Journal: American Journal of Respiratory and Critical Care Medicine

Article Title: Lactic Acid Is Elevated in Idiopathic Pulmonary Fibrosis and Induces Myofibroblast Differentiation via pH-Dependent Activation of Transforming Growth Factor-β

doi: 10.1164/rccm.201201-0084oc

Figure Lengend Snippet: Figure 4. Lactic acid induces myofibroblast differentiation in a pH-dependent manner. (A) Human lung fibroblasts were cultured with transforming growth factor (TGF)-b, media containing lactic acid in which pH was adjusted after the addition of lactic acid or media containing lactic acid in which the lactic acid solution was pH adjusted before its addition to the media (n ¼ 3 each). Myofibroblast differentiation was assessed by Western blot for expression of a smooth muscle actin (a SMA) (*analysis of variance [ANOVA], P , 0.05 compared with TGF-b and lactic acid). (B) Human lung fibroblasts were cultured in serum free media (SF) and media containing 1%, 5%, and 10% fetal bovine serum (FBS) (n ¼ 3 each). Protein lysates were analyzed by Western blot for aSMA (*ANOVA, P , 0.05 compared with untreated). (C) Mv1Lu cells were cultured in the presence of 5 ng/mL TGF-b or media containing 10 mM or 20 mM lactic acid for 22 hours (n ¼ 4 each). Cell proliferation was determined by BrdU colorimetric analysis. Diminished BrdU incorporation was indicative of decreased proliferation correlating to enhanced TGF-b bioactivity (*P , 0.05 compared with untreated controls). (D, E) Primary human lung fibroblasts were cocultured with and without SB431542, a specific TGF-b1 receptor inhibitor, 5 ng/mL TGF-b, and 20 mM lactic acid (n ¼ 3 each). Myofibroblast differentiation was assessed by Western blot for expression of aSMA (*ANOVA, P , 0.05 compared with TGF-b and 20 mM lactic acid). (F) Primary human lung fibroblasts were cultured with 5 ng/mL TGF-b or 20 mM lactic acid (n ¼ 3 each). p-Smad 2 and total Smad 2/3 expression were measured by Western blot of whole cell lysates (*ANOVA, P , 0.05 compared with untreated). GAPDH = glyceraldehyde 3-phosphate dehydrogenase; ns ¼ not significant; Untx ¼ untreated.

Article Snippet: TGF-b Bioassay Mv1Lu mink lung epithelial cells (American Type Culture Collection CCl-64, Manassas, VA) were cultured as previously described (30).

Techniques: Cell Culture, Western Blot, Expressing, BrdU Incorporation Assay

Journal: Cell Metabolism

Article Title: A purine metabolic checkpoint that prevents autoimmunity and autoinflammation

doi: 10.1016/j.cmet.2021.12.009

Figure Lengend Snippet:

Article Snippet: Influenza viral load was determined in total RNA extracted from lungs (RNeasy kit, QIAGEN, 74104), via qRT-PCR using primers for influenza M protein transcript, 5′-GGACTGCAGCGTTAGACGCTT-3′; and 5′-CATCCTGTTGTATATGAGGCCCAT-3′ ( ).

Techniques: Virus, Recombinant, Protease Inhibitor, Bicinchoninic Acid Protein Assay, Staining, Cell Isolation, Enzyme-linked Immunosorbent Assay, In Situ, Red Blood Cell Lysis, Control, Software

Journal: Journal of Cancer

Article Title: The Effect of Tumor Microenvironment on Autophagy and Sensitivity to Targeted Therapy in EGFR-Mutated Lung Adenocarcinoma

doi: 10.7150/jca.11187

Figure Lengend Snippet: Cytokine production and autophagy induction in both MRC-5 and HCC827 cells in the tumor microenvironment. (A) After staining for EpCAM, the co-cultured cells were divided into 2 populations. (B) The IL-6 and IL-8 mRNA expression in sorted cells was significantly increased compared with corresponding homotypical cells. (C) Autophagy was induced in sorted cells compared with their homotypical counterparts, as evidenced by p62 degradation. Western blot data were quantified by fold-change compared with homotypical MRC-5 cells or homotypical HCC827 cells. Comparisons are made between sorted cells with respective homotypical cells. Statistical significance (*: p <0.05, **: p <0.01) indicates comparison with control.

Article Snippet: Human lung ADC HCC827 cells (EGFR exon 19 deletion) and human lung fibroblast MRC-5 cells (American Type Culture Collection, Manassas, VA, USA) were obtained.

Techniques: Staining, Cell Culture, Expressing, Western Blot, Comparison, Control

Journal: Journal of Cancer

Article Title: The Effect of Tumor Microenvironment on Autophagy and Sensitivity to Targeted Therapy in EGFR-Mutated Lung Adenocarcinoma

doi: 10.7150/jca.11187

Figure Lengend Snippet: (A) Apoptosis in HCC827 or co-cultured HCC827 cells after erlotinib ± chloroquine treatment. In homotypical HCC827 cells, combination of erlotinib with chloroquine significantly increased apoptotic cell death compared with erlotinib alone. In co-culture conditions, the sensitivity to erlotinib was preserved and synergistic combination of erlotinib/chloroquine was evident. Statistical significance (*: p <0.05, **: p <0.01) indicates comparison with control. (B) Antitumor effect of erlotinib, chloroquine (CQ) or combined erlotinib/CQ in a HCC827 xenograft model in nude mice. Longitudinal tumor size with different treatment groups. The relative tumor volume was calculated as fold-change to the baseline tumor volume prior to treatment. Progressive tumor growth was noted in both control and CQ treatment group, while significant tumor growth suppression was evident in erlotinib treatment and combination group. Statistical significance (*: p <0.05, **: p <0.01) indicates comparison between erlotinib with control and combination with erlotinib.

Article Snippet: Human lung ADC HCC827 cells (EGFR exon 19 deletion) and human lung fibroblast MRC-5 cells (American Type Culture Collection, Manassas, VA, USA) were obtained.

Techniques: Cell Culture, Co-Culture Assay, Comparison, Control