Effect of human umbilical cord mesenchymal stem cells on the development of the drug resistance of imipenem-resistant Pseudomonas aeruginosa

doi:10.3877/cma.j.issn.1674-1358.2016.02.025

Abstract

Abstract:

Objective

To investigate the effect of human umbilical cord mesenchymal stem cells (hUCMSCs) in vitro on the development of the drug resistance of imipenem-resistant Pseudomonas aeruginosa and on OprD2 gene.

Methods

This experiment included three groups: experiment group (hUCMSCs group), control group (cells control group, namely normal human lung fibroblast, NHLF group) and blank control group. During the conductive process of Pseudomonas aeruginosa (PA) drug resistance induced by sub-inhibitory concentrations of broth, the hUCMSCs group and the NHLF group were added with the supernatants obtained from their joint cultivation with PA, respectively; while the blank control group was administrated with cell culture medium. The numbers of generations required by induced drug resistance in the three groups and the sizes of inhibition zones were observed. The sensitivity of PA to common antibacterial agents and the expression quantity of OprD2 gene were determined by K-B method and real-time PCR method.

Results

Through the induction of sub-inhibitory concentration of imipenem, the occurrence of PA drug resistance in the hUCMSCs group was relatively later than that of the blank control group. Imipenem drug resistance appeared in passage 17 of PA induced in the NHLF group and the blank control group, while it occurred in passage 19 in the hUCMSCs group. The results of real-time PCR showed that after the induction of drug resistance, the expression quantity of OprD2 in PA decreased or disappeared compared with that before the induction. The expression quantity of OprD2 in PA in the hUCMSCs group was reduced to 10.96% of the induction, while there were no OprD2 expression in the NHLF and the blank control group, OprD2 gene deletion occurred after the induction.

Conclusions

hUCMSCs could delay the formation of PA’s drug resistance. This mechanism is probably through reducing the inhibition of the expression of OprD2 by secreting antimicrobial peptide LL-37 and human β-defensin 2 (HBD-2), and the decrease or deletion of the expression of outer membrane protein OprD2 is the cause for PA’s drug resistance to imipenem.

Key words: Pseudomonas aeruginosa, Drug resistance, Human umbilical cord mesenchymal stem cells (hUCMSCs), Antimicrobial peptides

Xuan’er Zheng, Jie Yang, Xiaoping Mu, Weiming Lai, Fang Xu, Xiaohong Liu, Haoming Yang, Shumei Yang. Effect of human umbilical cord mesenchymal stem cells on the development of the drug resistance of imipenem-resistant Pseudomonas aeruginosa[J]. Chinese Journal of Experimental and Clinical Infectious Diseases(Electronic Edition), 2016, 10(02): 248-253.

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References 25

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基因	序列	产物大小（bp）
16S RNA	F：TAGGTGGTTCAGCAAGTTGGATGTGAAATC	174
	R：TGTCAGTATCAGTCCAGGTGGTCGC
OprD2	F：ACCTAGCCTCCTATGGCGTTCCC	150
	R：CGAGGTTGGTTTCGTGGTGCTT

基因	序列	产物大小（bp）
16S RNA	F：TAGGTGGTTCAGCAAGTTGGATGTGAAATC	174
	R：TGTCAGTATCAGTCCAGGTGGTCGC
OprD2	F：ACCTAGCCTCCTATGGCGTTCCC	150
	R：CGAGGTTGGTTTCGTGGTGCTT

组别	株数	Cathelicidin/LL-37	HBD-2
hUCMSCs实验组	5	5.90 ± 0.51	1.77 ± 0.25
NHLF对照组	5	3.98 ± 0.34^a	0.72 ± 0.25^a
空白对照组	5	3.09 ± 0.20^b	0.63 ± 0.20^b

组别	株数	Cathelicidin/LL-37	HBD-2
hUCMSCs实验组	5	5.90 ± 0.51	1.77 ± 0.25
NHLF对照组	5	3.98 ± 0.34^a	0.72 ± 0.25^a
空白对照组	5	3.09 ± 0.20^b	0.63 ± 0.20^b

组别	IPM	AK	CAZ	TZP	MEM	ATM
诱导前原始菌株	25	26	25	30	26	27
诱导后hUCMSCs组	13^*	27	26	29	18	25
诱导后NHLF对照组	12^*	26	27	29	13^*	26
诱导后空白对照组	12^*	26	25	28	14^*	25

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