摘要: Objectives The correlations between long non-coding RNAs (lncRNAs) and diverse mammal diseases have been clarified by many researches, but the cognition about bovine mastitis-related lncRNAs remains limited. This study aimed to investigate the potential role of lncRNA X-inactive specific transcript (XIST) in the inflammatory response of bovine mammary epithelial cells. Materials and methods Two inflammatory bovine mammary alveolar cell-T (MAC-T) models were established by infecting the cells with Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). The expressions of pro-inflammatory cytokines were measured, and the proliferation, viability and apoptosis of the inflammatory cells were evaluated after XIST was knocked down by an siRNA. The relationship among XIST, NF-kappa B pathway and NOD-like receptor protein 3 (NLRP3) inflammasome was investigated using an inhibitor of NF-kappa B signal pathway. Results The expression of XIST was abnormally increased in bovine mastitic tissues and inflammatory MAC-T cells. Silencing of XIST significantly increased the expression of E. coli or S. aureus-induced pro-inflammatory cytokines. Additionally, knockdown of XIST could inhibit cell proliferation, suppress cell viability and promote cell apoptosis under inflammatory conditions. Furthermore, XIST inhibited E. coli or S. aureus-induced NF-kappa B phosphorylation and the production of NLRP3 inflammasome. Conclusions The expression of XIST was promoted by activated NF-kappa B pathway and, in turn, XIST generated a negative feedback loop to regulate NF-kappa B/NLRP3 inflammasome pathway for mediating the process of inflammation.

摘要: With the development of synthetic biology, synthetic gene circuits have shown great applied potential in medicine, biology, and as commodity chemicals. An ultimate challenge in the construction of gene circuits is the lack of effective, programmable, secure and sequence-specific gene editing tools. The clustered regularly interspaced short palindromic repeat (CRISPR) system, a CRISPR-associated RNA-guided endonuclease Cas9 (CRISPR-associated protein 9)-targeted genome editing tool, has recently been applied in engineering gene circuits for its unique properties-operability, high efficiency and programmability. The traditional single-targeted therapy cannot effectively distinguish tumour cells from normal cells, and gene therapy for single targets has poor anti-tumour effects, which severely limits the application of gene therapy. Currently, the design of gene circuits using tumour-specific targets based on CRISPR/Cas systems provides a new way for precision cancer therapy. Hence, the application of intelligentized gene circuits based on CRISPR technology effectively guarantees the safety, efficiency and specificity of cancer therapy. Here, we assessed the use of synthetic gene circuits and if the CRISPR system could be used, especially artificial switch-inducible Cas9, to more effectively target and treat tumour cells. Moreover, we also discussed recent advances, prospectives and underlying challenges in CRISPR-based gene circuit development.

摘要: Objectives The evolutionary conserved JNK pathway plays crucial role in cell death, yet factors that modulate this signalling have not been fully disclosed. In this study, we aim to identify additional factors that regulate JNK signalling in cell death, and characterize the underlying mechanisms. Materials and Methods Drosophila were raised on standard media, and cross was carried out at 25 degrees C. The Gal4/UAS system was used to express proteins or RNAi in a specific temporal and spatial pattern. Gene expression was revealed by GFP fluorescence, X-gal staining or immunostaining of 3rd instar larval eye and wing discs. Cell death was visualized by acridine orange (AO) staining. Images of fly eyes and wings were taken by OLYMPUS microscopes. Results We found that licorne (lic) encoding the Drosophila MKK3 is an essential regulator of JNK-mediated cell death. Firstly, loss of lic suppressed ectopic Egr-triggered JNK activation and cell death in eye and wing development. Secondary, lic is necessary for loss-of-cell polarity-induced, physiological JNK-dependent cell death in wing development. Thirdly, Lic overexpression is sufficient to initiate JNK-mediated cell death in developing eyes and wings. Furthermore, ectopic Lic activates JNK signalling by promoting JNK phosphorylation. Finally, genetic epistatic analysis confirmed that Lic acts in parallel with Hep in the Egr-JNK pathway. Conclusions This study not only identified Lic as a novel component of the JNK signalling, but also disclosed the crucial roles and mechanism of Lic in cell death.

摘要: Objectives Breast cancer stem-like cells (BrCSCs) are the major reason for tumour generation, resistance and recurrence. The turbulence of their self-renewal ability could help to constrain the stem cell expansion. The way BrCSCs divided was related to their self-renewal capacity, and the symmetric division contributed to a higher ability. Non-coding long RNA of H19 was involved in multiple malignant procedures; the role and mechanistic proof of non-coding long RNA of H19 in controlling the divisions of BrCSCs were barely known. Materials and Methods Indicative functions of H19 in preclinical study were analysed by using the TCGA data base. Division manners were defined by using fluorescence staining. Results We identified the stimulation of H19 on symmetric division of BrCSCs, which subsequently resulted in self-renewing increasing. H19 inhibited the Let-7c availability by acting as its specific molecular sponge, and with Let-7c inhibition, oestrogen receptor activated Wnt signalling was unconstrained. Similarly, restoring Let-7c constrained oestrogen receptor activated Wnt factors, which sequentially inhibited the H19 decreasing of Let-7 bioavailability. Let-7c is reactivated in vitro where H19 was knockdown, and later inhibited the symmetric division of BrCSCs. Reciprocally, Wnt pathway activation leads to H19 increasing, which in turn decreased Let-7c bioavailability. Conclusions Our results revealed a previously undescribed double negative feedback loop between sponge H19 and targeted Let-7c through oestrogen activated Wnt signalling that dominated in stem cells' division.

摘要: Objectives The role of p62 in cancer is controversial. Evidence has shown that p62 is upregulated in different cancers and promotes tumour growth, such as in liver cancer and lung cancer. However, a recent study showed that the downregulation of p62 in hepatic stellate cells (HSCs) promotes hepatocellular carcinoma (HCC) development. How p62 is regulated in colorectal cancer (CRC) remains largely unknown. In this study, we aimed to investigate the roles and molecular mechanisms of p62 in CRC. Materials and Methods The expression levels of p62 in CRC tissues and adjacent non-tumour tissues were determined by immunohistochemistry (IHC). Stable p62-overexpression HCT116 cells and p62-knockdown SW480 cells were established with lentiviral vectors. The role of p62 in CRC was investigated in in vitro and in vivo functional studies. The relationship between p62 and the vitamin D receptor (VDR) was investigated by coimmunoprecipitation (Co-IP) assays. Results p62 was significantly upregulated in CRC, and a high p62 level was an independent risk factor for a poor prognosis in CRC patients. p62 promoted CRC migration and invasion by inhibiting apoptosis and promoting cell proliferation in vitro, and p62 aggravated tumour growth and metastasis in vivo. Co-IP assays indicated that p62 interacts with the VDR and may target the NRF2-NQO1 axis. Conclusions Our study suggested that p62 functions as an oncogene in CRC through inhibiting apoptosis and promoting cell proliferation by interacting with the VDR.