Label-free and noninvasive analysis of microorganism surface epistructures at the single-cell level.

Cell surface properties of microorganisms provide abundant information for their physiological status and fate choice. However, current methods for analyzing cell surface properties require labeling or fixation, which can alter the cell activity. This study establishes a label-free, rapid, noninvasive, and quantitative analysis of cell surface properties, including the presence and the dimension of epistructure, down to the single-cell level and at the nanometer scale. Simultaneously, electrorotation provides dielectric properties of intracellular contents. With the combined information, the growth phase of microalgae cells can be identified. The measurement is based on electrorotation of single cells, and an electrorotation model accounting for the surface properties is developed to properly interpret experimental data. The epistructure length measured by electrorotation is validated by scanning electron microscopy. The measurement accuracy is satisfactory in particular in the case of microscale epistructures in the exponential phase and nanoscale epistructures in the stationary phase. However, the measurement accuracy for nanoscale epistructures on cells in the exponential phase is offset by the effect of a thick double layer. Lastly, a diversity in epistructure length distinguishes exponential phase from stationary phase.

Mapping the salinity gradient in a microfluidic device with schlieren imaging.

This work presents the use of the schlieren imaging to quantify the salinity gradients in a microfluidic device. By partially blocking the back focal plane of the objective lens, the schlieren microscope produces an image with patterns that correspond to spatial derivative of refractive index in the specimen. Since salinity variation leads to change in refractive index, the fluid mixing of an aqueous salt solution of a known concentration and water in a T-microchannel is used to establish the relation between salinity gradients and grayscale readouts. This relation is then employed to map the salinity gradients in the target microfluidic device from the grayscale readouts of the corresponding micro-schlieren image. For saline solution with salinity close to that of the seawater, the grayscale readouts vary linearly with the salinity gradient, and the regression line is independent of the flow condition and the salinity of the injected solution. It is shown that the schlieren technique is well suited to quantify the salinity gradients in microfluidic devices, for it provides a spatially resolved, non-invasive, full-field measurement.

High-voltage, Long-duration Pulsed Radiofrequency to the Dorsal Root Ganglion Provides Improved Pain Relief for Herpes Zoster Neuralgia in the Subacute Stage.

BACKGROUND: Postherpetic neuralgia (PHN) is pain persisting beyond 3 months from rash onset and is the most common complication of herpes zoster (HZ); it is commonly refractory to medication treatment. Available evidence indicates that high-voltage, long-duration pulsed radiofrequency (PRF) to the dorsal root ganglion (DRG) is a novel and effective treatment for this complication. Nevertheless, the effects of this intervention on refractory HZ neuralgia less than 3 months have not been evaluated. OBJECTIVE: The objective of this study was to assess the therapeutic efficacy and safety of high-voltage, long-duration PRF to the DRG for patients with subacute HZ neuralgia compared with that of patients with PHN. STUDY DESIGN: A retrospective comparative research. SETTING: Hospital department in China. METHODS: Sixty-four patients with HZ neuralgia in different stages receiving high-voltage, long-duration PRF to the DRG were included. According to the days from zoster onset to PRF implementation, they were divided into the subacute (one to 3 months) or PHN group (more than 3 months). The therapeutic effect was evaluated by pain relief using the Numeric Rating Scale at one day, one week, one month, 3 months, and 6 months post-PRF. The five-point Likert scale measured patient satisfaction. Post-PRF side effects were also recorded to determine the safety of the intervention. RESULTS: The intervention significantly reduced pain in all patients, but pain relief at one month, 3 months, and 6 months post-PRF was better in the subacute group than in the PHN group. Furthermore, the success rate of PRF was significantly increased in the subacute group compared with the PHN group (81.3% vs 56.3%, P = 0.031). There was no significant difference in patient satisfaction at 6 months between groups. LIMITATIONS: This is a single-center retrospective study with a small sample size. CONCLUSIONS: High-voltage, long-duration PRF to the DRG is effective and safe for HZ neuralgia in different stages, and can provide an improved pain relief for HZ neuralgia in the subacute stage.

Avian reovirus triggers autophagy in primary chicken fibroblast cells and Vero cells to promote virus production.

