Down-regulation of microRNA-23b aggravates LPS-induced inflammatory injury in chondrogenic ATDC5 cells by targeting PDCD4
Authors: Huading Lu
Yuxing Tang
Guoyong Xu
Qing Zhao
Zhongmeng Yang
Journal: Iranian journal of basic medical sciences (2018)
Discipline: PHARMACOLOGY & TOXICOLOGY
Keywords: Inflammatory injury; NF-κB/Notch; Osteoarthritis; PDCD4; microRNA-23b.
Affiliations: Department of Orthopedics, the Fifth Affiliated Hospital of Sun Yat-Sen University, Zhuhai, Guangdong 519000, China.

Abstract:  Objectives: Osteoarthritis (OA), characterized by degradation of articular cartilage, is a leading cause of disability. As the only cell type present in cartilage, chondrocytes play curial roles in the progression of OA. In our study, we aimed to explore the roles of miR-23b in the lipopolysaccharide (LPS)-induced inflammatory injury. Materials and methods: LPS-induced cell injury of ATDC5 cells was evaluated by the loss of cell viability, enhancement of cell apoptosis, alteration of apoptosis-associated proteins, and release of inflammatory cytokines. Then, miR-23b level after LPS treatment was assessed by qRT-PCR. Next, the effects of aberrantly expressed miR-23b on the LPS-induced inflammatory injury were explored. The possible target genes of miR-23b were virtually screened by informatics and verified by luciferase assay. Subsequently, whether miR-23b functioned through regulating the target gene was validated. The involved signaling pathways were investigated finally. Results: Cell viability was decreased but cell apoptosis, as well as release of inflammatory cytokines, was enhanced by LPS treatment. MiR-23b was down-regulated by LPS and its overexpression alleviated LPS-induced inflammatory injury. PDCD4, negatively regulated by miR-23b expression, was verified as a target gene of miR-23b. Following experiments showed miR-23b alleviated LPS-induced cell injury through down-regulating PDCD4 expression. Phosphorylated levels of key kinases in the NF-κB pathway, as well as expressions of key kinases in the Notch pathways, were increased by PDCD4 overexpression. Conclusion: MiR-23b was down-regulated after LPS treatment, and its overexpression ameliorated LPS-induced inflammatory injury in ATDC5 cells by targeting PDCD4, which could activate the NF-κB/Notch pathways. 

Retracted: No

PubPeer comments:
 ![file] Partial duplication of images. Images with same region used as of different age groups. , 
 ![file] Same image (MBP) used as of different age groups in fig.6C, but labeled with different antibody staining (NF) in fig.6D. Magnification and contrast also adjusted. , 
 ![file] Same panel of images used as of different age groups. only brightness and contrast adjusted , 
 ![file] Partial duplication of images. Images with same region used as of different age groups. , 
 ![file] Recycling of beta actin blot for different age group samples. , 
 Here are the MM-24 and MM-60 rows from Fig 6C, with overlaps indicated. As far as I can tell, within the MM-24 row, the red contribution to the MERGED panel is in fact only the upper right section of the MBP stain, enlarged (marked by grey boxes). The same for the green component from the TUNEL stain... so the use of the same indicator arrows in TUNEL and MERGED panels is just bizarre. The blue component of MERGED overlaps with the Hoechst stain but requires major rescaling to align them. ![file] In contrast, the MM-60 panels line up horizontally to create an internally-consistent, plausible MERGED panel. Meanwhile the MBP stains of the MM-24 and MM-60 rows overlap, but the latter is darker (with a few bright points). The TUNEL stains overlap, more-or-less to the same extent, except that the MM-60 TUNEL has gained many apoptopic cells. There is a marginal overlap between MM-24 and MM-60 Hoechst stains, marked with yellow boxes. , 
 #3 For this comparison of Fig 6C (MM-6 row) and 6D (MM-24 row), I have marked overlapping sections of the corresponding panels. Then I rescaled the MM-60 row to bring the overlapping sections up to the same size, to facilitate comparisons. ![] It is clear that the Hoechst stains are the same in both versions, which is at least consistent, although they are supposed to be from rats sacrificed after 24 days and 60 days of metal exposure. In contrast, the TUNEL stain has gained a number of apoptopic cells in the MM-60 version (absent from the MM-24 version). Could the authors comment on this? ,