VTechWorks Repository :: Browsing by Author "Galindo, Cristi L."

Browsing by Author "Galindo, Cristi L."

Now showing 1 - 9 of 9

Brucella melitensis VjbR and C12-HSL regulons: contributions of the N-dodecanoyl homoserine lactone signaling molecule and LuxR homologue VjbR to gene expression
Weeks, Jenni N.; Galindo, Cristi L.; Drake, Kenneth L.; Adams, L. Garry; Garner, Harold R.; Ficht, Thomas A. (2010-06-08)
Background Quorum sensing is a communication system that regulates gene expression in response to population density and often regulates virulence determinants. Deletion of the luxR homologue vjbR highly attenuates intracellular survival of Brucella melitensis and has been interpreted to be an indication of a role for QS in Brucella infection. Confirmation for such a role was suggested, but not confirmed, by the demonstrated in vitro synthesis of an auto-inducer (AI) by Brucella cultures. In an effort to further delineate the role of VjbR to virulence and survival, gene expression under the control of VjbR and AI was characterized using microarray analysis. Results Analyses of wildtype B. melitensis and isogenic ΔvjbR transciptomes, grown in the presence and absence of exogenous N-dodecanoyl homoserine lactone (C12-HSL), revealed a temporal pattern of gene regulation with variances detected at exponential and stationary growth phases. Comparison of VjbR and C12-HSL transcriptomes indicated the shared regulation of 127 genes with all but 3 genes inversely regulated, suggesting that C12-HSL functions via VjbR in this case to reverse gene expression at these loci. Additional analysis using a ΔvjbR mutant revealed that AHL also altered gene expression in the absence of VjbR, up-regulating expression of 48 genes and a luxR homologue blxR 93-fold at stationary growth phase. Gene expression alterations include previously un-described adhesins, proteases, antibiotic and toxin resistance genes, stress survival aids, transporters, membrane biogenesis genes, amino acid metabolism and transport, transcriptional regulators, energy production genes, and the previously reported fliF and virB operons. Conclusions VjbR and C12-HSL regulate expression of a large and diverse number of genes. Many genes identified as virulence factors in other bacterial pathogens were found to be differently expressed, suggesting an important contribution to intracellular survival of Brucella. From these data, we conclude that VjbR and C12-HSL contribute to virulence and survival by regulating expression of virulence mechanisms and thus controlling the ability of the bacteria to survive within the host cell. A likely scenario occurs via QS, however, operation of such a mechanism remains to be demonstrated.
Comparative Analyses of Transcriptional Profiles in Mouse Organs Using a Pneumonic Plague Model after Infection with Wild-Type Yersinia pestis CO92 and Its Braun Lipoprotein Mutant
Galindo, Cristi L.; Moen, Scott T.; Kozlova, Elena V.; Sha, Jian; Garner, Harold R.; Agar, Stacy L.; Chopra, Ashok K. (Hindawi, 2010-01-20)
We employed Murine GeneChips to delineate the global transcriptional profiles of the livers, lungs, and spleens in a mouse pneumonic plague infection model with wild-type (WT) Y. pestis CO92 and its Braun lipoprotein () mutant with reduced virulence. These organs showed differential transcriptional responses to infection with WT Y. pestis, but the overall host functional processes affected were similar across all three tissues. Gene expression alterations were found in inflammation, cytokine signaling, and apoptotic cell death-associated genes. Comparison of WT and mutant-infected mice indicated significant overlap in lipopolysaccharide- (LPS-) associated gene expression, but the absence of Lpp perturbed host cell signaling at critical regulatory junctions resulting in altered immune response and possibly host cell apoptosis. We generated a putative signaling pathway including major inflammatory components that could account for the synergistic action of LPS and Lpp and provided the mechanistic basis of attenuation caused by deletion of the lpp gene from Y. pestis in a mouse model of pneumonic plague.
