A needle-free technique for interstitial fluid sample acquisition using a lorentz-force actuated jet injector.

We present a novel method of quickly acquiring dermal interstitial fluid (ISF) samples using a Lorentz-force actuated needle-free jet injector. The feasibility of the method is first demonstrated on post-mortem porcine tissue. The jet injector is used to first inject a small volume of physiological saline to breach the skin, and the back-drivability of the actuator is utilized to create negative pressure in the ampoule and collect ISF. The effect of the injection and extraction parameters on sample dilution and extracted volumes is investigated. A simple finite element model is developed to demonstrate why this acquisition method results in faster extractions than conventional sampling methods. Using this method, we are able to collect a sample that contains up to 3.5% ISF in 3.1s from post-mortem skin. The trends revealed from experimentation on post-mortem skin are then used to identify the parameters for a live animal study. The feasibility of the acquisition process is successfully demonstrated using live rats; the process is revealed to extract samples that have been diluted by a factor of 111-125.

Needle-free interstitial fluid acquisition using a Lorentz-force actuated jet injector.

The feasibility of a new method of quickly acquiring interstitial fluid (ISF) samples using a Lorentz-force actuated needle-free jet injector is demonstrated on ex vivo porcine tissue. The jet injector is used to first inject a small volume of physiological saline to breach the skin, and the back-drivability of the actuator is utilized to create a vacuum in the ampoule and collect ISF. Injection and extraction parameters are tested and optimized for minimal acquired sample dilution and extracted volume consistency. Using this method, we are able to collect a sample that contains up to 3.5% ISF in 3.1 s.

A network integration approach to predict conserved regulators related to pathogenicity of influenza and SARS-CoV respiratory viruses.

Respiratory infections stemming from influenza viruses and the Severe Acute Respiratory Syndrome corona virus (SARS-CoV) represent a serious public health threat as emerging pandemics. Despite efforts to identify the critical interactions of these viruses with host machinery, the key regulatory events that lead to disease pathology remain poorly targeted with therapeutics. Here we implement an integrated network interrogation approach, in which proteome and transcriptome datasets from infection of both viruses in human lung epithelial cells are utilized to predict regulatory genes involved in the host response. We take advantage of a novel "crowd-based" approach to identify and combine ranking metrics that isolate genes/proteins likely related to the pathogenicity of SARS-CoV and influenza virus. Subsequently, a multivariate regression model is used to compare predicted lung epithelial regulatory influences with data derived from other respiratory virus infection models. We predicted a small set of regulatory factors with conserved behavior for consideration as important components of viral pathogenesis that might also serve as therapeutic targets for intervention. Our results demonstrate the utility of integrating diverse 'omic datasets to predict and prioritize regulatory features conserved across multiple pathogen infection models.

Implication of inflammatory macrophages, nuclear receptors, and interferon regulatory factors in increased virulence of pandemic 2009 H1N1 influenza A virus after host adaptation.

While pandemic 2009 H1N1 influenza A viruses were responsible for numerous severe infections in humans, these viruses do not typically cause corresponding severe disease in mammalian models. However, the generation of a virulent 2009 H1N1 virus following serial lung passage in mice has allowed for the modeling of human lung pathology in this species. Genetic determinants of mouse-adapted 2009 H1N1 viral pathogenicity have been identified, but the molecular and signaling characteristics of the host response following infection with this adapted virus have not been described. Here we compared the gene expression response following infection of mice with A/CA/04/2009 (CA/04) or the virulent mouse-adapted strain (MA-CA/04). Microarray analysis revealed that increased pathogenicity of MA-CA/04 was associated with the following: (i) an early and sustained inflammatory and interferon response that could be driven in part by interferon regulatory factors (IRFs) and increased NF-κB activation, as well as inhibition of the negative regulator TRIM24, (ii) early and persistent infiltration of immune cells, including inflammatory macrophages, and (iii) the absence of activation of lipid metabolism later in infection, which may be mediated by inhibition of nuclear receptors, including PPARG and HNF1A and -4A, with proinflammatory consequences. Further investigation of these signatures in the host response to other H1N1 viruses of various pathogenicities confirmed their general relevance for virulence of influenza virus and suggested that lung response to MA-CA/04 virus was similar to that following infection with lethal H1N1 r1918 influenza virus. This study links differential activation of IRFs, nuclear receptors, and macrophage infiltration with influenza virulence in vivo.

Self-assembled, nanostructured polypyrrole films grown in a high-gravity environment.

