MiR-20b promotes osteocyte apoptosis in rats with steroid-induced necrosis of the femoral head through BMP signaling pathway
Authors: Z-Y Wang
G-Q Li
Journal: European review for medical and pharmacological sciences (2019)
Discipline: PHARMACOLOGY & TOXICOLOGY
Keywords: miR-20b; osteocyte apoptosis; steroid-induced necrosis of the femoral head; BMP signaling pathway; rats
Affiliations: Department of Orthopaedics, Caoxian People's Hospital, Heze, China. .

Abstract:  Objective: To study the effect of micro-ribonucleic acid (miR)-20b on osteocyte apoptosis in rats with steroid-induced necrosis of the femoral head (SNFH) and to analyze whether the bone morphogenetic protein (BMP) signaling pathway is involved in the regulation. Materials and methods: A total of 36 Sprague-Dawley rats were randomly divided into control group (n=12), model group (n=12) and intervention group (n=12). The rat model of SNFH was established in the model and intervention groups, while the rats in the intervention group were intraperitoneally injected with the bone morphogenetic protein (BMP) signaling pathway inhibitor. After modeling, the femoral head in each group was taken, and the morphology of osteocytes was observed via hematoxylin-eosin (HE) staining. The apoptosis level of femoral head cells was detected via terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining. The miR-20b expression level in femoral head cells in each group was detected via quantitative Polymerase Chain Reaction (qPCR). The expression levels of inflammatory factors in femoral head cells in each group were detected via enzyme-linked immunosorbent assay (ELISA). The expression levels of apoptotic proteins and BMP signaling pathway-related proteins in femoral head cells in each group were detected via Western blotting. Results: Compared with those in the control group, the bone trabecula was sparse, the number of osteocytes significantly declined and the number of apoptotic osteocytes markedly increased (p&lt;0.01); the expression level of miR-20b in bone tissues remarkably increased (p&lt;0.01), the content of interleukin-1β (IL-1β), IL-6 and tumor necrosis factor-α (TNF-α) in bone tissues increased (p&lt;0.01), the content of IL-10 significantly declined (p&lt;0.01), the expression level of cleaved caspase-3 protein in bone tissues markedly increased (p&lt;0.01), the Bcl-2/Bax expression level evidently declined (p&lt;0.01) and the expression levels of anaplastic lymphoma kinase3 (ALK3), GATA4 and NKX2.5 in bone tissues remarkably increased (p&lt;0.01) in the model group. Compared with those in the model group, the necrosis of bone tissues significantly decreased, the apoptosis level of osteocytes remarkably declined (p&lt;0.01), the content of IL-1β, IL-6 and TNF-α in bone tissues markedly decreased (p&lt;0.01), the content of IL-10 increased (p&lt;0.01), the expression level of cleaved caspase-3 protein in bone tissues significantly declined (p&lt;0.01), the B-cell lymphoma 2/BCL2-Associated X (Bcl-2/Bax) expression level markedly increased (p&lt;0.01) and the expression levels of ALK3, GATA4 and NKX2.5 in bone tissues significantly decreased (p&lt;0.01) in the intervention group. Conclusions: SNFH will significantly increase the expression level of miR-20b in bone tissues, thereby activating the BMP signaling pathway, promoting the release of inflammatory factors and leading to osteocyte apoptosis. Inhibiting the BMP signaling pathway can effectively reduce the osteocyte apoptosis level. 

Retracted: No

PubPeer comments:
 Figure 4.  Osteocyte apoptosis level detected via TUNEL staining. A, TUNEL staining. ... The number of TUNEL-positive cells in bone tissues is markedly larger in the model group than that in the control group, while it is significantly smaller in the intervention group than that in the model group . The Control also appears as the  Control model panels of Fig 4 in [Ding et al (2019)] , while the  Model image overlaps with that figure s  treatment image. ![file] ![file] No authors in common. , 
 Another observation Fig 3 of [Sun et al (2019)] . ![] , 
 Figure 6.  Expression levels of BMP signaling pathway-related proteins detected via Western blotting. A, Protein band. Compare Alk3 with (at right) Bcl-2, from Fig 5 of [Ding et al (2019)] . ![file] , 
 #2 Fig 4 again. ![file]  Treatment panel also appears in Fig 5A,  Apoptosis level of cortical tissues of each group detected via TUNEL staining. A, Micrograph , from [Shi, Wang Wei (2019)] . ![file] , 
 And another version. Compare *[below]* the present Fig 4 (after rotations) with *[above]* Figure 4.  Apoptosis level in the hippocampus of mice in each group detected via TUNEL staining. A, Micrograph , from [Dai et al (2020)] . ![file] , 
 And another. *[below]* Figure 4; *[above]* Figure 3 from [Yu et al (2010)] .  Apoptosis of TUNEL-stained hepatocytes. A, TUNEL staining microscopic image . ![file] , 
 And another. Below, Fig 4. Above, Figure 2 from [Yang et al (2018)] .  Detection of apoptosis of astrocytes in the cerebral cortex of rats in each group via TUNEL staining. A, Micrograph . ![file] , 
 And another. Below, Fig 4. Above, Figure 4 from [Wang et al (2020)] .  Neuronal apoptosis level in each group detected through TUNEL staining. (A) Micrograph . ![file] , 
 At left here, Figure 5A from [Wang et al (2020)] .  Expression levels of apoptosis-related proteins in CI tissues in each group. (A) Protein bands . Middle, Figure 6A from [Yu et al (2019)] .  Expressions of apoptosis-related proteins in each group measured by Western blotting . At right, Figure 6A from [Li Wang (2019)] :  Expression levels of BMP signaling pathway-related proteins detected via Western blotting. A, Protein band . ![file] Unexpected similarities. , 
 #9 Recolored. - *[left]* Figure 5A from [Wang et al (2020)] .  Expression levels of apoptosis-related proteins in CI tissues in each group. (A) Protein bands . - *[middle]* Figure 6A from [Yu et al (2019)] .  Expressions of apoptosis-related proteins in each group measured by Western blotting . - [right]* Figure 6A from [Li Wang (2019)] :  Expression levels of BMP signaling pathway-related proteins detected via Western blotting. A, Protein band . , 
 #3 *[left]* Fig 6, and *[right]* Fig 5 from  MiR-21 regulates pulmonary hypertension in rats via TGF-u03b21 Smad2 signaling pathway ([Ding et al 2019] ) **[retracted]**. ![file] ------------------------------------------- #9 Recolored. - *[left]* Figure 5A from [Wang et al (2020)] .  Expression levels of apoptosis-related proteins in CI tissues in each group. (A) Protein bands . - *[middle]* Figure 6A from [Yu et al (2019)] .  Expressions of apoptosis-related proteins in each group measured by Western blotting . - [right]* Figure 6A from [Li Wang (2019)] :  Expression levels of BMP signaling pathway-related proteins detected via Western blotting. A, Protein band . ![file] Curiously, some of these bands reappeared in Figs 5a, 6a of  Protective Mechanism of Trimetazidine in Myocardial Cells in Myocardial Infarction Rats through ERK Signaling Pathway ([Wu et al 2021] ). ![file] ,