Journal: Endocrinology and Metabolism

Article Title: Fancd2os Reduces Testosterone Production by Inhibiting Steroidogenic Enzymes and Promoting Cellular Apoptosis in Murine Testicular Leydig Cells

doi: 10.3803/enm.2022.1431

Figure Lengend Snippet: Fig. 4. Fancd2 opposite-strand (Fancd2os) reduces testosterone production and steroidogenic enzyme expression in TM3 cells. The testoster one levels in Fancd2os-overexpressing (A) or knockdown TM3 cells (B) were detected by enzyme-linked immunosorbent assays (n=3 per group). (C, D, E) Relative quantities of mRNA expression of steroidogenic acute regulatory protein (StAR), P450 cholesterol side-chain cleavage (P450scc), and 3β-hydroxysteroid dehydrogenase (3β-HSD) in Fancd2os-overexpressing TM3 cells or Fancd2os knockdown TM3 cells (F, G, H) were determined real-time polymerase chain reaction using β-actin as a housekeeping gene. Each bar represents the mean± standard deviation from three separate experiments. Significant difference compared to TM3, vector/TM3 or NC/TM3. aP<0.05; bP<0.01.

Article Snippet: The testosterone concentration of both serum and cell supernatants was measured using a testosterone ELISA Kit (Elabscience, Houston, TX, USA) according to the manufacturer’s protocol.

Techniques: Expressing, Knockdown, Real-time Polymerase Chain Reaction, Standard Deviation, Plasmid Preparation

Journal: Endocrinology and Metabolism

Article Title: Fancd2os Reduces Testosterone Production by Inhibiting Steroidogenic Enzymes and Promoting Cellular Apoptosis in Murine Testicular Leydig Cells

doi: 10.3803/enm.2022.1431

Figure Lengend Snippet: Fig. 6. Higher Fancd2 opposite-strand (Fancd2os) levels in older mouse Leydig cells result in cellular apoptosis and lower serum testosterone production. (A) The serum testosterone levels from mice of different ages were measured using enzyme-linked immunosorbent assays. (B, C) The testis tissues from different aged mice were sliced and then Fancd2os protein expression and apoptosis were analyzed using immuno chemistry and the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) assay, respectively. ST represents seminiferous tubule. Black arrows and red arrows indicate Fancd2os-positive cells and TUNEL-positive cells, respectively. Scale bar repre sents 25 μm. The results are given as the mean±standard deviation (n=3). (D) The correlations between Fancd2os expression (B) and the TUNEL-positive staining rate (C) were analyzed using Pearson correlation coefficients. An r≥0.5 was considered to indicate a strong correla tion, and P<0.05 was considered statistically significant. aP<0.05 compared with juvenile mice; bP<0.05 compared with young mice; cP<0.05 compared with middle-aged mice.

Article Snippet: The testosterone concentration of both serum and cell supernatants was measured using a testosterone ELISA Kit (Elabscience, Houston, TX, USA) according to the manufacturer’s protocol.

Techniques: Expressing, End Labeling, TUNEL Assay, Standard Deviation, Staining

Journal: Ecotoxicology and environmental safety

Article Title: Ameliorative effect of betulinic acid against zearalenone exposure triggers testicular dysfunction and oxidative stress in mice via p38/ERK MAPK inhibition and Nrf2-mediated antioxidant defense activation.

doi: 10.1016/j.ecoenv.2022.113561

Figure Lengend Snippet: Fig. 2. BA protected the aggravation of male reproduction injury in ZEA-induced mice. The morphology of the sperm was photographed using an optical microscope (A). The sperm motility was used to evaluate the male repro duction of mice, including sperm survival rate (B), sperm malformation rate (C), and sperm mortality rate (D). Protein and mRNA levels of ERα in testis were detected by immunoblotting and RT-PCR analysis, respectively, and the pro tein and mRNA levels were normalized to β-actin (E-G). The content of testosterone in serum was detected by ELISA kit (H). The mRNA expression of CLDN11 (I), CDH2 (J), and Vim (K) were measured by RT-PCR. Mean ± SEM, *P < 0.05 and **P < 0.01 represented a significant differ ence compared to the control group, while #P < 0.05 and ##P < 0.01 represented a signifi cant difference compared to the ZEA group.

