Targeted disruption of the BCR-ABL fusion gene by Cas9/dual-sgRNA inhibits proliferation and induces apoptosis in chronic myeloid leukemia cells.

The BCR-ABL fusion gene, formed by the fusion of the breakpoint cluster region protein ( BCR) and the Abl Oncogene 1, Receptor Tyrosine Kinase ( ABL) genes, encodes the BCR-ABL oncoprotein, which plays a crucial role in leukemogenesis. Current therapies have limited efficacy in patients with chronic myeloid leukemia (CML) because of drug resistance or disease relapse. Identification of novel strategies to treat CML is essential. This study aims to explore the efficiency of novel CRISPR-associated protein 9 (Cas9)/dual-single guide RNA (sgRNA)-mediated disruption of the BCR-ABL fusion gene by targeting BCR and cABL introns. A co-expression vector for Cas9 green fluorescent protein (GFP)/dual-BA-sgRNA targeting BCR and cABL introns is constructed to produce lentivirus to affect BCR-ABL expression in CML cells. The effects of dual-sgRNA virus-mediated disruption of BCR-ABL are analyzed via the use of a genomic sequence and at the protein expression level. Cell proliferation, cell clonogenic ability, and cell apoptosis are assessed after dual sgRNA virus infection, and phosphorylated BCR-ABL and its downstream signaling molecules are detected. These effects are further confirmed in a CML mouse model via tail vein injection of Cas9-GFP/dual-BA-sgRNA virus-infected cells and in primary cells isolated from patients with CML. Cas9-GFP/dual-BA-sgRNA efficiently disrupts BCR-ABL at the genomic sequence and gene expression levels in leukemia cells, leading to blockade of the BCR-ABL tyrosine kinase signaling pathway and disruption of its downstream molecules, followed by cell proliferation inhibition and cell apoptosis induction. This method prolongs the lifespan of CML model mice. Furthermore, the effect is confirmed in primary cells derived from patients with CML.

Melatonin protects sheep endometrial epithelial cells against lipopolysaccharide-induced inflammation in vitro.

Melatonin has known anti-inflammatory effects. Yet, how melatonin protects sheep endometrial epithelial cells from inflammation remains unknown. In this study, we investigated the melatonin synthetase AANAT and HIOMT and melatonin membrane receptors MT1 and MT2 distribution in sheep uterus. Using lipopolysaccharide (LPS)-stimulated sheep endometrial epithelial cells as an in vitro inflammation model. The results showed that melatonin attenuated the expression of inflammatory factors in a concentration-response manner. Melatonin also inhibited the LPS-stimulated phosphorylation of ERK1/2, JNK and NF-κB p65. This attenuation was partially blocked by luzindole (a non-specific MT1 and MT2 inhibitor) or 4P-PDOT (specific MT2 inhibitor). In addition, the above inhibition of melatonin was abolished by the PI3K/AKT pathway inhibitor LY294002. It was concluded that melatonin had an inhibitory effect on LPS-induced endometrial epithelial cell inflammation in sheep, which was mediated by the activation of the PI3K/AKT pathway via melatonin receptors.

[Quantitatively evaluate the effects of precipitation and vegetation variation to flood under the condition of single rainstorm: A case study of Pengchongjian small watershed, Jiangxi, China]. / 场次暴雨条件下降雨和植被变化对洪水影响的定量评估­­ä»¥å½­å†²æ¶§å°æµåŸŸä¸ºä¾‹.

Precipitation and vegetation are two key factors affecting floods in the watershed. Quantitative evaluating their contribution to flood is of great scientific significance to vegetation construction and water resources management. Based on the geographic information data and hydrometeorological data from 1983 to 2014, we analyzed the characteristics of rainstorm and flood in Pengchongjian small watershed in red soil region of South China by Mann-Kendall test method, cumulative anomaly method and HEC-HMS model, and quantitatively evaluated the contribution rate of precipitation and vegetation variation to flood. The results showed that the rainstorm volume and total flood volume in Pengchongjian small watershed from 1983 to 2014 showed a non-significant upward/downward trend, respectively. HEC-HMS model had good simulation effect on single rainstorm flood, with the evaluation results being within the error range. The contribution rates of precipitation and vegetation variation of different rainstorm floods were different with respect to the total flood volume or to the peak flow. The average contribution rate of precipitation and vegetation variation to the total flood volume was 66.5% and 33.5%, while to the peak flow was 58.9% and 41.1%, respectively. Our results could provide scientific basis for flood evaluation, vegetation construction and comprehensive control of soil erosion in small watershed.