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PromoCell primary human bone marrow derived mscs

Fig. 1. Screening of cytokines and small molecules for LSEC differentiation. (A) The schematic representation of BM-MSC differentiation. (B) qPCR analysis of the expression levels of LYVE1 and CD36 in BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of <t>BM-MSCs.</t> All data are presented as mean ± SD (n ¼ 3). Signiﬁcant differences were evaluated using a one-way ANOVA followed by Dunnett's post-hoc test (*p < 0.05, ***p < 0.001 compared with BFTc [BMP4, FGF8b, TGF-b inhibitor, and cAMP]). Abbreviations: B; BMP4, F; FGF8b, T; TGF-b signal inhibitor, c; cAMP.

Primary Human Bone Marrow Derived Mscs, supplied by PromoCell, used in various techniques. Bioz Stars score: 96/100, based on 200 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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1) Product Images from "Generation of functional liver sinusoidal endothelial-like cells from human bone marrow-derived mesenchymal stem cells."

Article Title: Generation of functional liver sinusoidal endothelial-like cells from human bone marrow-derived mesenchymal stem cells.

Journal: Regenerative therapy

doi: 10.1016/j.reth.2023.07.006

Figure Legend Snippet: Fig. 1. Screening of cytokines and small molecules for LSEC differentiation. (A) The schematic representation of BM-MSC differentiation. (B) qPCR analysis of the expression levels of LYVE1 and CD36 in BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of BM-MSCs. All data are presented as mean ± SD (n ¼ 3). Signiﬁcant differences were evaluated using a one-way ANOVA followed by Dunnett's post-hoc test (*p < 0.05, ***p < 0.001 compared with BFTc [BMP4, FGF8b, TGF-b inhibitor, and cAMP]). Abbreviations: B; BMP4, F; FGF8b, T; TGF-b signal inhibitor, c; cAMP.

Techniques Used: Expressing, Derivative Assay

Fig. 2. Analysis of the expression of LSEC-related markers. (A) The schematic representation of BM-MSC differentiation to LSEC-like cells. (B) Brightﬁeld images of the BM-MSCs and BM-MSC-derived cells. Scale bar ¼ 500 mm. (C) qPCR analysis of gene expression of LYVE1, CD36, CD32b, F8, PLVAP, CD31, and VEGFR2 in the BM-MSCs and BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of BM-MSCs. Data are presented as mean ± SD (n ¼ 6, sum of two independent experiments, n ¼ 3 for each experiment). (D) Immunocytochemistry analysis of LYVE1 (green) and CD36 (red) in the BM-MSCs and BM-MSC-derived cells. The nuclei were counterstained with DAPI (blue). Scale bar ¼ 50 mm. (E) The expression level of CD32 in the BM-MSCs and BM-MSC-derived cells was measured using ﬂow cytometry analysis. The values of mean ﬂuorescence intensity (MFI) of CD32 expression were normalized by the MFI of isotype control. Data are presented as mean ± SD (n ¼ 3). Signiﬁcant differences were evaluated using an unpaired two-tailed Student's t-test (***p < 0.001). (F) The percentage of LYVE1- or CD36-positive cells in the BM-MSC-derived cells was measured using ﬂow cytometry analysis.

Figure Legend Snippet: Fig. 2. Analysis of the expression of LSEC-related markers. (A) The schematic representation of BM-MSC differentiation to LSEC-like cells. (B) Brightﬁeld images of the BM-MSCs and BM-MSC-derived cells. Scale bar ¼ 500 mm. (C) qPCR analysis of gene expression of LYVE1, CD36, CD32b, F8, PLVAP, CD31, and VEGFR2 in the BM-MSCs and BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of BM-MSCs. Data are presented as mean ± SD (n ¼ 6, sum of two independent experiments, n ¼ 3 for each experiment). (D) Immunocytochemistry analysis of LYVE1 (green) and CD36 (red) in the BM-MSCs and BM-MSC-derived cells. The nuclei were counterstained with DAPI (blue). Scale bar ¼ 50 mm. (E) The expression level of CD32 in the BM-MSCs and BM-MSC-derived cells was measured using ﬂow cytometry analysis. The values of mean ﬂuorescence intensity (MFI) of CD32 expression were normalized by the MFI of isotype control. Data are presented as mean ± SD (n ¼ 3). Signiﬁcant differences were evaluated using an unpaired two-tailed Student's t-test (***p < 0.001). (F) The percentage of LYVE1- or CD36-positive cells in the BM-MSC-derived cells was measured using ﬂow cytometry analysis.

