Short-term waterlogging-induced autophagy in root cells of wheat can inhibit programmed cell death.

Autophagy is a pathway for the degradation of cytoplasmic components in eukaryotes. In wheat, the mechanism by which autophagy regulates programmed cell death (PCD) is unknown. Here, we demonstrated that short-term waterlogging-induced autophagy inhibited PCD in root cells of wheat. The waterlogging-tolerant wheat cultivar Huamai 8 and the waterlogging-sensitive wheat cultivar Huamai 9 were used as experimental materials, and their roots were waterlogged for 0-48 h. Waterlogging stress increased the number of autophagic structures, the expression levels of autophagy-related genes (TaATG), and the occurrence of PCD in root cells. PCD manifested as morphological changes in the cell nucleus, significant enhancement of DNA laddering bands, and increases in caspase-like protease activity and the expression levels of metacaspase genes. The autophagy promoter rapamycin (RAPA) reduced PCD levels, whereas the autophagy inhibitor 3-methyladenine (3-MA) enhanced them. The expression levels of TaATG genes and the number of autophagic structures were lower in cortex cells than in stele cells, but the levels of PCD were higher in cortex cells. The number of autophagic structures was greater in Huamai 8 than in Huamai 9, but the levels of PCD were lower. In summary, our results showed that short-term waterlogging induced autophagy which could inhibit PCD. Mechanisms of response to waterlogging stress differed between cortex and stele cells and between two wheat cultivars of contrasting waterlogging tolerance.

Mutual regulation of ROS accumulation and cell autophagy in wheat roots under hypoxia stress.

Here, we explored the mutual regulation of radical oxygen species (ROS) and autophagy in wheat (Triticum aestivum L.) roots under hypoxia stress. We also analyzed differences between the responses of the stele and the cortex in the two wheat cultivars Huamai 8 (waterlogging-tolerant) and Huamai 9 (waterlogging-sensitive) to hypoxia stress. In situ detection and ultracytochemical localization analysis in wheat roots showed that hypoxia stress caused greater increases in ROS levels and the expression levels of alternative oxidase (AOX) and antioxidant enzymes in the stele than in the cortex. The analysis of exogenous ROS addition and the inhibition of its production revealed the pivotal roles played by ROS in autophagy. Moreover, transmission electron microscopy and qRT-PCR analysis indicated that the stele had a higher level of autophagy than the cortex and that the two wheat cultivars primarily differed in the type and number of autophagosomes. Additional research revealed that autophagy could remove excess ROS, as pre-treatment with the autophagy inhibitor 3-methyladenine increased ROS levels in roots and the addition of the autophagy inducer rapamycin reduced root ROS levels. In conclusion, hypoxia stress induced ROS accumulation in wheat roots where ROS acted as an autophagy signal. Furthermore, higher levels of autophagy and antioxidant enzyme expression in the stele facilitated the elimination of oxidative damage caused by excessive ROS and thereby increased cell survival; in the cortex, a large number of cells died and formed aerenchyma.

Use of a locking intramedullary nail for the treatment of initial varus proximal humeral fracture: a prospective pilot study.

OBJECTIVE: To evaluate the feasibility of locked intramedullary nailing, rather than locking plate fixation combined with fibular allograft augmentation, for initial varus proximal humeral fractures. METHODS: This prospective pilot study enrolled patients with initial varus proximal humeral fractures that were treated with a locking intramedullary nail. Radiography was performed to evaluate fracture healing. Data about the visual analogue scale (VAS) pain score, Constant Shoulder Score (CSS), Disabilities of the Arm, Shoulder and Hand (DASH) score, American Shoulder and Elbow Surgeons (ASES) score and shoulder range of motion (ROM) were recorded. RESULTS: Twenty patients, including eight with Neer two-part and 12 with three-part fractures, were followed-up, with a mean time of 12.3 months. All patients sustained fractures that healed without re-varus. During the last follow-up, the shoulder function of the patients had recovered well, with a mean VAS pain score of 1.4, a mean CSS of 83.1, a mean DASH score of 80.8, a mean ASES score of 84.0 and a satisfactory ROM. In one patient, the proximal locking screw came out and was removed via a second surgery. CONCLUSIONS: The use of a locking intramedullary nail alone for initial varus proximal humeral two-/three-part fractures was feasible. This treatment has advantages, such as preventing re-varus and causing milder surgical trauma, than that seen with a locking plate.

The release of cytochrome c and the regulation of the programmed cell death progress in the endosperm of winter wheat (Triticum aestivum L.) under waterlogging.

It has been shown in mammalian systems that the mitochondria can play a key role in the regulation of apoptosis by releasing intermembrane proteins (such as cytochrome c) into the cytosol. Cytochrome c released from the mitochondria to the cytoplasm activates proteolytic enzyme cascades, leading to specific nuclear DNA degradation and cell death. This pathway is considered to be one of the important regulatory mechanisms of apoptosis. Previous studies have shown that endosperm cell development in wheat undergoes specialized programmed cell death (PCD) and that waterlogging stress accelerates the PCD process; however, little is known regarding the associated molecular mechanism. In this study, changes in mitochondrial structure, the release of cytochrome c, and gene expression were studied in the endosperm cells of the wheat (Triticum aestivum L.) cultivar "huamai 8" during PCD under different waterlogging durations. The results showed that waterlogging aggravated the degradation of mitochondrial structure, increased the mitochondrial permeability transition (MPT), and decreased mitochondrial transmembrane potential (ΔΨm), resulting in the advancement of the endosperm PCD process. In situ localization and western blotting of cytochrome c indicated that with the development of the endosperm cell, cytochrome c was gradually released from the mitochondria to the cytoplasm, and waterlogging stress led to an advancement and increase in the release of cytochrome c. In addition, waterlogging stress resulted in the increased expression of the voltage-dependent anion channel (VDAC) and adenine nucleotide translocator (ANT), suggesting that the mitochondrial permeability transition pore (MPTP) may be involved in endosperm PCD under waterlogging stress. The MPTP inhibitor cyclosporine A effectively suppressed cell death and cytochrome c release during wheat endosperm PCD. Our results indicate that the mitochondria play important roles in the PCD of endosperm cells and that the increase in mitochondrial damage and corresponding release of cytochrome c may be one of the major causes of endosperm PCD advancement under waterlogging.

Both non-covalent and covalent interactions were involved in the mechanism of detoxifying effects of persimmon tannin on Chinese cobra PLA2.

Persimmon tannin (PT) has been shown to inhibit snake venom activities and toxicities both in vitro and in vivo. To clarify the detoxifying mechanism of PT on snake venom, the interaction of characteristic structural elements of PT (EGCG, ECG, EGCG dimer and ECG dimer) and Chinese cobra phospholipase A2 (PLA2) was studied. The results revealed that except non-covalent bonds like hydrogen bonds, hydrophobic bonds and iron bonds were formed between PT and PLA2, covalent interaction was also occurred. PT could bind with the key active residues of PLA2, such as lysine, histidine, tryptophan and tyrosine, restraining their activity and disturbing the structure of PLA2, thus showing detoxifying effects on snake venom.