Rickettsia rickettsiiInfection of Human Macrovascular and Microvascular Endothelial Cells Reveals Activation of Both Common and Cell Type-Specific Host Response Mechanisms

Abstract

Although inflammation and altered barrier functions of the vasculature, due predominantly to the infection of endothelial cell lining of small and medium-sized blood vessels, represent salient pathological features of human rickettsioses, the interactions between pathogenic rickettsiae and microvascular endothelial cells remain poorly understood. We have investigated the activation of nuclear transcription factor-kappa B (NF-B) and p38 mitogen-activated protein (MAP) kinase, expression of heme oxygenase 1 (HO-1) and cyclooxygenase 2 (COX-2), and secretion of chemokines and prostaglandins after Rickettsia rickettsii infection of human cerebral, dermal, and pulmonary microvascular endothelial cells in comparison with pulmonary artery cells of macrovascular origin. NF-B and p38 kinase activation and increased HO-1 mRNA expression were clearly evident in all cell types, along with relatively similar susceptibility to R. rickettsii infection in vitro but considerable variations in the intensities/kinetics of the aforementioned host responses. As expected, the overall activation profiles of macrovascular endothelial cells derived from human pulmonary artery and umbilical vein were nearly identical. Interestingly, cerebral endothelial cells displayed a marked refractoriness in chemokine production and secretion, while all other cell types secreted various levels of interleukin-8 (IL-8) and monocyte chemoattractant protein 1 (MCP-1) in response to infection. A unique feature of all microvascular endothelial cells was the lack of induced COX-2 expression and resultant inability to secrete prostaglandin E 2 after R. rickettsii infection. Comparative evaluation thus yields the first experimental evidence for the activation of both common and unique cell type-specific host response mechanisms in macrovascular and microvascular endothelial cells infected with R. rickettsii, a prototypical species known to cause Rocky Mountain spotted fever in humans.The obligately intracellular Alphaproteobacteria belonging to genus Rickettsia are now classified into four subgroups, which include an ancestral group and a transitional group in addition to two major antigenically defined groups comprised of spotted fever and typhus species (12). Pathogenic rickettsiae display tropism for vascular endothelial cell lining of small and mediumsized blood vessels in their mammalian hosts. As a consequence, the predominant pathological features of resultant clinical syndromes-for example, Rocky Mountain spotted fever due to Rickettsia rickettsii-are generally attributed to altered vascular functions due to preferential infection of the endothelium (27, 36). However, despite critical contributions of endothelial injury, inflammation, and dysfunction to the pathogenesis of vasculotropic rickettsioses, the biological basis of rickettsial interactions with microvascular endothelium of vertebrate hosts still remains poorly understood.An initial breakthrough in the laboratory investigations of the interplay between host cells and pathogenic rickettsiae wa...

Small Regulatory RNAs of Rickettsia conorii

“…Cells were infected with R. conorii at an MOI of 20 following our established protocol62. Briefly, cell monolayers were incubated with R. conorii in a minimum volume of culture medium to ensure efficient adhesion and internalization.…”

Endothelial Cell Proteomic Response to Rickettsia conorii Infection Reveals Activation of the Janus Kinase (JAK)-Signal Transducer and Activator of Transcription (STAT)-Inferferon Stimulated Gene (ISG)15 Pathway and Reprogramming Plasma Membrane Integrin/Cadherin Signaling

“…examined (36). Downstream of IFN, activation of the STAT1 signaling pathway triggers production of ISG, MX1, SOCS, and other proteins important in host defense (37).…”

Regulation of inducible heme oxygenase and cyclooxygenase isozymes in a mouse model of spotted fever group rickettsiosis

“…However, the involvement of HO-1 as an anti-oxidant host protective mechanism has only been established for in vitro responses of endothelial cells of different origin to R. rickettsii [13,17]. To fill this critical gap in the knowledge, we first investigated HO-1 mRNA expression in different tissues (brain, liver, lungs, and spleen) after infection with three different doses of R. conorii ranging from 2.25×10 3 to 2.25×10 5 pfu per mouse.…”

“…Equal amounts of RNA (6-10 μg) from infected mouse tissues and corresponding controls were subjected to Northern blot analysis using radioactively labeled cDNA probes as described earlier [13,16,17]. Murine HO-1 probe was synthesized by PCR amplification of a 155 bp fragment using “Super Array” primer set (Frederick, MD).…”

“…Our recent studies, however, further suggest that there exist differences in the COX-2 response of different types of host endothelial cells to R. rickettsii infection. Whereas infection of primary cultures of macrovascular HUVECs (derived from a large vessel) results in an apparent biphasic pattern of induced expression, microvascular EC (HMECs) are largely refractory to prominent changes in the regulation of COX-2 expression at the levels of both mRNA and protein [15-17]. On the other hand, induction of HO-1 in both macro- as well as microvascular endothelium is relatively identical [17].…”