Techniques Used: Expressing, Derivative Assay, Gene Expression, Immunocytochemistry, Cytometry, Control, Two Tailed Test

Fig. 3. Evaluation of endothelial cell-related functions. (A) Tube-formation assay of the BM-MSCs, BM-MSC-derived cells, and HUVECs. Scale bar ¼ 200 mm. (B) acLDL-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated acLDL to measure their ability to take up acLDL. Scale bar ¼ 20 mm. Abbreviation: LDL, low-density lipoprotein.

Figure Legend Snippet: Fig. 3. Evaluation of endothelial cell-related functions. (A) Tube-formation assay of the BM-MSCs, BM-MSC-derived cells, and HUVECs. Scale bar ¼ 200 mm. (B) acLDL-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated acLDL to measure their ability to take up acLDL. Scale bar ¼ 20 mm. Abbreviation: LDL, low-density lipoprotein.

Techniques Used: Tube Formation Assay, Derivative Assay, Cell Culture

Fig. 4. Evaluation of LSEC-related functions. (A) HA-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing ﬂuoresceinamine- conjugated HA to measure their ability to take up HA. (B) IgG-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated IgG to measure their ability to endocytose IgG. Scale bar ¼ 20 mm. Abbreviation: HA, hyaluronic acid.

Figure Legend Snippet: Fig. 4. Evaluation of LSEC-related functions. (A) HA-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing ﬂuoresceinamine- conjugated HA to measure their ability to take up HA. (B) IgG-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated IgG to measure their ability to endocytose IgG. Scale bar ¼ 20 mm. Abbreviation: HA, hyaluronic acid.

Techniques Used: Derivative Assay, Cell Culture

Fig. 5. Evaluation of the differentiation capacity of primary human BM-MSCs into LSEC-like cells (A) The schematic representation of the differentiation of primary human BM- MSCs into LSEC-like cells. (B) qPCR analysis of gene expression of LYVE1, CD36, CD32b, and F8 in primary human BM-MSCs and primary human BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of primary human BM-MSCs. Data are presented as mean ± SD (n 8, sum of two independent experiments, n 3 for each experiment.). Signiﬁcant differences were evaluated using an unpaired two-tailed Student's t-test (*p < 0.05, ***p < 0.001). (C) The percentage of LYVE1- or CD36- positive cells in the primary human BM-MSC-derived cells was measured using ﬂow cytometry analysis. Data are presented as mean ± SD (n ¼ 3). (D) acLDL-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated acLDL to measure their ability to take up acLDL. Scale bar ¼ 20 mm. (E) HA- uptake assay. Primary human BM-MSCs and primary human BM-MSC-derived cells were cultured in a medium containing ﬂuoresceinamine-conjugated HA to measure their ability to take up HA. Scale bar ¼ 20 mm. Abbreviation: LDL, low-density lipoprotein; HA, hyaluronic acid.

Figure Legend Snippet: Fig. 5. Evaluation of the differentiation capacity of primary human BM-MSCs into LSEC-like cells (A) The schematic representation of the differentiation of primary human BM- MSCs into LSEC-like cells. (B) qPCR analysis of gene expression of LYVE1, CD36, CD32b, and F8 in primary human BM-MSCs and primary human BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of primary human BM-MSCs. Data are presented as mean ± SD (n 8, sum of two independent experiments, n 3 for each experiment.). Signiﬁcant differences were evaluated using an unpaired two-tailed Student's t-test (*p < 0.05, ***p < 0.001). (C) The percentage of LYVE1- or CD36- positive cells in the primary human BM-MSC-derived cells was measured using ﬂow cytometry analysis. Data are presented as mean ± SD (n ¼ 3). (D) acLDL-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated acLDL to measure their ability to take up acLDL. Scale bar ¼ 20 mm. (E) HA- uptake assay. Primary human BM-MSCs and primary human BM-MSC-derived cells were cultured in a medium containing ﬂuoresceinamine-conjugated HA to measure their ability to take up HA. Scale bar ¼ 20 mm. Abbreviation: LDL, low-density lipoprotein; HA, hyaluronic acid.

