Journal: Journal of Translational Medicine

Article Title: CCN5 negatively regulates TGF-β-induced endometriosis associated fibrosis through Wnt/β-catenin signaling via Smad3-dependent mechanism

doi: 10.1186/s12967-025-06804-9

Figure Lengend Snippet: Expression of CCN5 is significantly increased in endometriosis. A The mRNA expression of CCN5 in ectopic lesions of patients with endometriosis and control eutopic endometrial tissues measured by RT-qPCR (n = 15, **** p < 0.0001). B Protein expression of CCN5 in ectopic lesions of patients with endometriosis and control eutopic endometrial tissues determined by western blotting. C IHC assay of CCN5 protein expression in control eutopic endometrial tissues and ectopic lesions of patients with endometriosis. Scale bar: 100 μm; 50 μm. D Immunofluorescent staining assay of CCN5 protein expression in control eutopic endometrial tissues and ectopic lesions of patients with endometriosis. Scale bar: 50 μm; 20 μm. Data are presented as mean ± SD. Statistical analysis was performed using an unpaired student’s t-test, **** p < 0.0001

Article Snippet: The antibody for CCN5 (sc-514070) was purchased from Santa Cruz, and p-β-catenin (80,067–1-RR), β-actin (81,115–1-RR), LaminB1 (12,987–1-AP), c-Myc (67,447–1-Ig) and Cyclin D1 (60,186–1-Ig) were purchased from Proteintech, whilst α-SMA (#19,245), Collagen I (#72,026), Smad3 (#9523), p-Smad3 (#9520), β-catenin (#8480) were purchased from Cell Signaling Technology.

Techniques: Expressing, Control, Quantitative RT-PCR, Western Blot, Staining

Journal: Journal of Translational Medicine

Article Title: CCN5 negatively regulates TGF-β-induced endometriosis associated fibrosis through Wnt/β-catenin signaling via Smad3-dependent mechanism

doi: 10.1186/s12967-025-06804-9

Figure Lengend Snippet: Decreased CCN5 promotes TGF- β -induced proliferation and pro-fibrotic markers expression of primary HESCs in vitro. A The proliferation ability of shCCN5-HESCs and shNC-HESCs treated with TGF-β or not by CCK-8 assay. Data are presented as mean ± SD. Statistical analysis was conducted using one-way ANOVA for the optical density (OD) values at 72 h. * p < 0.05. B EdU assay was performed to reveal the effects of CCN5 knockdown on the proliferation of CCN5-knockdown HESCs and WT-HESCs treated with TGF-β or without TGF-β. Scale bar, 100 μm. C The mRNA expression levels of CCN5, COL1A1 and α-SMA in shCCN5-HESCs and shNC-HESCs treated with TGF-β or without TGF-β. Data are presented as the mean ± SEM. Statistical analysis was performed using one-way ANOVA. * p < 0.05, ** p < 0.01. D Expression of CCN5, α-SMA, Collagen Ι in shCCN5-HESCs and shNC-HESCs treated with TGF-β or without TGF-β. E Immunofluorescence for α-SMA in shCCN5-HESCs and shNC-HESCs treated with TGF-β or without TGF-β. Scale bar, 25 μm (red α-SMA, blue DAPI)

Article Snippet: The antibody for CCN5 (sc-514070) was purchased from Santa Cruz, and p-β-catenin (80,067–1-RR), β-actin (81,115–1-RR), LaminB1 (12,987–1-AP), c-Myc (67,447–1-Ig) and Cyclin D1 (60,186–1-Ig) were purchased from Proteintech, whilst α-SMA (#19,245), Collagen I (#72,026), Smad3 (#9523), p-Smad3 (#9520), β-catenin (#8480) were purchased from Cell Signaling Technology.

