hadmscs procell  (Procell Inc)

Journal: Cell Transplantation

Article Title: LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1

doi: 10.1177/0963689720968090

Figure Lengend Snippet: Identification of a model that induced hAdMSCs to adipocytes differentiation. (A) Expression of adipogenic marker proteins in different stages (D0, D3, D6, D9, and D12) was detected by Western blot, and GAPDH was used as a loading control in each sample. Expression of (B) C/EBPα, (C) PPARγ2, (D) AdipoQ, (E) FABP4, and (F) LPL were quantified using Image J software. The levels of lipogenesis enzymes, such as (G) ACC and (H) FAS were detected by spectrophotometry. (I) Expression of HCG11 in different stages (D0, D3, D6, D9, and D12) was detected by RT-qPCR. Statistical significance was determined using an independent sample t -test. Values were expressed as mean ± SEM, n = 3. ** P < 0.01 and *** P < 0.001 versus D0. ACC: acetyl coenzyme A carboxylase; AdipoQ: adiponectin; C/EBPα: CCAAT-enhancer-binding protein α; FABP4: fatty acid-binding protein 4; FAS: fatty acid synthase; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; hAdMSCs: human adipose-derived mesenchymal stem cells; HCG11: human leukocyte antigen complex group 11; LPL: lipoprotein lipase; PPARγ: peroxisome proliferator-activated receptor gamma; RT-qPCR: real-time quantitative polymerase chain reaction.

Article Snippet: Human embryonic kidney 293T (HEK-293T) cells (Shanghai, China) and hAdMSCs (Procell, Wuhan, Hubei, China) were resuspended in Dulbecco’s modified Eagle medium (Thermo Fisher Scientific, Waltham, MA, USA) mixture supplemented with 10% fetal bovine serum (FBS, GIBCO, Carlsbad, CA, USA), 100 U/ml penicillin, and 100 μg/ml streptomycin (Thermo Fisher Scientific) in a humidified incubator containing 5% carbon dioxide (CO 2 ) at 37 °C. hAdMSCs at passage five were grown for 2 days, followed by differentiation into adipocyte.

Techniques: Expressing, Marker, Western Blot, Control, Software, Spectrophotometry, Quantitative RT-PCR, Binding Assay, Derivative Assay, Real-time Polymerase Chain Reaction

Journal: Cell Transplantation

Article Title: LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1

doi: 10.1177/0963689720968090

Figure Lengend Snippet: Differentiation of adipocytes overexpressing lncRNA HCG11. (A) Expression of HCG11 in different stages (D0, D6, and D12) of hAdMSCs transfected with pcDNA-HCG11 was detected by RT-qPCR. (B) Expression of adipogenic marker protein was detected by Western blot, and GAPDH was used as a loading control in each sample. The expression of (C) C/EBPα, (D) PPARγ2, (E) AdipoQ, (F) FABP4, and (G) LPL in different stages (D0, D6, and D12) of hAdMSCs transfected with pcDNA-HCG11 and control was quantified using Image J software. The levels of lipogenesis enzymes such as (H) ACC and (I) FAS were detected by spectrophotometry. Statistical significance was determined using an independent sample t -test. Values were expressed as mean ± SEM, n = 3. * P < 0.05 and ** P < 0.01 versus control; # P < 0.05 and ## P < 0.01 versus vector. ACC: acetyl coenzyme A carboxylase; AdipoQ: adiponectin; C/EBPα: CCAAT-enhancer-binding protein α; FABP4: fatty acid-binding protein 4; FAS: fatty acid synthase; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; hAdMSCs: human adipose-derived mesenchymal stem cells; HCG11: human leukocyte antigen complex group 11; LPL: lipoprotein lipase; PPARγ: peroxisome proliferator-activated receptor gamma; RT-qPCR: real-time quantitative polymerase chain reaction.

Article Snippet: Human embryonic kidney 293T (HEK-293T) cells (Shanghai, China) and hAdMSCs (Procell, Wuhan, Hubei, China) were resuspended in Dulbecco’s modified Eagle medium (Thermo Fisher Scientific, Waltham, MA, USA) mixture supplemented with 10% fetal bovine serum (FBS, GIBCO, Carlsbad, CA, USA), 100 U/ml penicillin, and 100 μg/ml streptomycin (Thermo Fisher Scientific) in a humidified incubator containing 5% carbon dioxide (CO 2 ) at 37 °C. hAdMSCs at passage five were grown for 2 days, followed by differentiation into adipocyte.

