Non-thermal irreversible electroporation (NTIRE) is a biophysical phenomenon in which certain electric fields delivered across the cell membrane in tissue, cause cell death, without affecting the extracellular matrix. "Minimally invasive regenerative surgery" is a new medical modality for treatment of end-stage organ or tissue failure in which exogenous cells are implanted in a decellularized niche in tissue, formed by the delivery of NTIRE electric fields across a targeted volume of tissue. We anticipate that the success of the procedure will depend on the time of implantation relative to the application of NTIRE. This study was performed to elucidate the histological and molecular events that occur within 24 h after NTIRE, in the context of optimal criteria for the time of implantation. To this end, we examined the histology of NTIRE treated rat liver with H&E, Masson trichrome and TUNEL staining. Western blot was used to examine pro and cleaved caspase-3 (marker for apoptosis), pro and cleaved caspase-1 and gasdermin D (markers for pyroptosis), and RIP3 and MLKL (markers for necroptosis). The key findings are that, complete hepatocytes disintegration within an intact extracellular matrix is seen at 6 h and, new hepatocytes are seen in the treated region at 24 h, after NTIRE. There is no evidence of apoptotic cell death from NTIRE, contrary to commonly made claims in the NTIRE literature. However, molecular pathways of pyroptosis and necroptosis, programed necrosis associated with inflammation, are activated at 6 h after NTIRE and are not evident at 24 h after NTIRE. These are fundamental new findings of basic value to the field of NTIRE in all its applications. Taken together the results suggest the hypothesis that an optimal time for implantation is about 24 h after NTIRE. Future studies in which exogenous cells are implanted at different times after NTIRE are required to examine this hypothesis.
Tiller angle is a key factor determining rice plant architecture, planting density, light interception, photosynthetic efficiency, disease resistance, and grain yield. However, the mechanisms underlying tiller angle control are far from clear. In this study, we identified a mutant, termed bta1–1, with an enlarged tiller angle throughout its life cycle. A detailed analysis reveals that BTA1 has multiple functions because tiller angle, shoot gravitropism, and tolerance to drought stress are changed in bta1–1 plants. Moreover, BTA1 is a positive regulator of shoot gravitropism in rice. Shoot responses to gravistimulation are disrupted in bta1-1 under both light and dark conditions. Gene cloning reveals that bta1-1 is a novel mutant allele of LA1 renamed la1-SN. LA1 is able to rescue the tiller angle and shoot gravitropism defects observed in la1-SN. The nuclear localization signal of LA1 is disrupted by la1-SN, causing changes of its subcellular localization. LA1 is required to regulate the expression of auxin transporters and signaling factors that control shoot gravitropism and tiller angle. High-throughput mRNA sequencing is performed to elucidate the molecular and cellular functions of LA1. The results show that LA1 may be involved in the nucleosome and chromatin assembly, and protein-DNA interactions to control gene expression, shoot gravitropism, and tiller angle. Our results provide new insight into the mechanisms whereby LA1 controls shoot gravitropism and tiller angle in rice.
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