Journal: Frontiers in Cell and Developmental Biology

Article Title: Melatonin and agomelatine alleviate ivermectin-induced mouse spermatogonia apoptosis via suppression of oxidative stress and calcium overload

doi: 10.3389/fcell.2025.1713124

Figure Lengend Snippet: Effects of melatonin and agomelatine on proliferation loss in GC-1 spermatogonia (spg) treated with ivermectin (IVM) for 24 h (A) Chemical structure of IVM isomers. (B) Chemical structures of melatonin and its analogs, agomelatine and pinoline. (C) Schematic overview of the cell culture and experimental design. (D) Brightfield microscopy images showing cytoplasmic morphology. (E) Immunofluorescence images of Ki67 nuclear translocation. (F) Quantification of relative cell proliferation (n = 3). (G) Quantification of Ki67-positive nuclei (n = 3) in GC-1 spg treated with either control, melatonin, agomelatine, or pinoline with or without 16 μM IVM. IVM treatment conditions are denoted as positive (+) or negative (−). Scale bars: 100 µm. Data are presented as means ± SEM. Significant differences are indicated by different letters (a–d) at p < 0.05.

Article Snippet: Mouse type B GC-1 spermatogonial (spg) cells (CRL-2053, American Type Culture Collection, Manassas, VA, USA) were cultured in complete media using Dulbecco’s modified Eagle’s medium (DMEM, L0103-500; Biowest, Nuaillé, France) with 10% fetal bovine serum (FBS, S1480; Biowest) and penicillin/streptomycin (15140122, Gibco, Waltham, MA, USA) in 5% CO 2 at 37 °C.

Techniques: Cell Culture, Microscopy, Immunofluorescence, Translocation Assay, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Melatonin and agomelatine alleviate ivermectin-induced mouse spermatogonia apoptosis via suppression of oxidative stress and calcium overload

doi: 10.3389/fcell.2025.1713124

Figure Lengend Snippet: Inhibition of intracellular reactive oxygen species (ROS) generation by melatonin and agomelatine in GC-1 spermatogonia (spg) treated with ivermectin (IVM) for 3 h (A) Fluorescence microscopy images of GC-1 spg labeled with DCFDA. (B) Semi-quantification of DCFDA fluorescence intensity in GC-1 spg treated with either control, melatonin, agomelatine, or pinoline with or without 16 μM IVM (n = 5). IVM treatment conditions are denoted as positive (+) or negative (−). Scale bars: 100 µm. Data are shown as means ± SEM. Significant differences are indicated by different letters (a–d) at p < 0.05. Mel, melatonin; Ago, agomelatine; Pin, pinoline.

Article Snippet: Mouse type B GC-1 spermatogonial (spg) cells (CRL-2053, American Type Culture Collection, Manassas, VA, USA) were cultured in complete media using Dulbecco’s modified Eagle’s medium (DMEM, L0103-500; Biowest, Nuaillé, France) with 10% fetal bovine serum (FBS, S1480; Biowest) and penicillin/streptomycin (15140122, Gibco, Waltham, MA, USA) in 5% CO 2 at 37 °C.

Techniques: Inhibition, Fluorescence, Microscopy, Labeling, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Melatonin and agomelatine alleviate ivermectin-induced mouse spermatogonia apoptosis via suppression of oxidative stress and calcium overload

doi: 10.3389/fcell.2025.1713124

Figure Lengend Snippet: Melatonin and agomelatine inhibit cytoplasmic Ca 2+ accumulation in GC-1 spermatogonia (spg) treated with ivermectin (IVM) for 3 h (A) Fluorescence microscopy images of GC-1 spg cells labeled with Fluo-4, AM. (B) Semi-quantification of Fluo-4, AM fluorescence intensity in GC-1 spg treated with either control, melatonin, agomelatine, or pinoline with or without 16 μM IVM (n = 5). IVM treatment conditions are denoted as positive (+) or negative (−). Scale bars: 100 µm. Data are presented as means ± SEM. Significant differences are denoted by different letters (a–c) at p < 0.05. Mel, melatonin; Ago, agomelatine; Pin, pinoline.

Article Snippet: Mouse type B GC-1 spermatogonial (spg) cells (CRL-2053, American Type Culture Collection, Manassas, VA, USA) were cultured in complete media using Dulbecco’s modified Eagle’s medium (DMEM, L0103-500; Biowest, Nuaillé, France) with 10% fetal bovine serum (FBS, S1480; Biowest) and penicillin/streptomycin (15140122, Gibco, Waltham, MA, USA) in 5% CO 2 at 37 °C.