Techniques: Expressing, Transfection, Quantitative RT-PCR, Marker, Western Blot, Control, Software, Spectrophotometry, Plasmid Preparation, Binding Assay, Derivative Assay, Real-time Polymerase Chain Reaction

Journal: Cell Transplantation

Article Title: LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1

doi: 10.1177/0963689720968090

Figure Lengend Snippet: Differentiation of adipocytes interfering with sh-HCG11. (A) Expression of HCG11 in different stages (D0, D6, and D12) hAdMSCs transfected with sh-HCG11 was detected by RT-qPCR. (B) Expression of adipogenic marker protein such as (C) C/EBPα, (D) PPARγ2, (E) AdipoQ, (F) FABP4, and (G) LPL was detected by Western blot and quantified using Image J software. The levels of lipogenesis enzymes such as (H) ACC and (I) FAS were detected by spectrophotometry. Statistical significance was determined using an independent sample t -test. Values were expressed as mean ± SEM, n = 3. * P < 0.05 and ** P < 0.01 versus control; # P < 0.05 and ## P < 0.01 versus vector. ACC: acetyl coenzyme A carboxylase; AdipoQ: adiponectin; C/EBPα: CCAAT-enhancer-binding protein α; FABP4: fatty acid-binding protein 4; FAS: fatty acid synthase; hAdMSCs: human adipose-derived mesenchymal stem cells; HCG11: human leukocyte antigen complex group 11; LPL: lipoprotein lipase; PPARγ: peroxisome proliferator-activated receptor gamma; RT-qPCR: real-time quantitative polymerase chain reaction.

Article Snippet: Human embryonic kidney 293T (HEK-293T) cells (Shanghai, China) and hAdMSCs (Procell, Wuhan, Hubei, China) were resuspended in Dulbecco’s modified Eagle medium (Thermo Fisher Scientific, Waltham, MA, USA) mixture supplemented with 10% fetal bovine serum (FBS, GIBCO, Carlsbad, CA, USA), 100 U/ml penicillin, and 100 μg/ml streptomycin (Thermo Fisher Scientific) in a humidified incubator containing 5% carbon dioxide (CO 2 ) at 37 °C. hAdMSCs at passage five were grown for 2 days, followed by differentiation into adipocyte.

Techniques: Expressing, Transfection, Quantitative RT-PCR, Marker, Western Blot, Software, Spectrophotometry, Control, Plasmid Preparation, Binding Assay, Derivative Assay, Real-time Polymerase Chain Reaction

Journal: Cell Transplantation

Article Title: LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1

doi: 10.1177/0963689720968090

Figure Lengend Snippet: HCG11 directly targeted to miR-204-5p. (A) Online database StarBase showed the binding sites of HCG11 and miR-204-5p. (B) The luciferase reporter gene assay was performed in HEK-293T cells to validate the binding of HCG11 and miR-204-5p. Firefly and Renilla luciferase activities were determined. (C) HEK-293T cells were transfected with biotinylated miR-204-5p (Bio-204-5p-wt) or its mutant form (Bio-204-5p-mut), and then a biotin-based pull-down assay was performed to detect HCG11 expression and normalized to a biotinylated mimic control (Bio-NC). (D) Expression of miR-204-5p in hAdMSCs transfected with pcDNA-HCG11 or sh-HCG11 was detected by RT-qPCR. Statistical significance was determined using an independent sample t -test. Values were expressed as mean ± SEM, n = 3. ** P < 0.01 and *** P < 0.001 versus control; # P < 0.05 versus scramble. hAdMSCs: human adipose-derived mesenchymal stem cells; HCG11: human leukocyte antigen complex group 11; HEK-293T: human embryonic kidney 293T; RT-qPCR: real-time quantitative polymerase chain reaction.

