody (ImmunoLogic, WellMed, MS Arnhem, the Netherlands) using the ImmPACT ® Vector ® Red Substrate kit (Vector Laboratories, Newark, CA, USA).

Techniques: In Vitro, Cell Culture, Control, Immunohistochemistry, Activity Assay, Staining, Negative Staining, Gene Expression, MANN-WHITNEY

Journal: Cellular & Molecular Biology Letters

Article Title: Dichotomous SMAD2/3 regulation and selective antihypertrophic activity of heparin during in vitro chondrogenesis of mesenchymal stromal cells

doi: 10.1186/s11658-026-00899-8

Figure Lengend Snippet: Effect of soluble heparin on MSC in vitro chondrogenesis. MSCs were cultured as pellets for 28 days in standard chondrogenic medium (including 10 ng/mL TGF-β1 and 6.25 µg/mL insulin) supplemented with 0, 10, 100, or 700 µg/mL soluble heparin. A Paraffin microsections of day-28 samples were assessed for type II collagen and aggrecan via immunohistochemistry (scale bar: overview = 200 µm, magnification = 50 µm). Representative pictures of n = 5 experiments with independent MSCs are shown. B Type II collagen content per pellet was measured by ELISA ( n = 4). C Proteoglycan content per pellet was assessed using DMMB assay and normalized to DNA amounts per pellet ( n = 5). D Cell culture supernatants were collected from seven to nine pellets at weekly intervals and analyzed for PGE2 levels using ELISA. Box plots were built as described in the statistics section and analyzed using Mann–Whitney U test. No statistically significant differences were found between the groups ( p \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$>$$\end{document} > 0.05). Line graphs show data as mean \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\pm $$\end{document} ± standard error of the mean (SEM). # p \documentclass[12pt]{minimal} \usepackage{amsmath} \usepackage{wasysym} \usepackage{amsfonts} \usepackage{amssymb} \usepackage{amsbsy} \usepackage{mathrsfs} \usepackage{upgreek} \setlength{\oddsidemargin}{-69pt} \begin{document}$$\le $$\end{document} ≤ 0.05 versus control, paired Student’s t -test

Article Snippet: Human type II collagens, type X collagens, or aggrecans were visualized after incubation with a mouse anti-type II collagen antibody (1:1000, same as for western blotting), a mouse anti-type X collagen antibody (1:1, as above), and a mouse anti-aggrecan antibody (1:25, clone HAG7D4 (7D4), no. SM1353, Acris/OriGene, Rockville, MD, USA), respectively, and an ALP-coupled goat anti-mouse immunoglobulin G secondary antibody (ImmunoLogic, WellMed, MS Arnhem, the Netherlands) using the ImmPACT ® Vector ® Red Substrate kit (Vector Laboratories, Newark, CA, USA).

Techniques: In Vitro, Cell Culture, Immunohistochemistry, Enzyme-linked Immunosorbent Assay, Dimethylmethylene Blue Assay, MANN-WHITNEY, Control

Journal: Frontiers in Neuroanatomy

Article Title: Transmitter and ion channel profiles of saccadic omnipause neurons and cholinergic non-omnipause neurons in human nucleus raphe interpositus

doi: 10.3389/fnana.2025.1670220

Figure Lengend Snippet: Immunoperoxidase-based histochemical classification of omnipause neurons (OPN) and cholinergic non-OPNs in human nucleus raphe interpositus (RIP). (A) Combined immunoperoxidase labeling of perineuronal net (PN) marker aggrecan (ACAN, black) and non-phosphorylated neurofilament marker SMI32 (brown) reveals OPNs that are arranged as two columns around the midline in human RIP. The box indicates the area illustrated at higher magnification in B. (B) Close-up demonstrating OPNs co-immunolabeled with ACAN and SMI32 (red arrows) Note the rather dispersed arrangement of OPNs around the midline compared to OPNs in monkey (C) . Also note several neurons that are not ensheathed by PN marker ACAN but express SMI32-immunolabelling (green arrows). (C) In monkey RIP, combined peroxidase labeling of PN marker ACAN (black) and ChAT (brown) demonstrates the absence of such cholinergic neurons and tightly organized OPN columns around the midline. (D) Combined peroxidase labeling of perineuronal net (PN) marker aggrecan (ACAN, black) and non-phosphorylated neurofilament marker SMI32 (brown) and PN marker CSPG (black) choline acetyltransferase (ChAT, brown) reveal the cholinergic non-OPN population (green arrow) that are not ensheathed by PNs. Scale bar represents 1 mm A , 100 μm B,C and 50 μm in D .

Article Snippet: ACAN , Mouse/Monoclonal , Aggrecan , Acris Antibodies GmbH, 32052 Herford, GERMANY , AB_972582 , 1:75.

