MicroRNA-146a regulates the production of cytokines in lymphocytes stimulated by Porphyromonas gingivalis lipopolysaccharide

doi:10.7518/hxkq.2021.01.004

Abstract

Abstract: Objective

This study aimed to investigate the effects of microRNA-146a (miR-146a) on the production of cytokines in lymphocytes stimulated by Porphyromonas gingivalis (P.gingivalis) lipopolysaccharide (LPS).

Methods

Lymphocytes were harvested from mouse spleen and cultured in vitro. The cells were treated with P. gingivalis LPS, miR-146a mimic, or miR-146a inhibitor. Scramble RNA served as the negative control of mimic and inhibitor. The production of inflammatory cytokines was detected by quantitative real-time polymerase chain reaction and enzyme-linked immunosorbent assay.

Results

Compared with non-LPS-stimulated group, P. gingivalis LPS could increase the levels of interleukin (IL)-1β, IL-6, receptor activator NF-κB ligand (RANKL), and IL-10 (P<0.05) and decrease the mRNA level of osteoprotectin (OPG) (P<0.05). However, it did not significantly change the secretion of OPG. Compared with the negative control group, miR-146a mimic upregulated the levels of IL-10 and OPG (P<0.05), downregulated IL-1β, IL-6, and RANKL (P<0.05). Meanwhile, miR-146a inhibitor had a reverse effect on these cytokines (P<0.05) in P.gingivalis LPS-treated-lymphocytes.

Conclusion

MiR-146a can provide a suitable microenvironment for bone formation by preventing the inflammatory effects of P.gingivalis LPS through the inhibition of IL-1β, IL-6, and RNAKL, thereby enhancing IL-10 and OPG.

Key words: periodontitis, microRNA-146a, cytokine, lymphocytes, lipopolysaccharide

CLC Number:

Q 257

Si Yuting, Song Jinhua, Fang Zhen, Han Xiaozhe, Jiang Shaoyun. MicroRNA-146a regulates the production of cytokines in lymphocytes stimulated by Porphyromonas gingivalis lipopolysaccharide[J]. West China Journal of Stomatology, 2021, 39(1): 26-31.

Figures/Tables 7

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基因	引物序列
IL-1β	F：5’-ATGCCTTCCCCAGGGCATGT-3’ R：5’-CTGAGCGACCTGTCTTGGCCG-3’
IL-6	F：5’-TCCAGTTGCCTTCTTGGGAC-3’ R：5’-GTACTCCAGAAGACCAGAGG-3’
IL-10	F：5’-GACCAGCTGGACAACATACTGCTAA-3’ R：5’-GATAAGGCTTGGCAACCCAAGTAA-3’
OPG	F：5’-AGCAGGAGTGCAACCGCACC-3’ R：5’-TTCCAGCTTGCACCACGCCG-3’
RANKL	F：5’-CATGTGCCACTGAGAACCTTGAA-3’ R：5’-CAGGTCCCAGCGCAATGTAAC-3’
GAPDH	F：5’-CCCCAGCAAGGACACTGAGCAA-3’ R：5’-GTGGGTGCAGCGAACTTTATTGATG-3’
miR-146a	F：5’-GGGTGAGAACTGAATTCCA-3’ R：5’-CAGTGCGTGTCGTGGAGT-3’
U6 small nuclear RNA	F：5’-GCTTCGGCAGCACATATACTAAAAT-3’ R：5’-CGCTTCACGAATTTGCGTGTCAT-3’

细胞因子	P.gingivalis LPS 未刺激组	P.gingivalis LPS 刺激组	t值	P值
IL-1β	1.03±0.26	1.52±0.02	3.199	0.032 9
IL-6	1.03±0.28	2.60±0.77	3.325	0.029 2
IL-10	1.01±0.20	2.37±0.10	10.59	0.000 4
RANKL	1.00±0.03	3.85±1.07	4.604	0.010 0
OPG	1.01±0.20	0.59±0.03	3.631	0.022 1

细胞因子	P.gingivalis LPS未刺激组/pg·mL^-1	P.gingivalis LPS刺激组/pg·mL^-1	t值	P值
IL-1β	3.14±0.31	21.66±3.00	5.319	0.006 0
IL-6	349.09±26.10	498.26±20.52	7.781	0.001 5
IL-10	305.56±27.78	531.96±34.47	8.858	0.000 1
RANKL	16.36±0.91	48.20±0.66	24.50	<0.000 1
OPG	12.70±1.07	12.16±0.62	0.756	0.491 8

细胞因子	P.gingivalis LPS/ 模拟物阴性 RNA对照组	P.gingivalis LPS/miR-146a模拟物组	t值	P值
IL-1β	1.84±0.25	0.64±0.04	8.130	0.001 2
IL-6	2.32±0.28	1.60±0.28	3.111	0.035 8
IL-10	2.51±0.18	3.26±0.29	3.799	0.019 1
RANKL	2.94±0.46	0.90±0.27	6.620	0.002 7
OPG	0.63±0.05	3.25±0.29	15.71	<0.000 1

细胞因子	P.gingivalis LPS/ 模拟物阴性RNA对照组/pg·mL^-1	P.gingivalis LPS/miR-146a模拟物组/pg·mL^-1	t值	P值
IL-1β	18.20±2.60	6.06±1.50	3.50	0.024 9
IL-6	506.52±25.84	417.85±17.40	4.026	0.015 8
IL-10	566.21±53.20	684.93±24.70	3.506	0.024 8
RANKL	47.78±0.84	14.66±1.85	14.10	0.000 1
OPG	11.70±0.40	17.60±0.23	11.00	0.000 4