Extracellular Matrix-Derived Biophysical Cues Mediate Interstitial Flow-Induced Sprouting Angiogenesis.

Sprouting angiogenesis is orchestrated by an intricate balance of biochemical and mechanical cues in the local tissue microenvironment. Interstitial flow has been established as a potent regulator of angiogenesis. Similarly, extracellular matrix (ECM) physical properties, such as stiffness and microarchitecture, have also emerged as important mediators of angiogenesis. However, the interplay between interstitial flow and ECM physical properties in the initiation and control of angiogenesis is poorly understood. Using a three-dimensional (3D) microfluidic tissue analogue of angiogenic sprouting with defined interstitial flow superimposed over ECM with well-characterized physical properties, we found that the addition of hyaluronan (HA) to collagen-based matrices significantly enhances sprouting induced by interstitial flow compared to responses in collagen-only hydrogels. We confirmed that both the stiffness and matrix pore size of collagen-only hydrogels were increased by the addition of HA. Interestingly, interstitial flow-potentiated sprouting responses in collagen/HA matrices were not affected when functionally blocking the HA receptor CD44. In contrast, enzymatic depletion of HA in collagen/HA matrices with hyaluronidase (HAdase) resulted in decreased stiffness, pore size, and interstitial flow-mediated sprouting to the levels observed in collagen-only matrices. Taken together, these results suggest that HA enhances interstitial flow-mediated angiogenic sprouting through its alterations to collagen ECM stiffness and pore size.

Microfluidic Collective Cell Migration Assay for Study of Endothelial Cell Proliferation and Migration under Combinations of Oxygen Gradients, Tensions, and Drug Treatments.

Proliferation and migration of endothelial cells play an important role in many biological activities, and they can be regulated by various microenvironmental factors. In this paper, a novel microfluidic collective cell migration assay is developed to study endothelial cell migration and proliferation under combinations of three oxygen conditions: normoxia, oxygen gradient, and hypoxia and three medium compositions: normal growth medium, the medium with cytochalasin-D for actin polymerization inhibition, and with YC-1 for hypoxia-inducible factor (HIF) inhibition. The microfluidic device designed in the paper allows cell patterns formed with consistent dimensions using laminar flow patterning. In addition, stable oxygen gradients can be generated within the device by a spatially confined chemical reaction method. The device can be operated in conventional cell incubators with minimal chemical reagents and instrumentation for practical applications. The results show directional collective cell migration of the endothelial cells under the oxygen gradients for all the medium compositions. The directional behavior has never been discussed before, and indicates critical roles of oxygen gradients in guiding endothelial cell migration during various biological activities. The developed assay provides a practical yet powerful tool for further in vitro study of endothelial cell behaviors under various physiological microenvironments.

Directly observed therapy reduces tuberculosis-specific mortality: a population-based follow-up study in Taipei, Taiwan.

OBJECTIVES: To determine the effect of directly observed therapy (DOT) on tuberculosis-specific mortality and non-TB-specific mortality and identify prognostic factors associated with mortality among adults with culture-positive pulmonary TB (PTB). METHODS: All adult Taiwanese with PTB in Taipei, Taiwan were included in a retrospective cohort study in 2006-2010. Backward stepwise multinomial logistic regression was used to identify risk factors associated with each mortality outcome. RESULTS: Mean age of the 3,487 patients was 64.2 years and 70.4% were male. Among 2471 patients on DOT, 4.2% (105) died of TB-specific causes and 15.4% (381) died of non-TB-specific causes. Among 1016 patients on SAT, 4.4% (45) died of TB-specific causes and 11.8% (120) died of non-TB-specific causes. , After adjustment for potential confounders, the odds ratio for TB-specific mortality was 0.45 (95% CI: 0.30-0.69) among patients treated with DOT as compared with those on self-administered treatment. Independent predictors of TB-specific and non-TB-specific mortality included older age (ie, 65-79 and ≥80 years vs. 18-49 years), being unemployed, a positive sputum smear for acid-fast bacilli, and TB notification from a general ward or intensive care unit (reference: outpatient services). Male sex, end-stage renal disease requiring dialysis, malignancy, and pleural effusion on chest radiography were associated with increased risk of non-TB-specific mortality, while presence of lung cavities on chest radiography was associated with lower risk. CONCLUSIONS: DOT reduced TB-specific mortality by 55% among patients with PTB, after controlling for confounders. DOT should be given to all TB patients to further reduce TB-specific mortality.

Electrofluidic pressure sensor embedded microfluidic device: a study of endothelial cells under hydrostatic pressure and shear stress combinations.

Various microfluidic cell culture devices have been developed for in vitro cell studies because of their capabilities to reconstitute in vivo microenvironments. However, controlling flows in microfluidic devices is not straightforward due to the wide varieties of fluidic properties of biological samples. Currently, flow observations mainly depend on optical imaging and macro scale transducers, which usually require sophisticated instrumentation and are difficult to scale up. Without real time monitoring, the control of flows can only rely on theoretical calculations and numerical simulations. Consequently, these devices have difficulty in being broadly exploited in biological research. This paper reports a microfluidic device with embedded pressure sensors constructed using electrofluidic circuits, which are electrical circuits built by fluidic channels filled with ionic liquid. A microfluidic device culturing endothelial cells under various shear stress and hydrostatic pressure combinations is developed to demonstrate this concept. The device combines the concepts of electrofluidic circuits for pressure sensing, and an equivalent circuit model to design the cell culture channels. In the experiments, human umbilical vein endothelial cells (HUVECs) are cultured in the device with a continuous medium perfusion, which provides the combinatory mechanical stimulations, while the hydrostatic pressures are monitored in real time to ensure the desired culture conditions. The experimental results demonstrate the importance of real time pressure monitoring, and how both mechanical stimulations affect the HUVEC culture. This developed microfluidic device is simple, robust, and can be easily scaled up for high-throughput experiments. Furthermore, the device provides a practical platform for an in vitro cell culture under well-controlled and dynamic microenvironments.

