Journal: Cancers

Article Title: Differential Protumoral Mechanisms Induced by CAFs in Cervical Cancer Cells Occur Independently of 17β-Estradiol Stimulation

doi: 10.3390/cancers18101509

Figure Lengend Snippet: CAFs decrease the mitochondrial metabolism and enhance the ROS production in SiHa cells. Mitochondrial metabolism was assessed by MTT assay, and ROS by H2DCFDA. SiHa and HeLa cells were cultured with LG or HG CAFs’ supernatant prior to metabolism or ROS measurement. ( A ) Mitochondrial metabolism of SiHa and HeLa cells stimulated with LG and HG CAFs, and metabolic activity was normalized against the control. ( B ) Production of ROS by SiHa and HeLa cells stimulated with LG and HG CAFs. Statistical analysis was first performed with a normality test using the Shapiro–Wilk test and later with Kruskal–Wallis or one-way ANOVA. Data is presented as mean ± SD. * p < 0.05, ** p < 0.01, **** p < 0.0001. LG CAF: Cancer-associated fibroblasts from low-grade lesion, HG CAF: Cancer-associated fibroblasts from high-grade lesion, Basal: Cells cultured with DMEM, CAF basal: Cells cultured with CAF’s supernatant without E2 stimulation, CAF E2: Cells cultured with CAF’s supernatant with previous stimulation with E2.

Article Snippet: CC cell lines SiHa and HeLa were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: MTT Assay, Cell Culture, Activity Assay, Control

Journal: Cancers

Article Title: Differential Protumoral Mechanisms Induced by CAFs in Cervical Cancer Cells Occur Independently of 17β-Estradiol Stimulation

doi: 10.3390/cancers18101509

Figure Lengend Snippet: CAFs promote cell migration in HeLa cells. Migration was assessed using a wound-healing assay and quantified as the percentage of wound closure. ( A ) Migration of SiHa cultured with LG and HG cervical cancer lesion CAFs’ supernatant. ( B ) Migration of HeLa cultured with LG and HG cervical cancer lesion CAFs’ supernatant. In photographs, the yellow color is used to highlight the measured % of area in the assay. Statistical analysis was performed first with a normality test using the Shapiro–Wilk test and later using Kruskal–Wallis for panel A and one-way ANOVA for panel B. Data are presented as mean ± SD. * p < 0.05, *** p < 0.001, **** p < 0.0001. LG CAF: Cancer-associated fibroblasts from low-grade lesion, HG CAF: Cancer-associated fibroblasts from high-grade lesion, Basal: Cells cultured with DMEM, CAF: Cells cultured with CAF’s supernatant without E2 stimulation, CAF E2: Cells cultured with CAF’s supernatant with previous stimulation with E2.

Article Snippet: CC cell lines SiHa and HeLa were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Migration, Wound Healing Assay, Cell Culture

Journal: Cancers

Article Title: Differential Protumoral Mechanisms Induced by CAFs in Cervical Cancer Cells Occur Independently of 17β-Estradiol Stimulation

doi: 10.3390/cancers18101509

Figure Lengend Snippet: CAFs do not affect apoptosis in CC cell lines. Apoptosis was assessed by Annexin V staining for phosphatidylserine exposure, and cell death by PI staining, with results expressed as percentages. ( A ) Apoptosis and death of SiHa cultured with low- and high-grade cervical cancer lesion CAFs’ supernatant. ( B ) apoptosis and death of HeLa cultured with low- and high-grade cervical cancer lesion CAFs’ supernatant. Statistical analysis was first performed with a normality test using the Shapiro–Wilk test, followed by Kruskal–Wallis or one-way ANOVA. Data is presented as mean ± SD. ns: Not significant. C-: Negative control, only cultured with DMEM, LG: Cancer-associated fibroblasts from low-grade lesion, LGE2: Cancer-associated fibroblasts from low-grade lesion with previous E2 stimulation, CIS: Cisplatin 80 μg/mL, LG CIS: Cancer-associated fibroblasts from low-grade lesion + cisplatin 80 μg/mL, LGE2 CIS: Cancer-associated fibroblasts from low-grade lesion with previous E2 stimulation + cisplatin 80 μg/mL, HG: Cancer-associated fibroblasts from high-grade lesion, HGE2: Cancer-associated fibroblasts from high-grade lesion with previous E2 stimulation, HG CIS: Cancer-associated fibroblasts from high-grade lesion + cisplatin 80 μg/mL, HGE2 CIS: Cancer-associated fibroblasts from high-grade lesion with previous E2 stimulation + cisplatin 80 μg/mL.

Article Snippet: CC cell lines SiHa and HeLa were obtained from the American Type Culture Collection (ATCC, Manassas, VA, USA).

