Metformin strengthens uroepithelial immunity against E. coli infection

Rakesh Kumar Majhi,Hanna Brauner,Annelie Brauner,John Kerr White,Andrea Scheffschick,Witchuda Kamolvit,Soumitra Mohanty

doi:10.1038/s41598-021-98223-1

Rakesh Kumar Majhi, Hanna Brauner + Show 5 more

Open Access

https://doi.org/10.1038/s41598-021-98223-1

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Abstract

Urinary tract infection frequently caused by E. coli is one of the most common bacterial infections. Increasing antibiotic resistance jeopardizes successful treatment and alternative treatment strategies are therefore mandatory. Metformin, an oral antidiabetic drug, has been shown to activate macrophages in the protection against certain infecting microorganisms. Since epithelial cells often form the first line of defense, we here investigated the effect on uroepithelial cells during E. coli infection. Metformin upregulated the human antimicrobial peptides cathelicidin LL-37 and RNase7 via modulation of the TRPA1 channel and AMPK pathway. Interestingly, metformin stimulation enriched both LL-37 and TRPA1 in lysosomes. In addition, metformin specifically increased nitric oxide and mitochondrial, but not cytosolic ROS. Moreover, metformin also triggered mRNA expression of the proinflammatory cytokines IL1B, CXCL8 and growth factor GDF15 in human uroepithelial cells. The GDF15 peptide stimulated macrophages increased LL-37 expression, with increased bacterial killing. In conclusion, metformin stimulation strengthened the innate immunity of uroepithelial cells inducing enhanced extracellular and intracellular bacterial killing suggesting a favorable role of metformin in the host defense.

Highlights

Bladder epithelial cells form the first line of defense against invading pathogens and deploy multiple intrinsic mechanisms to clear invading bacteria during urinary tract infection[1]
Metformin treatment upregulated the expression of the antimicrobial peptides (AMPs) CAMP and RNASE7 mRNA and corresponding intracellular proteins LL-37 and RNase[7], compared to untreated uroepithelial cells, TERT-NHUC (Fig. 1A–F; Fig. S1A–D) and 5637 (Fig. S1E–J). 4 mM metformin treatment was found to be optimal based on the dose response profiles of CAMP and RNASE7 expression in both TERT-NHUC and 5637 (Fig. S1A,B,E,H)
Metformin treatment resulted in increased localization of LL-37, but not RNase[7] in the lysosomes of TERT-NHUC (Fig. 1G) and 5637 cells (Fig. S1K), as shown by confocal microscopy

Summary

Introduction

Bladder epithelial cells form the first line of defense against invading pathogens and deploy multiple intrinsic mechanisms to clear invading bacteria during urinary tract infection[1]. Metformin was shown to be effective against Mycobacterium tuberculosis and Legionella pneumonia, by inducing mitochondrial ROS production in macrophages[13,14]. This has led to the possible use of metformin complementing antibacterial drugs by stimulating host cells[15]. In this work we assessed the efficacy of metformin in strengthening the uroepithelial cells against E. coli infection and explored the role of the nociceptive cationic channel transient receptor potential ankyrin 1 (TRPA1) and 5’ adenosine monophosphate-activated protein kinase (AMPK) pathway in metformin stimulated expression of the antimicrobial peptides LL-37 and RNase[7], respectively. Our data show that metformin treatment stimulates multiple host-protective responses, resulting in increased intra- and extracellular E. coli killing

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Conclusion