Strategic placement of an obstacle suppresses droplet break up in the hopper flow of a microfluidic soft crystal.

When granular materials, colloidal suspensions, and even animals and crowds exit through a narrow outlet, clogs can form spontaneously when multiple particles or entities attempt to exit simultaneously, thereby obstructing the outlet and ultimately halting the flow. Counterintuitively, the presence of an obstacle upstream of the outlet has been found to suppress clog formation. For soft particles such as emulsion drops, clogging has not been observed in the fast flow limit due to their deformability and vanishing interparticle friction. Instead, they pinch off each other and undergo break up when multiple drops attempt to exit simultaneously. Similar to how an obstacle reduces clogging in a rigid particle system, we hypothesize and demonstrate that an obstacle could suppress break up in the two-dimensional hopper flow of a microfluidic crystal consisting of dense emulsion drops by preventing the simultaneous exit of multiple drops. A regime map plotting the fraction of drops that undergo break up in a channel with different obstacle sizes and locations delineates the geometrical constraints necessary for effective break up suppression. When optimally placed, the obstacle induced an unexpected ordering of the drops, causing them to alternate and exit the outlet one at a time. Droplet break up is suppressed drastically by almost three orders of magnitude compared to when the obstacle is absent. This result can provide a simple, passive strategy to prevent droplet break up and can find use in improving the robustness and integrity of droplet microfluidic biochemical assays as well as in extrusion-based three-dimensional printing of emulsion or foam-based materials.

Confinement size determines the architecture of Ran-induced microtubule networks.

The organization of microtubules (MTs) is critical for cells during interphase and mitosis. During mitotic spindle assembly, MTs are made and organized around chromosomes in a process regulated by RanGTP. The role of RanGTP has been explored in Xenopus egg extracts, which are not limited by a cell membrane. Here, we investigated whether cell-sized confinements affect the assembly of RanGTP-induced MT networks in Xenopus egg extracts. We used microfluidics to encapsulate extracts within monodisperse extract-in-oil droplets. Importantly, we find that the architecture of Ran-induced MT networks depends on the droplet diameter and the Ran concentration, and differs from structures formed in bulk extracts. Our results highlight that both MT nucleation and physical confinement play critical roles in determining the spatial organization of the MT cytoskeleton.

Microfluidic guillotine for single-cell wound repair studies.

Wound repair is a key feature distinguishing living from nonliving matter. Single cells are increasingly recognized to be capable of healing wounds. The lack of reproducible, high-throughput wounding methods has hindered single-cell wound repair studies. This work describes a microfluidic guillotine for bisecting single Stentor coeruleus cells in a continuous-flow manner. Stentor is used as a model due to its robust repair capacity and the ability to perform gene knockdown in a high-throughput manner. Local cutting dynamics reveals two regimes under which cells are bisected, one at low viscous stress where cells are cut with small membrane ruptures and high viability and one at high viscous stress where cells are cut with extended membrane ruptures and decreased viability. A cutting throughput up to 64 cells per minute-more than 200 times faster than current methods-is achieved. The method allows the generation of more than 100 cells in a synchronized stage of their repair process. This capacity, combined with high-throughput gene knockdown in Stentor, enables time-course mechanistic studies impossible with current wounding methods.

Spatiotemporal periodicity of dislocation dynamics in a two-dimensional microfluidic crystal flowing in a tapered channel.

When a many-body system is driven away from equilibrium, order can spontaneously emerge in places where disorder might be expected. Here we report an unexpected order in the flow of a concentrated emulsion in a tapered microfluidic channel. The velocity profiles of individual drops in the emulsion show periodic patterns in both space and time. Such periodic patterns appear surprising from both a fluid and a solid mechanics point of view. In particular, when the emulsion is considered as a soft crystal under extrusion, a disordered scenario might be expected based on the stochastic nature of dislocation dynamics in microscopic crystals. However, an orchestrated sequence of dislocation nucleation and migration is observed to give rise to a highly ordered deformation mode. This discovery suggests that nanocrystals can be made to deform more controllably than previously thought. It can also lead to novel flow control and mixing strategies in droplet microfluidics.

Puerarin Attenuates Anoxia/Reoxygenation Injury Through Enhancing Bcl-2 Associated Athanogene 3 Expression, a Modulator of Apoptosis and Autophagy.

