The Impact of Different Antibiotic Regimens on the Emergence of Antimicrobial-Resistant Bacteria

Backgroud: The emergence and ongoing spread of antimicrobial-resistant bacteria is a major public health threat. Infections caused by antimicrobial-resistant bacteria are associated with substantially higher rates of morbidity and mortality compared to infections caused by antimicrobial-susceptible bacteria. The emergence and spread of these bacteria is complex and requires incorporating numerous interrelated factors which clinical studies cannot adequately address. Methods/Principal Findings: A model is created which incorporates several key factors contributing to the emergence and spread of resistant bacteria including the effects of the immune system, acquisition of resistance genes and antimicrobial exposure. The model identifies key strategies which would limit the emergence of antimicrobial-resistant bacterial strains. Specifically, the simulations show that early initiation of antimicrobial therapy and combination therapy with two antibiotics prevents the emergence of resistant bacteria, whereas shorter courses of therapy and sequential administration of antibiotics promote the emergence of resistant strains. Conclusions/Significance: The principal findings suggest that (i) shorter lengths of antibiotic therapy and early interruption of antibiotic therapy provide an advantage for the resistant strains, (ii) combination therapy with two antibiotics prevents the emergence of resistance strains in contrast to sequential antibiotic therapy, and (iii) early initiation of antibiotics is among the most important factors preventing the emergence of resistant strains. These findings provide new insights into strategies aimed at optimizing the administration of antimicrobials for the treatment of infections and the prevention of the emergence of antimicrobial resistance

Human Papillomavirus-16 E7 Interacts with Glutathione S-Transferase P1 and Enhances Its Role in Cell Survival

Background:Human Papillomavirus (HPV)-16 is a paradigm for "high-risk" HPVs, the causative agents of virtually all cervical carcinomas. HPV E6 and E7 viral genes are usually expressed in these tumors, suggesting key roles for their gene products, the E6 and E7 oncoproteins, in inducing malignant transformation.Methodology/Principal Findings:By protein-protein interaction analysis, using mass spectrometry, we identified glutathione S-transferase P1-1 (GSTP1) as a novel cellular partner of the HPV-16 E7 oncoprotein. Following mapping of the region in the HPV-16 E7 sequence that is involved in the interaction, we generated a three-dimensional molecular model of the complex between HPV-16 E7 and GSTP1, and used this to engineer a mutant molecule of HPV-16 E7 with strongly reduced affinity for GSTP1.When expressed in HaCaT human keratinocytes, HPV-16 E7 modified the equilibrium between the oxidized and reduced forms of GSTP1, thereby inhibiting JNK phosphorylation and its ability to induce apoptosis. Using GSTP1-deficient MCF-7 cancer cells and siRNA interference targeting GSTP1 in HaCaT keratinocytes expressing either wild-type or mutant HPV-16 E7, we uncovered a pivotal role for GSTP1 in the pro-survival program elicited by its binding with HPV-16 E7.Conclusions/Significance:This study provides further evidence of the transforming abilities of this oncoprotein, setting the groundwork for devising unique molecular tools that can both interfere with the interaction between HPV-16 E7 and GSTP1 and minimize the survival of HPV-16 E7-expressing cancer cells. © 2009 Mileo et al

Neurotrophic requirements of human motor neurons defined using amplified and purified stem-cell derived cultures

Neurotrophic requirements of human motor neurons defined using amplified and purified stem-cell derived culturesHuman motor neurons derived from embryonic and induced pluripotent stem cells (hESCs and hiPSCs) are a potentially important tool for studying motor neuron survival and pathological cell death. However, their basic survival requirements remain poorly characterized. Here, we sought to optimize a robust survival assay and characterize their response to different neurotrophic factors. First, to increase motor neuron yield, we screened a small-molecule collection and found that the Rho-associated kinase (ROCK) inhibitor Y-27632 enhances motor neuron progenitor proliferation up to 4-fold in hESC and hiPSC cultures. Next, we FACS-purified motor neurons expressing the Hb9::GFP reporter from Y-27632-amplified embryoid bodies and cultured them in the presence of mitotic inhibitors to eliminate dividing progenitors. Survival of these purified motor neurons in the absence of any other cell type was strongly dependent on neurotrophic support. GDNF, BDNF and CNTF all showed potent survival effects (EC(50) 1-2 pM). The number of surviving motor neurons was further enhanced in the presence of forskolin and IBMX, agents that increase endogenous cAMP levels. As a demonstration of the ability of the assay to detect novel neurotrophic agents, Y-27632 itself was found to support human motor neuron survival. Thus, purified human stem cell-derived motor neurons show survival requirements similar to those of primary rodent motor neurons and can be used for rigorous cell-based screening.This work was funded by Project A.L.S., P2ALS and NYSTEM grant number CO24415. The work of N.J.L. was supported by the Portuguese Foundation for Science and Technology SFRH/BD/33421/2008 and the Luso-American Development Foundation. B.J.-K. was supported by the National Institute of Neurological Disorders and Stroke (NINDS). L.R. was supported by the Swedish Brain Foundation/Hjarnfonden. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript