Journal: International Journal of Biological Sciences

Article Title: MiR-206 suppresses the deterioration of intrahepatic cholangiocarcinoma and promotes sensitivity to chemotherapy by inhibiting interactions with stromal CAFs

doi: 10.7150/ijbs.62602

Figure Lengend Snippet: Decreased miR-206 expression promoted NF reprogramming into the CAF phenotype and enhanced IL-6 secretion by targeting Anxa2. (A) Expression of alpha-SMA and miR-206 in NFs was evaluated by FISH assay to observe the response of miR-206 expression to TGF-beta1 treatment (20 ng/mL for 12 hours). Scale bar=50 µm. (B) Relative changes in the alpha-SMA and miR-206 mRNA levels in response to TGF-beta1 treatment were detected by qPCR. (C) The protein expression of CAF markers, alpha-SMA and FAP in miR-206-mimic/CAFs and miR-206-inhibitor/NFs was detected by WB assay. (D) The binding site of miR-206 in the Anxa2 3' UTR was predicted, and the binding site was confirmed by dual-luciferase reporter assay. (E) Relative expression of Anxa2 in 3 pairs of NFs and CAFs was detected by qPCR. (F) Regulation of alpha-SMA and Anxa2 expression by miR-206 was analyzed by WB assay. (G) The correlation between miR-206 expression and Anxa2 expression in tissues was analyzed ( P <0.05, R 2 =0.3726). (H) Anxa2 overexpression was confirmed by WB assay. (I) Alpha-SMA protein levels were evaluated after Anxa2 expression was upregulated in miR-206-mimic/CAFs. (J-M) The effects of Anxa2 in miR-206-mimic/CAFs on CAF proliferation, motility, collagen contraction and vascular formation were studied. (N) Fold changes in the IL-1beta, IL-6, IL-8, VEGF-alpha and CXCL12 levels in conditioned medium from miR-206-mimic/CAFs with Anxa2 upregulated expression. The data are shown as the mean ± SD, * P <0.05, ** P <0.01, *** P < 0.001.

Article Snippet: Primary antibodies against Vimentin, alpha-SMA and FAP were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Expressing, Binding Assay, Luciferase, Reporter Assay, Over Expression

Journal: International Journal of Biological Sciences

Article Title: MiR-206 suppresses the deterioration of intrahepatic cholangiocarcinoma and promotes sensitivity to chemotherapy by inhibiting interactions with stromal CAFs

doi: 10.7150/ijbs.62602

Figure Lengend Snippet: Overexpression of miR-206 suppressed the mutual promotion of malignant behaviors and gemcitabine resistance in the CCA-CAF environment. (A-B) The proliferation and migration abilities of HUCCT1 cells cocultured with miR-206-mimic/CAFs were explored by colony formation and Transwell assays. (C) The effects of miR-206-mimic/CAFs on HUCCT1 cell resistance to gemcitabine were evaluated by CCK-8 and colony formation assays. (D) The cycle distribution of HUCCT1 cells cocultured with miR-206-mimic/CAFs was analyzed by FCM. (E) Sphere formation by HUCCT1 cells cocultured with miR-206-mimic/CAFs were detected and assessed. Scale bar=100 µm. (F) Expression of Nang, Sox2 and Oct4 in HUCCT1 cells was detected by qPCR. (G) MiR-206-mimic/CAFs combined with HUCCT1 cells were subcutaneously coinjected into mice. After gemcitabine treatment for 3-4 weeks, the mice were sacrificed, subcutaneous tumors were imaged, and tumor volume was calculated. (H) The mRNA level of TGF-beta1 in HUCCT1 cells cocultured with miR-206-mimic/CAFs was detected by qPCR. (I) Alpha-SMA and FAP expression in NFs cocultured with miR-206-mimic/HUCCT1 cells was evaluated by IF assay. NFs culture alone as negative control. Scale bar=100 µm. (J) The mRNA levels of IL-1beta, IL-6, IL-8, VEGF-alpha and CXCL12 in NFs cocultured with miR-206-mimic/HUCCT1 cells were detected by qPCR. NFs culture alone as negative control. The data are shown as the mean ± SD, * P <0.05, ** P <0.01, *** P < 0.001.

