Journal: Food Science & Nutrition

Article Title: Luteolin Protects Against Acrylamide‐Induced Cellular Toxicity in Mouse Leydig Cells

doi: 10.1002/fsn3.70621

Figure Lengend Snippet: LUT improved Acr‐inhibited cell viability in Leydig cells. (A) Effects of cell viability Lut and/or Acr on Leydig cells; (B) Heat map showing the dose–response relationship of Lut and/or Acr according to % viability; (C) Lut and Acr synergy scoring (Bliss score) using minimal dose–response matrices. Data denote the mean ± SEM from three different experiments in triplicates ( n = 9). *Indicate statistically different values in treated cells compared to the control group (**** p < 0.0001) and # Statistically different values in groups versus the Acr‐alone group ( ### p < 0.001).

Article Snippet: The TM3 mouse Leydig cell line (Product code: CRL‐1714) was obtained from the American‐Type Culture Collection and transferred to our laboratory.

Techniques: Control

Journal: Food Science & Nutrition

Article Title: Luteolin Protects Against Acrylamide‐Induced Cellular Toxicity in Mouse Leydig Cells

doi: 10.1002/fsn3.70621

Figure Lengend Snippet: Evidence of membrane damage by LDH release in Leydig cells after treatment with Lut and/or Acr. Data denote the mean ± SEM from three different experiments in triplicates ( n = 9). *Statistically significant differences compared to control (*** p < 0.001) and # Statistically significant differences compared to Acr‐only treated groups ( ### p < 0.001).

Article Snippet: The TM3 mouse Leydig cell line (Product code: CRL‐1714) was obtained from the American‐Type Culture Collection and transferred to our laboratory.

Techniques: Membrane, Control

Journal: Food Science & Nutrition

Article Title: Luteolin Protects Against Acrylamide‐Induced Cellular Toxicity in Mouse Leydig Cells

doi: 10.1002/fsn3.70621

Figure Lengend Snippet: Effects of Lut and/or Acr on total ROS production in Leydig cells. (A) Representative images of Leydig cells labeled with a DCFH‐DA probe and visualized by fluorescence microscopy; (B) Total ROS levels determined by relative fluorescence intensity and expressed in arbitrary units. Experiments were repeated three times in duplicate. The values are means ± SEM ( n = 6). *Statistically significant differences compared to control (**** p < 0.0001) and # Statistically significant differences compared to Acr‐only treated groups ( ### p < 0.001).

Article Snippet: The TM3 mouse Leydig cell line (Product code: CRL‐1714) was obtained from the American‐Type Culture Collection and transferred to our laboratory.

Techniques: Labeling, Fluorescence, Microscopy, Control

Journal: Food Science & Nutrition

Article Title: Luteolin Protects Against Acrylamide‐Induced Cellular Toxicity in Mouse Leydig Cells

doi: 10.1002/fsn3.70621

Figure Lengend Snippet: Lut prevents Acr‐induced apoptosis. (A) Double‐fluorescent labeling images with Ho342 and PI dyes are presented to show apoptosis in Leydig cells after Lut and/or Acr treatments. Arrow: live cells, write arrowhead: apoptotic cells, empty arrowhead: dead cells. Scale bar: 100 μm. (B) Effects of Lut on the apoptotic cell rate induced by Acr. Experiments were repeated three times in duplicate. The values are means ± SEM ( n = 6). *Statistically significant differences compared to control (**** p < 0.0001) and # Statistically significant differences compared to Acr‐only treated groups ( ### p < 0.001).

Article Snippet: The TM3 mouse Leydig cell line (Product code: CRL‐1714) was obtained from the American‐Type Culture Collection and transferred to our laboratory.

Techniques: Labeling, Control

Journal: Food Science & Nutrition

Article Title: Luteolin Protects Against Acrylamide‐Induced Cellular Toxicity in Mouse Leydig Cells

doi: 10.1002/fsn3.70621

Figure Lengend Snippet: Effects of Lut and/or Acr on mRNA expression of apoptotic and antiapoptotic genes in Leydig cells. mRNA expression of (A) Trp53 , (B) Casp3 , (C) Bax , and (D) Bcl2 . Experiments were repeated three times in duplicate. The values are means ± SEM ( n = 6). *: Statistically significant differences compared to control (*** p < 0.001) and # statistically significant differences compared to Acr‐only treated groups ( # p < 0.05, ### p < 0.001).