Techniques: Expressing, In Vitro, CCK-8 Assay, EdU Assay, Knockdown, Immunofluorescence

Journal: Journal of Translational Medicine

Article Title: CCN5 negatively regulates TGF-β-induced endometriosis associated fibrosis through Wnt/β-catenin signaling via Smad3-dependent mechanism

doi: 10.1186/s12967-025-06804-9

Figure Lengend Snippet: Overexpression of CCN5 attenuates TGF-β-induced proliferation and pro-fibrotic markers expression of primary HESCs in vitro. A The proliferation ability of CCN5-overexpression HESCs (CCN5) and control-HESCs (Vector) treated with TGF-β or without TGF-β by CCK-8 assay. Data are presented as mean ± SD. Statistical analysis was conducted using one-way ANOVA for the optical density values at 72 h. * p < 0.05. B EdU assay was performed to reveal the effects of CCN5 knockdown on the proliferation of CCN5-overexpression HESCs and control-HESCs treated with TGF-β or without TGF-β. Scale bar, 100 μm. C The mRNA expression levels of CCN5 , COL1A1 and α-SMA in CCN5-overexpression HESCs and control-HESCs treated with TGF-β or without TGF-β. Data are presented as the mean ± SEM. Statistical analysis was performed using one-way ANOVA for each target gene. * p < 0.05, ** p < 0.01. D Expression of CCN5, α-SMA, Collagen Ι in CCN5-overexpression HESCs and control-HESCs treated with TGF-β or without TGF-β. E. Immunofluorescence for α-SMA in CCN5-overexpression HESCs and control-HESCs treated with TGF-β or without TGF-β. Scale bar, 25 μm (red α-SMA, blue DAPI)

Article Snippet: The antibody for CCN5 (sc-514070) was purchased from Santa Cruz, and p-β-catenin (80,067–1-RR), β-actin (81,115–1-RR), LaminB1 (12,987–1-AP), c-Myc (67,447–1-Ig) and Cyclin D1 (60,186–1-Ig) were purchased from Proteintech, whilst α-SMA (#19,245), Collagen I (#72,026), Smad3 (#9523), p-Smad3 (#9520), β-catenin (#8480) were purchased from Cell Signaling Technology.

Techniques: Over Expression, Expressing, In Vitro, Control, Plasmid Preparation, CCK-8 Assay, EdU Assay, Knockdown, Immunofluorescence

Journal: Journal of Translational Medicine

Article Title: CCN5 negatively regulates TGF-β-induced endometriosis associated fibrosis through Wnt/β-catenin signaling via Smad3-dependent mechanism

doi: 10.1186/s12967-025-06804-9

Figure Lengend Snippet: CCN5 regulates activation of TGF-β/Smad and Wnt/β-catenin signaling pathways. A , B Luciferase reporter assays in primary HESCs transfected with the indicated plasmids and treated with the vehicle, TGF-β or prifenidone. FLuc/RLuc activity was determined as mean ± SEM. Statistical analysis was performed using one-way ANOVA. * p < 0.05, ** p < 0.01. C, D. The mRNA expression of downstream target genes of TGF-β/Smad and Wnt/β-catenin signaling pathways in CCN5-overexpression HESCs ( C ) or CCN5-knockdown HESCs ( D ) treated with TGF-β or without TGF-β. Data are presented as the mean ± SEM. Statistical analysis was performed using one-way ANOVA for each target gene. * p < 0.05, ** p < 0.01. E, F. The protein expression of p-Smad3, Smad3, p-β-catenin and β-catenin in CCN5-overexpression HESCs ( E ) or CCN5-knockdown HESCs ( F ) treated with TGF-β or without TGF-β. G. The protein expression of β-catenin in cytoplasmic and nuclear part of CCN5-overexpression HESCs or CCN5-knockdown HESCs was measured by western blotting

Article Snippet: The antibody for CCN5 (sc-514070) was purchased from Santa Cruz, and p-β-catenin (80,067–1-RR), β-actin (81,115–1-RR), LaminB1 (12,987–1-AP), c-Myc (67,447–1-Ig) and Cyclin D1 (60,186–1-Ig) were purchased from Proteintech, whilst α-SMA (#19,245), Collagen I (#72,026), Smad3 (#9523), p-Smad3 (#9520), β-catenin (#8480) were purchased from Cell Signaling Technology.

