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. 2023 Jan 10:2023:9380398.
doi: 10.1155/2023/9380398. eCollection 2023.

lncRNA DLX6-AS1 Promotes Myocardial Ischemia-Reperfusion Injury through Mediating the miR-204-5p/FBXW7 Axis

Fanshun Wang  1 , Yuan Wu  2
Affiliations

lncRNA DLX6-AS1 Promotes Myocardial Ischemia-Reperfusion Injury through Mediating the miR-204-5p/FBXW7 Axis

Fanshun Wang et al. Mediators Inflamm. .

Abstract

Myocardial ischemia-reperfusion (IR) injury is the restoration of blood flow post ischemia, which threatens the human life. Long noncoding RNA distal-less homeobox 6 antisense 1 (DLX6-AS1) has been found to take part in the IR-induced cerebral injury. Here, we determined the functional role of DLX6-AS1 in IR-induced myocardial injury. We ligated the left anterior descending coronary artery of rats to induce IR injury. IR injury rats exhibited severe tissue damage and increase of infraction size. The levels of lactate dehydrogenase (LDH), creatine kinase (CK), proinflammatory factors including MCP-1, IL-6, and IL-1β, and cell apoptosis were also enhanced in IR rats, indicating that IR induced significant myocardial injury in rats. DLX6-AS1 expression was elevated in the myocardial tissues of IR injury rats, while DLX6-AS1 deficiency alleviated IR-induced myocardial injury in rats by reducing inflammatory response and cell apoptosis. Moreover, rat embryonic cardiomyocyte cell line H9c2 was subjected to hypoxia reoxygenation (HR). DLX6-AS1 was upregulated in the HR-treated H9c2 cells, and DLX6-AS1 enhanced the expression of F-box and WD40 repeat domain-containing 7 (FBXW7) by sponging miR-204-5p. Inhibition of DLX6-AS1 inhibited inflammatory response and cell apoptosis in H9c2 cells via miR-204-5p/FBXW7 axis. In conclusion, this work demonstrates that DLX6-AS1 accelerates myocardial IR injury through regulating miR-204-5p/FBXW7 axis. Thus, this work provides a novel ceRNA DLX6-AS1/miR-204-5p/FBXW7 axis in myocardial IR injury, and DLX6-AS1 may be a potential target for the treatment of myocardial IR injury.

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Conflict of interest statement

The authors state that there are no conflicts of interest to disclose.

Figures

Figure 1
Figure 1
DLX6-AS1 expression was increased in the myocardial tissues of IR injury rats. IR rats were established by LAD ligation. Sham-operated rats were served as control. (a) The pathological changes of myocardial tissues were analyzed by HE staining. (b) Infarction size of myocardial tissues was examined by TTC staining. (c) The levels of LDH and CK in the serum were detected by ELISA. (d) The levels of MCP-1, IL-6, and IL-1β in the serum were detected by ELISA. (e) Apoptosis in myocardial tissues was detected by TUNEL staining. (f) The expression of Bax and Bcl-2 in myocardial tissues was analyzed by WB. (g) The DLX6-AS1 mRNA expression was examined through qRT-PCR. ∗∗P vs. Sham.
Figure 2
Figure 2
DLX6-AS1 downregulation alleviated myocardial infarction and myocardial damage in IR rats. IR rats were established by LAD ligation, which were then transfected with sh-DLX6-AS1 or Scramble. (a) The expression of DLX6-AS1 in the myocardial tissues was examined by qRT-PCR. (b and c) Infarction size of myocardial tissues was examined by TTC staining. (d) The levels of LDH and CK in the serum were assessed by ELISA. ∗∗P vs. IR + Scramble.
Figure 3
Figure 3
DLX6-AS1 downregulation repressed inflammatory response and cell apoptosis in IR rats. IR rats were established by LAD ligation, which were then transfected with sh-DLX6-AS1 or Scramble. (a) The levels of MCP-1, IL-6, and IL-1β in the serum were detected by ELISA. (b) Apoptosis in myocardial tissues was detected by TUNEL staining. (c) The expression of Bax and Bcl-2 in myocardial tissues was analyzed by WB. ∗∗P vs. IR + Scramble.
Figure 4
Figure 4
DLX6-AS1 interacted with miR-204-5p and repressed miR-204-5p expression. (a) The predicted binding sites between DLX6-AS1 and miR-204-5p were presented. (b) The interaction between DLX6-AS1 and miR-204-5p in H9c2 cells was verified by luciferase reporter assay. (c) IR rats were established by LAD. Sham-operated rats were served as control. H9c2 cells were subjected to HR. Normal H9c2 cells were served as control. The expression of miR-204-5p and DLX6-AS1 in the myocardial tissues and H9c2 cells was examined by qRT-PCR. (d) RIP assay was carried out utilizing an anti-Ago2 antibody in H9c2 cells transfected with pc-DLX6-AS1 to detect the expression of DLX6-AS1 and miR-204-5p through qRT-PCR. (e) The qRT-PCR was carried out to assess the expression of miR-204-5p in H9c2 cells transfected with si-NC, si-DLX6-AS1, pc-DNA, or pc-DLX6-AS1.∗∗P vs. NC-mimic, Sham, Control, IgG, si-NC; ##P vs. pc-DNA.
Figure 5
Figure 5
DLX6-AS1 promoted FBXW7 expression by sponging miR-204-5p. (a) The predicted binding sites between miR-204-5p and FBXW7 were presented. (b) The interaction between miR-204-5p and FBXW7 in H9c2 cells was verified by luciferase reporter assay. (c) H9c2 cells were transfected with pc-DLX6-AS1 or pc-DNA and miR-204-5p-mimic or NC-mimic. FBXW7 expression in the H9c2 cells was examined by WB. ∗∗P vs. NC-mimic, pc-DNA + NC-mimic; ##P vs. pc-DNA + miR-204-5p-mimic; &&P vs. miR-204-5p-mimic + pc-DLX6-AS1.
Figure 6
Figure 6
DLX6-AS1 deficiency repressed inflammatory response and cell apoptosis in HR-treated H9c2 cells by regulating miR-204-5p. H9c2 cells were transfected with si-DLX6-AS1 or si-NC and miR-204-5p inhibitor or inh-NC, followed by HR treatment. (a) The expression of MCP-1, IL-6, and IL-1β in the H9c2 cells was detected by qRT-PCR. (b) The expression of Bax and Bcl-2 in the H9c2 cells was assessed by WB. (c) Apoptosis of H9c2 cells was measured through flow cytometry. ∗∗P vs. si-NC + inh-NC; ##P vs. si-NC + miR-204-5p inhibitor; &&P vs. miR-204-5p inhibitor + si-DLX6-AS1.
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