Journal: International Journal of Biological Sciences

Article Title: SLC25A33-mediated mitochondrial DNA synthesis plays a critical role in the inflammatory response of M1 macrophages by contributing to mitochondrial ROS and VDAC oligomerization

doi: 10.7150/ijbs.96563

Figure Lengend Snippet: SLC25A33 is upregulated in LPS/IFN-γ-induced M1 macrophages and CD14+ monocytes from septic patients. (A) Volcano plot illustrating gene expression differences between untreated PMs and LPS/IFN-γ-treated PMs. Red asterisks denote differentially expressed genes in the SLC25 family, and green asterisks highlight highly upregulated genes within the same family. (B-D) Relative mRNA expression of SLC25A33 , SLC25A22 , and SLC25A37 (B) and pro-inflammatory cytokines (C) in CD14+ monocytes from septic patients with liver abscesses (n=17) compared with healthy controls (n=10), and (D) changes in relative mRNA expression pre- and post-recovery in these septic patients. (E-F) Relative iNOS and SLC25A33 mRNA expression (E) and their protein levels (F) in LPS/IFN-γ-treated PMs. (G) An ex vivo experimental protocol for the isolation and harvesting of PMs from C57BL/6 mice treated with LPS. (H-J) Relative mRNA expression of iNOS and SLC25A33 (H), their corresponding protein expression levels (I), and relative mRNA expression of pro-inflammatory cytokines (J) in PMs isolated from mice following intraperitoneal LPS injection. In all in vitro experiments, PMs were treated with LPS (100 ng/mL) and IFN-γ (10 ng/mL) for 24 h. For ex vivo experiments, mice were injected intraperitoneally with LPS (2 mg/kg) and PMs were isolated after 24 h. All experimental data were verified in at least three independent experiments. Data are presented as the mean ± SEM. N.S., not significant, *p < 0.05, **p < 0.01 and ***p < 0.001. iNOS: inducible nitric oxide synthase; PMs: peritoneal macrophages.

Article Snippet: Cells were transfected with scramble siRNA, siSLC25A33, siMyD88, siPI3K, simTOR, siTAK1, siTBK1, siATF4 (Bioneer), SLC25A33 ORF Clone (Origene) or ATF4 ORF Clone (GeneCopoeia) using Lipofectamine TM 3000 (Thermo Fisher Scientific).

Techniques: Gene Expression, Expressing, Ex Vivo, Isolation, Injection, In Vitro

Journal: International Journal of Biological Sciences

Article Title: SLC25A33-mediated mitochondrial DNA synthesis plays a critical role in the inflammatory response of M1 macrophages by contributing to mitochondrial ROS and VDAC oligomerization

doi: 10.7150/ijbs.96563

Figure Lengend Snippet: SLC25A33 inhibition reduces LPS/IFN-γ-induced inflammatory response and mtROS of M1 macrophages. (A-B) Effect of SLC25A33 -targeting siRNA on iNOS mRNA expression (A) and protein levels of iNOS, CD80, and CD86 (B) in LPS/IFN-γ-treated PMs. (C-D) Effect of SLC25A33 -targeting siRNA on relative mRNA expression (C) and secretion (D) of pro-inflammatory cytokines in the LPS/IFN-γ-treated PMs. (E-F) Effect of PLP on iNOS mRNA expression (E) and protein levels of iNOS, CD80, and CD86 (F) in the LPS/IFN-γ-treated PMs. (G-H) Effect of PLP on the relative mRNA expression (G) and secretion (H) of pro-inflammatory cytokines in the PMs treated with LPS/IFN-γ. (I-J) Representative image of immunofluorescence staining for MitoSOX Red (left panel) and quantification of fluorescence intensity (right panel) in the LPS/IFN-γ-treated PMs following treatment with SLC25A33 -targeting siRNA (I) and PLP (J). In all experiments, PMs were treated with LPS (100 ng/mL) and IFN-γ (10 ng/mL) for 24 h, with PLP (400 μM, 24 h), MitoSOX Red (5 μM, 15 minutes), and SLC25A33 -targeting siRNA (40 pM; 48 h). All experimental data were verified in at least three independent experiments. Data are presented as the mean ± SEM. Scale bar represents 10 μm. *p < 0.05, **p < 0.01, and ***p < 0.001. iNOS: inducible nitric oxide synthase; PMs: peritoneal macrophages; PLP: pyridoxal 5'-phosphate.