Article Snippet: The TM3 mouse Leydig cell line (Product code: CRL‐1714) was obtained from the American‐Type Culture Collection and transferred to our laboratory.

Techniques: Expressing, Control

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Cordycepin and radiation induce morphological changes and reduce cell growth density in TM3 cell cultures. TM3 cells were treated (A–F) with 0, 10, 25, 50, 100, or 200 μM cordycepin for 24 h; (G–K) with 0, 2, 4, 6, or 8 Gy radiation for 24 h; or (L–P) with the combination of cordycepin and radiation under 0 μM cordycepin plus 0 Gy radiation, 10 μM cordycepin plus 2 Gy radiation, 25 μM cordycepin plus 4 Gy radiation, 50 μM cordycepin plus 6 Gy radiation, or 100 μM cordycepin plus 8 Gy radiation conditions for 24 h, respectively. Cell morphology and growth density were observed using phase contrast microscopy and images were captured by digital camera (scale bar, 100 μm). The experiment was repeated three times, and a representative result is presented.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Microscopy

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Cordycepin and radiation decrease cell viability of TM3 cells in a dose-dependent manner. TM3 cells were treated (A) with 0, 10, 25, 50, 100, and 200 μM cordycepin for 24 h, (B) with 0, 2, 4, 6, and 8 Gy radiation for 24 h, or (C) with the combination of cordycepin and radiation under 0 μM cordycepin plus 0 Gy radiation, 10 μM cordycepin plus 2 Gy radiation, 25 μM cordycepin plus 4 Gy radiation, 50 μM cordycepin plus 6 Gy radiation, and 100 μM cordycepin plus 8 Gy radiation conditions for 24 h, respectively. Cell viabilities were determined by Trypan blue exclusion assay. Data are presented as mean ± SEM of three independent experiments in percentages of cell viability relative to the control group. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001 indicate significant statistical difference compared to the control group. (D) Combination index (CI) was calculated by using CalcuSyn software. The dashed line represents the CI equal to 1. A CI of less than, equal to, and more than 1 indicates synergistic, additive, and antagonistic interaction of two treatments, respectively.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Trypan Blue Exclusion Assay, Control, Software

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Cordycepin and radiation regulate cell cycle distribution in TM3 cells. TM3 cells were treated without or with 50 μM cordycepin, 6 Gy radiation, or 50 μM cordycepin plus 6 Gy radiation for 3, 6, 9, 12, and 24 h, respectively. Cells were fixed and stained with propidium iodide, and cell cycle was measured using flow cytometry. The experiment was repeated three times, and (A) a representative result is presented. The horizontal axis represents DNA content, and the vertical axis shows the cell counts. (B–E) The distribution of cells in sub-G1, G1, S and G2/M phase of the TM3 cells treated without or with 50 μM cordycepin, 6 Gy radiation, or 50 μM cordycepin plus 6 Gy radiation at different time points are illustrated. (F–I) The curves depicting the changes in the percentage of cells over time in the Sub-G1, G1, S, and G2/M phases for different treatment groups are shown. Data are presented as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001 indicate significant statistical difference compared to the control group.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Staining, Flow Cytometry, Control

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Cordycepin and radiation induce autophagy in TM3 cells. TM3 cells were treated without or with 50 μM cordycepin, 6 Gy radiation, or 50 μM cordycepin plus 6 Gy radiation for 12 and 24 h, respectively, and stained with acridine orange (AO). The green and red fluorescence intensity of AO-stained cells were measured by flow cytometry. The experiment was repeated three times, and (A) the original density plots of a representative result are presented. The horizontal axis represents the green fluorescence intensity, and the vertical axis shows the red fluorescence intensity. The red numbers represent the percentage of autophagic cells calculated from the sum of the upper left and upper right quadrants. (B) The percentages of autophagic cells in different treatment groups from three independent experiments were calculated, and data are presented as mean ± SEM. *P < 0.05, **P < 0.01, and ****P < 0.0001 indicate significant statistical difference compared to the control group.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Staining, Fluorescence, Flow Cytometry, Control