Techniques: Labeling, Marker, Immunolabeling

Journal: Frontiers in Neuroanatomy

Article Title: Transmitter and ion channel profiles of saccadic omnipause neurons and cholinergic non-omnipause neurons in human nucleus raphe interpositus

doi: 10.3389/fnana.2025.1670220

Figure Lengend Snippet: Immunoperoxidase-based histochemical classification of omnipause neurons (OPN) and cholinergic non-OPNs in human nucleus raphe interpositus (RIP). (A) Combined immunoperoxidase labeling of perineuronal net (PN) marker aggrecan (ACAN, black) and non-phosphorylated neurofilament marker SMI32 (brown) reveals OPNs that are arranged as two columns around the midline in human RIP. The box indicates the area illustrated at higher magnification in B. (B) Close-up demonstrating OPNs co-immunolabeled with ACAN and SMI32 (red arrows) Note the rather dispersed arrangement of OPNs around the midline compared to OPNs in monkey (C) . Also note several neurons that are not ensheathed by PN marker ACAN but express SMI32-immunolabelling (green arrows). (C) In monkey RIP, combined peroxidase labeling of PN marker ACAN (black) and ChAT (brown) demonstrates the absence of such cholinergic neurons and tightly organized OPN columns around the midline. (D) Combined peroxidase labeling of perineuronal net (PN) marker aggrecan (ACAN, black) and non-phosphorylated neurofilament marker SMI32 (brown) and PN marker CSPG (black) choline acetyltransferase (ChAT, brown) reveal the cholinergic non-OPN population (green arrow) that are not ensheathed by PNs. Scale bar represents 1 mm A , 100 μm B,C and 50 μm in D .

Article Snippet: ACAN , Mouse/Monoclonal , Aggrecan , Acris Antibodies GmbH, 32052 Herford, GERMANY , AB_972582 , 1:75.

Techniques: Labeling, Marker, Immunolabeling

Journal: IBRO Neuroscience Reports

Article Title: Chronic fluoxetine modulates perineuronal nets and inhibitory neuronal function in relation to behavioral outcomes

doi: 10.1016/j.ibneur.2025.10.012

Figure Lengend Snippet: Immunohistochemical analysis of parvalbumin (PV)-, AB1031-, and Cat-315-positive elements in the primary somatosensory cortex (S1BF). Images from the saline-treated group (A–C) and fluoxetine-treated group (A′–C′): PV-positive neurons (A, A′), AB1031-positive perineuronal nets (PNNs; B, B′), and Cat-315-positive PNNs (C, C′). Cortical layers (L) 2/3, L4, and L5/6 are indicated. The scale bar (100 µm), shown in panel C′, applies to all images (A–C and A′–C′). (G) Density of PV-positive neurons in L2/3, L4, and L5/6. (H) Density of AB1031-positive PNNs in L2/3, L4, and L5/6. (I) Density of Cat-315-positive PNNs in L2/3, L4, and L5/6. Data are presented as box-and-whisker plots with individual data points (n = 5–6 mice per group). Statistical analysis was performed using two-way ANOVA. p * < 0.05, p ** < 0.01; ns = not significant.

Article Snippet: The following primary antibodies were used for immunostaining: mouse anti-aggrecan (Cat-315; MAB1581, Merck; 1:1000), rabbit anti-aggrecan (AB1031, Millipore, Tokyo, Japan; 1:200), and mouse anti-parvalbumin (clone PARV-19, P3088; Sigma-Aldrich Japan, Tokyo, Japan; 1:1000).

Techniques: Immunohistochemical staining, Saline, Whisker Assay

Journal: IBRO Neuroscience Reports

Article Title: Chronic fluoxetine modulates perineuronal nets and inhibitory neuronal function in relation to behavioral outcomes

doi: 10.1016/j.ibneur.2025.10.012

Figure Lengend Snippet: Immunohistochemical analysis of parvalbumin (PV)-, AB1031-, and Cat-315-positive elements in the hippocampal CA1 and CA3 regions. Images from the CA1 region in the saline-treated group (A–C) and fluoxetine-treated group (A′–C′): PV-positive neurons (A, A′), AB1031-positive perineuronal nets (PNNs; B, B′), and Cat-315-positive PNNs (C, C′). Images from the CA3 region in the saline-treated group (D–F) and fluoxetine-treated group (D′–F′): PV-positive neurons (D, D′), AB1031-positive PNNs (E, E′), and Cat-315-positive PNNs (F, F′). Hippocampal subregions are indicated: so = stratum oriens, sp = stratum pyramidale, sr = stratum radiatum. The scale bar (100 µm), shown in panel F′, applies to all images (A–F and A′–F′). (G–I) Quantification of PV-positive neurons (G), AB1031-positive PNNs (H), and Cat-315-positive PNNs (I) in the CA1 region. (J–L) Quantification of PV-positive neurons ( J ), AB1031-positive PNNs (K), and Cat-315-positive PNNs ( L ) in the CA3 region. Data are presented as box-and-whisker plots with individual data points (n = 5–6 mice per group). Statistical analysis was performed using Student’s t -test. p * < 0.05, p ** < 0.01; ns = not significant.

Article Snippet: The following primary antibodies were used for immunostaining: mouse anti-aggrecan (Cat-315; MAB1581, Merck; 1:1000), rabbit anti-aggrecan (AB1031, Millipore, Tokyo, Japan; 1:200), and mouse anti-parvalbumin (clone PARV-19, P3088; Sigma-Aldrich Japan, Tokyo, Japan; 1:1000).

Techniques: Immunohistochemical staining, Saline, Whisker Assay