Journal: Immunity, inflammation and disease

Article Title: Novel CLC3 transcript variants in blood eosinophils and increased CLC3 expression in nasal lavage and blood eosinophils of asthmatics.

doi: 10.1002/iid3.36

Figure Lengend Snippet: Figure 1. (A) Constructs of the respective primer pairs to identify CLC3 transcript variants. (B)–(D) showing fold change in the mRNA levels of CLC3, CLC3b, and CLC3e, respectively among healthy (N ¼ 10), mild-to-moderate (N ¼ 10), and moderate-to-severe (N ¼ 10) asthmatic peripheral blood eosinophils. P < 0.05, P < 0.01, P < 0.001, P < 0.0001.

Article Snippet: GFP-tagged ORF gene clone of CLC3b (Accession number: NM_001829.2) and RFP-tagged ORF clone of CLC3e (Accession number: NM_173872.2) were obtained from Origene (Rockville, MD).

Techniques: Construct

Journal: Immunity, inflammation and disease

Article Title: Novel CLC3 transcript variants in blood eosinophils and increased CLC3 expression in nasal lavage and blood eosinophils of asthmatics.

doi: 10.1002/iid3.36

Figure Lengend Snippet: Figure 4. Co-localization of CLC3 transcript variants. HEK 293 cells transfected with CLC3b-GFP and CLC3e-RFP showing homo- and hetero- dimerization. (A) Confocal micrographs showing co-localization of CLC3b-GFP and CLC3e-RFP. (B) Z-stack of the confocal micrograph showing segregation of the transcript variants to apical and basal plane of the cells. (C) qPCR showing change in mRNA levels among respective transfected groups with specific primers designed to detect the CLC3 transcript variants.

Article Snippet: GFP-tagged ORF gene clone of CLC3b (Accession number: NM_001829.2) and RFP-tagged ORF clone of CLC3e (Accession number: NM_173872.2) were obtained from Origene (Rockville, MD).

Techniques: Transfection

Journal: Cell Death Discovery

Article Title: CLC3 regulates V-ATPase to enhance lysosomal degradation and cisplatin resistance in cervical cancer cells

doi: 10.1038/s41420-025-02876-0

Figure Lengend Snippet: A Relative CLC3 mRNA expression levels in various CVC cell lines (SiHa, MS751, HeLa, ME180, Hela229, CaSKi) compared to normal cervical epithelial cells (NCCs). * P < 0.05, ** P < 0.001. vs. NCCs. B Western blot analysis showing CLC3 protein expression in NCCs and different CVC cell lines (SiHa, MS751, HeLa, ME180, Hela229, CaSKi), with GAPDH as a loading control. * P < 0.05, ** P < 0.001. vs. NCCs. C CLC3 mRNA expression in CVC cell lines (SiHa, MS751, HeLa, ME180, Hela229, and CaSKi) following CLC3 overexpression (CLC3 High ) or knockdown (CLC3 Low ) compared to control. * P < 0.05, ** P < 0.001. vs. Control. D Western blot analysis of CLC3 protein levels in CLC3 High , CLC3 Low , and control cells across the same cell lines, with quantification shown in the accompanying graphs. * P < 0.05, ** P < 0.001. vs. Control. E IC50 values for cisplatin, paclitaxel, and 5-FU in CLC3 High , CLC3 Low , and control groups in SiHa and CaSKi cells, showing CLC3 modulation’s effect on chemoresistance. * P < 0.05 vs. Control.

Article Snippet: For immunoprecipitation, equal amounts of total protein (500–800 μg) were incubated overnight at 4 °C with 2–3 μg of anti-CLC3 antibody or normal rabbit IgG as a negative control, followed by incubation with protein A/G agarose beads (Santa Cruz Biotechnology).

Techniques: Expressing, Western Blot, Control, Over Expression, Knockdown

Journal: Cell Death Discovery

Article Title: CLC3 regulates V-ATPase to enhance lysosomal degradation and cisplatin resistance in cervical cancer cells

doi: 10.1038/s41420-025-02876-0

Figure Lengend Snippet: A Fluorescence images and bar graphs showing cytosolic pH in SiHa and CaSKi cells with CLC3 overexpression (CLC3 High ), knockdown (CLC3 Low ), and control, using SypHer3s as the pH probe at 60- and 120-min post-treatment. B Representative images and quantification of lysosomal pH in SiHa and CaSKi cells with CLC3 modulation, analyzed using LysoSensor Yellow/Blue DND-160. C Lysosomal pH in cells treated with V-ATPase inhibitor Baf A1 combined with CLC3 High and CLC3 Low modulation. D Immunofluorescence images showing the fluorescence intensity of cathepsin B (B MR) and cathepsin L (L MR) in cells treated with Baf A1 and CLC3-modulated conditions, indicating lysosomal degradation capacity. * P < 0.05, ** P < 0.001. vs. Control.

Article Snippet: For immunoprecipitation, equal amounts of total protein (500–800 μg) were incubated overnight at 4 °C with 2–3 μg of anti-CLC3 antibody or normal rabbit IgG as a negative control, followed by incubation with protein A/G agarose beads (Santa Cruz Biotechnology).

