3-TYP (3-(1H-1,2,3-triazol-4-yl) pyridine) is a selective SIRT3 inhibitor.

SIRT1:88 nM, SIRT3:16 nM, SIRT2:92 nM

3-TYP显著减弱了在Cd暴露的HepG2细胞中，褪黑激素诱导的去乙酰化SOD2表达和SOD2活性的增加。3-TYP抑制了褪黑激素增强的SIRT3活性，但并不影响SIRT3蛋白表达。3-TYP预处理逆转了褪黑激素对镉(Cd)引起的线粒体来源的O2•−产生和自噬细胞死亡的保护效果。

3-TYP显著降低SIRT3活性，增加了SOD2的乙酰化水平，但不影响SIRT3的表达。3-TYP通过减少LVEF和LVFS，降低了褪黑激素的心脏保护效应，此效应在24小时再灌注后显著。另外，与IR+Mel组相比，3-TYP增加了梗塞面积、血清LDH水平和凋亡比率。3-TYP（50 mg/kg, i.p.）与Sham组相比，对LVEF、LVFS、梗塞面积、血清LDH水平、凋亡及氧化应激的影响不明显。此外，与Sham组相比，3-TYP对gp91phox、Nrf2、NQO 1、Bax、Bcl-2、Caspase-3及活化Caspase-3的表达水平影响甚微。

Cell viability is analyzed using Cell Counting Kit-8. 1×104 cells are inoculated into 96-well plates. After being treated, 90 μL of medium and 10 μL of CCK-8 solution are added to each well. The cells are then incubated at 37°C for 2 h. After incubation, the absorption at 450 nm is measured using an Infinite? M200 Microplate Reader. The results are expressed as a percentage of the control. The cell death is also evaluated using the trypan blue assay. HepG2 cells are plated in the 6-well plates (5×105 cells per well) and incubated for 24 h. After being treated with Cd or melatonin, the cells are detached with 300 μL trypsin-EDTA solution. The mixture of detached cells is centrifugated at 300 g for 5 min. Then, the residue is combined with 800 μL trypan blue solution and dispersed. After 3 min staining, cells are counted using an automated cell counter. The dead cells are stained with the blue color. Cell mortality (%) is expressed as percentage of the dead cell number/the total cell number.

3-TYP is formulated in 1% ethanol.Male C57BL/6 mice are anesthetized with 2% isoflurane, and myocardial ischemia is produced by temporarily exteriorizing the heart via a left thoracic incision and placing a 6-0 silk suture slipknot around the left anterior descending coronary artery. After 30 minutes of myocardial ischemia, the slipknot is released, and the myocardium is reperfused for 3 hour (for western blot analysis and oxidative stress measurement) or 24 hour (for cardiac function, apoptotic index and infarct size determination). Sham-operated mice undergo the same surgical procedures except the suture placed under the left coronary artery is not tied. Ten minutes before reperfusion, mice are randomized to receive either vehicle (1% ethanol) or melatonin (20 mg/kg) by intraperitoneal injection. C57BL/6 mice are randomly divided into the following groups: (i) Sham group: mice underwent the sham operation and are treated with vehicle (1% ethanol); (ii) Mel group: mice are treated with melatonin (20 mg/kg via intraperitoneal injection); (iii) IR+V group: mice underwent the MI/R operation and are treated with vehicle (1% ethanol); (iv) IR+Mel group: mice underwent the MI/R operation and are treated with melatonin (20 mg/kg via intraperitoneal injection 10 minutes before reperfusion); (v) IR+Mel+3-TYP group: mice are pretreated with 3-TYP (3-TYP is intraperitoneally injected at a dose of 50 mg/kg every 2 days for a total of three doses prior to the MI/R surgery), subjected to the MI/R operation, and treated with melatonin (20 mg/kg via intraperitoneal injection 10 minutes before reperfusion); and (vi) IR+3-TYP group: mice are pretreated with 3-TYP and then subjected to the MI/R operation.