Article Snippet: Primary antibodies against Vimentin, alpha-SMA and FAP were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Over Expression, Migration, CCK-8 Assay, Expressing, Negative Control

Journal: International Journal of Biological Sciences

Article Title: MiR-206 suppresses the deterioration of intrahepatic cholangiocarcinoma and promotes sensitivity to chemotherapy by inhibiting interactions with stromal CAFs

doi: 10.7150/ijbs.62602

Figure Lengend Snippet: Exosome-delivered miR-206 eliminated the CCA-CAF mutually promoting environment and suppressed malignancy. (A) The expression level of miR-206 in -derived exosomes transfected with the miR-206-mimic was detected by qPCR. (B-E) Effects of miR-206-mimic/exo (50 μg/10 5 cells) on HUCCT1 cell proliferation and migration were detected. The CCK-8, colony formation, cell cycle progression and Transwell migration (scale bar=20 μm) assay results are presented. (F) Levels of LASP1, Nanog, and p-STAT3 in HUCCT1 cells after miR-206-mimic/exo treatment were assessed. (G-H) The expression of CAF markers (alpha-SMA and FAP) and Anxa2 in CAFs was evaluated. (I) HUCCT1 cells and CAFs were cocultured in 6-well plates, and both cell types were treated with gemcitabine or miR-206-mimic/exo. The cells were analyzed compared to those cultured alone. Cell colony number and fold change in the gemcitabine and miR-206-mimic/exo treatment groups were compared and are presented in the right panel. (J) The Bcl-2/Bax ratio and fold change in the Bcl-2/Bax ratio in the gemcitabine and miR-206-mimic/exo groups were also evaluated and analyzed. * P <0.05, ** P <0.01, *** P < 0.001.

Article Snippet: Primary antibodies against Vimentin, alpha-SMA and FAP were purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Expressing, Derivative Assay, Transfection, Migration, CCK-8 Assay, Cell Culture

Journal: Food Science & Nutrition

Article Title: Effects of Miao sour soup on hyperlipidemia in high‐fat diet‐induced obese rats via the AMPK signaling pathway

doi: 10.1002/fsn3.2394

Figure Lengend Snippet: Effects of MSS on Lipogenesis‐Associated Genes and Proteins in the Liver of HFD‐fed Rats. Gene expression in the liver was analyzed via quantitative real‐time PCR (qRT‐PCR), and protein extracts from the liver were analyzed via Western blotting (WB). (a) qRT‐PCR analyses of live lipid‐related molecules such as AMP‐activated protein kinase (AMPK), sterol regulatory element‐binding protein‐1c (SREBP‐1c), fatty acid synthase (FAS), and acetyl‐CoA carboxylase alpha (ACCα). (b) Western blot analysis of lipid‐related molecules (AMPKα, SREBP‐1c, ACCα, and FAS). Results are presented as the mean ± SD ( n = 8). (∗) p < .05, versus ND1 group, (#) p < .05, versus the HFD1 group

Article Snippet: Zhibituo, a drug used in the clinical treatment of hyperlipidemia, was obtained from the Chengdu Diao Jiuhong Pharmaceutical Co., Ltd. Antibodies against AMPKα, ACC, and FAS were purchased from Cell Signaling Technology, and SREBP‐1c and β‐actin antibodies were purchased from Affinity Biosciences.