Techniques: Activation Assay, Protein-Protein interactions, Luciferase, Transfection, Activity Assay, Expressing, Over Expression, Knockdown, Western Blot

Journal: Journal of Translational Medicine

Article Title: CCN5 negatively regulates TGF-β-induced endometriosis associated fibrosis through Wnt/β-catenin signaling via Smad3-dependent mechanism

doi: 10.1186/s12967-025-06804-9

Figure Lengend Snippet: CCN5 regulates TGF-β induced proliferation and pro-fibrotic transition in HESCs through phosphorylation of Smad3. A Proliferative capability of primary HESCs in shCCN5-HESCs or shNC-HESCs treated with or without SIS3 assessed by CCK8 assay. Data are presented as mean ± SD. Statistical analysis was conducted using one-way ANOVA for the optical density values at 72 h. * p < 0.05. B The mRNA expression of downstream target genes of TGF-β/Smad and Wnt/β-catenin signaling in shCCN5-HESCs or shNC-HESCs treated with SIS3 or without SIS3. Data are presented as the mean ± SEM. Statistical analysis was performed using one-way ANOVA for each target gene. ns, non-significant, * p < 0.05, ** p < 0.01. C The protein expression of α-SMA and Collagen Ι in shCCN5-HESCs or shNC-HESCs treated with SIS3 or without SIS3. D The HE, Masson and Sirius red staining of the endometriosis tissues from the endometriosis mice constructed based on CCN5 knockout mice and combined SIS3 treatment, along with the expression of α-SMA in the endometriosis tissues. Scale bar, 100 μm

Article Snippet: The antibody for CCN5 (sc-514070) was purchased from Santa Cruz, and p-β-catenin (80,067–1-RR), β-actin (81,115–1-RR), LaminB1 (12,987–1-AP), c-Myc (67,447–1-Ig) and Cyclin D1 (60,186–1-Ig) were purchased from Proteintech, whilst α-SMA (#19,245), Collagen I (#72,026), Smad3 (#9523), p-Smad3 (#9520), β-catenin (#8480) were purchased from Cell Signaling Technology.

Techniques: Phospho-proteomics, CCK-8 Assay, Expressing, Staining, Construct, Knock-Out

Journal: Journal of Translational Medicine

Article Title: CCN5 negatively regulates TGF-β-induced endometriosis associated fibrosis through Wnt/β-catenin signaling via Smad3-dependent mechanism

doi: 10.1186/s12967-025-06804-9

Figure Lengend Snippet: CCN5 prohibited Smad3 shuttle into nucleus by forming complex in cytosol. A Immunofluorescence for CCN5 and Smad3 in HESCs treated with TGF-β or without TGF-β. Scale bar, 20 μm (green CCN5, red Smad3, blue DAPI). B The distribution of Smad3 in the nucleus and cytoplasm in CCN5-overexpression HESCs (CCN5) and control-HESCs treated with TGF-β or without TGF-β was detected by WB. C Co-immunoprecipitation assay of endogenous CCN5 and Smad3 in HESCs treated with or without TGF-β specific inhibitor TGF-β-IN-1. D Co-immunoprecipitation assay of CCN5 and Smad3 in cytoplasm or nucleus in HESCs treated with or without TGF-β specific inhibitor TGF-β-IN-1. E. Molecular docking of CCN5 with Smad3

Article Snippet: The antibody for CCN5 (sc-514070) was purchased from Santa Cruz, and p-β-catenin (80,067–1-RR), β-actin (81,115–1-RR), LaminB1 (12,987–1-AP), c-Myc (67,447–1-Ig) and Cyclin D1 (60,186–1-Ig) were purchased from Proteintech, whilst α-SMA (#19,245), Collagen I (#72,026), Smad3 (#9523), p-Smad3 (#9520), β-catenin (#8480) were purchased from Cell Signaling Technology.