Techniques Used: Gene Expression, Derivative Assay, Expressing, Two Tailed Test, Cytometry, Cell Culture

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Mesenchymal stromal cells enhances stemness and chemoresistance of gastric cancer cells. A-E: The MKN45 and AGS cells were co-cultured with mesenchymal stromal cells (MSCs); The stemness was analyzed using sphere formation assay (A); The mRNA expression of SOX2, Oct4, LIN28, and CD133 was measured by qPCR (B); The levels of SOX2, Oct4, LIN28, and CD133 was detected by Western blot analysis (C); The CD44+ and CD133+ cells were identified by flow cytometry analysis (D and E); F and G: The MKN45 and AGS cells were co-cultured with MSCs and treated with oxaliplatin or 5-Fu; The cell apoptosis was determined by flow cytometry analysis (F); The cell proliferation was examined by colony formation assay (G). a P < 0.01; MSC: Mesenchymal stromal cell.

Journal: World Journal of Gastrointestinal Oncology

Article Title: Mesenchymal stem cell-derived lncRNAs NKILA contributes to stemness and chemoresistance by fatty acid oxidation in gastric cancer via miR-485-5p/STAT3

doi: 10.4251/wjgo.v17.i8.105006

Figure Lengend Snippet: Mesenchymal stromal cells enhances stemness and chemoresistance of gastric cancer cells. A-E: The MKN45 and AGS cells were co-cultured with mesenchymal stromal cells (MSCs); The stemness was analyzed using sphere formation assay (A); The mRNA expression of SOX2, Oct4, LIN28, and CD133 was measured by qPCR (B); The levels of SOX2, Oct4, LIN28, and CD133 was detected by Western blot analysis (C); The CD44+ and CD133+ cells were identified by flow cytometry analysis (D and E); F and G: The MKN45 and AGS cells were co-cultured with MSCs and treated with oxaliplatin or 5-Fu; The cell apoptosis was determined by flow cytometry analysis (F); The cell proliferation was examined by colony formation assay (G). a P < 0.01; MSC: Mesenchymal stromal cell.

Article Snippet: Primary human bone marrow-derived MSCs (HUXMA-01001, Procell, China) were cultured in α-MEM medium (Hyclone, United States) supplemented with 10% FBS and 1% penicillin/streptomycin.

Techniques: Cell Culture, Tube Formation Assay, Expressing, Western Blot, Flow Cytometry, Colony Assay

Effect of different donor-derived human peripheral blood mononuclear cells (PBMCs) on the in vitro immunomodulatory response of bone marrow (BM)- mesenchymal stem cells (MSCs). (a) Schematic of the protocol for lymphocyte proliferation assays with mitogen as an immunomodulatory potency assay for MSCs. (b) Proliferation of 10 donor-derived PBMCs upon phytohemagglutinin (PHA; 0.2, 0.5, and 1.0 μg/mL) stimulation was assessed through BrdU uptake. BrdU uptake was indicated by luminescence signal (counts per second, CPS). Two-way ANOVA showed that PBMCs proliferation was significantly affected by the stimulating PHA concentration, the presence or absence of MSCs, and their interaction (p < 0.0001). All values indicate the mean ± standard deviation of mean of three biological replicates (n = 3) (∗P < 0.05, ∗∗P < 0.01, ∗∗∗∗P < 0.0001, two-way analysis of variance, followed by Bonferroni's multiple comparison test). (c) Effect of PBMCs derived from 10 donors on the in vitro immunomodulatory response of BM-MSCs. The rate of suppression by BM-MSCs on PBMC proliferation stimulated with three PHA concentrations (0.2, 0.5, and 1.0 μg/mL) is shown as the immunosuppression rate; CPS(PBMC + MSC)/CPS(PBMC).

Journal: Regenerative Therapy

Article Title: Increasing robustness of in vitro assay for immnosuppressive effect of mesenchymal stromal/stem cells: The role of inflammatory cytokine production by peripheral blood mononuclear cells

