Journal: Frontiers in pharmacology

Article Title: Hydrogen Sulfide Promotes Thyroid Hormone Synthesis and Secretion by Upregulating Sirtuin-1.

doi: 10.3389/fphar.2022.838248

Figure Lengend Snippet: FIGURE 1 | H2S generation and related enzymes in human thyroid. (A) H2S generation in human primary thyrocytes was measured by an in situ fluorescent H2S probe. (B) H2S levels in supernatants were detected by H2S-selective sensor in human primary thyrocytes with different culture days. (C) The enzymes that generated H2S were detected by immunohistochemistry staining in human thyroid tissues. All of them were located in the cytoplasm of thyrocytes. CBS (a), CSE (b), 3-MPST (c). (D) The enzymes that generated H2S in human primary thyrocytes were detected by real-time PCR. The level of CBS mRNA expression is the highest. *p < .05, **p < .01. CBS, cystathionine β-synthase; CSE, cystathionine γ-lyase; 3-MPST, 3-mercaptosulfurtransferase; H2S, hydrogen sulfide. Data are expressed as the mean ± SEM, and all experiments were performed independently three times.

Article Snippet: After blocking with 3% bovine serum albumin (BSA, Sigma–Aldrich), tissue slides were incubated with the following primary antibodies at 4°C overnight: CBS antibody (1:200), CSE antibody (1:25), and 3-MPST antibody (1:25) (all from Santa Cruz, California, United States).

Techniques: In Situ, Generated, Immunohistochemistry, Staining, Real-time Polymerase Chain Reaction, Expressing

Journal: PLoS ONE

Article Title: Serotonin and Dopamine Protect from Hypothermia/Rewarming Damage through the CBS/ H 2 S Pathway

doi: 10.1371/journal.pone.0022568

Figure Lengend Snippet: ( A and B ) Immunostaining demonstrating the expression of CBS in DDT-1 (A) and SMAC (B). Magnification 80×. ( C ) Downregulation of CBS by siRNA precludes protection of SMAC from hypothermic cell death by serotonin (30 µM), dopamine (20 µM) and PLP (50 µM). NaHS protects cells against hypothermia even in the presence of AOAA (1 mM) an inhibitor of CBS. The hypothermia protocol consisted of 24 h at 3°C for, followed by rewarming to 37°C for 3 h. Cell survival was assessed by adding MTS to the cells upon rewarming and spectrophotometrical formazon measurement. Inset: Silencing RNA substantially decreases the expression of CBS in SMAC cells. con: untreated cells, mock: negative control siRNA. ( D ) The increase in production of H 2 S as measured in cell medium of hypothermic cells (24 h at 3°C) treated with serotonin (30 µM) or dopamine (20 µM) was abrogated by pretreatment of the cells with the inhibitor of CBS, amino-oxyacetic acid (+AOAA, 1 mM, 15 min at 37°C+24 hr at 3°C ). ( E ) Treatment with serotonin (30 µM, 15 min at 37°C+24 hr at 3°C )) upregulates CBS expression, which is prevented by pretreatment with rapamycin (rap, 30 nM, 15 min at 37°C+24 hr at 3°C). Inset: western blot with time points as indicated. ( F ) Cooling SMAC induces the production of ROS in cells which is reduced by dopamine and serotonin treatment and aggravated by CBS siRNA transfection. ROS formation is measured by the level of Fluorescin fluorescence in SMAC in dopamine and serotonin treated cells compared to controls at 3°C and 37°C individually. Experiments consist of n≥3. Means ± SEM. # indicates significant difference to untreated control and * indicates significant difference to control within each treatment group.

Article Snippet: The expression of CBS in SMAC was reduced by applying a predesigned siRNA (sc-60336, Santa Cruz) and compared to a silencer negative control (Ambion, AM4644).

Techniques: Immunostaining, Expressing, Negative Control, Western Blot, Transfection, Fluorescence, Control