Techniques: Immunofluorescence, Over Expression, Control, Co-Immunoprecipitation Assay

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: Expression kinetics of Ccn5 , Ccn2 , and Col Iα1 in isolated liver cells. ( A ) mRNA expression of Ccn5, Ccn2 , and Col Iα1 in hepatocytes, hepatic stellate cells (HSC), myofibroblasts (MFB) and portal myofibroblasts (pMF) in passage 2 (p2) cultured for the indicated time intervals was determined by RT-qPCR. Ccn5 and Col Iα1 expression is undetectable in hepatocytes, while in contrast Ccn2 mRNA increases during prolonged culture times in hepatocytes. In activated HSC and transdifferentiated MFB Ccn5 mRNA is up-regulated with the activation state of the cells. This up-regulation is even more pronounced than the expression of Ccn2 and parallels the induction of Col Iα1 in the course of HSC to MFB transition. Activated pMF displaying a MFB-like phenotype express large quantities of Ccn5 , Ccn2 , and Col Iα1 mRNAs. ( B ) Protein analysis confirms the absence of CCN5 in hepatocytes and the expression in HSC/MFB and PMF. In this analysis, Vimentin and α-SMA served as markers to demonstrate the mesenchymal phenotype of HSC, MFB and pMF. LCN2 was used as a marker for hepatocytes. Fibronectin and Col Iα1 expression confirmed elevated expression of extracellular matrix proteins during transition of HSC to MFB, while the discrimination between pMF and HSC/MFB was done by probing for Desmin

Article Snippet: The two expression vectors RC204636 and MR203197 encoding full-length human or mouse Myc-DDK-tagged CCN5/WISP2 were obtained from Origene.

Techniques: Expressing, Isolation, Cell Culture, Quantitative RT-PCR, Activation Assay, Marker

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: Ccn5 expression in the inflammatory/fibrogenic bile duct obstruction model. ( A ) Mice were subjected to bile duct ligation (BDL) for 2 weeks. Hepatic expression of indicated genes was compared to sham-operated control animals by RT-qPCR showing that Col Iα1 , Acta2 , Lcn2 , and Tgfb1 are strongly upregulated during hepatic injury. Similarly, the expression of Ccn1 and Ccn2 in fibrotic liver tissue is increased, while the expression of Ccn3 is comparable in healthy and diseased livers. Interestingly, the expression of Ccn5 is significantly upregulated in diseased livers. Differences between the groups reaching significance are marked by asterisks (* p < 0.05, ** p < 0.01, *** p < 0.001). ( B ) Protein analysis confirms the increased expression of α-SMA and Col Iα1 and the elevated expression of CCN5 in livers of animals subjected to BDL. ( C ) Densitometric analysis of Western blot data depicted in ( B )

Article Snippet: The two expression vectors RC204636 and MR203197 encoding full-length human or mouse Myc-DDK-tagged CCN5/WISP2 were obtained from Origene.

Techniques: Expressing, Ligation, Control, Quantitative RT-PCR, Western Blot

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: Effects of adenoviral overexpression of CCN5 in portal myofibroblasts. ( A ) Cultured portal myofibroblasts were infected for indicated time intervals with adenoviral expression vectors directing expression of murine CCN5, human CCN5 or Luciferase as a control. Mock infected cells served as a further control. Total RNA was isolated and the expression of mCcn5 , hccn5 , Col Iα1 , and Acta2 determined by RT-qPCR. ( B ) Protein analysis confirms the prominent overexpression of the transgenes (CCN5, Luciferase). Both CCN5 transgenes cause a reduction in Col Iα1 expression after infection for 48 h, 72 h, and 96 h, as evaluated by densitometric analysis of the depicted Western blot results. On the other hand, the effect on α-SMA expression is much less, leading to a lower expression only at 72 h and 96 h. ( C ) Western blot analysis demonstrates that overexpression of CCN5 results in reduced expression of the TGF-β type I receptor (ALK5, TβRI) and TGF-β type II receptor (TβRII). As a mutual consequence of the lowered receptor expression, Smad2 gets less activated from 48 h on, while at 72 h and 96 h after overexpression of CCN5 the genuine BMP or „alternative “ TGF-β1/Smad pathway as assessed by increased Smad1/5/8 phosphorylation is activated. In addition, Jun B is strongly up-regulated in cells overexpressing CCN5. GAPDH expression was used in Western blot analysis to document equal protein loading. Depicted is a representative experiment of an analysis that was done twice. The densitometric analysis of the Western blot is shown in Suppl. Figure 6

Article Snippet: The two expression vectors RC204636 and MR203197 encoding full-length human or mouse Myc-DDK-tagged CCN5/WISP2 were obtained from Origene.

