Journal: International Journal of Nanomedicine

Article Title: Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

doi: 10.2147/IJN.S76062

Figure Lengend Snippet: Effect of AgNPs on the viability of TM3 and TM4 cells. Notes: TM3 and TM4 cells were treated with various concentrations of 10 nm ( A ) and 20 nm ( B ) AgNPs for 24 hours, and cytotoxicity was determined using the CCK-8 (top panel) and MTS (bottom panel) assays. The data are expressed as the mean ± SD of three independent experiments performed in triplicate; ** P <0.01. Abbreviations: AgNPs, silver nanoparticles; CCK, cell counting kit-8; IC 50 , half maximal inhibitory concentration; MTS, 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt; SD, standard deviation; TM3, Leydig; TM4, Sertoli.

Article Snippet: TM3 (KCLB No 21714) and TM4 (KCLB No 21715) cell lines were obtained from Korean cell line bank (Seoul, South Korea).

Techniques: CCK-8 Assay, Cell Counting, Concentration Assay, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

doi: 10.2147/IJN.S76062

Figure Lengend Snippet: Effect of AgNPs on LDH activity and ROS generation in TM3 and TM4 cells. Notes: TM3 and TM4 cells were treated with two different sizes of AgNPs: 10 nm ( A ) and 20 nm ( B ); LDH activity was assessed in the supernatants of cells treated or not with AgNPs and NAC using the LDH cytotoxicity detection kit. The y axis represents the absorbance as percentage of the controls, and the x axis shows IC 50 of AgNPs; ( C ) ROS generation in AgNPs (IC 50 )-treated TM3 and TM4 cells; DCF formation from H2DCF-DA was measured by fluorescence (excitation at 480 nm, emission at 530 nm). The data are expressed as the mean ± SD of three independent experiments performed in triplicate; ** P <0.01. Abbreviations: AgNPs, silver nanoparticles; DCF, 2′,7′-dichlorofluorescein; DMSO, dimethyl sulfoxide; H2DCF-DA, 2,7-dichlorodihydrofluorecein diacetate; H 2 O 2 , hydrogen peroxide; IC 50 , half maximal inhibitory concentration; LDH, lactate dehydrogenase; NAC, N-acetyl L-cysteine; ROS, reactive oxygen species; SD, standard deviation; TM3, Leydig; TM4, Sertoli.

Article Snippet: TM3 (KCLB No 21714) and TM4 (KCLB No 21715) cell lines were obtained from Korean cell line bank (Seoul, South Korea).

Techniques: Activity Assay, Fluorescence, Concentration Assay, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

doi: 10.2147/IJN.S76062

Figure Lengend Snippet: Cellular uptake of AgNPs induces accumulation of autophagosomes and autolysosomes. Notes: TM3 and TM4 cells were treated with AgNPs at IC 50 for 24 hours and analyzed by transmission electron microscopy (TEM). TEM images showed TM3 ( A – C , G – J ) and TM4 ( D , F , K – Q ) cells: not treated with AgNPs ( A , D ) and AgNP-treated showing crescent-like vacuoles ( B , E ), autophagic vacuoles ( C , F ), and accumulation of autophagosomes, autolysosomes, and damaged mitochondria ( G – J and K – Q ). Red arrows indicate the formation of autolysosomes. Autophagic vacuoles (AV) were classified into three types based on morphological criteria: AV-initial (AVi), intermediated (Avi/d), and degradative (Avd). All experiments were carried out in triplicate and repeated at least three times. Abbreviations: AgNPs, silver nanoparticles; Cy, cytoplasm; IC 50 , half maximal inhibitory concentration; N, nucleus; TM3, Leydig; TM4, Sertoli; TAV, typical autophagic vacuoles; DM, damaged mitochondria; MVB, multivesicular bodies.

Article Snippet: TM3 (KCLB No 21714) and TM4 (KCLB No 21715) cell lines were obtained from Korean cell line bank (Seoul, South Korea).

Techniques: Transmission Assay, Electron Microscopy, Concentration Assay

Journal: International Journal of Nanomedicine

Article Title: Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

doi: 10.2147/IJN.S76062

Figure Lengend Snippet: AgNPs accelerate expression of autophagy genes. Notes: TM3 ( A ) and TM4 ( B ) cells were treated with AgNPs for 24 hours and analyzed for the expression of autophagy-related genes; for each treatment condition and time point, mRNA expression of AgNP-treated cells was normalized to that of untreated cells. The data are expressed as the mean relative gene expression ± SD of three independent experiments performed in triplicate; ** P <0.01. Abbreviations: AgNPs, silver nanoparticles; mRNA, messenger RNA; SD, standard deviation; TM3, Leydig; TM4, Sertoli.

Article Snippet: TM3 (KCLB No 21714) and TM4 (KCLB No 21715) cell lines were obtained from Korean cell line bank (Seoul, South Korea).

Techniques: Expressing, Gene Expression, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

doi: 10.2147/IJN.S76062

Figure Lengend Snippet: AgNPs promote ROS-induced apoptosis. Notes: TM3 and TM4 cells were pretreated with 10 mM NAC for 1 hour and then treated with AgNPs for 6 hours ( A , B ); TM3 ( A ) and TM4 ( B ) cells were stained with Annexin V (x axis) and PI (y axis) and analyzed for cell death by flow cytometry using a FACSCanto TMII; the percentages of apoptotic cells are shown in each quadrant ( C , D ); TM3 ( C ) and TM4 ( D ) cells were exposed to the respective IC 50 of AgNPs for 6 hours and analyzed for protein expression by western blotting using the indicated antibodies; ( E , F ) protein expression relative to β-actin used as the loading control; ( G , H ) relative mRNA expression levels of apoptotic genes analyzed by RT-PCR. The data are expressed as the mean relative gene expression ± SD of three independent experiments performed in triplicate; * P <0.05, ** P <0.01. Abbreviations: AgNPs, silver nanoparticles; IC 50 , half maximal inhibitory concentration; mRNA, messenger RNA; NAC, N-acetyl L-cysteine; PI, propidium iodide; ROS, reactive oxygen species; RT-PCR, real-time polymerase chain reaction; SD, standard deviation; TM3, Leydig; TM4, Sertoli.

Article Snippet: TM3 (KCLB No 21714) and TM4 (KCLB No 21715) cell lines were obtained from Korean cell line bank (Seoul, South Korea).

Techniques: Staining, Flow Cytometry, Expressing, Western Blot, Control, Reverse Transcription Polymerase Chain Reaction, Gene Expression, Concentration Assay, Real-time Polymerase Chain Reaction, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

doi: 10.2147/IJN.S76062

Figure Lengend Snippet: Effect of AgNPs on gene expression in TM3 and TM4 cells. Notes: ( A ) Quantitative (q) RT-PCR for testosterone biosynthesis-related genes in TM3 cells; ( B ) qRT-PCR for the BTB genes Gdnf , Tgfβ1 , Ar , and Amh in TM4 cells. The expression of all genes was normalized to that of Gapdh . The data are expressed as the mean relative gene expression ± SD of three independent experiments performed in triplicate; * P <0.05, ** P <0.01. Abbreviations: AgNPs, silver nanoparticles; BTB, blood–testes barrier; mRNA, messenger RNA; RT-PCR, real-time polymerase chain reaction; SD, standard deviation; TM3, Leydig; TM4, Sertoli; qRT-PCR, quantitative RT-PCR.

Article Snippet: TM3 (KCLB No 21714) and TM4 (KCLB No 21715) cell lines were obtained from Korean cell line bank (Seoul, South Korea).

Techniques: Gene Expression, Reverse Transcription Polymerase Chain Reaction, Quantitative RT-PCR, Expressing, Real-time Polymerase Chain Reaction, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Differential nanoreprotoxicity of silver nanoparticles in male somatic cells and spermatogonial stem cells

doi: 10.2147/IJN.S76062

Figure Lengend Snippet: Effects of AgNPs on the proliferation and differentiation of SSCs. Notes: ( A ) FACS analysis of SSCs using CD49f antibody; ( B ) the percentage of SSCs cultured on TM3 or TM4 as feeder cells; ( C ) expression of the genes involved in meiosis, self-renewal, and differentiation assessed by quantitative RT-PCR and normalized to that of Gapdh. Incubation of SSCs on TM3 and TM4 cells treated with AgNP (10 μg/mL) caused a significant decrease in mRNA levels of the analyzed genes. The results are presented as mean ± SEM of triplicate measurements; ** P <0.01. Abbreviations: AgNPs, silver nanoparticles; FACS, fluorescence-activated cell sorting; RT-PCR, real-time polymerase chain reaction; SEM, standard error of the mean; SSCs, spermatogonial stem cells; TM3, Leydig; TM4, Sertoli; FSC-H, forward scatter-height.

Article Snippet: TM3 (KCLB No 21714) and TM4 (KCLB No 21715) cell lines were obtained from Korean cell line bank (Seoul, South Korea).

Techniques: Cell Culture, Expressing, Quantitative RT-PCR, Incubation, Fluorescence, FACS, Reverse Transcription Polymerase Chain Reaction, Real-time Polymerase Chain Reaction

Journal: International Journal of Environmental Research and Public Health

Article Title: Effects and Mechanisms of Phthalates’ Action on Reproductive Processes and Reproductive Health: A Literature Review

doi: 10.3390/ijerph17186811

Figure Lengend Snippet: The function of some genes regulating male and female reproductive system development [ 242 ].

Article Snippet: In vitro studies demonstrated that DEHP exposure at 40, 80 and 160 μM and dibutyl phthalate (DBP) exposure at 10 and 100 mg/L caused the apoptosis of TM3 Leydig cells and Sertoli cells of Male Sprague-Dawley rats, respectively [ , ].

Techniques: Expressing, Inhibition