Article Snippet: Cells were transfected with scramble siRNA, siSLC25A33, siMyD88, siPI3K, simTOR, siTAK1, siTBK1, siATF4 (Bioneer), SLC25A33 ORF Clone (Origene) or ATF4 ORF Clone (GeneCopoeia) using Lipofectamine TM 3000 (Thermo Fisher Scientific).

Techniques: Inhibition, Expressing, Immunofluorescence, Staining, Fluorescence

Journal: International Journal of Biological Sciences

Article Title: SLC25A33-mediated mitochondrial DNA synthesis plays a critical role in the inflammatory response of M1 macrophages by contributing to mitochondrial ROS and VDAC oligomerization

doi: 10.7150/ijbs.96563

Figure Lengend Snippet: MyD88-PI3K-mTORC1 signaling and ATF4 are essential drivers of SLC25A33 expression in M1 macrophages. (A) Relative mRNA expression of SLC25A33 in PMs treated with LPS, IFN-γ, or their combination. (B-D) Effect of the MyD88 inhibitor ST2825 and the TBK1 inhibitor MRT67307 on the levels of indicated proteins (B), as well as on relative mRNA expression (C) and protein levels (D) of SLC25A33 in the LPS/IFN-γ-treated PMs. (E-G) Effect of the PI3K inhibitor LY294002 or the TAK1 inhibitor Takinib on the levels of the indicated proteins (E) and their relative mRNA expression (F) and the protein levels (G, LY294002 only) of SLC25A33 in the LPS/IFN-γ-treated PMs. (H) Effect of rapamycin on the relative mRNA expression of SLC25A33 in the LPS/IFN-γ-treated PMs. (I) Effect of rapamycin or LY294002 on protein levels of phosphorylated-S6K, S6K, SLC25A33, and ATF4 in the LPS/IFN-γ -treated PMs. (J) Effect of MyD88 , PI3K , mTOR , TAK1 or TBK1 -targeting siRNAs on protein levels of ATF4 and SLC25A33 in the LPS/IFN-γ-treated PMs. (K-M) Effect of ATF4 -targeting siRNA on the relative mRNA expression of ATF4 (K), SLC25A33 (L) and protein levels of ATF4 and SLC25A33 in the LPS/IFN-γ-treated PMs (M). (N) Effect of ATF4 -targeting siRNA on the relative luciferase activity of the Gluc reporter in the LPS/IFN-γ-treated PMs. (O) ChIP assay of ATF4 binding to its putative site within the SLC25A33 promoter in LPS/IFN-γ-treated PMs. (P) Effect of mutation of the putative ATF4 binding site within the SLC25A33 promoter on the relative luciferase activity of the Gluc reporter in the LPS/IFN-γ-treated PMs. In all experiments, PMs were treated with LPS (100 ng/mL) and IFN-γ (10 ng/mL) for 24 h, with ST2825 (10 μM, 24 h), MRT67307 (1 μM, 24 h), LY294002 (25 nM, 24 h), Takinib (10 μM, 24 h), rapamycin (50 nM, 24 h), and siRNAs against each gene (40 pM, 48 h). All experimental data were verified in at least three independent experiments. Data are presented as the mean ± SEM. N.S., not significant, *p < 0.05, **p < 0.01, and ***p < 0.001. ATF4: activating transcription factor 4; Gluc: Gaussia luciferase; PMs: peritoneal macrophages.

Article Snippet: Cells were transfected with scramble siRNA, siSLC25A33, siMyD88, siPI3K, simTOR, siTAK1, siTBK1, siATF4 (Bioneer), SLC25A33 ORF Clone (Origene) or ATF4 ORF Clone (GeneCopoeia) using Lipofectamine TM 3000 (Thermo Fisher Scientific).

