Degradation of AF1Q by chaperone-mediated autophagy.

AF1Q, a mixed lineage leukemia gene fusion partner, is identified as a poor prognostic biomarker for pediatric acute myeloid leukemia (AML), adult AML with normal cytogenetic and adult myelodysplastic syndrome. AF1Q is highly regulated during hematopoietic progenitor differentiation and development but its regulatory mechanism has not been defined clearly. In the present study, we used pharmacological and genetic approaches to influence chaperone-mediated autophagy (CMA) and explored the degradation mechanism of AF1Q. Pharmacological inhibitors of lysosomal degradation, such as chloroquine, increased AF1Q levels, whereas activators of CMA, including 6-aminonicotinamide and nutrient starvation, decreased AF1Q levels. AF1Q interacts with HSPA8 and LAMP-2A, which are core components of the CMA machinery. Knockdown of HSPA8 or LAMP-2A increased AF1Q protein levels, whereas overexpression showed the opposite effect. Using an amino acid deletion AF1Q mutation plasmid, we identified that AF1Q had a KFERQ-like motif which was recognized by HSPA8 for CMA-dependent proteolysis. In conclusion, we demonstrate for the first time that AF1Q can be degraded in lysosomes by CMA.

Nerve and conduction tissue injury caused by contact with BioGlue.

BACKGROUND: BioGlue-a surgical adhesive composed of bovine albumin and glutaraldehyde-is commonly used in cardiovascular operations. The objectives of this study were to determine whether BioGlue injures nerves and cardiac conduction tissues, and whether a water-soluble gel barrier protects against such injury. MATERIALS AND METHODS: In 18 pigs, diaphragmatic excursion during direct phrenic nerve stimulation was measured at baseline and at 3 and 30 min after nerve exposure to albumin (n = 3), glutaraldehyde (n = 3), BioGlue (n = 6), or water-soluble gel followed by BioGlue (n = 6). Additionally, BioGlue was applied to the cavoatrial junction overlying the sinoatrial node (SAN), either alone (n = 12) or after application of gel (n = 6). RESULTS: Mean diaphragmatic excursions in the BioGlue and glutaraldehyde groups were lower at 3 min and 30 min than in the albumin group (P < 0.05). Mean excursions in the gel group were similar to those of the albumin group (P = 0.9). Five BioGlue pigs (83%) and one gel pig (17%) had diaphragmatic paralysis by 30 min (P < 0.05 and P = 0.3 versus albumin, respectively). Coagulation necrosis extended into the myocardium at the cavoatrial junction in all 12 BioGlue pigs but only two gel pigs (33%, P < 0.01). Two BioGlue pigs (17%), but no gel pigs, had focal SAN degeneration and persistent bradycardia (P < 0.01). CONCLUSIONS: BioGlue causes acute nerve injury and myocardial necrosis that can lead to SAN damage. A water-soluble gel barrier is protective.

The structural and cellular viability in cryopreserved rabbit carotid arteries.

OBJECTIVE: We investigated the histological and mechanical changes in addition to viable cellular recovery in cryopreserved blood vessels. MATERIALS AND METHODS: Rabbit carotids were cryopreserved in a cryoprotective medium containing 1.5 M of 1,2-propanediol (PD) and then were thawed slowly in an ice bag that had been precooled in liquid nitrogen. Fresh carotids were used as the control. The fresh and freeze-thawed arteries were cultured for the growth of vascular smooth muscle cells (VSMCs). The freeze-thawed arterial tissues were perfused in vitro for 6, 12, or 24 h, respectively, to assess the integrity of carotid walls and the mechanical properties. RESULTS: The results showed that it took almost the same time (24 approximately 36 h) for the VSMCs of the PD-cryopreserved arteries to regenerate as those from the fresh arteries. Their growing speeds also were similar. On the contrary, Me2SO-cryopreserved (1.5 M) arteries were unable to regenerate VSMCs in culture. After freeze-thawing, the mechanical properties decreased significantly (P < 0.003 for elastic modulus and P < 0.001 for fracture strength). After in vitro perfusion of the freeze-thawed carotid arteries, all of the survived endothelial cells fell off, and some of the VSMCs denaturalized or necrosed. The internal elastic fibers and collagen showed various degrees of cracking. The mechanical properties were decreased (P < 0.05). CONCLUSION: Our findings demonstrate that the PD-containing cryoprotective medium can preserve regenerative capacity of VSMCs, which makes it a useful technique for viable VSMC recovery. However, the freeze-thawing process and the in vitro perfusion caused serious disruption in the arterial mechanical properties, rendering the cryopreserved blood vessels less useful for vessel reconstruction.