Comparative Global Gene Expression Profiles of Wild-Type Yersinia pestis CO92 and Its Braun Lipoprotein Mutant at Flea and Human Body Temperatures
Galindo, Cristi L.; Sha, Jian; Moen, Scott T.; Agar, Stacy L.; Kirtley, Michelle L.; Foltz, Sheri M.; McIver, Lauren J.; Kozlova, E. V.; Garner, Harold R.; Chopra, Ashok K. (Hindawi, 2010-05-19)
Braun/murein lipoprotein (Lpp) is involved in inflammatory responses and septic shock. We previously characterized a lpp mutant of Yersinia pestis CO92 and found that this mutant was defective in surviving in macrophages and was attenuated in a mouse inhalation model of plague when compared to the highly virulent wild-type (WT) bacterium. We performed global transcriptional profiling of WT Y. pestis and its lpp mutant using microarrays. The organisms were cultured at 26 and 37 degrees Celsius to simulate the flea vector and mammalian host environments, respectively. Our data revealed vastly different effects of lpp mutation on the transcriptomes of Y. pestis grown at 37 versus . While the absence of Lpp resulted mainly in the downregulation of metabolic genes at , the Y. pestislpp mutant cultured at exhibited profound alterations in stress response and virulence genes, compared to WT bacteria. We investigated one of the stress-related genes (htrA) downregulated in the lpp mutant relative to WT Y. pestis. Indeed, complementation of the lpp mutant with the htrA gene restored intracellular survival of the Y. pestislpp mutant. Our results support a role for Lpp in Y. pestis adaptation to the host environment, possibly via transcriptional activation of htrA.
Dynamic innate immune responses of human bronchial epithelial cells to severe acute respiratory syndrome-associated coronavirus infection
Yoshikawa, Tomoki; Hill, Terence E.; Yoshikawa, Naoko; Popov, Vsevolod L.; Galindo, Cristi L.; Garner, Harold R.; Peters, C. J.; Tseng, Chien-Te (Kent) (Public Library of Science, 2010-01-15)
Human lung epithelial cells are likely among the first targets to encounter invading severe acute respiratory syndromeassociated coronavirus (SARS-CoV). Not only can these cells support the growth of SARS-CoV infection, but they are also capable of secreting inflammatory cytokines to initiate and, eventually, aggravate host innate inflammatory responses, causing detrimental immune-mediated pathology within the lungs. Thus, a comprehensive evaluation of the complex epithelial signaling to SARS-CoV is crucial for paving the way to better understand SARS pathogenesis. Based on microarraybased functional genomics, we report here the global gene response of 2B4 cells, a cloned bronchial epithelial cell line derived from Calu-3 cells. Specifically, we found a temporal and spatial activation of nuclear factor (NF)kB, activator protein (AP)-1, and interferon regulatory factor (IRF)-3/7 in infected 2B4 cells at 12-, 24-, and 48-hrs post infection (p.i.), resulting in the activation of many antiviral genes, including interferon (IFN)-b, -ls, inflammatory mediators, and many IFN-stimulated genes (ISGs). We also showed, for the first time, that IFN-b and IFN-ls were capable of exerting previously unrecognized, non-redundant, and complementary abilities to limit SARS-CoV replication, even though their expression could not be detected in infected 2B4 bronchial epithelial cells until 48 hrs p.i. Collectively, our results highlight the mechanics of the sequential events of antiviral signaling pathway/s triggered by SARS-CoV in bronchial epithelial cells and identify novel cellular targets for future studies, aiming at advancing strategies against SARS.
Inhibitory Role of Notch1 in Calcific Aortic Valve Disease
Acharya, Asha; Hans, Chetan P.; Koenig, Sara N.; Nichols, Haley A.; Galindo, Cristi L.; Garner, Harold R.; Merrill, Walter H.; Hinton, Robert B.; Garg, Vidu (PLoS ONE, 2011-11-16)
Aortic valve calcification is the most common form of valvular heart disease, but the mechanisms of calcific aortic valve disease (CAVD) are unknown. NOTCH1 mutations are associated with aortic valve malformations and adult-onset calcification in families with inherited disease. The Notch signaling pathway is critical for multiple cell differentiation processes, but its role in the development of CAVD is not well understood. The aim of this study was to investigate the molecular changes that occur with inhibition of Notch signaling in the aortic valve. Notch signaling pathway members are expressed in adult aortic valve cusps, and examination of diseased human aortic valves revealed decreased expression of NOTCH1 in areas of calcium deposition. To identify downstream mediators of Notch1, we examined gene expression changes that occur with chemical inhibition of Notch signaling in rat aortic valve interstitial cells (AVICs). We found significant downregulation of Sox9 along with several cartilage-specific genes that were direct targets of the transcription factor, Sox9. Loss of Sox9 expression has been published to be associated with aortic valve calcification. Utilizing an in vitro porcine aortic valve calcification model system, inhibition of Notch activity resulted in accelerated calcification while stimulation of Notch signaling attenuated the calcific process. Finally, the addition of Sox9 was able to prevent the calcification of porcine AVICs that occurs with Notch inhibition. In conclusion, loss of Notch signaling contributes to aortic valve calcification via a Sox9-dependent mechanism.
