ROS‐generating oxidases Nox1 and Nox4 contribute to oncogenic Ras‐induced premature senescence

Activated oncogenes induce premature cellular senescence, a permanent state of proliferative arrest in primary rodent and human fibroblasts. Recent studies suggest that generation of reactive oxygen species (ROS) is involved in oncogenic Ras‐induced premature senescence. However, the signaling mechanism controlling this oxidant‐mediated irreversible growth arrest is not fully understood. Here, we show that through the Ras/MEK pathway, Ras oncogene up‐regulated the expression of superoxide‐generating oxidases, Nox1 in rat REF52 cells and Nox4 in primary human lung TIG‐3 cells, leading to an increase in intracellular level of ROS. Ablation of Nox1 and Nox4 by small interfering RNAs (siRNAs) blocked the RasV12 senescent phenotype including β‐galactosidase activity, growth arrest and accumulation of tumor suppressors such as p53 and p16^Ink4a. This suggests that Nox‐generated ROS transduce senescence signals by activating the p53 and p16^Ink4a pathway. Furthermore, Nox1 and Nox4 siRNAs inhibited both Ras‐induced DNA damage response and p38MAPK activation, whereas overexpression of Nox1 and Nox4 alone was able to induce senescence. The involvement of Nox1 in Ras‐induced senescence was also confirmed with embryonic fibroblasts derived from Nox1 knockout mice. Together, these findings suggest that Nox1‐ and Nox4‐generated ROS play an important role in Ras‐induced premature senescence, which may involve DNA damage response and p38MAPK signaling pathways.     

Varicose bleeding after liver transplantation in a patient with severe portosystemic shunts

Recipients for liver transplantation often have portosystemic shunts due to portal hypertension. It is an important problem whether such shunts should be ligated during operations. Ligating the shunts seems of benefit for increasing portal blood flow to the liver, but it is sometimes difficult technically, and it is invasive to the patient. We experienced a recipient with huge portosystemic shunts and no esophageal varices before living-related liver transplantation. Some shunts were ligated during operation to increase portal blood flow to the graft. Unfortunately, the patient suffered severe bleeding from esophagogastric varices after he underwent retransplantation owing to accidental liver failure. Based on our experience, extreme care should be exercised to avoid varicose bleeding after ligating the portosystemic shunts of liver transplantation patients.     

SNARE mediates autophagosome–lysosome fusion

Autophagy is an intracellular bulk degradation/recycling system that turns over cellular constituents. This system has also been shown to play a crucial role in host defense, termed antimicrobial autophagy (xenophagy), in which it functions to degrade intracellular foreign microbial invaders. Xenophagosomes undergo a stepwise maturation process that consists of fusion with lysosomes, after which the cargo undergoes degradation. We have previously shown that intracellular group A Streptococcus (GAS) is captured by xenophagosomes termed GAS-containing autophagosome-like vacuoles (GcAVs), where GAS organisms are degraded following fusion with lysosomes. Our recent investigations have shown that endocytic soluble N-ethylmaleimide-sensitive factor attachment protein receptors (SNAREs) are involved in the fusion of xenophagosomes and lysosomes. Confocal microscopic analysis has shown that SNAREs, including VAMP8 and Vti1b, colocalize with GFP-LC3 in GcAVs. Our findings also suggested that Vti1b is derived from autophagic compartments, whereas VAMP8 originates from lysosomes. Knockdown of the combinational SNARE proteins VAMP8 and Vti1b with siRNAs disturbed the autophagic fusion of xenophagosomes with lysosomes, and cellular bactericidal efficiency significantly diminished. Furthermore, knockdown of these SNAREs inhibited the fusion of canonical autophagosomes with lysosomes. These findings strongly suggest that a combinatorial SNARE complex with VAMP8 and Vti1b mediates the fusion of antimicrobial and canonical autophagosomes with lysosomes, an essential event for autophagic degradation.     

Interferential Electric Stimulation Applied to the Neck Increases Swallowing Frequency

Swallowing disorders are a common complaint among the elderly. Recently, surface electrical stimulation applied to the neck region has received increased attention as a new modality to treat pharyngeal dysphagia. Previous reports used pulsed current at a frequency range of 1-120 Hz. Kilohertz-frequency alternating currents (ACs) have not been tested for treating dysphagia. Therefore, we evaluated the effects of interferential currents (IFCs), the most popular modality of amplitude-modulated kilohertz-frequency ACs in clinical practice, on the swallowing reflex in healthy subjects. We found that IFC stimulation at the sensory threshold with 50-Hz modulation significantly increased the number of swallows without any discomfort, but pure AC stimulation at the carrier frequency did not have a significant effect. There was no statistically significant difference in the time course of the number of swallows among 1,000-, 2,000-, 4,000-, and 6,000-Hz carrier frequencies. The number of swallows remained increased during the 15-min IFC stimulation, suggesting that IFC stimulation facilitates the swallowing reflex without adaptation, at least during this stimulation period. We suggest that an IFC stimulation or a low-frequency, modulated kilohertz AC stimulation, which would be more comfortable than pulsed currents, is an alternative stimulation mode for treating pharyngeal dysphagia.