JNK activation and translocation to mitochondria mediates mitochondrial dysfunction and cell death induced by VDAC opening and sorafenib in hepatocarcinoma cells.

The multikinase inhibitor sorafenib, and opening of voltage dependent anion channels (VDAC) by the erastin-like compound X1 promotes oxidative stress and mitochondrial dysfunction in hepatocarcinoma cells. Here, we hypothesized that X1 and sorafenib induce mitochondrial dysfunction by increasing reactive oxygen species (ROS) formation and activating c-Jun N-terminal kinases (JNKs), leading to translocation of activated JNK to mitochondria. Both X1 and sorafenib increased production of ROS and activated JNK. X1 and sorafenib caused a drop in mitochondrial membrane potential (ΔΨ), a readout of mitochondrial metabolism, after 60 min. Mitochondrial depolarization after X1 and sorafenib occurred in parallel with JNK activation, increased superoxide (O2•-) production, decreased basal and oligomycin sensitive respiration, and decreased maximal respiratory capacity. Increased production of O2•- after X1 or sorafenib was abrogated by JNK inhibition and antioxidants. S3QEL 2, a specific inhibitor of site IIIQo, at Complex III, prevented depolarization induced by X1. JNK inhibition by JNK inhibitors VIII and SP600125 also prevented mitochondrial depolarization. After X1, activated JNK translocated to mitochondria as assessed by proximity ligation assays. Tat-Sab KIM1, a peptide selectively preventing the binding of JNK to the outer mitochondrial membrane protein Sab, blocked the depolarization induced by X1 and sorafenib. X1 promoted cell death mostly by necroptosis that was partially prevented by JNK inhibition. These results indicate that JNK activation and translocation to mitochondria is a common mechanism of mitochondrial dysfunction induced by both VDAC opening and sorafenib.

Calculation of estrogen excretion rates from urinary estrogen to creatinine ratios.

1. The excretion of estrogens by 42 menstruant and 7 pregnant (1st trimester) women ranged from 2 - 550 mug/24hr. 2. These changes were followed by measuring estrogen to creatinine ratios in small samples of urine. 3. Values of the ratio changed little in each 24 hr, and correlated closely with daily estrogen output (r = 0.971; P less than 0.001; estrogens (mug/24hr) =(see article. 4. The data suggests that ovarian activity in women, including those given gonadotropins to induce ovulation, may be followed by measuring estrogen to creatinine ratios in small samples of urine; the collection of 24 hr samples is unnecessary.

Use of small samples of urine to monitor testicular response to human chorionic gonadotropin.

The increased urinary excretion of androsterone and aetiocholanolone, 17-oxosteroids, and oestrogens which occurs after the administration of human chorionic gonadotropin to normal men, was followed by measuring steroid to creatinine ratios in small samples of urine. For androsterone + aetiocholanolone, values of the ratio correspond closely with 24 h output (correlation coefficients: 0.91, 0.92, and 0.91 for first morning, mid-morning, and mid-afternoon urine specimens.) Comparable correlation coefficients for the 17-oxosteroids in urine were 0.90, 0.95, and 0.91; and for oestrogens, 0.88, 0.87, and 0.86.