Techniques: Expressing, Luciferase, Activity Assay, Binding Assay, Mutagenesis

Journal: International Journal of Biological Sciences

Article Title: SLC25A33-mediated mitochondrial DNA synthesis plays a critical role in the inflammatory response of M1 macrophages by contributing to mitochondrial ROS and VDAC oligomerization

doi: 10.7150/ijbs.96563

Figure Lengend Snippet: SLC25A33 inhibition reduces mitochondrial DNA release and modulates the cGAS-STING pathway in M1 macrophages. (A-B) Effects of SLC25A33 -targeting siRNA (A) and PLP (B) on relative mtDNA levels in LPS/IFN-γ-treated PMs. (C-D) Representative immunofluorescence images using 6- O -Propynyl-dG (PdG) and click chemistry (left panel) and quantification of PdG (right panel) in the LPS/IFN-γ-treated PMs following treatment with SLC25A33 -targeting siRNA (C) and PLP (D). (E) Representative immunofluorescence images of dsDNA and mitochondria (left panel) and quantification of cytosolic dsDNA foci (right panel) in the LPS/IFN-γ-treated PMs with SLC25A33 -targeting siRNA. (F) Effect of SLC25A33 -targeting siRNA on relative cytosolic mtDNA levels in LPS/IFN-γ-treated PMs, measured by qPCR. (G) Representative immunofluorescence images of dsDNA and mitochondria (left panel) and quantification of cytosolic dsDNA foci (right panel) in the LPS/IFN-γ-treated PMs with PLP. (H) Effect of PLP on the relative cytosolic mtDNA levels in LPS/IFN-γ-treated PMs, measured by qPCR. (I-J) Effects of SLC25A33 -targeting siRNA (I) and PLP (J) on phosphorylation status of STING, p65, and IRF3 in the LPS/IFN-γ-treated PMs. (K) Effect of ATF4 overexpression on protein levels of SLC25A33 and phosphorylation status of IKKα/β, IκBα, and p65 in SLC25A33 knock-downed LPS/IFN-γ-treated PMs. (L) Effect of SLC25A33 overexpression on phosphorylation of cGAS-STING pathway proteins in ATF4 knock-downed LPS/IFN-γ-treated PMs. In all experiments, PMs were treated with LPS (100 ng/mL) and IFN-γ (10 ng/mL) for 24 h, with PLP (400 μM, 24 h), MitoTracker (250 nM, 30 minutes), PdG (10 μM, 1 h), and siRNAs against each gene (40 pM, 48 h). All experimental data were verified in at least three independent experiments. Data are presented as the mean ± SEM. Scale bar represents 10 μm. *p < 0.05, **p < 0.01, and ***p < 0.001. dsDNA: double-stranded DNA; PLP: pyridoxal 5'-phosphate; IRF3: interferon resgulatory factor 3; STING: stimulator of interferon genes.

Article Snippet: Cells were transfected with scramble siRNA, siSLC25A33, siMyD88, siPI3K, simTOR, siTAK1, siTBK1, siATF4 (Bioneer), SLC25A33 ORF Clone (Origene) or ATF4 ORF Clone (GeneCopoeia) using Lipofectamine TM 3000 (Thermo Fisher Scientific).

Techniques: Inhibition, Immunofluorescence, Phospho-proteomics, Over Expression

Journal: International Journal of Biological Sciences

Article Title: SLC25A33-mediated mitochondrial DNA synthesis plays a critical role in the inflammatory response of M1 macrophages by contributing to mitochondrial ROS and VDAC oligomerization