Article Snippet: Human embryonic kidney 293T (HEK-293T) cells (Shanghai, China) and hAdMSCs (Procell, Wuhan, Hubei, China) were resuspended in Dulbecco’s modified Eagle medium (Thermo Fisher Scientific, Waltham, MA, USA) mixture supplemented with 10% fetal bovine serum (FBS, GIBCO, Carlsbad, CA, USA), 100 U/ml penicillin, and 100 μg/ml streptomycin (Thermo Fisher Scientific) in a humidified incubator containing 5% carbon dioxide (CO 2 ) at 37 °C. hAdMSCs at passage five were grown for 2 days, followed by differentiation into adipocyte.

Techniques: Binding Assay, Luciferase, Reporter Gene Assay, Transfection, Mutagenesis, Pull Down Assay, Expressing, Control, Quantitative RT-PCR, Derivative Assay, Real-time Polymerase Chain Reaction

Journal: Cell Transplantation

Article Title: LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1

doi: 10.1177/0963689720968090

Figure Lengend Snippet: Overexpression of miR-204-5p promoted cell proliferation and adipocytes differentiation in hAdMSCs. The hAdMSCs were transfected with NC mimic (20 nM), miR-204-5p mimic (20 nM), NC inhibitor (20 nM), and miR-204-5p inhibitor (20 nM) for 48 h. (A) Expression of miR-204-5p was detected by RT-qPCR. (B) The expression of adipogenic marker protein and inflammatory factor was detected by Western blot. Expression of (C) C/EBPα, (D) PPARγ2, (E) AdipoQ, (F) FABP4, (G) LPL, (H) IL-6, and (I) TNF-α was quantified using Image J software. The levels of lipogenesis enzymes such as ACC (J) and FAS (K) were detected by spectrophotometry. (L) Cell proliferation ability was tested by CCK-8. Statistical significance was determined using an independent sample t -test. Values were expressed as mean ± SEM, n = 3. * P < 0.05 and ** P < 0.01 versus control. ACC: acetyl coenzyme A carboxylase; AdipoQ: adiponectin; C/EBPα: CCAAT-enhancer-binding protein α; CCK: cell counting kit; FABP4: fatty acid-binding protein 4; FAS: fatty acid synthase; hAdMSCs: human adipose-derived mesenchymal stem cells; IL-6: interleukin-6; LPL: lipoprotein lipase; NC: normal control; PPARγ: peroxisome proliferator-activated receptor gamma; TNF-α: tumor necrosis factor-alpha.

Article Snippet: Human embryonic kidney 293T (HEK-293T) cells (Shanghai, China) and hAdMSCs (Procell, Wuhan, Hubei, China) were resuspended in Dulbecco’s modified Eagle medium (Thermo Fisher Scientific, Waltham, MA, USA) mixture supplemented with 10% fetal bovine serum (FBS, GIBCO, Carlsbad, CA, USA), 100 U/ml penicillin, and 100 μg/ml streptomycin (Thermo Fisher Scientific) in a humidified incubator containing 5% carbon dioxide (CO 2 ) at 37 °C. hAdMSCs at passage five were grown for 2 days, followed by differentiation into adipocyte.

Techniques: Over Expression, Transfection, Expressing, Quantitative RT-PCR, Marker, Western Blot, Software, Spectrophotometry, CCK-8 Assay, Control, Binding Assay, Cell Counting, Derivative Assay