BACKGROUND: Puerarin has protective effects on ischemia-reperfusion injury, but the underlying mechanisms are not fully revealed. This study explored the effect of puerarin on the expression of Bcl-2 associated athanogene 3 (BAG3) in an in vitro model of anoxia/reoxygenation injury (A/RI) in neonate rat primary cardiomyocytes and the functions of BAG3 in A/RI. MATERIAL/METHODS: BAG3 expression in cardiomyocytes with or without puerarin pre-treatment was quantified using qRT-PCR and Western blot analysis. The effects of BAG3 on A/RI were studied by measuring the activity of lactate dehydrogenase (LDH) and creatine phosphate kinase (CPK), the concentration of malondialdehyde (MDA), superoxide dismutase (SOD), and glutathione peroxidase (GSH-Px). The effects of BAG3 on autophagy and apoptosis of the cardiomyocytes after A/RI were further studied. RESULTS: Puerarin significantly promoted BAG3 expression in the rat primary cardiomyocytes after A/RI. Enforced BAG3 expression presented similar effects as puerarin pre-treatment in attenuating A/RI in terms of CPK, LDH, MDA, SOD, GSH-Px, ROS generation, and cell viability. BAG3 overexpression significantly stimulated autophagy in cardiomyocytes after A/RI, which presented protective effects on A/RI in terms of cell viability and apoptosis. Autophagy inhibition partly abrogated the protective effects of BAG3. CONCLUSIONS: Puerarin can directly increase BAG3 transcription and translation in cardiomyocytes after A/RI. The elevated BAG3 expression presents protective effects on A/RI at least through enhancing autophagy and reducing apoptosis, which is a novel protective mechanism of puerarin in ARI.

[Anti-atherosclerosis role of N-oleoylethanolamine in CB2].

To observe a PPAR-alpha agonist effect of N-oleoylethanolamine (OEA) on CB2 (cannabinoid receptor 2), an anti-inflammatory receptor in vascular endothelial cell, healthy HUVECs and TNF-alpha induced HUVECs were used to establish a human vascular endothelial cell inflammatory model. Different doses of OEA (10, 50 and 100 micromol x L(-1)) had been given to HUVECs, cultured at 37 degrees C for 7 h and then collected the total protein and total mRNA. CB2 protein expression was detected by Western blotting and CB2 mRNA expression was assayed by real-time PCR. As the results shown, OEA (10 and 50 micromol x L(-1)) could induce the CB2 protein and mRNA expression, but not 100 micromol x L(-1). To detect if anti-inflammation effect of OEA is partly through CB2, CB2 inhibitor AM630 was used to inhibit HUVEC CB2 expression, then the VCAM-1 expression induced by TNF-alpha was detected, or THP-1 adhere to TNF-alpha induced HUVECs was examined. OEA (50 micromol x L(-1)) could inhibit TNF-alpha induced VCAM-1 expression and THP-1 adhere to HUVECs, these effects could be partly inhibited by a CB2 inhibitor AM630. The anti-inflammation effect of OEA is induced by PPAR-alpha and CB2, suggesting that CB2 signaling could be a target for anti-atherosclerosis, OEA have wide effect in anti-inflammation, it may have better therapeutic potential in anti-inflammation in HUVECs, thus achieving anti-atherosclerosis effect.

One new 12, 8-guaianolide sesquiterpene lactone with antihyperglycemic activity from the roots of Cichorium intybus.

Three 12, 8-guaianolide sesquiterpene lactones, including a new compound intybusin F (1), and a new natural product cichoriolide I (2), along with six known 12, 6-guaianolide compounds (4-9) were isolated from the roots of Cichorium intybus L. Their structures were determined by extensive spectroscopic analysis. The absolute configurations of new compounds were elucidated based on analysis of the experimental and calculated electronic circular dichroism spectra. Compounds 1, 2, 4, 7, 8 showed significant effects on facilitating the glucose uptake in oleic acid plus high glucose-stimulated HepG2 cells at 50 µM. In addition, compounds 1, 2, 3, 6, 7 exhibited obvious inhibitory effects against NO production, of them, compounds 1, 2, 7 can significantly decrease the secretion of inflammatory cytokines (TNF-α, IL-6 and COX-2) levels in this hyperglycemic HepG2 cell model.

Chicoric Acid Attenuated Renal Tubular Injury in HFD-Induced Chronic Kidney Disease Mice through the Promotion of Mitophagy via the Nrf2/PINK/Parkin Pathway.

