Silencing circular ANRIL protects HK-2 cells from lipopolysaccharide-induced inflammatory injury through up-regulating microRNA-9
Authors: Wenyan Deng
Kai Chen
Yingying Wang
Shuxia Liu
Journal: Artificial Cells Nanomedicine and Biotechnology (2019)
Discipline: BIOLOGY & BIOCHEMISTRY
Keywords: Chronic kidney disease; circular ANRIL; inflammatory response; lipopolysaccharide; microRNA-9.
Affiliations: Department of Nephrology, Jining No.1 People's Hospital , Jining , Shandong , China.

Abstract:  Circular antisense non-coding RNA in the INK4 locus (cANRIL) participated in inflammation of endothelial cells. However, whether cANRIL is associated with inflammatory injury of HK-2 cells, thereby affecting chronic kidney disease has not been investigated. We tested the hypothesis that cANRIL participated in inflammatory response in vitro. HK-2 cells were stimulated by lipopolysaccharides (LPS). RT-qPCR was executed for cANRIL expression assessment. After transfection, cell viability, apoptosis, inflammatory cytokines and ROS generation were appraised to evaluate the impact of silencing cANRIL on LPS-induced inflammatory injury. The regulatory relationship between cANRIL and microRNA-9 (miR-9) was verified. In addition, whether miR-9 affected LPS-induced inflammatory injury was measured after miR-9 inhibitor transfection. Western blot was utilized to detect NF-κB and JNK/p38 pathway-related proteins. The results showed that LPS promoted cANRIL expression and cell injuries in HK-2 cells. Furthermore, silencing cANRIL alleviated inflammatory injuries by promoting viability, suppressing apoptosis, inflammatory cytokines and ROS generation in HK-2 cells. In addition, miR-9 expression was accelerated by silencing cANRIL. Meanwhile, miR-9 down-regulation invalidated the effect of silencing cANRIL on inflammation and NF-κB and JNK/p38 pathways. The study clarified that silencing cANRIL hindered NF-κB and JNK/p38 pathways by positively regulating miR-9, thereby protecting HK-2 cells from LPS-induced injury. 

Retracted: No

PubPeer comments:
 This article of Vantomme and Averous in Green Chemistry shows questionable data. ![file] This figure is supposed to show the stress relaxation plots of the vitrimer at different temperature obtained by a rheometer. One can be surprised that the curves appear to have no noise and the apparatus being able to measure null G. At best these curves are the fitted curved following Maxwell model but if so it has to be mentioned and the experimental curve have to be added in the supplementary data. Using these data authors are able to calculate an activation energy of 64 kJ mol but no error bar are given contrary to most of the vitrimer literature. ![file] Then the authors performed Creep experiment. Again no apparent noise but the strangest thing on this figure is that the creep experiment plot are performed between -10u00b0C to 40u00b0C but the authors appears to be able to extract the data for the Arrhenius plot between 10u00b0C to 50u00b0C. Why the authors did not use the -10u00b0C and 0u00b0C creep experiment results? And where is the creep test at 50u00b0C? Moreover, the data extracted for the creep experiment appears to be inconsistent with the Arrhenius plot. Indeed at 40u00b0C, creep rate is around 6% meaning the -ln(u03b5u2019) should be equal to 2.8 and not 5 as reported in the Arrhenius plot. Similarly, at 30u00b0C, u03b5u2019u22482 so -ln(u03b5u2019) should be equal to 3.9 and not 5,8. And so one. Roughly using the experimental data that can be estimated from the creep experiment an activation energy of 80kJ mol can be calculated not 60 kJ mol like reported in the article (still no error bar given). ![file] Concerning the mechanical properties, the authors reported extremely good results with unchanged propertied between pristine and recycled materials. However, the stress-strain curves given for the pristine material shows a maximum stress of 0.85 MPa which is not in agreement to the result reported in the table 2 with a maximum stress that should be included between 0.66 and 0.72 MPa. , 
 The authors report that the priamine 1071 (batch 0000487627) used is composed of 75% of dimer and 25% of trimer. This looks pretty odd compared to the Croda usual purity (84% in dimer) and no information is given on the remaining 16%. ![file] Moreover the authors synthesize the vitrimer network using an excess of amines compare to aldehyde (r=0.8). The average functionality can be calculated equal to f=2.1. Then using the Peebles, Schaefgen, Flory equation, it can be determined that even at full conversion (p=1) of aldehyde only oligomers should be formed. Then the extremely low soluble fractions reported (1%) are unexpected. ![file] ![file] ,