Article Snippet: Testosterone enzyme linked immunosorbent assay (ELISA) kit (CSB-E05101m) was obtained from Cusabio Biotech Co. Ltd. (Wuhan, China).

Techniques: Microscopy, Western Blot, Reverse Transcription Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay, Expressing, Control

Journal: Molecular Medicine

Article Title: MicroRNA regulation of the proliferation and apoptosis of Leydig cells in diabetes

doi: 10.1186/s10020-021-00370-8

Figure Lengend Snippet: Effects of diabetes on testicular function and apoptosis. Eight weeks after diabetes was established, the right testis of each rat was removed and separately photographed ( A ) and the testis index {(testis weight/body weight) × 100%} was calculated ( B ). Concentrations of serum ( C ) and testicular ( D ) testosterone detected by ELISA in each group. Representative hematoxylin & eosin (H&E) and TUNEL staining of rat testicular tissues from ND (first 2 panels) and DM (last 2 panels) groups. For a better comparison, the second panel in each group is a partially enlarged panel (black box) of the first panel. Scale bar = 100 μm (first panel) and 40 μm (second panel) ( E ). Data are presented as mean ± SD.*p < 0.05 **p < 0.01 compared with the ND group

Article Snippet: Total testosterone was measured using the Rat or Human Testosterone ELISA kit (Cusabio, Wuhan, China) according to the manufacturer’s instructions.

Techniques: Enzyme-linked Immunosorbent Assay, TUNEL Assay, Staining, Comparison

Journal: Molecular Medicine

Article Title: MicroRNA regulation of the proliferation and apoptosis of Leydig cells in diabetes

doi: 10.1186/s10020-021-00370-8

Figure Lengend Snippet: Effect of glucose concentration on miRNAs and apoptosis. Expression of miR-504 ( A ) and miR-935 ( B ) in R2C cells at 24 h after culturing in a glucose concentration gradient (basal glucose for R2C cell was 5 mM and stimulated concentrations were 15 mM and 30 mM). Data were normalised to U6 RNA, used as an internal control. Expression of MEK5 ( C ) and MEF2C ( D ) determined using RT-qPCR analysis. β-actin was used as an internal control. Representative immunoblotting ( E ) and cumulative quantification of the protein levels of MEK5 ( F ) and MEF2C ( G ) in R2C cells. Media were collected and assayed for concentration of testosterone using ELISA ( H ). Detection of apoptotic cells using FACS analysis with FITC-labelled annexin V and PI staining ( I ). Bar graphs represent the percentage of apoptotic cells in each group ( J ). *p < 0.05, **p < 0.01, ***p < 0.001. n = 3

Article Snippet: Total testosterone was measured using the Rat or Human Testosterone ELISA kit (Cusabio, Wuhan, China) according to the manufacturer’s instructions.

Techniques: Concentration Assay, Expressing, Control, Quantitative RT-PCR, Western Blot, Enzyme-linked Immunosorbent Assay, Staining

Journal: Molecular Medicine

Article Title: MicroRNA regulation of the proliferation and apoptosis of Leydig cells in diabetes

doi: 10.1186/s10020-021-00370-8

Figure Lengend Snippet: Modulation of proliferation and apoptosis of Leydig cells by mRNA targets of miR-504. Expression of miR-504 in miR-504 mimic-or miR-504 inhibitor-infected R2C cells at 24 h after culturing in normal or high glucose (HG). Data were normalised to U6 RNA, used as an internal control ( A ). Expression of MEK5 and MEF2C determined by RT-qPCR analysis. β-actin was used as an internal control ( B , C ). Representative immunoblotting ( D ) and cumulative quantification ( E , F ) of the protein levels of MEK5 and MEF2C in R2C cells transfected with miR-504 mimic, miR-504 inhibitor, mimic NC, or inhibitor NC. Media were collected and assayed for concentration of testosterone using ELISA ( G ). Cell proliferation was assayed using CCK8 ( H ). Detection of apoptotic cells by FACS analysis with FITC-labelled annexin V and PI staining ( I ). Bar graphs represent the percentage of apoptotic cells in each group (J). *p < 0.05, **p < 0.01, ***p < 0.001. n = 3

Article Snippet: Total testosterone was measured using the Rat or Human Testosterone ELISA kit (Cusabio, Wuhan, China) according to the manufacturer’s instructions.