Techniques: Over Expression, Cell Culture, Infection, Expressing, Luciferase, Control, Isolation, Quantitative RT-PCR, Western Blot

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: Impact of CCN5 on TGF-β signaling. Portal myofibroblasts were infected with adenoviral vectors expressing either human CCN5 (hCCN5) or the luciferase reporter gene (Luc). After 48 h, the cells were stimulated with the indicated concentrations of recombinant TGF-β1. Protein extracts and supernatants were prepared and tested for expression of Col Iα1, Fibronectin, α-SMA, CHOP, GAPDH ( left panel, cell lysate ), Col Iα1, Fibronectin, hCCN5 ( middle panel, culture supernatant ), and GRP94, GRP78, CCN2, and CCN5 ( right panel, cell lysate ). The expression of β-actin was used to document equal protein loading in each lane. Of note, there was no significant effect on the extracellular matrix components Col Iα1, Fibronectin and the activation marker α-SMA, but a strong TGF-β1 independent up-regulation of the UPR related proteins GRP94, GRP78 and CHOP. The densitometric analysis of the three Western blots is shown in Suppl. Figure 7

Article Snippet: The two expression vectors RC204636 and MR203197 encoding full-length human or mouse Myc-DDK-tagged CCN5/WISP2 were obtained from Origene.

Techniques: Infection, Expressing, Luciferase, Recombinant, Activation Assay, Marker, Western Blot

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: CCN5 induces endoplasmic reticulum stress in portal myofibroblasts. ( A ) Portal myofibroblasts were infected with indicated adenoviral constructs for indicated time intervals. Subsequently, total RNA was isolated, cDNA synthesized. The expression of unspliced (u) and spliced (s) Xbp1 expression after infection with adenoviral vectors expressing murine or human CCN5 was analyzed by semi-quantitative PCR. Spliced Xbp1 (Xbp1 (s)) transcripts were found in the presence of overexpressed mCCN5 and hCCN5, while cells that were infected with a control virus (Ad-Luc) or cells that were left uninfected showed no spliced Xbp1 . In this analysis, GAPDH was taken as a loading control. ( B ) Relative mRNA expression of Grp94 , Bip and Chop as assessed by RT-qPCR in samples taken from ( A ). ( C ) Western blot analysis of GRP94, GRP78/BIP, IRE1α, ATF6, peIF2α, eIF2α, ATF4, CHOP, pJNK, JNK2, cytochrome c, cleaved caspase-9, and cleaved caspase-3 in protein samples taken from portal myofibroblasts that were infected with indicated adenoviral vectors for indicated time intervals. Expression of GAPDH was used to demonstrate equal protein loading in each lane. The densitometric analysis of the Western blot is given in Suppl. Figure 8

Article Snippet: The two expression vectors RC204636 and MR203197 encoding full-length human or mouse Myc-DDK-tagged CCN5/WISP2 were obtained from Origene.

Techniques: Infection, Construct, Isolation, Synthesized, Expressing, Real-time Polymerase Chain Reaction, Control, Virus, Quantitative RT-PCR, Western Blot

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: Effects of adenoviral overexpression of CCN5 in portal myofibroblasts. ( A ) Cultured portal myofibroblasts were infected for indicated time intervals with adenoviral expression vectors directing expression of murine CCN5, human CCN5 or Luciferase as a control. Mock infected cells served as a further control. Total RNA was isolated and the expression of mCcn5 , hccn5 , Col Iα1 , and Acta2 determined by RT-qPCR. ( B ) Protein analysis confirms the prominent overexpression of the transgenes (CCN5, Luciferase). Both CCN5 transgenes cause a reduction in Col Iα1 expression after infection for 48 h, 72 h, and 96 h, as evaluated by densitometric analysis of the depicted Western blot results. On the other hand, the effect on α-SMA expression is much less, leading to a lower expression only at 72 h and 96 h. ( C ) Western blot analysis demonstrates that overexpression of CCN5 results in reduced expression of the TGF-β type I receptor (ALK5, TβRI) and TGF-β type II receptor (TβRII). As a mutual consequence of the lowered receptor expression, Smad2 gets less activated from 48 h on, while at 72 h and 96 h after overexpression of CCN5 the genuine BMP or „alternative “ TGF-β1/Smad pathway as assessed by increased Smad1/5/8 phosphorylation is activated. In addition, Jun B is strongly up-regulated in cells overexpressing CCN5. GAPDH expression was used in Western blot analysis to document equal protein loading. Depicted is a representative experiment of an analysis that was done twice. The densitometric analysis of the Western blot is shown in Suppl. Figure 6