doi: 10.7150/ijbs.96563

Figure Lengend Snippet: SLC25A33 increases mtDNA release via VDAC1 oligomerization and mtROS in M1 macrophages. (A-C) Effect of ddC (A), SLC25A33 -targeting siRNA (B), and PLP (C) on VDAC1 oligomerization, as indicated by immunoblotting positions for monomers (Mono), trimers (Tri), tetramers (Tetra), and multimers (Multi). Asterisk indicates a nonspecific band. (D) Representative image of immunofluorescence staining for dsDNA and mitochondria (left panel) and quantification of cytosolic dsDNA foci (right panel) in LPS/IFN-γ-treated PMs with VBIT-4. (E) Effect of VBIT-4 on the protein expression level of iNOS, CD80, and CD86 in the LPS/IFNγ-treated PMs. (F) Effect of VBIT-4 on the mRNA expression of iNOS and pro-inflammatory cytokines in the LPS/IFNγ-treated PMs. (G) Representative image of immunofluorescence staining for MitoSOX Red in the LPS/IFN-γ-treated PMs with VBIT4. (H) Effect of MitoTEMPO on VDAC1 oligomerization. Asterisk indicates a nonspecific band. (I) Representative image of immunofluorescence staining for dsDNA and mitochondria (left panel) and quantification of cytosolic dsDNA foci (right panel) in the LPS/IFN-γ-treated PMs with MitoTEMPO. In all experiments, PMs were treated with LPS (100 ng/mL) and IFN-γ (10 ng/mL) for 24 h, with ddC (40 μM, 5 days), PLP (400 μM, 24 h), VBIT-4 (10 μM, 48 h), MitoTEMPO (500 μM, 24 h), MitoSOX Red (5 μM, 15 minutes), and SLC25A33 -targeting siRNAs (40 pM, 48 h). All experimental data were verified in at least three independent experiments. Data are presented as the mean ± SEM. Scale bar represents 10 μm. *p < 0.05, **p < 0.01, and ***p < 0.001. ddC: 2′,3′-dideoxycytidine; dsDNA: double-stranded DNA; EGS: ethylene glycol-bis(succinimidyl succinate); VDAC1: voltage-dependent anion channel 1; PLP: pyridoxal 5'-phosphate.

Article Snippet: Cells were transfected with scramble siRNA, siSLC25A33, siMyD88, siPI3K, simTOR, siTAK1, siTBK1, siATF4 (Bioneer), SLC25A33 ORF Clone (Origene) or ATF4 ORF Clone (GeneCopoeia) using Lipofectamine TM 3000 (Thermo Fisher Scientific).

Techniques: Western Blot, Immunofluorescence, Staining, Expressing

Journal: International Journal of Biological Sciences

Article Title: SLC25A33-mediated mitochondrial DNA synthesis plays a critical role in the inflammatory response of M1 macrophages by contributing to mitochondrial ROS and VDAC oligomerization

doi: 10.7150/ijbs.96563

Figure Lengend Snippet: Reconstitution with SLC25A33 -silenced BMDMs mitigates inflammation in clodronate liposome (CL) treated mice during LPS-induced sepsis. (A) Schematic representation of the procedure for the isolation of BMDMs from C57BL/6 male mice, transduction of SLC25A33 -targeting shRNA, and their reconstitution into macrophage-depleted mice treated with 200 μL CL. (B) SLC25A33 protein levels in SLC25A33 -silenced BMDMs before their introduction into CL-treated mice. (C) Quantification of macrophages in mice 2 days post CL treatment. (D-E) Effect of SLC25A33 -silenced BMDMs reconstitution on the survival rate (n=8) (D) and pro-inflammatory cytokines (E) in LPS (30 mg/kg) induced septic mice. (F) Representative images of H&E staining (upper panel) (Scale bar represents 100 μm), immunohistochemical staining with anti-CD86 antibodies (lower panel) (Scale bar represents 20 μm), and the CD86 expression scores (right panel) determined using ImageJ in lung tissue sections from LPS-induced septic mice reconstituted with or without the SLC25A33 -silenced BMDMs. All experimental data were verified in at least three independent experiments for each animal group. Data represent the mean ± SEM from independent mice. *p < 0.05, **p < 0.01, and ***p < 0.001. BMDMs: bone marrow-derived macrophages.

Article Snippet: Cells were transfected with scramble siRNA, siSLC25A33, siMyD88, siPI3K, simTOR, siTAK1, siTBK1, siATF4 (Bioneer), SLC25A33 ORF Clone (Origene) or ATF4 ORF Clone (GeneCopoeia) using Lipofectamine TM 3000 (Thermo Fisher Scientific).

Techniques: Isolation, Transduction, shRNA, Staining, Immunohistochemical staining, Expressing, Derivative Assay