Techniques: Fluorescence, Over Expression, Knockdown, Control, Immunofluorescence

Journal: Cell Death Discovery

Article Title: CLC3 regulates V-ATPase to enhance lysosomal degradation and cisplatin resistance in cervical cancer cells

doi: 10.1038/s41420-025-02876-0

Figure Lengend Snippet: A Western blot analysis of autophagy and apoptosis markers, including P62, LC3 (LC3I and LC3II), and cleaved caspases-3 and -8 in CLC3 High , CLC3 Low , and control cells. Bar graphs quantify the relative expression levels of these proteins, showing increased autophagic flux and apoptotic activity in CLC3 Low cells. B Immunofluorescence staining of LC3 and P62 in CLC3-modulated cells, with quantification of relative fluorescence intensity in CLC3 High , CLC3 Low , and control groups. * P < 0.05, ** P < 0.001. vs. Control. Data are presented as mean ± SD. Statistical significance was determined using one-way ANOVA with post hoc tests; p < 0.05, p < 0.01, p < 0.001.

Article Snippet: For immunoprecipitation, equal amounts of total protein (500–800 μg) were incubated overnight at 4 °C with 2–3 μg of anti-CLC3 antibody or normal rabbit IgG as a negative control, followed by incubation with protein A/G agarose beads (Santa Cruz Biotechnology).

Techniques: Western Blot, Control, Expressing, Activity Assay, Immunofluorescence, Staining, Fluorescence

Journal: Cell Death Discovery

Article Title: CLC3 regulates V-ATPase to enhance lysosomal degradation and cisplatin resistance in cervical cancer cells

doi: 10.1038/s41420-025-02876-0

Figure Lengend Snippet: A Heatmap showing the expression of various V-ATPase subunits and companion genes across control, a2v-mAb-treated, CLC3 Low with a2v-mAb, and CLC3 High with a2v-mAb groups in CVC cells. B Lysosomal pH analysis in SiHa and CaSKi cells under the same experimental conditions, with images from LysoSensor Yellow/Blue DND-160 assays and quantitative analysis of pH changes. * P < 0.05, ** P < 0.001. vs. Control. # P < 0.05, ## P < 0.001. vs. a2v-mAb.

Article Snippet: For immunoprecipitation, equal amounts of total protein (500–800 μg) were incubated overnight at 4 °C with 2–3 μg of anti-CLC3 antibody or normal rabbit IgG as a negative control, followed by incubation with protein A/G agarose beads (Santa Cruz Biotechnology).

Techniques: Expressing, Control

Journal: Cell Death Discovery

Article Title: CLC3 regulates V-ATPase to enhance lysosomal degradation and cisplatin resistance in cervical cancer cells

doi: 10.1038/s41420-025-02876-0

Figure Lengend Snippet: A Cell viability assays in SiHa and CaSKi cells under various treatment conditions, including a2v-mAb treatment, CLC3 High , and CLC3 Low . B Colony formation assay results, quantifying cell colony numbers to evaluate the proliferative capacity of each treatment group. C Flow cytometry analysis of apoptosis rates in SiHa and CaSKi cells with CLC3 modulation and a2v-mAb treatment, with quantification of apoptotic cells. D RT-PCR analysis of tumor stemness markers (NANOG, OCT4, KLF4, ABCG2, BMI1, MYC, TIP30, MAD2, STAT3) and CLC3 expression levels in CLC3-modulated cells treated with a2v-mAb, demonstrating the effect of CLC3 on maintaining stemness characteristics. E IC50 values for cisplatin, paclitaxel, and 5-FU in SiHa and CaSKi cells under each treatment condition. * P < 0.05 vs. Control. # P < 0.05 vs. a2v-mAb.

Article Snippet: For immunoprecipitation, equal amounts of total protein (500–800 μg) were incubated overnight at 4 °C with 2–3 μg of anti-CLC3 antibody or normal rabbit IgG as a negative control, followed by incubation with protein A/G agarose beads (Santa Cruz Biotechnology).

Techniques: Colony Assay, Flow Cytometry, Reverse Transcription Polymerase Chain Reaction, Expressing, Control

Journal: Cell Death Discovery

Article Title: CLC3 regulates V-ATPase to enhance lysosomal degradation and cisplatin resistance in cervical cancer cells

doi: 10.1038/s41420-025-02876-0

Figure Lengend Snippet: A Tumor volume and weight measurements in mice treated with cisplatin, CLC3 Low with cisplatin, CLC3 High with cisplatin, and control. B Hematoxylin and eosin (H&E) staining, along with immunohistochemical (IHC) staining for CLC3, cyclin D1, cyclin D3, and Ki67 in tumor sections from each group, with quantification of positive cells. C TUNEL assay showing apoptosis levels in tumors from each treatment condition, with quantification of TUNEL-positive cells. D Histological analysis of lung tissue for metastases, comparing the number of metastatic lung nodules across different treatment groups, highlighting the anti-metastatic effect of CLC3 knockdown with cisplatin. * P < 0.05, ** P < 0.001. vs. Control. # P < 0.05, ## P < 0.001. vs. Cisplatin.

Article Snippet: For immunoprecipitation, equal amounts of total protein (500–800 μg) were incubated overnight at 4 °C with 2–3 μg of anti-CLC3 antibody or normal rabbit IgG as a negative control, followed by incubation with protein A/G agarose beads (Santa Cruz Biotechnology).

Techniques: Control, Staining, Immunohistochemical staining, Immunohistochemistry, TUNEL Assay, Knockdown