Improved consistency in dosing anti-tuberculosis drugs in Taipei, Taiwan.

BACKGROUND: It was reported that 35.5% of tuberculosis (TB) cases reported in 2003 in Taipei City had no recorded pre-treatment body weight and that among those who had, inconsistent dosing of anti-TB drugs was frequent. Taiwan Centers for Disease Control (CDC) have taken actions to strengthen dosing of anti-TB drugs among general practitioners. Prescribing practices of anti-TB drugs in Taipei City in 2007-2010 were investigated to assess whether interventions on dosing were effective. METHODOLOGY/PRINCIPAL FINDINGS: Lists of all notified culture positive TB cases in 2007-2010 were obtained from National TB Registry at Taiwan CDC. A medical audit of TB case management files was performed to collect pretreatment body weight and regimens prescribed at commencement of treatment. Dosages prescribed were compared with dosages recommended. The proportion of patients with recorded pre-treatment body weight was 64.5% in 2003, which increased to 96.5% in 2007-2010 (p<0.001). The proportion of patients treated with consistent dosing of a 3-drug fixed-dose combination (FDC) increased from 73.9% in 2003 to 87.7% in 2007-2010 (p<0.001), and that for 2-drug FDC from 76.0% to 86.1% (p = 0.024), for rifampicin (RMP) from 62.8% to 85.5% (p<0.001), and for isoniazid from 87.8% to 95.3% (p<0.001). In 2007-2010, among 2917 patients treated with either FDCs or RMP in single-drug preparation, the dosage of RMP was adequate (8-12 mg/kg) in 2571(88.1%) patients, too high in 282(9.7%), too low in 64(2.2%). In multinomial logistic regression models, factors significantly associated with adequate dosage of RMP were body weight and preparations of RMP. Patients weighting <40 kg (relative risk ratio (rrr) 6010.5, 95% CI 781.1-46249.7) and patients weighting 40-49 kg (rrr 1495.3, 95% CI 200.6-11144.6) were more likely to receive higher-than-recommended dose of RMP. CONCLUSIONS/SIGNIFICANCE: Prescribing practice in the treatment of TB in Taipei City has remarkably improved after health authorities implemented a series of interventions.

Risk factors for unfavorable outcome of pulmonary tuberculosis in adults in Taipei, Taiwan.

This study was undertaken to identify factors associated with unfavorable outcomes in patients with pulmonary tuberculosis (PTB) in Taipei, Taiwan in 2007-2008. Taiwanese adults with culture-positive PTB diagnosed in Taipei during the study period were included in this retrospective cohort study. Unfavorable outcomes were classified as treatment default, death, treatment failure, or transfer. Of 1616 eligible patients, 22.6% (365) had unfavorable outcomes, mainly death. After controlling for patient sociodemographic factors, clinical findings, and underlying disease, independent risk factors for unfavorable outcomes included advanced age, unemployment, end-stage renal disease requiring dialysis, malignancy, acid-fast bacilius smear-positivity, multidrug-resistant TB, and notification from ordinary ward or intensive care unit. In contrast, patients receiving directly observed treatment, and with a high school or higher education were significantly less likely to have unfavorable outcomes. This study advanced our understanding by revealing that a high school or higher education might lower the risk of an unfavorable outcome. Our results also confirmed the risk factors for unfavorable outcomes shown in previous research. Future TB control programmes in Taiwan should target particularly high-risk patients including those who had lower educational levels.

Generation of oxygen gradients in microfluidic devices for cell culture using spatially confined chemical reactions.

This paper reports a microfluidic device capable of generating oxygen gradients for cell culture using spatially confined chemical reactions with minimal chemical consumption. The microfluidic cell culture device is constructed by single-layer polydimethylsiloxane (PDMS) microfluidic channels, in which the cells can be easily observed by microscopes. The device can control the oxygen gradients without the utilization of bulky pressurized gas cylinders, direct addition of oxygen scavenging agents, or tedious gas interconnections and sophisticated flow control. In addition, due to the efficient transportation of oxygen within the device using the spatially confined chemical reactions, the microfluidic cell culture device can be directly used in conventional cell incubators without altering their gaseous compositions. The oxygen gradients generated in the device are numerically simulated and experimentally characterized using an oxygen-sensitive fluorescence dye. In this paper, carcinomic human alveolar basal epithelial (A549) cells have been cultured in the microfluidic device with a growth medium and an anti-cancer drug (Tirapazamine, TPZ) under various oxygen gradients. The cell experiment results successfully demonstrate the hyperoxia-induced cell death and hypoxia-induced cytotoxicity of TPZ. In addition, the results confirm the great cell compatibility and stable oxygen gradient generation of the developed device. Consequently, the microfluidic cell culture device developed in this paper is promising to be exploited in biological labs with minimal instrumentation to study cellular responses under various oxygen gradients.