doi: 10.1016/j.reth.2024.12.016

Figure Lengend Snippet: Effect of different donor-derived human peripheral blood mononuclear cells (PBMCs) on the in vitro immunomodulatory response of bone marrow (BM)- mesenchymal stem cells (MSCs). (a) Schematic of the protocol for lymphocyte proliferation assays with mitogen as an immunomodulatory potency assay for MSCs. (b) Proliferation of 10 donor-derived PBMCs upon phytohemagglutinin (PHA; 0.2, 0.5, and 1.0 μg/mL) stimulation was assessed through BrdU uptake. BrdU uptake was indicated by luminescence signal (counts per second, CPS). Two-way ANOVA showed that PBMCs proliferation was significantly affected by the stimulating PHA concentration, the presence or absence of MSCs, and their interaction (p < 0.0001). All values indicate the mean ± standard deviation of mean of three biological replicates (n = 3) (∗P < 0.05, ∗∗P < 0.01, ∗∗∗∗P < 0.0001, two-way analysis of variance, followed by Bonferroni's multiple comparison test). (c) Effect of PBMCs derived from 10 donors on the in vitro immunomodulatory response of BM-MSCs. The rate of suppression by BM-MSCs on PBMC proliferation stimulated with three PHA concentrations (0.2, 0.5, and 1.0 μg/mL) is shown as the immunosuppression rate; CPS(PBMC + MSC)/CPS(PBMC).

Article Snippet: Primary human bone marrow-derived MSCs (BM-MSC) were purchased from Lonza (Walkersville, MD, USA) (PT-2501).

Techniques: Derivative Assay, In Vitro, Potency Assay, Concentration Assay, Standard Deviation, Comparison

Relationship between the production of cytokines by PHA-stimulated PBMCs and the suppression rate of PBMC proliferation by MSCs in the immunomodulatory potency assay. Spearman correlation analysis between the concentration of each cytokine in the culture supernatant when PBMCs were stimulated with 1.0 μg/mL PHA and the rate of proliferation inhibition by BM-MSCs in the immunomodulatory potency assay is shown with linear regression lines and their 95 % confidential intervals. r, correlation coefficient.

Journal: Regenerative Therapy

doi: 10.1016/j.reth.2024.12.016

Figure Lengend Snippet: Relationship between the production of cytokines by PHA-stimulated PBMCs and the suppression rate of PBMC proliferation by MSCs in the immunomodulatory potency assay. Spearman correlation analysis between the concentration of each cytokine in the culture supernatant when PBMCs were stimulated with 1.0 μg/mL PHA and the rate of proliferation inhibition by BM-MSCs in the immunomodulatory potency assay is shown with linear regression lines and their 95 % confidential intervals. r, correlation coefficient.

Article Snippet: Primary human bone marrow-derived MSCs (BM-MSC) were purchased from Lonza (Walkersville, MD, USA) (PT-2501).

Techniques: Potency Assay, Concentration Assay, Inhibition

Inter-test variability in the rate of proliferation inhibition of PBMCs by repeated one-way MLR assay.

Journal: Regenerative Therapy

doi: 10.1016/j.reth.2024.12.016

Figure Lengend Snippet: Inter-test variability in the rate of proliferation inhibition of PBMCs by repeated one-way MLR assay.

Article Snippet: Primary human bone marrow-derived MSCs (BM-MSC) were purchased from Lonza (Walkersville, MD, USA) (PT-2501).

Techniques: Inhibition, Mlr Assay

Figure summarizing the flow of the reversed effects of MSCs depending on the cytokine production concentration of PHA-stimulated PBMCs in the immunomodulatory potency assay. The illustration of MSC in the figure was done using BioRender: scientific image and illustration software.

Journal: Regenerative Therapy

doi: 10.1016/j.reth.2024.12.016

Figure Lengend Snippet: Figure summarizing the flow of the reversed effects of MSCs depending on the cytokine production concentration of PHA-stimulated PBMCs in the immunomodulatory potency assay. The illustration of MSC in the figure was done using BioRender: scientific image and illustration software.

Article Snippet: Primary human bone marrow-derived MSCs (BM-MSC) were purchased from Lonza (Walkersville, MD, USA) (PT-2501).

Techniques: Concentration Assay, Potency Assay, Software

Endocytosis of EVs by HMSCs (A) Representative confocal micrograph of fluorescently labeled fresh and lyophilized BMP2 EVs (green) endocytosed by naïve HMSCs at 37°C. Scale bar = 20 μm. (B) Dose-dependent endocytosis of fresh and lyophilized BMP2 EVs by HMSCs. Data points represent mean percentage fluorescence ± SD (n = 6). (C) Graph showing the inhibition of EV endocytosis after pre-treatment of the EVs with heparin to block interaction with the cell surface HSPGs. The reduction of EV endocytosis at 4°C was also measured compared to 37°C. Data represent mean percentage fluorescence with respect to control ± SD (n = 6). *: statistical significance with respect to No blocking control, #: statistical significance with respect to BMP2 EV group as measured by Student’s t-test (P< 0.05).

