Systems Biology Analysis of Temporal In vivo Brucella melitensis and Bovine Transcriptomes Predicts host:Pathogen Protein-Protein Interactions.

To date, fewer than 200 gene-products have been identified as Brucella virulence factors, and most were characterized individually without considering how they are temporally and coordinately expressed or secreted during the infection process. Here, we describe and analyze the in vivo temporal transcriptional profile of Brucella melitensis during the initial 4 h interaction with cattle. Pathway analysis revealed an activation of the "Two component system" providing evidence that the in vivo Brucella sense and actively regulate their metabolism through the transition to an intracellular lifestyle. Contrarily, other Brucella pathways involved in virulence such as "ABC transporters" and "T4SS system" were repressed suggesting a silencing strategy to avoid stimulation of the host innate immune response very early in the infection process. Also, three flagellum-encoded loci (BMEII0150-0168, BMEII1080-1089, and BMEII1105-1114), the "flagellar assembly" pathway and the cell components "bacterial-type flagellum hook" and "bacterial-type flagellum" were repressed in the tissue-associated B. melitensis, while RopE1 sigma factor, a flagellar repressor, was activated throughout the experiment. These results support the idea that Brucella employ a stealthy strategy at the onset of the infection of susceptible hosts. Further, through systems-level in silico host:pathogen protein-protein interactions simulation and correlation of pathogen gene expression with the host gene perturbations, we identified unanticipated interactions such as VirB11::MAPK8IP1; BtaE::NFKBIA, and 22 kDa OMP precursor::BAD and MAP2K3. These findings are suggestive of new virulence factors and mechanisms responsible for Brucella evasion of the host's protective immune response and the capability to maintain a dormant state. The predicted protein-protein interactions and the points of disruption provide novel insights that will stimulate advanced hypothesis-driven approaches toward revealing a clearer understanding of new virulence factors and mechanisms influencing the pathogenesis of brucellosis.

Concordant changes of plasma and kidney microRNA in the early stages of acute kidney injury: time course in a mouse model of bilateral renal ischemia-reperfusion.

BACKGROUND: Acute kidney injury (AKI) is a syndrome characterized by the rapid loss of the kidney excretory function and is strongly associated with increased early and long-term patient morbidity and mortality. Early diagnosis of AKI is challenging; therefore we profiled plasma microRNA in an effort to identify potential diagnostic circulating markers of renal failure. The goal of the present study was to investigate the dynamic relationship of circulating and renal microRNA profiles within the first 24 hours after bilateral ischemia-reperfusion kidney injury in mice. METHODOLOGY/PRINCIPAL FINDINGS: Bilateral renal ischemia was induced in C57Bl/6 mice (n = 10 per group) by clamping the renal pedicle for 27 min. Ischemia-reperfusion caused highly reproducible, progressive, concordant elevation of miR-714, miR-1188, miR-1897-3p, miR-877*, and miR-1224 in plasma and kidneys at 3, 6 and 24 hours after acute kidney injury compared to the sham-operated mice (n = 5). These dynamics correlated with histologic findings of kidney injury and with a conventional plasma marker of renal dysfunction (creatinine). Pathway analysis revealed close association between miR-1897-3p and Nucks1 gene expression, which putative downstream targets include genes linked to renal injury, inflammation and apoptosis. CONCLUSIONS/SIGNIFICANCE: Systematic profiling of renal and plasma microRNAs in the early stages of experimental AKI provides the first step in advancing circulating microRNAs to the level of promising novel biomarkers.

Longitudinal in vivo analysis of the region-specific efficacy of parathyroid hormone in a rat cortical defect model.

PTH has been shown to enhance fracture repair; however, exactly when and where PTH acts in this process remains to be elucidated. Therefore, we conducted a longitudinal, region-specific analysis of bone regeneration in mature, osteopenic rats using a cortical defect model. Six-month-old rats were ovariectomized, and allowed to lose bone for 2 months, before being subjected to bilateral 2-mm circular defects in their femoral diaphyses. They were then treated for 5 wk with hPTH1-38 at doses of 0, 3, 10, or 30 microg/kg . d and scanned weekly by in vivo quantitative computed tomography. Quantitative computed tomography analyses showed temporal, dose-dependent increases in mineralization in the defects, intramedullary (IM) spaces, and whole diaphyses at the defect sites. Histomorphometry confirmed PTH stimulation of primarily woven bone in the defects and IM spaces, but not the periosteum. After necropsy, biomechanical testing identified an increase in strength at the highest PTH dose. Serum procollagen type I N-terminal propeptide concentration showed a transient increase due to drilling, but procollagen type I N-terminal propeptide also increased with PTH treatment, whereas tartrate-resistant acid phosphatase unexpectedly decreased. Analyses of lumber vertebra confirmed systemic efficacy of PTH at a nonfracture site. In summary, PTH dose dependently induced new bone formation within defects, at endocortical surfaces, and in IM spaces, resulting in faster and greater bone healing, as well as efficacy at other skeletal sites. The effects of PTH were kinetic, region specific, and most apparent at high doses that may not be entirely clinically relevant; therefore, clinical studies are necessary to clarify the therapeutic utility of PTH in bone healing.