MiR-139-5p regulates the Notch/RBP-J/Hes1 axis to promote homing of bone mesenchymal stem cells in bronchial asthma

doi:10.12122/j.issn.1673-4254.2024.12.03

Abstract

Abstract:

Objective To observe the role of miR-139-5p and Notch1 signaling pathway in regulation of homing of bone mesenchymal stem cells (BMSCs) of asthmatic rats. Methods Normal rat BMSCs were co-cultured with bronchial epithelial cells from normal or asthmatic rats, followed by transfection with miR-139-5p mimics or a negative control sequence. The changes in cell viability and cell cycle were analyzed, and the cellular expressions of CXCR4 and SDF-1 were detected using immunofluorescence staining. The changes of BMSC homing after the transfection were observed, and the expressions of Notch1, RBP-J, and Hes1 mRNAs and proteins and Th1/Th2 cytokines were detected with RT-qPCR, Western blotting or ELISA. Results The co-cultures of BMSCs and asthmatic bronchial epithelial cells showed significantly decreased expressions of miR-139-5p, IL-2 and IL-12 and increased expressions of CXCR4, SDF-1, IL-5, IL-9, Notch1, RBP-J, and Hes1. Transfection with miR-139-5p mimics significantly increased the expressions of miR-139-5p, IL-2, CXCR4 and SDF-1 and lowered the expression levels of IL-5, IL-9, Notch1, activated Notch1, and Hes1 in the co-cultured cells. Correlation analysis showed that BMSC homing was positively correlated with miR-139-5p and IL-12 and negatively correlated with IL-5 expression. The expression of CXCR4 was negatively correlated with activated Notch1, and SDF-1 was positively correlated with miR-139-5p but negatively correlated with Notch1 expression. Conclusion High expression of miR-139-5p promotes homing of BMSCs in asthma by targeting the Notch1 signaling pathway to regulate the expressions of Th1/Th2 cytokines, thereby alleviating airway inflammation.

Key words: asthma, bone mesenchymal stem cells, miR-139-5p, Notch1 signaling pathway

Kun WANG, Haoxiang FANG, Xiaomei CAO, Ziheng ZHU. MiR-139-5p regulates the Notch/RBP-J/Hes1 axis to promote homing of bone mesenchymal stem cells in bronchial asthma[J]. Journal of Southern Medical University, 2024, 44(12): 2283-2290.

Figures/Tables 11

References 30

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Primer	Sequence (5' to 3')	Length (bp)
Notch1-F	CCAGGAAAGAGGGCAGC	198
Notch1-R	GGAGCATCTCAAGCCTCTT	198
RBP-J-F	CAGGCATTCTACAGGAAGTTTG	99
RBP-J-R	TACTGTTTGATCCCCTCGTTC	99
Hes1-F	AAACCAAAGACAGCCTCTGA	184
Hes1-R	GGTGCTTCACTGTCATTTCC	184
miR139-5p-F	ACACTCCAGCTGGGTCTACAGTGCACGTGT	66
miR139-5p-R	TGGTGTCGTGGAGTCG	66
β-actin-F	CCCATCTATGAGGGTTACGC	150
β-actin-R	TTTAATGTCACGCACGATTTC	150

Primer	Sequence (5' to 3')	Length (bp)
Notch1-F	CCAGGAAAGAGGGCAGC	198
Notch1-R	GGAGCATCTCAAGCCTCTT	198
RBP-J-F	CAGGCATTCTACAGGAAGTTTG	99
RBP-J-R	TACTGTTTGATCCCCTCGTTC	99
Hes1-F	AAACCAAAGACAGCCTCTGA	184
Hes1-R	GGTGCTTCACTGTCATTTCC	184
miR139-5p-F	ACACTCCAGCTGGGTCTACAGTGCACGTGT	66
miR139-5p-R	TGGTGTCGTGGAGTCG	66
β-actin-F	CCCATCTATGAGGGTTACGC	150
β-actin-R	TTTAATGTCACGCACGATTTC	150

Group	Cell viability	G1(%)	S (%)	G2(%)
NC	1.000±0.014	56.74±2.15	8.80±0.72	34.46±0.77
MC	0.558±0.033^Δ	73.99±2.48 ^ΔΔ	4.24±0.63^ΔΔ	21.78±0.66^ΔΔ
miR-139-5p mimics	0.892±0.046*^▲	62.61±1.89*^▲	8.26±0.49*^▲	29.13±0.72*^▲
miR-139-5p mimics-NC	0.544±0.122^Δ	72.94±2.49	5.27±0.62	21.79±0.94

Group	Cell viability	G1(%)	S (%)	G2(%)
NC	1.000±0.014	56.74±2.15	8.80±0.72	34.46±0.77
MC	0.558±0.033^Δ	73.99±2.48 ^ΔΔ	4.24±0.63^ΔΔ	21.78±0.66^ΔΔ
miR-139-5p mimics	0.892±0.046*^▲	62.61±1.89*^▲	8.26±0.49*^▲	29.13±0.72*^▲
miR-139-5p mimics-NC	0.544±0.122^Δ	72.94±2.49	5.27±0.62	21.79±0.94

Group	Ave CT	2^-△△CT
NC	18.58	1.00±0.04
MC	19.08	0.54±0.05^Δ
miR-139-5p mimics	18.77	0.82±0.04*^▲
miR-139-5p mimics-NC	19.11	0.55±0.02^Δ