As the main factor in the pathogenesis of chronic kidney disease (CKD), the excessive apoptosis of renal tubular epithelial cells (RTECs) and its underlying mechanism of action are worth further investigation. Chicoric acid (CA), a major active constituent of the Uyghur folk medicine chicory, was recorded to possess a renal protective effect. The precise effect of CA on renal tubular injury in obesity-related CKD remains unknown. In the current study, CA was proven to ameliorate metabolic disorders including overweight, hyperglycemia, hyperlipidemia, and hyperuricemia in high fat diet (HFD)-fed mice. Furthermore, the reverse effect of CA on renal histological changes and functional damage was confirmed. In vitro, the alleviation of lipid accumulation and cell apoptosis was observed in palmitic acid (PA)-exposed HK2 cells. Treatment with CA reduced mitochondrial damage and oxidative stress in the renal tubule of HFD-fed mice and PA-treated HK2 cells. Finally, CA was observed to activate the Nrf2 pathway; increase PINK and Parkin expression; and regulate LC3, SQSTM1, Mfn2, and FIS1 expression; therefore, it would improve mitochondrial dynamics and mitophagy to alleviate mitochondrial damage in RTECs of obesity-related CKD. These results may provide fresh insights into the promotion of mitophagy in the prevention and alleviation of obesity-related CKD.

Andalucin from Artemisia lannta suppresses the neuroinflammation via the promotion of Nrf2-mediated HO-1 levels by blocking the p65-p300 interaction in LPS-activated BV2 microglia.

BACKGROUND: Neuroinflammation plays an important role in many neurodegenerative conditions such as Alzheimer's disease (AD) and Parkinson disease (PD). Andalucin (ADL), a sesquiterpene lactone from Artemisia lannta, has been reported to exhibit NO inhibition in vitro. However, the effect of ADL on microglia-mediated neuroinflammation has not been investigated. PURPOSE: This study was designed to determine the anti-neuroinflammatory effect of ADL against LPS-activated BV2 microglial cells and to explore the underlying mechanisms. METHODS: The production of pro-inflammatory mediators and cytokines were measured by ELISA. The relevant mechanisms were analyzed by qRT-PCR, Luciferase assay, Western blot and Co-immunoprecipitation Assay. RESULTS: ADL inhibited the LPS-induced release of NO, PGE2, TNF-α, IL-6 and IL-1ß. In addition, ADL reduced the mRNA and protein levels of iNOS and COX-2. Mechanism studies found that ADL activated Nrf2/HO-1 signaling pathway and suppressed NF-κB signaling pathway. Further investigation showed that the stimulative effect of ADL on Nrf2 transcriptional activity and the inhibitory effect of ADL on RelA transcriptional activity were due to its regulation on p300-Nrf2/p65 interaction. CONCLUSION: ADL displayed anti-neuroinflammatory activity in LPS-activated BV2 cells. The mechanism concerns its regulatory effect on the crosstalk between Nrf2 and p65.

Amphiphilic nanoparticles suppress droplet break-up in a concentrated emulsion flowing through a narrow constriction.

This paper describes the break-up behavior of a concentrated emulsion comprising drops stabilized by amphiphilic silica nanoparticles flowing in a tapered microchannel. Such geometry is often used in serial droplet interrogation and sorting processes in droplet microfluidics applications. When exposed to high viscous stresses, drops can undergo break-up and compromise their physical integrity. As these drops are used as micro-reactors, such compromise leads to a loss in the accuracy of droplet-based assays. Here, we show droplet break-up is suppressed by replacing the fluoro-surfactant similar to the one commonly used in current droplet microfluidics applications with amphiphilic nanoparticles as droplet stabilizer. We identify parameters that influence the break-up of these drops and demonstrate that break-up probability increases with increasing capillary number and confinement, decreasing nanoparticle size, and is insensitive to viscosity ratio within the range tested. Practically, our results reveal two key advantages of nanoparticles with direct applications to droplet microfluidics. First, replacing surfactants with nanoparticles suppresses break-up and increases the throughput of the serial interrogation process to 3 times higher than that in surfactant system under similar flow conditions. Second, the insensitivity of break-up to droplet viscosity makes it possible to process samples having different composition and viscosities without having to change the channel and droplet geometry in order to maintain the same degree of break-up and corresponding assay accuracy.

Confinement and viscosity ratio effect on droplet break-up in a concentrated emulsion flowing through a narrow constriction.

This paper describes the dimensionless groups that determine the break-up probability of droplets in a concentrated emulsion during its flow in a tapered microchannel consisting of a narrow constriction. Such channel geometry is commonly used in droplet microfluidics to investigate the content of droplets from a concentrated emulsion. In contrast to solid wells in multi-well plates, drops are metastable, and are prone to break-up which compromises the accuracy and the throughput of the assay. Unlike single drops, the break-up process in a concentrated emulsion is stochastic. Analysis of the behavior of a large number of drops (N > 5000) shows that the probability of break-up increases with applied flow rate, the size of the drops relative to the size of the constriction, and the viscosity ratio of the emulsion. This paper shows that the break-up probability collapses into a single curve when plotted as a function of the product of capillary number, viscosity ratio, and confinement factor defined as the un-deformed radius of the drop relative to the hydraulic radius of the constriction. Fundamentally, the results represent a critical step towards the understanding of the physics governing instability in concentrated emulsions. Practically, the results provide a direct guide for the rational design of microchannels and the choice of operation parameters to increase the throughput of the droplet interrogation step while preserving droplet integrity and assay accuracy.