Journal: Cell Transplantation

Article Title: LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1

doi: 10.1177/0963689720968090

Figure Lengend Snippet: MiR-204-5p directly targeted SIRT1. (A) Online database StarBase showed the sequence alignment between miR-204-5p and SIRT1. (B) The luciferase reporter gene assay was performed in HEK-293T cells to validate the binding of miR-204-5p and SIRT1. Firefly and Renilla luciferase activities were determined. (C) Expression of SIRT1 in hAdMSCs transfected with miR-204-5p mimic, inhibitor, and their control was detected by Western blotting. (D) The expression of adipogenic marker protein and inflammatory factor was detected by Western blot. Expression of (E) C/EBPα, (F) PPARγ2, (G) AdipoQ, (H) FABP4, (I) LPL, (J) IL-6, and (K) TNF-α in hAdMSCs transfected with miR-204-5p mimic, Res, and pcDNA-SIRT1. The levels of lipogenesis enzymes such as ACC (L) and FAS (M) were detected by spectrophotometry. (N) Cell proliferation ability was tested by CCK-8. Statistical significance was determined using an independent sample t -test. Values were expressed as mean ± SEM, n = 3. * P < 0.05 and ** P < 0.01 versus control. ACC: acetyl coenzyme A carboxylase; AdipoQ: adiponectin; C/EBPα: CCAAT-enhancer-binding protein α; CCK: cell counting kit; FABP4: fatty acid-binding protein 4; FAS: fatty acid synthase; hAdMSCs: human adipose-derived mesenchymal stem cells; IL-6: interleukin-6; LPL: lipoprotein lipase; PPARγ: peroxisome proliferator-activated receptor gamma; TNF-α: tumor necrosis factor alpha.

Article Snippet: Human embryonic kidney 293T (HEK-293T) cells (Shanghai, China) and hAdMSCs (Procell, Wuhan, Hubei, China) were resuspended in Dulbecco’s modified Eagle medium (Thermo Fisher Scientific, Waltham, MA, USA) mixture supplemented with 10% fetal bovine serum (FBS, GIBCO, Carlsbad, CA, USA), 100 U/ml penicillin, and 100 μg/ml streptomycin (Thermo Fisher Scientific) in a humidified incubator containing 5% carbon dioxide (CO 2 ) at 37 °C. hAdMSCs at passage five were grown for 2 days, followed by differentiation into adipocyte.

Techniques: Sequencing, Luciferase, Reporter Gene Assay, Binding Assay, Expressing, Transfection, Control, Western Blot, Marker, Spectrophotometry, CCK-8 Assay, Cell Counting, Derivative Assay

Journal: Cell Transplantation

Article Title: LncRNA HCG11 Inhibits Adipocyte Differentiation in Human Adipose-Derived Mesenchymal Stem Cells by Sponging miR-204-5p to Upregulate SIRT1

doi: 10.1177/0963689720968090

Figure Lengend Snippet: Overexpression of miR-204-5p reversed the effect of HCG11 on hAdMSCs. (A) The expression of adipogenic marker protein and inflammatory factor was detected by Western blot. Expression of (B) C/EBPα, (C) PPARγ2, (D) AdipoQ, (E) FABP4, (F) LPL, (G) IL-6, and (H) TNF-α in hAdMSCs transfected with miR-204-5p mimic, pcDNA-HCG11, and their control was quantified using Image J software. The levels of lipogenesis enzymes such as (I) ACC and (J) FAS were detected by spectrophotometry. (K) Cell proliferation ability was tested by CCK-8. Statistical significance was determined using an independent sample t -test. Values were expressed as mean ± SEM, n = 3. * P < 0.05 and ** P < 0.01 versus control. ACC: acetyl coenzyme A carboxylase; AdipoQ: adiponectin; C/EBPα: CCAAT-enhancer-binding protein α; CCK: cell counting kit; FABP4: fatty acid-binding protein 4; FAS: fatty acid synthase; hAdMSCs: human adipose-derived mesenchymal stem cells; HCG11: human leukocyte antigen complex group 11; IL-6: interleukin-6; LPL: lipoprotein lipase; PPARγ: peroxisome proliferator-activated receptor gamma; TNF-α: tumor necrosis factor alpha.

Article Snippet: Human embryonic kidney 293T (HEK-293T) cells (Shanghai, China) and hAdMSCs (Procell, Wuhan, Hubei, China) were resuspended in Dulbecco’s modified Eagle medium (Thermo Fisher Scientific, Waltham, MA, USA) mixture supplemented with 10% fetal bovine serum (FBS, GIBCO, Carlsbad, CA, USA), 100 U/ml penicillin, and 100 μg/ml streptomycin (Thermo Fisher Scientific) in a humidified incubator containing 5% carbon dioxide (CO 2 ) at 37 °C. hAdMSCs at passage five were grown for 2 days, followed by differentiation into adipocyte.

Techniques: Over Expression, Expressing, Marker, Western Blot, Transfection, Control, Software, Spectrophotometry, CCK-8 Assay, Binding Assay, Cell Counting, Derivative Assay