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Functionality of lyophilized osteoinductive EVs: a mechanistic study

doi: 10.3389/fbioe.2024.1452428

Figure Lengend Snippet: Endocytosis of EVs by HMSCs (A) Representative confocal micrograph of fluorescently labeled fresh and lyophilized BMP2 EVs (green) endocytosed by naïve HMSCs at 37°C. Scale bar = 20 μm. (B) Dose-dependent endocytosis of fresh and lyophilized BMP2 EVs by HMSCs. Data points represent mean percentage fluorescence ± SD (n = 6). (C) Graph showing the inhibition of EV endocytosis after pre-treatment of the EVs with heparin to block interaction with the cell surface HSPGs. The reduction of EV endocytosis at 4°C was also measured compared to 37°C. Data represent mean percentage fluorescence with respect to control ± SD (n = 6). *: statistical significance with respect to No blocking control, #: statistical significance with respect to BMP2 EV group as measured by Student’s t-test (P< 0.05).

Article Snippet: Primary human bone marrow derived MSCs (HMSCs) were purchased from Lonza and cultured in αMEM basal media (Gibco) supplemented with 20% fetal bovine serum (FBS, Gibco), 1% (v/v) L-Glutamine (Gibco) and 1% (v/v) antibiotic-antimycotic solution (Gibco).

Techniques: Labeling, Fluorescence, Inhibition, Blocking Assay, Control

Osteoinductive property of the lyophilized BMP2 EVs (A) Osteoinductive gene expression levels in fresh and lyophilized BMP2 EV HMSCs at day 3 and 5 with respect to control (no EV) group. Data are represented as mean fold changes ± SD (n = 4). *: statistical significance with respect to BMP2 EV group as measured by student’s t-test (P< 0.05). (B) Graph representing relative luciferase activity in HMSCs transfected with a BMP2 reporter luciferase plasmid (n = 4). Recombinant BMP2 protein (BMP2 GF) was used as positive control. Note the significant increase in luciferase activity in fresh and lyophilized BMP2 EV treated groups compared to control. *: statistical significance with respect to control, #: statistical significance with respect to BMP2 GF group as measured by Tukey’s test post ANOVA (P< 0.05). (C) Graph representing relative alkaline phosphatase (ALP) activity in HMSCs treated with osteogenic differentiation medium (OS) in the presence/absence of fresh and lyophilized BMP2 EVs at 3 and 7 days (n = 4). *: statistical significance with respect to control, #: statistical significance with respect to OS + BMP2 EV group as measured by Student’s t-test (P< 0.05). (D) Quantitative in-cell western for the presence of phosphorylated SMAD 1/5/8 and the corresponding tubulin expression in a 96 well plate assay (n = 6). The graph represents quantitation of the fluorescence from the wells using the LICOR Odyssey imager normalized to tubulin. *: statistical significance with respect to control group as measured by Tukey’s test post ANOVA (P< 0.05).

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Functionality of lyophilized osteoinductive EVs: a mechanistic study

doi: 10.3389/fbioe.2024.1452428

Figure Lengend Snippet: Osteoinductive property of the lyophilized BMP2 EVs (A) Osteoinductive gene expression levels in fresh and lyophilized BMP2 EV HMSCs at day 3 and 5 with respect to control (no EV) group. Data are represented as mean fold changes ± SD (n = 4). *: statistical significance with respect to BMP2 EV group as measured by student’s t-test (P< 0.05). (B) Graph representing relative luciferase activity in HMSCs transfected with a BMP2 reporter luciferase plasmid (n = 4). Recombinant BMP2 protein (BMP2 GF) was used as positive control. Note the significant increase in luciferase activity in fresh and lyophilized BMP2 EV treated groups compared to control. *: statistical significance with respect to control, #: statistical significance with respect to BMP2 GF group as measured by Tukey’s test post ANOVA (P< 0.05). (C) Graph representing relative alkaline phosphatase (ALP) activity in HMSCs treated with osteogenic differentiation medium (OS) in the presence/absence of fresh and lyophilized BMP2 EVs at 3 and 7 days (n = 4). *: statistical significance with respect to control, #: statistical significance with respect to OS + BMP2 EV group as measured by Student’s t-test (P< 0.05). (D) Quantitative in-cell western for the presence of phosphorylated SMAD 1/5/8 and the corresponding tubulin expression in a 96 well plate assay (n = 6). The graph represents quantitation of the fluorescence from the wells using the LICOR Odyssey imager normalized to tubulin. *: statistical significance with respect to control group as measured by Tukey’s test post ANOVA (P< 0.05).

Techniques: Gene Expression, Control, Luciferase, Activity Assay, Transfection, Plasmid Preparation, Recombinant, Positive Control, In-Cell ELISA, Expressing, Quantitation Assay, Fluorescence

Binding of lyophilized EVs to ECM proteins (A) Representative confocal image showing the binding of fluorescently labeled lyophilized EVs (green) to the decellularized ECM of HMSCs. Counterstain was performed for type I collagen (red, scale bar = 5 μm). (B) Dose-dependent binding of fluorescently labeled lyophilized EVs to type I collagen coated assay plates (data points represent mean ± SD, n = 6). (C) Representative confocal image showing the binding of fluorescently labeled lyophilized EVs (green) to the decellularized ECM of HMSCs. Counterstain was performed for fibronectin (red, scale bar = 20 μm). (D) Dose-dependent binding of fluorescently labeled lyophilized EVs to fibronectin coated assay plates (data points represent mean ± SD, n = 6).

Journal: Frontiers in Bioengineering and Biotechnology

Article Title: Functionality of lyophilized osteoinductive EVs: a mechanistic study

doi: 10.3389/fbioe.2024.1452428

Figure Lengend Snippet: Binding of lyophilized EVs to ECM proteins (A) Representative confocal image showing the binding of fluorescently labeled lyophilized EVs (green) to the decellularized ECM of HMSCs. Counterstain was performed for type I collagen (red, scale bar = 5 μm). (B) Dose-dependent binding of fluorescently labeled lyophilized EVs to type I collagen coated assay plates (data points represent mean ± SD, n = 6). (C) Representative confocal image showing the binding of fluorescently labeled lyophilized EVs (green) to the decellularized ECM of HMSCs. Counterstain was performed for fibronectin (red, scale bar = 20 μm). (D) Dose-dependent binding of fluorescently labeled lyophilized EVs to fibronectin coated assay plates (data points represent mean ± SD, n = 6).

Techniques: Binding Assay, Labeling

Journal: Regenerative therapy

Article Title: Generation of functional liver sinusoidal endothelial-like cells from human bone marrow-derived mesenchymal stem cells.

doi: 10.1016/j.reth.2023.07.006

Figure Lengend Snippet: Fig. 1. Screening of cytokines and small molecules for LSEC differentiation. (A) The schematic representation of BM-MSC differentiation. (B) qPCR analysis of the expression levels of LYVE1 and CD36 in BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of BM-MSCs. All data are presented as mean ± SD (n ¼ 3). Signiﬁcant differences were evaluated using a one-way ANOVA followed by Dunnett's post-hoc test (*p < 0.05, ***p < 0.001 compared with BFTc [BMP4, FGF8b, TGF-b inhibitor, and cAMP]). Abbreviations: B; BMP4, F; FGF8b, T; TGF-b signal inhibitor, c; cAMP.

Article Snippet: Primary human bone marrow-derived MSCs (C-12974; PromoCell) were cultured in Mesenchymal Stem Cell Growth Medium 2 (C-28009; PromoCell).

Techniques: Expressing, Derivative Assay

Journal: Regenerative therapy

Article Title: Generation of functional liver sinusoidal endothelial-like cells from human bone marrow-derived mesenchymal stem cells.

doi: 10.1016/j.reth.2023.07.006

Figure Lengend Snippet: Fig. 2. Analysis of the expression of LSEC-related markers. (A) The schematic representation of BM-MSC differentiation to LSEC-like cells. (B) Brightﬁeld images of the BM-MSCs and BM-MSC-derived cells. Scale bar ¼ 500 mm. (C) qPCR analysis of gene expression of LYVE1, CD36, CD32b, F8, PLVAP, CD31, and VEGFR2 in the BM-MSCs and BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of BM-MSCs. Data are presented as mean ± SD (n ¼ 6, sum of two independent experiments, n ¼ 3 for each experiment). (D) Immunocytochemistry analysis of LYVE1 (green) and CD36 (red) in the BM-MSCs and BM-MSC-derived cells. The nuclei were counterstained with DAPI (blue). Scale bar ¼ 50 mm. (E) The expression level of CD32 in the BM-MSCs and BM-MSC-derived cells was measured using ﬂow cytometry analysis. The values of mean ﬂuorescence intensity (MFI) of CD32 expression were normalized by the MFI of isotype control. Data are presented as mean ± SD (n ¼ 3). Signiﬁcant differences were evaluated using an unpaired two-tailed Student's t-test (***p < 0.001). (F) The percentage of LYVE1- or CD36-positive cells in the BM-MSC-derived cells was measured using ﬂow cytometry analysis.