Role of SPI-1 Secreted Effectors in Acute Bovine Response to Salmonella enterica Serovar Typhimurium: A Systems Biology Analysis Approach
Lawhon, Sara D.; Khare, Sangeeta; Rossetti, Carlos A.; Everts, Robin E.; Galindo, Cristi L.; Luciano, Sarah A.; Figueiredo, Josely F.; Nunes, Jairo E. S.; Gull, Tamara; Davidson, George S.; Drake, Kenneth L.; Garner, Harold R.; Lewin, Harris A.; Baumler, Andreas J.; Adams, Leslie Garry (Public Library of Science, 2011-11-11)
Salmonella enterica Serovar Typhimurium (S. Typhimurium) causes enterocolitis with diarrhea and polymorphonuclear cell (PMN) influx into the intestinal mucosa in humans and calves. The Salmonella Type III Secretion System (T3SS) encoded at Pathogenicity Island I translocates Salmonella effector proteins SipA, SopA, SopB, SopD, and SopE2 into epithelial cells and is required for induction of diarrhea. These effector proteins act together to induce intestinal fluid secretion and transcription of C-X-C chemokines, recruiting PMNs to the infection site. While individual molecular interactions of the effectors with cultured host cells have been characterized, their combined role in intestinal fluid secretion and inflammation is less understood. We hypothesized that comparison of the bovine intestinal mucosal response to wild type Salmonella and a SipA, SopABDE2 effector mutant relative to uninfected bovine ileum would reveal heretofore unidentified diarrhea-associated host cellular pathways. To determine the coordinated effects of these virulence factors, a bovine ligated ileal loop model was used to measure responses to wild type S. Typhimurium (WT) and a ΔsipA, sopABDE2 mutant (MUT) across 12 hours of infection using a bovine microarray. Data were analyzed using standard microarray analysis and a dynamic Bayesian network modeling approach (DBN). Both analytical methods confirmed increased expression of immune response genes to Salmonella infection and novel gene expression. Gene expression changes mapped to 219 molecular interaction pathways and 1620 gene ontology groups. Bayesian network modeling identified effects of infection on several interrelated signaling pathways including MAPK, Phosphatidylinositol, mTOR, Calcium, Toll-like Receptor, CCR3, Wnt, TGF-β, and Regulation of Actin Cytoskeleton and Apoptosis that were used to model of host-pathogen interactions. Comparison of WT and MUT demonstrated significantly different patterns of host response at early time points of infection (15 minutes, 30 minutes and one hour) within phosphatidylinositol, CCR3, Wnt, and TGF-β signaling pathways and the regulation of actin cytoskeleton pathway.
Selective amplification of Brucella melitensis mRNA from a mixed host:pathogen total RNA
Rossetti, Carlos A.; Galindo, Cristi L.; Garner, Harold R.; Adams, L. Garry (2010-09-28)
Background Brucellosis is a worldwide anthropozoonotic disease caused by an in vivo intracellular pathogen belonging to genus Brucella. The characterization of brucelae transcriptome's during host-pathogen interaction has been limited due to the difficulty of obtaining an adequate quantity of good quality eukaryotic RNA-free pathogen RNA for downstream applications. Findings Here, we describe a combined protocol to prepare RNA from intracellular B. melitensis in a quantity and quality suitable for pathogen gene expression analysis. Initially, B. melitensis total RNA was enriched from a host:pathogen mixed RNA sample by reducing the eukaryotic RNA..Then, to increase the Brucella RNA concentration and simultaneously minimize the contaminated host RNA in the mixed sample, a specific primer set designed to anneal to all B. melitensis ORF allows the selective linear amplification of sense-strand prokaryotic transcripts in a previously enriched RNA sample. Conclusion The novelty of the method we present here allows analysis of the gene expression profile of B. melitensis when limited amounts of pathogen RNA are present, and is potentially applicable to both in vivo and in vitro models of infection, even at early infection time points.