Avian reovirus (ARV) is an important cause of disease in poultry. Although ARV is known to induce apoptosis in infected cells, the interaction between ARV and its target cells requires further elucidation. In this report, we show that the ARV isolate strain GX/2010/1 induces autophagy in both Vero and primary chicken embryonic fibroblast (CEF) cells based on the appearance of an increased number of double-membrane vesicles, the presence of GFP-microtubule-associated protein 1 light chain 3 (GFP-LC3) dot formation, and the elevated production of LC3II. We further demonstrate that the class I phosphoinositide 3-kinase (PI3K)/Akt/mTOR pathway contributes to autophagic induction by ARV infection. Moreover, treatment of ARV-infected cells with the autophagy inducer rapamycin increased viral yields, while inhibition of the autophagosomal pathway using chloroquine led to a decrease in virus production. Altogether, our studies strongly suggest that autophagy may play a critical role in determining viral yield during ARV infection.

Change in rheotactic behavior patterns of dinoflagellates in response to different microfluidic environments.

Plankton live in dynamic fluid environments. Their ability to change in response to different hydrodynamic cues is critical to their energy allocation and resource uptake. This study used a microfluidic device to evaluate the rheotactic behaviors of a model dinoflagellate species, Karlodinium veneficum, in different flow conditions. Although dinoflagellates experienced forced alignment in strong shear (i.e. "trapping"), fluid straining did not play a decisive role in their rheotactic movements. Moderate hydrodynamic magnitude (20 < |uf| < 40 µm s-1) was found to induce an orientation heading towards an oncoming current (positive rheotaxis), as dinoflagellates switched to cross-flow swimming when flow speed exceeded 50 µm s-1. Near the sidewalls of the main channel, the steric mechanism enabled dinoflagellates to adapt upstream orientation through vertical migration. Under oscillatory flow, however, positive rheotaxis dominated with occasional diversion. The varying flow facilitated upstream exploration with directional controlling, through which dinoflagellates exhibited avoidance of both large-amplitude perturbance and very stagnant zones. In the mixed layer where water is not steady, these rheotactic responses could lead to spatial heterogeneity of dinoflagellates. The outcome of this study helps clarify the interaction between swimming behaviors of dinoflagellates and the hydrodynamic environment they reside in.

Blockade of Erythropoietin-Producing Human Hepatocellular Carcinoma Receptor B1 in Spinal Dorsal Horn Alleviates Visceral Pain in Rats.

Objective: This experiment was designed to determine whether erythropoietin-producing human hepatocellular carcinoma (Eph) receptors were involved in the development of visceral pain. Methods: Adult male Sprague-Dawley rats were randomly divided into three groups receiving different treatments (n = 16 per group): intracolonic vehicle (control group), intracolonic 2, 4, 6-trinitrobenzene sulfonic acid (TNBS) (TNBS group), and intracolonic TNBS and intrathecal EphB1 receptor blocking reagent (TNBS + EphB2-Fc group). Visceral hyperalgesia was evaluated with quantification of visceral pain threshold induced by colorectal distention. The spinal expressions of EphB1 and ephrinB2 and levels of their phosphorylated forms (p-EphB1 and p-ephrinB2) were assessed by Western blotting and immunohistochemistry. Results: The TNBS-treated rats developed significant visceral hyperalgesia. The spinal expressions of EphB1, p-EphB1, ephrinB2, and p-ephrinB2 were significantly increased in the TNBS group compared with the control group, but visceral hyperalgesia and elevation of spinal EphB1 and p-EphB1 expressions were evidently alleviated by intrathecal administration of EphB2-Fc in the TNBS + EphB2-Fc group. The number of EphB1- and p-EphB1-immunopositive cells, the average optical (AO) value of EphB1, and its phosphorylated form in the spinal dorsal horn were significantly increased in the TNBS group than in the control group, but they were obviously reduced by intrathecal administration of EphB2-Fc. There were no significant differences in the number of ephrinB2- and p-ephrinB2-immunopositive cells and the AO value of ephrinB2 and its phosphorylated form between the TNBS and TNBS + EphB2-Fc groups. Conclusion: EphB1 receptors in the spinal dorsal horn play a pivotal role in the development of visceral pain and may be considered as a potential target for the treatment of visceral pain.

Electrorotation of single microalgae cells during lipid accumulation for assessing cellular dielectric properties and total lipid contents.