Article Snippet: Primary human bone marrow-derived MSCs (C-12974; PromoCell) were cultured in Mesenchymal Stem Cell Growth Medium 2 (C-28009; PromoCell).

Techniques: Expressing, Derivative Assay, Gene Expression, Immunocytochemistry, Cytometry, Control, Two Tailed Test

Journal: Regenerative therapy

Article Title: Generation of functional liver sinusoidal endothelial-like cells from human bone marrow-derived mesenchymal stem cells.

doi: 10.1016/j.reth.2023.07.006

Figure Lengend Snippet: Fig. 3. Evaluation of endothelial cell-related functions. (A) Tube-formation assay of the BM-MSCs, BM-MSC-derived cells, and HUVECs. Scale bar ¼ 200 mm. (B) acLDL-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated acLDL to measure their ability to take up acLDL. Scale bar ¼ 20 mm. Abbreviation: LDL, low-density lipoprotein.

Article Snippet: Primary human bone marrow-derived MSCs (C-12974; PromoCell) were cultured in Mesenchymal Stem Cell Growth Medium 2 (C-28009; PromoCell).

Techniques: Tube Formation Assay, Derivative Assay, Cell Culture

Journal: Regenerative therapy

Article Title: Generation of functional liver sinusoidal endothelial-like cells from human bone marrow-derived mesenchymal stem cells.

doi: 10.1016/j.reth.2023.07.006

Figure Lengend Snippet: Fig. 4. Evaluation of LSEC-related functions. (A) HA-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing ﬂuoresceinamine- conjugated HA to measure their ability to take up HA. (B) IgG-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated IgG to measure their ability to endocytose IgG. Scale bar ¼ 20 mm. Abbreviation: HA, hyaluronic acid.

Article Snippet: Primary human bone marrow-derived MSCs (C-12974; PromoCell) were cultured in Mesenchymal Stem Cell Growth Medium 2 (C-28009; PromoCell).

Techniques: Derivative Assay, Cell Culture

Journal: Regenerative therapy

Article Title: Generation of functional liver sinusoidal endothelial-like cells from human bone marrow-derived mesenchymal stem cells.

doi: 10.1016/j.reth.2023.07.006

Figure Lengend Snippet: Fig. 5. Evaluation of the differentiation capacity of primary human BM-MSCs into LSEC-like cells (A) The schematic representation of the differentiation of primary human BM- MSCs into LSEC-like cells. (B) qPCR analysis of gene expression of LYVE1, CD36, CD32b, and F8 in primary human BM-MSCs and primary human BM-MSC-derived cells. On the y-axis, the expression levels are shown as a relative value to those of primary human BM-MSCs. Data are presented as mean ± SD (n 8, sum of two independent experiments, n 3 for each experiment.). Signiﬁcant differences were evaluated using an unpaired two-tailed Student's t-test (*p < 0.05, ***p < 0.001). (C) The percentage of LYVE1- or CD36- positive cells in the primary human BM-MSC-derived cells was measured using ﬂow cytometry analysis. Data are presented as mean ± SD (n ¼ 3). (D) acLDL-uptake assay. BM-MSCs, BM-MSC-derived cells, and HUVECs were cultured in a medium containing Alexa Fluor 488-conjugated acLDL to measure their ability to take up acLDL. Scale bar ¼ 20 mm. (E) HA- uptake assay. Primary human BM-MSCs and primary human BM-MSC-derived cells were cultured in a medium containing ﬂuoresceinamine-conjugated HA to measure their ability to take up HA. Scale bar ¼ 20 mm. Abbreviation: LDL, low-density lipoprotein; HA, hyaluronic acid.

Article Snippet: Primary human bone marrow-derived MSCs (C-12974; PromoCell) were cultured in Mesenchymal Stem Cell Growth Medium 2 (C-28009; PromoCell).

Techniques: Gene Expression, Derivative Assay, Expressing, Two Tailed Test, Cytometry, Cell Culture

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