Systems Biology Analysis of Gene Expression during In Vivo Mycobacterium avium paratuberculosis Enteric Colonization Reveals Role for Immune Tolerance
Khare, Sangeeta; Lawhon, Sara D.; Drake, Kenneth L.; Nunes, Jairo E. S.; Figueiredo, Josely F.; Rossetti, Carlos A.; Gull, Tamara; Everts, Robin E.; Lewin, Harris A.; Galindo, Cristi L.; Garner, Harold R.; Adams, Leslie Garry (Public Library of Science, 2012-08-17)
Survival and persistence of Mycobacterium avium subsp. paratuberculosis (MAP) in the intestinal mucosa is associated with host immune tolerance. However, the initial events during MAP interaction with its host that lead to pathogen survival, granulomatous inflammation, and clinical disease progression are poorly defined. We hypothesize that immune tolerance is initiated upon initial contact of MAP with the intestinal Peyer's patch. To test our hypothesis, ligated ileal loops in neonatal calves were infected with MAP. Intestinal tissue RNAs were collected (0.5, 1, 2, 4, 8 and 12 hrs post-infection), processed, and hybridized to bovine gene expression microarrays. By comparing the gene transcription responses of calves infected with the MAP, informative complex patterns of expression were clearly visible. To interpret these complex data, changes in the gene expression were further analyzed by dynamic Bayesian analysis, and genes were grouped into the specific pathways and gene ontology categories to create a holistic model. This model revealed three different phases of responses: i) early (30 min and 1 hr post-infection), ii) intermediate (2, 4 and 8 hrs post-infection), and iii) late (12 hrs post-infection). We describe here the data that include expression profiles for perturbed pathways, as well as, mechanistic genes (genes predicted to have regulatory influence) that are associated with immune tolerance. In the Early Phase of MAP infection, multiple pathways were initiated in response to MAP invasion via receptor mediated endocytosis and changes in intestinal permeability. During the Intermediate Phase, perturbed pathways involved the inflammatory responses, cytokine-cytokine receptor interaction, and cell-cell signaling. During the Late Phase of infection, gene responses associated with immune tolerance were initiated at the level of T-cell signaling. Our study provides evidence that MAP infection resulted in differentially regulated genes, perturbed pathways and specifically modified mechanistic genes contributing to the colonization of Peyer's patch.
Transcriptional profile of isoproterenol-induced cardiomyopathy and comparison to exercise-induced cardiac hypertrophy and human cardiac failure
Galindo, Cristi L.; Skinner, Michael A.; Errami, Mounir; Olson, L Danielle; Watson, David A.; Li, Jing; McCormick, John F.; McIver, Lauren J.; Kumar, Neil M.; Pham, Thinh Q.; Garner, Harold R. (2009-12-09)
Background Isoproterenol-induced cardiac hypertrophy in mice has been used in a number of studies to model human cardiac disease. In this study, we compared the transcriptional response of the heart in this model to other animal models of heart failure, as well as to the transcriptional response of human hearts suffering heart failure. Results We performed microarray analyses on RNA from mice with isoproterenol-induced cardiac hypertrophy and mice with exercise-induced physiological hypertrophy and identified 865 and 2,534 genes that were significantly altered in pathological and physiological cardiac hypertrophy models, respectively. We compared our results to 18 different microarray data sets (318 individual arrays) representing various other animal models and four human cardiac diseases and identified a canonical set of 64 genes that are generally altered in failing hearts. We also produced a pairwise similarity matrix to illustrate relatedness of animal models with human heart disease and identified ischemia as the human condition that most resembles isoproterenol treatment. Conclusion The overall patterns of gene expression are consistent with observed structural and molecular differences between normal and maladaptive cardiac hypertrophy and support a role for the immune system (or immune cell infiltration) in the pathology of stress-induced hypertrophy. Cross-study comparisons such as the results presented here provide targets for further research of cardiac disease that might generally apply to maladaptive cardiac stresses and are also a means of identifying which animal models best recapitulate human disease at the transcriptional level.

Browsing by Author "Galindo, Cristi L."

Results Per Page

Sort Options