Journal: International Journal of Biological Sciences

Article Title: SLC25A33-mediated mitochondrial DNA synthesis plays a critical role in the inflammatory response of M1 macrophages by contributing to mitochondrial ROS and VDAC oligomerization

doi: 10.7150/ijbs.96563

Figure Lengend Snippet: PLP alleviates systemic inflammatory responses in LPS- and CLP-induced septic mice. (A) Effect of PLP (20 mg/kg) on the survival rate of septic mice induced by LPS (30 mg/kg, n=6). (B-C) Relative mRNA levels of pro-inflammatory cytokines in PMs isolated LPS-induced (B) and their relative cytokines levels in the serum of LPS-induced (C) septic mice treated with PLP. (D) Representative images of H&E staining (upper panel) and immunohistochemical staining (lower panel) with antibodies against CD86 in lung tissue in the LPS-induced septic mice (Scale bar represents 100 μm). (E) Representative immunofluorescence images for CD80 (red) and SLC25A33 (green) in lung tissue from the LPS-induced septic mice treated with PLP (Scale bar represents 30 μm). (F) Effect of PLP on the survival rate of septic mice induced by CLP (n=7). (G-H) Relative mRNA levels of pro-inflammatory cytokines in the colon of CLP-induced (G) and their relative protein levels in the serum of CLP-induced (H) septic mice treated with PLP. (I) Representative images of H&E staining (upper panel) (Scale bar represents 100 μm) and immunohistochemical staining (lower panel) (Scale bar represents 40 μm) with antibodies against CD86 in colon tissue in CLP-induced septic mice. (J) Representative immunofluorescence images for CD80 (red) and SLC25A33 (green) in colon tissue from CLP-induced septic mice treated with PLP (Scale bar represents 40 μm). All experimental data were verified in at least three independent experiments for each animal group. Data are presented as the mean ± SEM. *p < 0.05, **p < 0.01, and ***p < 0.001. CLP: cecal ligation and puncture; PLP: pyridoxal 5'-phosphate.

Article Snippet: Cells were transfected with scramble siRNA, siSLC25A33, siMyD88, siPI3K, simTOR, siTAK1, siTBK1, siATF4 (Bioneer), SLC25A33 ORF Clone (Origene) or ATF4 ORF Clone (GeneCopoeia) using Lipofectamine TM 3000 (Thermo Fisher Scientific).

Techniques: Isolation, Staining, Immunohistochemical staining, Immunofluorescence, Ligation

Journal: Scientific Reports

Article Title: SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGFβ-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells

doi: 10.1038/srep43164

Figure Lengend Snippet: Following lentiviral transduction with either SMAD2-shRNA, SMAD3-shRNA, SMAD4-shRNA or non-mammalian shRNA control (Ctrl-shRNA), human fetal BMSCs were cultured in chondrogenic medium for 1 day. Short hairpin RNA-mediated knockdown was evaluated by determining gene (RT-qPCR; ( a , c , e ) and protein (Western blot; ( b , d , f ) expression of SMAD2 ( a , b ), SMAD3 ( c , d ) and SMAD4 ( e , f ). Protein levels were normalized to GAPDH and expressed as relative to Ctrl-shRNA. Gene expression was normalized to the mean C T value of RPS27a and TBP . Data are expressed as % relative to normalized mRNA levels in Ctrl-shRNA. Bars represent mean + S.D. of triplicate pellets from 2 experiments. **p < 0.01; ***p < 0.001 compared to Ctrl-shRNA.

Article Snippet: Human fetal BMSCs (#SCC7500, Lot#6890, ScienCell Research Laboratories, Carlsbad, CA, USA) were expanded in Mesenchymal Stem Cell Growth Medium (MSCGMTM; Lonza, Basel, Switzerland) supplemented with 1% Penicillin-Streptomycin-Glutamine (Gibco, Carlsbad, CA, USA).

Techniques: Transduction, shRNA, Control, Cell Culture, Knockdown, Quantitative RT-PCR, Western Blot, Expressing, Gene Expression

Journal: Scientific Reports

Article Title: SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGFβ-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells

doi: 10.1038/srep43164

Figure Lengend Snippet: Human fetal BMSCs were transduced with adenovirus overexpressing SMAD2 (ad-SMAD2), ad-SMAD3, ad-SMAD4 or ad-LacZ as control, followed by culturing in chondrogenic medium for 1 day. To confirm overexpression, gene (RT-qPCR; ( a , c , e ) and protein (Western blot; ( b , d , f ) expression of SMAD2 ( a , b ), SMAD3 ( c , d ) and SMAD4 ( e , f ) were measured. Protein levels were normalized to GAPDH and expressed as relative to ad-LacZ. Gene expression data are presented as −ΔC T compared to the mean C T value of RPS27a and TBP . Bars represent mean + S.D. of triplicate pellets from 2 experiments. ***p < 0.001 compared to ad-LacZ.