Article Snippet: The vectors used for transfection were Myc-DDK-tagged pCMV6-Entry-mCCN5 (#MR203197, Origene, Herford, Germany) and Myc-DDK-tagged pCMV6-Entry hCCN5 (#RC204636, Origene).

Techniques: Over Expression, Cell Culture, Infection, Expressing, Luciferase, Control, Isolation, Quantitative RT-PCR, Western Blot

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: Impact of CCN5 on TGF-β signaling. Portal myofibroblasts were infected with adenoviral vectors expressing either human CCN5 (hCCN5) or the luciferase reporter gene (Luc). After 48 h, the cells were stimulated with the indicated concentrations of recombinant TGF-β1. Protein extracts and supernatants were prepared and tested for expression of Col Iα1, Fibronectin, α-SMA, CHOP, GAPDH ( left panel, cell lysate ), Col Iα1, Fibronectin, hCCN5 ( middle panel, culture supernatant ), and GRP94, GRP78, CCN2, and CCN5 ( right panel, cell lysate ). The expression of β-actin was used to document equal protein loading in each lane. Of note, there was no significant effect on the extracellular matrix components Col Iα1, Fibronectin and the activation marker α-SMA, but a strong TGF-β1 independent up-regulation of the UPR related proteins GRP94, GRP78 and CHOP. The densitometric analysis of the three Western blots is shown in Suppl. Figure 7

Article Snippet: The vectors used for transfection were Myc-DDK-tagged pCMV6-Entry-mCCN5 (#MR203197, Origene, Herford, Germany) and Myc-DDK-tagged pCMV6-Entry hCCN5 (#RC204636, Origene).

Techniques: Infection, Expressing, Luciferase, Recombinant, Activation Assay, Marker, Western Blot

Journal: Journal of Cell Communication and Signaling

Article Title: Expression and biological function of the cellular communication network factor 5 (CCN5) in primary liver cells

doi: 10.1007/s12079-023-00757-8

Figure Lengend Snippet: CCN5 induces endoplasmic reticulum stress in portal myofibroblasts. ( A ) Portal myofibroblasts were infected with indicated adenoviral constructs for indicated time intervals. Subsequently, total RNA was isolated, cDNA synthesized. The expression of unspliced (u) and spliced (s) Xbp1 expression after infection with adenoviral vectors expressing murine or human CCN5 was analyzed by semi-quantitative PCR. Spliced Xbp1 (Xbp1 (s)) transcripts were found in the presence of overexpressed mCCN5 and hCCN5, while cells that were infected with a control virus (Ad-Luc) or cells that were left uninfected showed no spliced Xbp1 . In this analysis, GAPDH was taken as a loading control. ( B ) Relative mRNA expression of Grp94 , Bip and Chop as assessed by RT-qPCR in samples taken from ( A ). ( C ) Western blot analysis of GRP94, GRP78/BIP, IRE1α, ATF6, peIF2α, eIF2α, ATF4, CHOP, pJNK, JNK2, cytochrome c, cleaved caspase-9, and cleaved caspase-3 in protein samples taken from portal myofibroblasts that were infected with indicated adenoviral vectors for indicated time intervals. Expression of GAPDH was used to demonstrate equal protein loading in each lane. The densitometric analysis of the Western blot is given in Suppl. Figure 8

Article Snippet: The vectors used for transfection were Myc-DDK-tagged pCMV6-Entry-mCCN5 (#MR203197, Origene, Herford, Germany) and Myc-DDK-tagged pCMV6-Entry hCCN5 (#RC204636, Origene).

Techniques: Infection, Construct, Isolation, Synthesized, Expressing, Real-time Polymerase Chain Reaction, Control, Virus, Quantitative RT-PCR, Western Blot