Techniques: Staining, Cell Culture, Negative Control

Journal: Biology Direct

Article Title: FAM83D facilitates EMT and metastasis of cervical cancer via interaction with GSK3β and inactivation of GSK3β/stabilization of Snail signaling

doi: 10.1186/s13062-026-00761-z

Figure Lengend Snippet: FAM83D was associated with metastasis of CC. ( A ) Western blotting was employed to examine the expression of FAM83D in HeLa and SiHa cells with stable overexpression or knockdown of FAM83D. ( B-C ) Colony formation assay was conducted to evaluate the clonogenic ability of HeLa and SiHa cells following the overexpression or knockdown of FAM83D. ( D ) CCK-8 assay was used to detect cell viability in FAM83D-silenced HeLa and SiHa cells treated with different concentrations of 5-fluorouracil, cisplatin, oxaliplatin, and paclitaxel for 48 h. The inhibition rate was subsequently calculated. ( E-I ) Subcutaneous injection of FAM83D-silenced and control HeLa cells into BALB/c Nude mice ( n = 6) was used to determine the tumorigenicity. The tumor growth curves, tumor volumes, and tumor weights were measured. ( J ) And then, the nude mice that developed liver metastases were counted. ( K ) Representative images of liver metastases developed from FAM83D-silenced and control HeLa cells were shown, while ( L ) H&E staining of liver metastases in nude mice was presented. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Human CC cell lines SiHa and HeLa were purchased from the American Type Culture Collection (ATCC).

Techniques: Western Blot, Expressing, Over Expression, Knockdown, Colony Assay, CCK-8 Assay, Inhibition, Injection, Control, Staining

Journal: Biology Direct

Article Title: FAM83D facilitates EMT and metastasis of cervical cancer via interaction with GSK3β and inactivation of GSK3β/stabilization of Snail signaling

doi: 10.1186/s13062-026-00761-z

Figure Lengend Snippet: FAM83D promoted the metastasis of CC cells in vitro and vivo. ( A-D ) Wound healing assay was conducted to determine the migratory ability, and ( E-H ) transwell assay was utilized to determine the migratory and invasive ability, respectively, in HeLa and SiHa cells exhibiting stable overexpression or silencing of FAM83D. ( I ) Lung metastasis in nude mice ( n = 6) by tail vein injection of HeLa cells with or without silencing of FAM83D. ( J-K ) The statistical data of lung metastasis occurring in the mouse model. ( L ) Representative images and H&E staining of lung metastasis in nude mice. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Human CC cell lines SiHa and HeLa were purchased from the American Type Culture Collection (ATCC).

Techniques: In Vitro, Wound Healing Assay, Transwell Assay, Over Expression, Injection, Staining

Journal: Biology Direct

Article Title: FAM83D facilitates EMT and metastasis of cervical cancer via interaction with GSK3β and inactivation of GSK3β/stabilization of Snail signaling

doi: 10.1186/s13062-026-00761-z

Figure Lengend Snippet: FAM83D promoted GSK3β/Snail signaling and EMT. ( A-D ) Western blotting for E-cadherin, N-cadherin, Vimentin, and FAM83D in HeLa and SiHa cells following stable overexpression or silencing of FAM83D. ( E-H ) Western blotting for Snail, β-catenin, GSK3β, and phosphorylated GSK3β (Ser9), and FAM83D protein changes in CC cells with stable overexpression or silencing of FAM83D. ( I-J ) Cycloheximide (CHX) chase assays for Snail stability in CC cells with stable overexpression of FAM83D. ( K-L ) Ubiquitination assay for Snail protein ubiquitination level in CC cells with stable overexpression or silencing of FAM83D in the presence or absence of LiCl. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Human CC cell lines SiHa and HeLa were purchased from the American Type Culture Collection (ATCC).

Techniques: Western Blot, Over Expression, Ubiquitin Proteomics

Journal: Biology Direct

Article Title: FAM83D facilitates EMT and metastasis of cervical cancer via interaction with GSK3β and inactivation of GSK3β/stabilization of Snail signaling

doi: 10.1186/s13062-026-00761-z

Figure Lengend Snippet: FAM83D interacted with GSK3β. ( A ) The possible interaction between FAM83D and GSK3β was screened from the IntACT database. ( B-D ) Co-IP assay for interaction between FAM83D and GSK3β in HeLa and SiHa cells with or without silencing of FAM83D. ( E-F ) Co-IP assay for interaction between FAM83D and phosphorylated GSK3β (Ser9) in HeLa and SiHa cells. ( G ) In vitro cell-free proteins binding assay for validation of interaction specificity between FAM83D and GSK3β. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Human CC cell lines SiHa and HeLa were purchased from the American Type Culture Collection (ATCC).

Techniques: Co-Immunoprecipitation Assay, In Vitro, Binding Assay, Biomarker Discovery

Journal: Biology Direct

Article Title: FAM83D facilitates EMT and metastasis of cervical cancer via interaction with GSK3β and inactivation of GSK3β/stabilization of Snail signaling

doi: 10.1186/s13062-026-00761-z

Figure Lengend Snippet: The role of GSK3β/Snail axis in FAM83D-regulated EMT and metastasis of CC. ( A-D ) Western blotting for expressions of E-cadherin, N-cadherin, Vimentin, Snail, GSK3β, p-GSK3β (Ser9), and FAM83D in CC cells following stable FAM83D knockdown, as well as treated with LiCl (10 mM and 15 mM in HeLa and SiHa cells, respectively) for 24 h. ( E-H ) Following the treatment of HeLa cells with LiCl (10 mM) for 24 h and SiHa cells with LiCl (15 mM) for 24 h, a transwell assay was conducted to assess the migratory and invasive capabilities of HeLa and SiHa cells with stable FAM83D knockdown. The counts of migratory and invasive cells were recorded and subjected to statistical analysis. * p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Human CC cell lines SiHa and HeLa were purchased from the American Type Culture Collection (ATCC).

Techniques: Western Blot, Knockdown, Transwell Assay