Photosynthetic microalgae not only perform fixation of carbon dioxide but also produce valuable byproducts such as lipids and pigments. However, due to the lack of effective tools for rapid and noninvasive analysis of microalgal cellular contents, the efficiency of strain screening and culture optimizing is usually quite low. This study applied single-cell electrorotation on Scenedesmus abundans to assess cellular dielectric properties during lipid accumulation and to promptly quantify total cellular contents. The experimental electrorotation spectra were fitted with the double-shell ellipsoidal model, which considered varying cell wall thickness, to obtain the dielectric properties of cellular compartments. When the amount of total lipids increased from 15.3 wt% to 33.8 wt%, the conductivity and relative permittivity of the inner core (composed of the cytoplasm, lipid droplets, and nucleus) decreased by 21.7% and 22.5%, respectively. These dielectric properties were further used to estimate the total cellular lipid contents by the general mixing formula, and the estimated values agreed with those obtained by weighing dry biomass and extracted lipids with an error as low as 0.22 wt%. Additionally, the conductivity and relative permittivity of cell wall increased during nitrogen-starvation conditions, indicating the thickening of cell wall, which was validated by the transmission electron microscopy.

Repetitive Transcranial Magnetic Stimulation at Different Frequencies for Postherpetic Neuralgia: A Double-Blind, Sham-Controlled, Randomized Trial.

BACKGROUND: Repetitive transcranial magnetic stimulation (rTMS) at 5 Hz and 10 Hz is effective in improving pain, sleep quality, and anxiety among patients with postherpetic neuralgia (PHN). But it has not been reported which frequency is more effective and which frequency is safer. OBJECTIVES: This study aimed to observe the efficacy and safety of rTMS at different high frequencies (5 Hz, 10 Hz) for PHN. STUDY DESIGN: The design of the study was a prospective randomized, controlled clinical trial. SETTING: The research was conducted within a department of pain management at a university hospital in China. METHODS: Sixty patients with PHN who were treated at the Department of Pain Management at Xuanwu Hospital of Capital Medical University were recruited. Using a computer-created number list, the cases were equally divided into 3 groups (n = 20), namely, the sham rTMS group, 5-Hz rTMS group, and 10-Hz rTMS group. The sham rTMS group received sham stimulation, and the other 2 groups received high-frequency (5-Hz and 10-Hz) rTMS, respectively. The primary motor cortex (M1) on the healthy side was stimulated with an intensity of 80% transcranial magnetic stimulation (RMT). For the 5-Hz rTMS group, each stimulation session consisted of a series of 300 one-second pulses with a frequency of 5 Hz and an interval of 2.5 seconds between each train, giving a total of 1500 pulses per session. For the 10-Hz rTMS group, each stimulation session consisted of a series of 300 0.5-second pulses with a frequency of 10 Hz and an interval of 3 seconds between each train, giving a total of 1500 pulses per session; the total time of stimulations was 17.5 minutes. rTMS was performed once daily for 10 days. The 3 groups received conventional medication therapy. Baseline data (gender, age, course of disease, affected side) were recorded in the 3 groups. At different time points (before treatment, T0; during treatment, T1-T10; 1 month after treatment, T11; and 3 months after treatment, T12), the patients were evaluated on the following scales: Visual Analog Scale (VAS), short-form McGill Pain Questionnaire (SF-MPQ), Quality of Life (QOL) scale, sleep quality (SQ) scale, Self-Rating Depression Scale (SDS), Patient Global Impression of Change (PGIC), and incidence of adverse events. RESULTS: Compared with the sham rTMS group, there was a significant reduction in VAS scores in the 5-Hz rTMS group and 10-Hz rTMS group at T2-T12 (P < .05). VAS scores in the 10-Hz rTMS group at T7-T12 were significantly lower compared with the 5-Hz rTMS group (P < .05). The average VAS reduction was significantly different between the 5-Hz and 10-Hz rTMS groups; 28.3% (95% confidence interval [CI],19.48%-49.35%), compared to 39.89% (95% CI, 22.47%-58.64%), with (F = 5.289, P = .022). The 3 groups did not differ significantly in general SF-MPQ, QOL, SQ, SDS, and PGIC scores. However, the QQL, SQ, and PGIC scores of the 5-Hz rTMS group and the 10-HZ rTMS group at T12 were significantly higher than that of the sham rTMS group. LIMITATIONS: The study's follow-up period was limited to 3 months. CONCLUSIONS: rTMS at either frequency, 5 Hz or 10 Hz, relieved PHN and improved the patients' quality of life. rTMS at 10 Hz was superior to rTMS at 5 Hz in terms of pain relief, quality of life, and improvement in sleep quality, though the latter had higher safety. rTMS at either 5 Hz or 10 Hz can be used as an adjuvant therapy for PHN. KEY WORDS: Repetitive transcranial magnetic stimulation, postherpetic neuralgia, pain evaluation.

Measurements of flow-induced birefringence in microfluidics.

In this study, we demonstrate the use of a microscopic circular polariscope to measure the flow-induced birefringence in a microfluidic device that represents the kinematics of fluid motion optically. Unlike the commercial birefringence microscope employed in the previous studies, our approach is able to provide direct measurement of retardance, which quantifies the difference in refractive index of the fluid experienced by the ordinary and extraordinary rays, from one single image frame. This capability facilitates unsteady full-field quantitation of flow-induced birefringence in microfluidics that has never been achieved before. At low flow rates, we find that the value of the retardance is independent of the microfluidic design and proportional to the nominal strain rates. This linearity bridges the measurement of birefringence and the deformation rate in the microflow environment, which yields the stress information of the fluid flow. In addition, the µPIV results confirm that both extensional and shear strain rates contribute to the flow-induced birefringence so that the retardance distribution can be used to represent the field of the principal strain rate in a microfluidic device. The outcome of this study proves that our approach provides a non-invasive method that enables an intuitive full-field representation of stress in the instantaneous flow field in a microfluidic device.

Analyzing Mixing Inhomogeneity in a Microfluidic Device by Microscale Schlieren Technique.

In this paper, we introduce the use of microscale schlieren technique to measure mixing inhomogeneity in a microfluidic device. The microscale schlieren system is constructed from a Hoffman modulation contrast microscope, which provides easy access to the rear focal plane of the objective lens, by removing the slit plate and replacing the modulator with a knife-edge. The working principle of microscale schlieren technique relies on detecting light deflection caused by variation of refractive index. The deflected light either escapes or is obstructed by the knife-edge to produce a bright or a dark band, respectively. If the refractive index of the mixture varies linearly with its composition, the local change in light intensity in the image plane is proportional to the concentration gradient normal to the optical axis. The micro-schlieren image gives a two-dimensional projection of the disturbed light produced by three-dimensional inhomogeneity. To accomplish quantitative analysis, we describe a calibration procedure that mixes two fluids in a T-microchannel. We carry out a numerical simulation to obtain the concentration gradient in the T-microchannel that correlates closely with the corresponding micro-schlieren image. By comparison, a relationship between the grayscale readouts of the micro-schlieren image and the concentration gradients presented in a microfluidic device is established. Using this relationship, we are able to analyze the mixing inhomogeneity from associate micro-schlieren image and demonstrate the capability of microscale schlieren technique with measurements in a microfluidic oscillator(4). For optically transparent fluids, microscale schlieren technique is an attractive diagnostic tool to provide instantaneous full-field information that retains the three-dimensional features of the mixing process.

Role of MK2 signaling pathway in the chronic compression of cervical spinal cord.

OBJECTIVE: In this study, chronic compression of cervical spinal cord was introduced into twy/twy mice and the role of MK2 signaling pathway was investigated in this disease. METHODS: twy/twy mice aged 6-24 weeks were used and the inflammatory response in the cervical spinal cord was observed. The Institute of Cancer Research (ICR) mice were used as controls. MK2 inhibitor (PF-3644022, 30 mg/kg) was administered intragastrically to twy/twy mice. The motor behavior was firstly observed in these three groups by Catwalk gait analysis. And the cervical spinal cord between C2 and C3 of vertebral segments was analyzed by MRI and Western blot assay. RESULTS: The stride length of paws and interlimb coordination reduced in twy/twy mice. However, at 4 weeks after PF-3644022 treatment, a marked improvement was observed in the motor function. The expressions of inflammation related factors (such as IL-1ß, NF-κB, TNF-α, MK2 and p-MK2) and apoptosis related proteins (such as cleaved caspase-8 and bax/bcl-2) in the spinal cord of twy/twy mice significantly increased as compared to controls, but 4-week treatment with PF-3644022 markedly reduced the expressions of these factors and apoptotic proteins in the cervical spinal cord. CONCLUSION: MK2 signaling pathway is involved in the chronic compression induced inflammation of the cervical spinal cord. Thus, to inhibit the MK2 pathway may used to improve the outcome and prevent the deterioration of neurological dysfunction.