Paired-end RNA Sequencing on human dermal microvascular endothelial cells (HMEC-1) infected or not with Borrelia burgdorferi (MOI= 200 bacteria/cell) examined at different times post-infection (namely 4, 24 or 48 h post-infection)

Purpose: When we infect human dermal microvascular endothelial cells (HMEC-1) infected or not with Borrelia burgdorferi (MOI= 200 bacteria/cell) and examine the host cell kinematics and dynamics over the course of 3 days post-infection we observe a change in those at early times post-infection, but at late times (> 20 h post-infection) cells from infected or not wells behave similarly. To get a sense of the biochemical signals that might be driving the changes in host cell mechanics, we performed RNA-Sequencing and gene expression profiling for uninfected HMEC-1 cells or cells infected for 4, 24 or 48 h with Borrelia burgdorferi at an MOI of 200 bacteria/cell. Methods and Results: By analyzing the differentially expressed genes between all conditions we find a significant number of up- or down-regulated genes infected and uninfected cells just at 4 h post-infection. Through pathway enrichemnet analysis we find that compared to cells not exposed to infection infected cells showed significant upregulation to genes related IL-17 and NF-kB and TNF related signaling pathways. Interestingly these changes are not present at 24 or 48 h post-infection where cells from infected wells are more similar gene expression-wise to uninfected cells. Conclusions: The changes in kinematics and dynamics of HMEC-1 cells exposed to Borrelia burgdorferi (MOI= 200 bacteria/cell) at early times post-infection are associated with changes in innate immune signaing of the host cells. At later times post-infection were these changes in gene expression are not present anymore, host cell kinematics and dynamics are identical between infected and uninfected cells. Thus it is possible that innate immune singaling might be underlying the changes that host cells undergo in their motion and traction forces at early infection. Overall design: Endothelial HMEC-1 cell mRNA profiles of cells previously residing in confluency on glass substrates coated with 0.25 mg/mL collagen I and being (1) uninfected; or (2) infected with Borrelia burgdorferi (MOI=200) for different amounts of time (4, 24 or 48 h) were generated in quadruplicate, using Illumina NovaSeq 6000 using the NovaSeq 6000 S4 Reagent Kit (300 cycles). Equipment: Illumina with 2x150 read length and 40 million reads per sample. ----------------------------------------------------------- Reasearch funded by Lyme disease association (LDA) research grant award, for the research proposal "Biomechanical alterations of endothelial cells infected with Borrelia burgdorferi".