Streptococcus canis interacts with smooth muscle cells in a 3D-co-cultivation system with endothelial cells under microfluidic conditions

Introduction: The opportunistic zoonotic bacterium Streptococcus canis (S. canis) is known as a bacterial pathogen of companion animals and of domesticated live stock. The range of infection severity extends from minor urinary tract infections to systemic diseases such as necrotizing fasciitis, and even sepsis. Consequently, S. canis can enter the blood circulation and adheres to the blood vessel endothelium. The inner lining of the vasculature is formed by endothelial cells (EC) embedded in a collagen-rich extracellular matrix supported by smooth muscle cells (SMC).

Goals: So far, the pathophysiology of the transcellular interaction between S. canis and each single component forming the blood vessel wall remains elusive. We aim to decipher bacterial and cellular factors mediating S. canis adherence, cell internalization and transmigration of the complex blood vessel wall throughout systemic bloodstream infection.

Materials & Methods: Cell culture-based reconstruction of vessel wall was achieved by co-cultivation of human primary EC and SMC within a 3D-collagen matrix. To mimick the physiological bloodstream, the co-cultivation was introduced to a microfluidic system. After optimization of cell-type-specific immunofluorescent staining procedure, bacterial infection with S. canis was conducted. Electron microscopic (EM) imaging visualized morphological response to bacterial infection.

Results: A 3D-co-cultivation system based on EC and SMC embedded within a collagen matrix was successfully established in a microfluidic system. Microscopic and EM visualization showed that S. canis adheres to EC and is also invasive. Furthermore, S. canis traverses even under high shear stress conditions the EC layer and the collagen matrix and enters deeper layers of the co-cultivation system, thereby attaching to underlaying SMC.

Summary: The established 3D-co-cultivation system provides a suitable infection model for the elucidation of pathomechanisms determining bacterial bloodstream infections.