Extracellular matrix stiffness modulates host-bacteria interactions and antibiotic therapy of bacterial internalization

Abstract

Pathogenic bacteria evolve multiple strategies to hijack host cells for intracellular survival and persistent infections. Previous studies have revealed the intricate interactions between bacteria and host cells at genetic, biochemical and even single molecular levels. Mechanical interactions and mechanotransduction exert a crucial impact on the behaviors and functions of pathogenic bacteria and host cells, owing to the ubiquitous mechanical microenvironments like extracellular matrix (ECM) stiffness. Nevertheless, it remains unclear whether and how ECM stiffness modulates bacterial infections and the sequential outcome of antibacterial therapy. Here we show that bacteria tend to adhere to and invade epithelial cells located on the regions with relatively high traction forces. ECM stiffness regulates spatial distributions of bacteria during the invasion through arrangements of F-actin cytoskeletons in host cells. Depolymerization of cytoskeletons in the host cells induced by bacterial infection decreases intracellular accumulation of antibiotics, thus preventing the eradication of invaded bacterial pathogens. These findings not only reveal the key regulatory role of ECM stiffness, but suggest that the coordination of cytoskeletons may provide alternative approaches to improve antibiotic therapy against multidrug resistant bacteria in clinic.

Introduction

The infection caused by pathogenic bacteria, particularly multi-drug resistant bacteria, is becoming one of the most common infections worldwide at an alarming rate [1,2]. Such a global threat created by infectious bacteria is a major public health concern due to the big gap from antibiotic development to clinical applications [3,4]. Alternatively, revitalization of existing antibiotics is a critical approach to substantially combat bacterial pathogens. To extend the lifetime of antibiotics, we need to explore the bacterial pathogenesis at the interface between bacteria and host cells instead of focusing solely on bacteria themselves, thus necessitating the design of therapeutic approaches to effectively subvert bacterial infections.

Both pathogenic bacteria and host cells essentially live in complex mechanical microenvironments [[5], [6], [7]]. There is increasing evidence indicating that mechanical aspects of microenvironments, e.g., fluid shear force, osmotic stress, mechanical stretch, interfacial adhesion force, as well as extracellular matrix (ECM) stiffness, play a crucial role in modulating physiological functions and behaviors of bacteria and cells [[8], [9], [10], [11]], as well as cellular adhesion, migration, proliferation and division [12,13]. For instance, it was reported that bacteria-induced asymmetric adhesion can cause mechanical tension and therefore constrain microcolony morphogenesis [8,9], whereas mechanical instability and interfacial energy are found to be key driving forces for mechano-morphogenesis of bacterial biofilms [11]. Experiments based on atomic force microscopy found that the increase in tensile stresses induced by local stress concentration and the decrease in material strength at the pre-cleavage furrow regulated by peptidoglycan hydrolases together play a pivotal role in controlling bacterial cell division [10]. Likewise, recent advances identified the importance of cellular biomechanics in mediating host-bacteria interaction and resulting infection, where they revealed that collective mechanical response of epithelial monolayers triggered by innate immune signals is able to lead to the extrusion of bacterially infected cells, thus limiting the spread of internalized bacteria along the basal monolayer [14].

Previous efforts have made great achievements on signal transduction of numerous biochemical information to modulate cellular homeostasis during the interactions between bacteria and host cells [15,16]. Recent progresses in mechanobiology revealed that mechanical factors of microsurroundings, e.g., ECM stiffness [17,18], enable cells/bacteria to trigger a cascade of mechanotransduction signal pathways [19,20], which then modulate the interaction between host cells and pathogens. For example, it is reported that ECM stiffness-regulated mechanical properties of host cells can either promote apoptosis [21] or lead to abundant actin filaments [22], essentially depending on rigidities of the host cells.

In fact, there always exist complicated mechanotransduction processes among pathogenic bacteria, host cells and ECM microenvironments in the bodies with clinical symptoms such as diarrhea and wound infection [6,23,24]. Yet little is known about whether and how ECM stiffness regulates bacteria-cell interactions and the outcome of antibacterial therapy. Here, we first report a new technique for the construction of in vitro infection model using elastic substrates with physiologically comparable rigidities, rather than rigid substrates or traditional cell culture dishes. By investigating the interactions between bacteria and host cell monolayers cultured on extracellular substrates with tissues-like rigidities, we reveal that the law that ECM stiffness mediates spatial distributions of invaded bacteria through cellular traction forces and cytoskeletons, and that there is a transition of bacterial invasion modes as ECM stiffness increases. Further, we show that accumulation of intracellular antibiotics and the corresponding efficacy are ECM stiffness dependent in essence.