Techniques: Expressing, Real-time Polymerase Chain Reaction, Quantitative RT-PCR, Western Blot, Binding Assay

Journal: Food Science & Nutrition

Article Title: Effects of Miao sour soup on hyperlipidemia in high‐fat diet‐induced obese rats via the AMPK signaling pathway

doi: 10.1002/fsn3.2394

Figure Lengend Snippet: Effects of MSS on lipogenesis‐related factors in the livers of obese rats. Gene expression in the liver was analyzed via qRT‐PCR, and protein extracts from the liver were measured via WB. (a) qRT‐PCR analyses of live lipid‐related molecules (AMPKα, SREBP‐1c, ACC, and FAS). (b) Western blot analysis of lipid‐related molecules (AMPKα, SREBP‐1c, ACC, and FAS). Results are presented as the mean ± SD ( n = 8). (∗) p < .05, versus ND2 group, (#) p < .05, versus HFD2 group

Article Snippet: Zhibituo, a drug used in the clinical treatment of hyperlipidemia, was obtained from the Chengdu Diao Jiuhong Pharmaceutical Co., Ltd. Antibodies against AMPKα, ACC, and FAS were purchased from Cell Signaling Technology, and SREBP‐1c and β‐actin antibodies were purchased from Affinity Biosciences.

Techniques: Expressing, Quantitative RT-PCR, Western Blot

Journal: PLoS ONE

Article Title: Airway epithelial specific deletion of Jun-N-terminal kinase 1 attenuates pulmonary fibrosis in two independent mouse models

doi: 10.1371/journal.pone.0226904

Figure Lengend Snippet: Primer sequences used in real time PCR analyses.

Article Snippet: Antibodies: Total JNK (#9252), phospho-JNK (#9251) antibodies were obtained from Cell Signaling Technology (Danvers, MA) (all anti Rabbit, 1:1000), antibodies for S100a4 (FSP1) (sc-19949, anti-Goat, 1:500) and actin (sc-8432, anti-mouse 1:5000) were from Santa Cruz Biotechnology (Santa Cruz, CA), antibodies for CCSP (anti goat 1:4000) were a kind gift from Dr. B. Stripp, Cedars-Sinai Medical Center, Los Angeles, CA).

Techniques: Real-time Polymerase Chain Reaction, Amplification

Journal: PLoS ONE

Article Title: Airway epithelial specific deletion of Jun-N-terminal kinase 1 attenuates pulmonary fibrosis in two independent mouse models

doi: 10.1371/journal.pone.0226904

Figure Lengend Snippet: Analysis of mesenchymal ( A ) or epithelial ( B ) mRNA expression in homogenized lung tissue from mice exposed to bleomycin for 3 weeks. Results were normalized to the housekeeping gene cyclophilin, and are expressed as fold expression changes (+/- SEM) compared to the WT vehicle control groups. (WT- Jnk1 : PBS n = 6, Bleo n = 12, ΔEpi Jnk1 : PBS n = 5, Bleo n = 8 respectively mice/group from 2 independent experiments). * p< 0.05 compared to the PBS control group. † p< 0.05 compared to the respective WT group. (ANOVA). C : Evaluation of mesenchymal proteins (α-SMA ( Acta2 ), Col1a1, FSP1 ( S100a4 )) in lungs from ΔEpiJNK1 mice, or control groups subjected to bleomycin-induced lung fibrosis, and the impact of ablation of epithelial JNK1. Homogenized lung tissues were subjected to Western blot analysis for the indicated proteins. β-actin: Loading control. Shown are results from individual mice.

Article Snippet: Antibodies: Total JNK (#9252), phospho-JNK (#9251) antibodies were obtained from Cell Signaling Technology (Danvers, MA) (all anti Rabbit, 1:1000), antibodies for S100a4 (FSP1) (sc-19949, anti-Goat, 1:500) and actin (sc-8432, anti-mouse 1:5000) were from Santa Cruz Biotechnology (Santa Cruz, CA), antibodies for CCSP (anti goat 1:4000) were a kind gift from Dr. B. Stripp, Cedars-Sinai Medical Center, Los Angeles, CA).

Techniques: Expressing, Western Blot