Article Snippet: Human fetal BMSCs (#SCC7500, Lot#6890, ScienCell Research Laboratories, Carlsbad, CA, USA) were expanded in Mesenchymal Stem Cell Growth Medium (MSCGMTM; Lonza, Basel, Switzerland) supplemented with 1% Penicillin-Streptomycin-Glutamine (Gibco, Carlsbad, CA, USA).

Techniques: Transduction, Control, Over Expression, Quantitative RT-PCR, Western Blot, Expressing, Gene Expression

Journal: Scientific Reports

Article Title: SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGFβ-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells

doi: 10.1038/srep43164

Figure Lengend Snippet: Human fetal BMSCs were transduced (lentivirus) either with SMAD2-shRNA, SMAD3-shRNA, SMAD4-shRNA or Ctrl-shRNA and subsequently pellet-cultured for 1, 7 or 14 days in chondrogenic medium with TGFβ. The effect of shRNA-mediated knockdown of SMAD2, SMAD3 and SMAD4 on chondrogenesis was determined by RT-qPCR analysis of the chondrogenesis-specific genes; ACAN ( a ), COL2A1 ( b ) and SOX9 ( c ), deposition of glycosaminoglycans (GAGs) per pellet ( d ) and per μg DNA ( e ), histological examination of GAGs by Safranin O (Saf.O) staining and collagen type II by immunohistochemistry in sections of 14 days-cultured pellets ( f ), and macroscopic evaluation of 14 days-cultured pellets ( g ). In ( f ) representative images of consecutive pellet sections per condition are shown and the scale bar represents 500 μm. Gene expression data are presented as −ΔC T compared to the mean C T value of RPS27a and TBP . Bars represent mean + S.D. of triplicate pellets from 2 experiments. *p < 0.05; **p < 0.01; ***p < 0.001 compared to Ctrl-shRNA.

Article Snippet: Human fetal BMSCs (#SCC7500, Lot#6890, ScienCell Research Laboratories, Carlsbad, CA, USA) were expanded in Mesenchymal Stem Cell Growth Medium (MSCGMTM; Lonza, Basel, Switzerland) supplemented with 1% Penicillin-Streptomycin-Glutamine (Gibco, Carlsbad, CA, USA).

Techniques: shRNA, Cell Culture, Knockdown, Quantitative RT-PCR, Staining, Immunohistochemistry, Gene Expression

Journal: Scientific Reports

Article Title: SMAD3 and SMAD4 have a more dominant role than SMAD2 in TGFβ-induced chondrogenic differentiation of bone marrow-derived mesenchymal stem cells

doi: 10.1038/srep43164

Figure Lengend Snippet: Human fetal BMSCs transduced either with adenoviral SMAD2 (ad-SMAD2), ad-SMAD3, ad-SMAD4 or ad-LacZ as control were pellet-cultured for 1, 7 or 14 days in chondrogenic medium with TGFβ. The effect of SMAD overexpression on chondrogenesis was determined by RT-qPCR analysis of the chondrogenesis-specific genes; ACAN ( a ), COL2A1 ( b ) and SOX9 ( c ), deposition of glycosaminoglycans (GAGs) per pellet ( d ) and per μg DNA ( e ), histological examination of GAGs by Safranin O (Saf.O) staining and collagen type II by immunohistochemistry in sections of 14 days-cultured pellets ( f ), and macroscopic evaluation of 14 days-cultured pellets ( g ). In ( f ) representative images of consecutive pellet sections per condition are shown and the scale bar represents 500 μm. Gene expression data are presented as −ΔC T compared to the mean C T value of RPS27a and TBP . Bars represent mean + S.D. of triplicate pellets from 2 experiments. *p < 0.05; **p < 0.01; ***p < 0.001 compared to ad-LacZ.

Article Snippet: Human fetal BMSCs (#SCC7500, Lot#6890, ScienCell Research Laboratories, Carlsbad, CA, USA) were expanded in Mesenchymal Stem Cell Growth Medium (MSCGMTM; Lonza, Basel, Switzerland) supplemented with 1% Penicillin-Streptomycin-Glutamine (Gibco, Carlsbad, CA, USA).