A simple, novel method of synthesizing self-assembled, nanostructured conducting polymer films has been developed. Applying an increased centrifugal force on the electrodes during the electrochemical deposition process yields high surface area, micro- or nanostructured polymer films. Scanning electron microscopy showed that as the applied g-force increased, the polymers progressed from having smooth, "cauliflower" morphologies, to intermediate microstructured surfaces, to finally dense nanostructured surfaces with pore sizes as small as 50 nm. Cyclic voltammetry revealed that films grown at higher centrifugal accelerations (higher than 500g) exhibited less degradation after electrochemical cycling and more capacitive behavior.

Development of a Lorentz-force actuated intravitreal jet injector.

Intravitreal injection is a common treatment in ophthalmology, but it can lead to numerous complications. Needle-free jet injection has been shown to successfully deliver fluid to various layers of skin, and, by its nature, may reduce intravitreal injection complications. From injection trials into ex vivo rabbit eyes, we find that needle-free jet injection can be used for intravitreal drug delivery. A custom-designed control scheme, characterized in this study, is crucial to this delivery. The system is capable of delivering 40 µL of fluid to the posterior vitreous humor, with an injection duration less than 100 ms and scleral entry site less than 350 µm in diameter.

Host regulatory network response to infection with highly pathogenic H5N1 avian influenza virus.

During the last decade, more than half of humans infected with highly pathogenic avian influenza (HPAI) H5N1 viruses have died, yet virus-induced host signaling has yet to be clearly elucidated. Airway epithelia are known to produce inflammatory mediators that contribute to HPAI H5N1-mediated pathogenicity, but a comprehensive analysis of the host response in this cell type is lacking. Here, we leveraged a system approach to identify and statistically validate signaling subnetworks that define the dynamic transcriptional response of human bronchial epithelial cells after infection with influenza A/Vietnam/1203/2004 (H5N1, VN1203). Importantly, we validated a subset of transcripts from one subnetwork in both Calu-3 cells and mice. A more detailed examination of two subnetworks involved in the immune response and keratinization processes revealed potential novel mediators of HPAI H5N1 pathogenesis and host response signaling. Finally, we show how these results compare to those for a less virulent strain of influenza virus. Using emergent network properties, we provide fresh insight into the host response to HPAI H5N1 virus infection and identify novel avenues for perturbation studies and potential therapeutic interventions for fatal HPAI H5N1 disease.

A superhydrophobic to superhydrophilic in situ wettability switch of microstructured polypyrrole surfaces.

We present an electrochemical layered system that allows for the fast, in situ wettability switch of microstructured PPy upon the application of an electric stimulus. We have eliminated the need for PPy to be immersed in an electrolyte to switch between wetting states, laying the groundwork for PPy to be used as a viable material in many applications, including microfluidics or smart textiles. The PPy surface was switched from the superhydrophobic state (contact angle=159) to the superhydrophilic state (contact angle=0) in 3 s. A wettability gradient was also created on a PPy surface using the layered system, causing a 3 µL droplet to travel approximately 2 mm in 0.8 s.

Lethal influenza virus infection in macaques is associated with early dysregulation of inflammatory related genes.

The enormous toll on human life during the 1918-1919 Spanish influenza pandemic is a constant reminder of the potential lethality of influenza viruses. With the declaration by the World Health Organization of a new H1N1 influenza virus pandemic, and with continued human cases of highly pathogenic H5N1 avian influenza virus infection, a better understanding of the host response to highly pathogenic influenza viruses is essential. To this end, we compared pathology and global gene expression profiles in bronchial tissue from macaques infected with either the reconstructed 1918 pandemic virus or the highly pathogenic avian H5N1 virus A/Vietnam/1203/04. Severe pathology was observed in respiratory tissues from 1918 virus-infected animals as early as 12 hours after infection, and pathology steadily increased at later time points. Although tissues from animals infected with A/Vietnam/1203/04 also showed clear signs of pathology early on, less pathology was observed at later time points, and there was evidence of tissue repair. Global transcriptional profiles revealed that specific groups of genes associated with inflammation and cell death were up-regulated in bronchial tissues from animals infected with the 1918 virus but down-regulated in animals infected with A/Vietnam/1203/04. Importantly, the 1918 virus up-regulated key components of the inflammasome, NLRP3 and IL-1beta, whereas these genes were down-regulated by A/Vietnam/1203/04 early after infection. TUNEL assays revealed that both viruses elicited an apoptotic response in lungs and bronchi, although the response occurred earlier during 1918 virus infection. Our findings suggest that the severity of disease in 1918 virus-infected macaques is a consequence of the early up-regulation of cell death and inflammatory related genes, in which additive or synergistic effects likely dictate the severity of tissue damage.