Techniques: Expressing, Infection, Control, Quantitative RT-PCR, Western Blot, Transfection, Concentration Assay, Enzyme-linked Immunosorbent Assay, Staining

Journal: Molecular Medicine

Article Title: MicroRNA regulation of the proliferation and apoptosis of Leydig cells in diabetes

doi: 10.1186/s10020-021-00370-8

Figure Lengend Snippet: Modulation of proliferation and apoptosis of Leydig cells by mRNA targets of miR-935. Expression of miR-935 in miR-935 mimic-or miR-935 inhibitor-infected R2C cells at 24 h after culturing in normal or high glucose (HG). Data were normalised to U6 RNA used as an internal control ( A ). Expression of MEF2C determined by RT-qPCR analysis. β-actin was used as an internal control ( B ). Representative immunoblotting ( C ) and cumulative quantification ( D ) of the protein levels of MEF2C in R2C cells transfected with miR-935 mimic, miR-935 inhibitor, mimic NC, or inhibitor NC. Media were collected and assayed for concentration of testosterone using ELISA ( E ). Cell proliferation was assayed using CCK8 ( F ). Detection of apoptotic cells by FACS analysis with FITC-labelled annexin V and PI staining ( G ). Bar graphs represent the percentage of apoptotic cells in each group (H). *p < 0.05, **p < 0.01, ***p < 0.001. n = 3

Article Snippet: Total testosterone was measured using the Rat or Human Testosterone ELISA kit (Cusabio, Wuhan, China) according to the manufacturer’s instructions.

Techniques: Expressing, Infection, Control, Quantitative RT-PCR, Western Blot, Transfection, Concentration Assay, Enzyme-linked Immunosorbent Assay, Staining

Journal: Sexual Medicine

Article Title: Icariin inhibits hyperglycemia-induced cell death in penile cavernous tissue and improves erectile function in type 1 diabetic rats

doi: 10.1093/sexmed/qfaf017

Figure Lengend Snippet: Effect of icariin on oxidative stress in the penile cavernous tissue of DM rats. (A) MDA levels, (B) GSH levels, (C) the GSH/GSSG ratio, and (D) SOD activity in each group were measured via ELISA. (E) Semiquantitative analysis and (G) representative images (40×) of ROS in each group. (F) Semiquantitative analysis and (H) representative images (15×) of Prussian blue staining showing nucleus (red areas) and iron deposition (blue areas). And P < .05 vs the control group. * P < .05 vs the control + ICA group. # P < .05 vs the DM group.

Article Snippet: Serum testosterone was detected according to the instructions of the rat serum testosterone ELISA kit (Elabscience Biotechnology, Wuhan, China; E-OSEL-R0003).

Techniques: Activity Assay, Enzyme-linked Immunosorbent Assay, Staining, Control

Journal: Sexual Medicine

Article Title: Icariin inhibits hyperglycemia-induced cell death in penile cavernous tissue and improves erectile function in type 1 diabetic rats

doi: 10.1093/sexmed/qfaf017

Figure Lengend Snippet: Effect of icariin on the fibrosis of penile cavernous tissue in DM rats. (A) Representative images (15×) of Masson's trichrome staining showing smooth muscle cells (SMCs) and collagen fibers. (B) Semiquantitative analysis of the SM/C ratio of each group. (C) NO levels in each group were measured using ELISA. (D) Western blot analysis of the expression of eNOS and peNOS in penile cavernous tissue from each group. (E, F) Semiquantitative analysis of eNOS and p-eNOS levels and the p-eNOS/eNOS ratio in penile cavernous tissue from each group. And P < .05 vs the control group. * P < .05 vs the control + ICA group. # P < .05 vs the DM group.

Article Snippet: Serum testosterone was detected according to the instructions of the rat serum testosterone ELISA kit (Elabscience Biotechnology, Wuhan, China; E-OSEL-R0003).

Techniques: Staining, Enzyme-linked Immunosorbent Assay, Western Blot, Expressing, Control