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Autophagy inhibition aggravates cordycepin and radiation-induced cell death in TM3 cells. TM3 cells were preincubated without or with 10 or 50 μM chloroquine (CQ) for 1 h and then treated without or with 50 μM cordycepin, 6 Gy radiation, or 50 μM cordycepin plus 6 Gy radiation for additional 24 h. Cell viabilities were determined by Trypan blue exclusion assay. Data are presented as mean ± SEM of three independent experiments in percentages of cell viability relative to the group of control without CQ-preincubation. *P < 0.05 indicates significant statistical difference compared to the 0 μM CQ-preincubation group under individual treatment conditions. ##P < 0.01, and ####P < 0.0001 indicate significant statistical difference compared to the 0 μM CQ-preincubation, untreated control group.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Inhibition, Trypan Blue Exclusion Assay, Control

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Cordycepin and radiation regulate autophagy-related protein expressions in TM3 cells. TM3 cells were treated without or with 50 μM cordycepin, 6 Gy radiation, or 50 μM cordycepin plus 6 Gy radiation for 3, 6, 9, 12, and 24 h, respectively. The protein expression levels of LC3 (LC3 I: 16 kDa; LC3 II: 14 kDa), Atg5 (55 kDa), Atg12-Atg5 (55 kDa), Beclin-1 (60 kDa), and β-actin (43 kDa) were detected by western blotting. The experiment was repeated three times, and (A) a representative result is presented. (B) The integrated optical densities of LC3 II protein expression were normalized with corresponding LC3 I proteins expression in each lane. The integrated optical densities of (C) Atg5, (D) Atg12-Atg5, and (E) Beclin-1 protein expressions were normalized with corresponding β-actin protein expression in each lane. Data are presented as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001 indicate significant statistical difference compared to the control group.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Expressing, Western Blot, Control

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Cordycepin and radiation induce p62 protein accumulation in TM3 cells. TM3 cells were treated without or with 50 μM cordycepin, 6 Gy radiation, or 50 μM cordycepin plus 6 Gy radiation for 12, 24, and 48 h, respectively. The protein expression levels of p62 (62 kDa), and β-actin (43 kDa) were detected by western blotting. The experiment was repeated three times, and (A) a representative result is presented. The integrated optical densities of p62 protein expression were normalized with corresponding β-actin protein expression in each lane (B). Data are presented as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, and ***P < 0.001 indicate significant statistical difference compared to the control group.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Expressing, Western Blot, Control

Journal: Journal of Food and Drug Analysis

Article Title: Combination treatment of cordycepin and radiation induces apoptosis accompanied by protective autophagy in TM3 mouse Leydig progenitor cells

doi: 10.38212/2224-6614.3567

Figure Lengend Snippet: Cordycepin and radiation induce apoptosis in TM3 cells. TM3 cells were treated without or with 50 μM cordycepin, 6 Gy radiation, or 50 μM cordycepin plus 6 Gy radiation for 24, 48, and 72 h, respectively. Cells were stained with annexin-V-FITC and propidium iodide (PI). (A) The fractions of viable cells (annexin V-FITC −, PI −; lower left quadrant), necrotic cells (annexin V-FITC −, PI +; upper left quadrant), early apoptotic cells (annexin V-FITC +, PI −; lower right quadrant), and late apoptotic cells (annexin V-FITC +, PI +; upper right quadrant) were determined by flow cytometry. The experiment was repeated three times, and the original density plots of a representative result are presented. The horizontal axis represents the annexin V-FITC fluorescence intensity, and the vertical axis shows the PI fluorescence intensity. (B) Percentages of apoptotic cells are calculated, and data are presented as mean ± SEM of three independent experiments. *P < 0.05, **P < 0.01, and ****P < 0.0001 indicate significant statistical difference compared to the control group.

Article Snippet: TM3 mouse Leydig progenitor cell line (cat. no. CRL-1714) was purchased from ATCC (Manassas, VA, USA).

Techniques: Staining, Flow Cytometry, Fluorescence, Control