Techniques: Control, Cell Culture, Over Expression, Quantitative RT-PCR, Staining, Immunohistochemistry, Gene Expression

Journal: Tissue Engineering. Part A

Article Title: Perfusion Enhances Hypertrophic Chondrocyte Matrix Deposition, But Not the Bone Formation

doi: 10.1089/ten.tea.2017.0356

Figure Lengend Snippet: Creation of hypertrophic chondrocyte constructs through application of variable flow rates. Bone cores were extracted from juvenile bovine wrists, decellularized, and sized to 4 mm diameter × 5 mm height. Human BMSCs were expanded and seeded into the decellularized bone cores (2.25 × 106 cells/construct). Seeded constructs were then placed into previously validated perfusion bioreactors21,31 and differentiated for 2 weeks in the chondrogenic medium, then matured to hypertrophic chondrocytes over an additional 3 weeks. Flow rates were determined by the tubing diameter, and flow rates approximating 0, 100, 400, and 1200 μm/s were chosen for experimentation. At the end of 5 weeks of culture, constructs were implanted into a 5 mm, critical-size defect made in the right femur of RNU nude male rats. Construct regeneration occurred for 8 weeks in vivo before harvest and construct evaluation. BMSC, bone marrow-derived stem cell; RNU, Rowett Nude; SD, standard deviation. Color images available online at www.liebertpub.com/tea

Article Snippet: Cell expansion and seeding Human BMSCs were obtained from Lonza (Basel, CH) and expanded until passage 5 in an expansion medium consisting of high glucose medium with L-glutamine, 10% fetal bovine serum, 1% penicillin/streptomycin (P/S), and 1 ng/mL basic fibroblast growth factor.

Techniques: Construct, In Vivo, Derivative Assay, Standard Deviation

Journal: Journal of Nanobiotechnology

Article Title: Exosomes as therapeutic and drug delivery vehicle for neurodegenerative diseases

doi: 10.1186/s12951-024-02681-4

Figure Lengend Snippet: Potential therapeutic effects and mechanisms of action of exosomes and their cargoes in SCI treatment

Article Snippet: Human BMSCs (lonza)-derived exosomes , PTEN siRNA , - , Adult female SD rats , ↑functional recovery in complete SCI rats/Axonal growth and neovascularization/ Reduced microgliosis , [ ] .

Techniques: Activity Assay, Isolation

Journal: Journal of Nanobiotechnology

Article Title: Exosomes as therapeutic and drug delivery vehicle for neurodegenerative diseases

doi: 10.1186/s12951-024-02681-4

Figure Lengend Snippet: Potential therapeutic effects and mechanisms of action of exosomes and their cargoes in MS treatment

Article Snippet: Human BMSCs (lonza)-derived exosomes , PTEN siRNA , - , Adult female SD rats , ↑functional recovery in complete SCI rats/Axonal growth and neovascularization/ Reduced microgliosis , [ ] .

Techniques:

Journal: Journal of Nanobiotechnology

Article Title: Exosomes as therapeutic and drug delivery vehicle for neurodegenerative diseases

doi: 10.1186/s12951-024-02681-4

Figure Lengend Snippet: Potential therapeutic effects and mechanisms of action of exosomes and their cargoes in AD treatment

Article Snippet: Human BMSCs (lonza)-derived exosomes , PTEN siRNA , - , Adult female SD rats , ↑functional recovery in complete SCI rats/Axonal growth and neovascularization/ Reduced microgliosis , [ ] .

Techniques: Clinical Proteomics, Transgenic Assay, Cell Culture

Journal: Journal of Nanobiotechnology

Article Title: Exosomes as therapeutic and drug delivery vehicle for neurodegenerative diseases

doi: 10.1186/s12951-024-02681-4

Figure Lengend Snippet: Potential therapeutic effects and mechanisms of action of exosomes and their cargoes in PD treatment

Article Snippet: Human BMSCs (lonza)-derived exosomes , PTEN siRNA , - , Adult female SD rats , ↑functional recovery in complete SCI rats/Axonal growth and neovascularization/ Reduced microgliosis , [ ] .

Techniques: Derivative Assay, Transgenic Assay, Expressing, Transfection, shRNA