Techniques: Fluorescence, Microscopy, Labeling, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Melatonin and agomelatine alleviate ivermectin-induced mouse spermatogonia apoptosis via suppression of oxidative stress and calcium overload

doi: 10.3389/fcell.2025.1713124

Figure Lengend Snippet: Melatonin’s and agomelatine’s recovery of ΔΨm and mitochondrial mass in GC-1 spermatogonia (spg) treated with 16 μM ivermectin (IVM) for 3 h (A) Fluorescence microscopy images of GC-1 spg stained with tetramethylrhodamine ethyl ester (TMRE) and/or MitoTracker and Hoechst 33,342. (B) Semi-quantification of MitoTracker fluorescence intensity (n = 5), and (C) ratio of TMRE to MitoTracker fluorescence intensity in GC-1 spg treated with either control, melatonin, agomelatine, or pinoline with or without 16 μM IVM (n = 5). Scale bars: 100 µm. IVM treatment conditions are denoted as positive (+) or negative (−). Data are shown as means ± SEM. Significant differences are denoted by different letters (a–c) at p < 0.05. Mel, melatonin; Ago, agomelatine; Pin, pinoline

Article Snippet: Mouse type B GC-1 spermatogonial (spg) cells (CRL-2053, American Type Culture Collection, Manassas, VA, USA) were cultured in complete media using Dulbecco’s modified Eagle’s medium (DMEM, L0103-500; Biowest, Nuaillé, France) with 10% fetal bovine serum (FBS, S1480; Biowest) and penicillin/streptomycin (15140122, Gibco, Waltham, MA, USA) in 5% CO 2 at 37 °C.

Techniques: Fluorescence, Microscopy, Staining, Control

Journal: Frontiers in Cell and Developmental Biology

Article Title: Melatonin and agomelatine alleviate ivermectin-induced mouse spermatogonia apoptosis via suppression of oxidative stress and calcium overload

doi: 10.3389/fcell.2025.1713124

Figure Lengend Snippet: Seahorse assay of GC-1 spermatogonia (spg) treated with or without melatonin analogs and/or ivermectin (IVM) for 3 h (A) Oxygen consumption rate (OCR) plot following sequential injections of oligomycin, FCCP (carbonyl cyanide-p-trifluoromethoxyphenylhydrazone), and rotenone/antimycin A (R/AA). (B) Basal respiration, (C) ATP-linked respiration, (D) maximal respiration capacity, and (E) reserve capacity calculated from OCR data (n = 3). Data are presented as means ± SEM. Significant differences are indicated by different letters (a–d) at p < 0.05. Mel, melatonin; Ago, agomelatine; Pin, pinoline.

Article Snippet: Mouse type B GC-1 spermatogonial (spg) cells (CRL-2053, American Type Culture Collection, Manassas, VA, USA) were cultured in complete media using Dulbecco’s modified Eagle’s medium (DMEM, L0103-500; Biowest, Nuaillé, France) with 10% fetal bovine serum (FBS, S1480; Biowest) and penicillin/streptomycin (15140122, Gibco, Waltham, MA, USA) in 5% CO 2 at 37 °C.

Techniques:

Journal: Frontiers in Cell and Developmental Biology

Article Title: Melatonin and agomelatine alleviate ivermectin-induced mouse spermatogonia apoptosis via suppression of oxidative stress and calcium overload

doi: 10.3389/fcell.2025.1713124

Figure Lengend Snippet: Attenuation of apoptosis by melatonin and agomelatine in GC-1 spermatogonia (spg) treated with ivermectin (IVM) for 3 h (A) Brightfield microscopy images showing morphological changes in GC-1 spg following treatment with IVM and/or melatonin analogs. Scale bars: 100 µm. (B) Western blot analysis of apoptosis-related proteins, including cleaved caspases, BCL-2, BAX, Cytochrome c, and α-tubulin (used as a loading control), under the indicated treatment conditions. Mel, melatonin; Ago, agomelatine; Pin, pinoline (C) Quantification of apoptosis-related proteins (n = 3). Data are shown as means ± SEM. Significant differences are denoted by different letters (a–c) at p < 0.05. Mel, melatonin; Ago, agomelatine; Pin, pinoline.

Article Snippet: Mouse type B GC-1 spermatogonial (spg) cells (CRL-2053, American Type Culture Collection, Manassas, VA, USA) were cultured in complete media using Dulbecco’s modified Eagle’s medium (DMEM, L0103-500; Biowest, Nuaillé, France) with 10% fetal bovine serum (FBS, S1480; Biowest) and penicillin/streptomycin (15140122, Gibco, Waltham, MA, USA) in 5% CO 2 at 37 °C.

Techniques: Microscopy, Western Blot, Control