Incidence of chemotherapy- and chemoradiotherapy-induced amenorrhea in premenopausal women with stage II/III colorectal cancer.

BACKGROUND: The incidence rates of colorectal cancer (CRC) in young individuals are increasing. There has been a significant improvement in overall survival in CRC because of advances in adjuvant chemotherapy and chemoradiotherapy over the past decades. However, these procedures may compromise the function of the reproductive system, and ovarian failure and premature menopause may occur. The objective of this analysis was to determine the incidence of long-term amenorrhea (≥ 12 months) in women with CRC aged 40 years and younger after adjuvant treatment. PATIENTS AND METHODS: The authors identified 162 premenopausal women with CRC aged 40 years or younger who were treated with adjuvant chemotherapy and chemoradiotherapy at Fudan University Shanghai Cancer Center from January 2008 to December 2012. One hundred twenty-three patients met all eligibility criteria and had sufficient follow-up for evaluation. The median age at diagnosis in patients with colon and rectal cancers was, respectively, 36 and 35 years (range, 17-40 and 24-40 years). RESULTS: All patients had regular menses before treatment; 3 patients with colon cancer (4.2%) experienced long-term amenorrhea, and 48 patients with rectal cancer (94.1%) experienced long-term amenorrhea. The incidence of amenorrhea was significantly lower in patients with colon cancer (4.2%; 3 of 72) than in patients with rectal cancer (94.1%; 48 of 51) (P < .01). CONCLUSION: In this retrospective series, the incidence of amenorrhea in patients with colon and rectal cancers was 4.2% and 94.1%, respectively. We believe our data support the fact that young female patients with CRC, especially those with rectal cancer who are scheduled to undergo pelvic irradiation, should be counseled regarding fertility preservation options, including ovarian transposition and cryopreservation of ovarian tissue, embryo, or oocyte.

Optofluidic ultrahigh-throughput detection of fluorescent drops.

This paper describes an optofluidic droplet interrogation device capable of counting fluorescent drops at a throughput of 254,000 drops per second. To our knowledge, this rate is the highest interrogation rate published thus far. Our device consists of 16 parallel microfluidic channels bonded directly to a filter-coated two-dimensional Complementary Metal-Oxide-Semiconductor (CMOS) sensor array. Fluorescence signals emitted from the drops are collected by the sensor that forms the bottom of the channel. The proximity of the drops to the sensor facilitates efficient collection of fluorescence emission from the drops, and overcomes the trade-off between light collection efficiency and field of view in conventional microscopy. The interrogation rate of our device is currently limited by the acquisition speed of CMOS sensor, and is expected to increase further as high-speed sensors become increasingly available.

Effect of propane-2-sulfonic acid octadec-9-enyl-amide on the expression of adhesion molecules in human umbilical vein endothelial cells.

Oleoylethanolamide (OEA), an endogenous agonist of PPARα, has been reported to have anti-atherosclerotic properties. However, OEA can be enzymatically hydrolyzed to oleic acid and ethanolamine and, thus, is not expected to be orally active. In the present study, we designed and synthesized an OEA analog, propane-2-sulfonic acid octadec-9-enyl-amide (N15), which is resistant to enzymatic hydrolysis. The purpose of this study was to investigate the effects of N15 on the expression of adhesion molecules in human umbilical vein endothelial cells (HUVECs). The results showed that N15 inhibited TNFα-induced production of vascular cell adhesion molecule-1 and intercellular adhesion molecule-1 and the adhesion of monocytes to TNFα-induced HUVECs. Furthermore, the protective effect of N15 on inflammation is dependent upon a PPAR-α/γ-mediated mechanism. In conclusion, N15 protects against TNFα-induced vascular endothelial inflammation. This anti-inflammatory effect of N15 is dependent on PPAR-α/γ dual targets.

[Multi-central randomized controlled study on electroacupuncture at Fenglong (ST 40) for regulating blood lipids].

OBJECTIVE: To investigate the clinical effects of electroacupuncture (EA) at Fenglong (ST 40) on blood lipids. METHODS: Two hundred and four patients of hyperlipidemia were randomly divided into a Fenglong group and a Xuezhikang group, 102 cases in each group. The patients in the Fenglong group were treated with electroacupuncture at Fenglong (ST 40). After arrival of qi, the needles were connected with acupoint nerve stimulator (LH 202 H type, HANS). The primary parameters of EA: for high triglycerides (TG) type, AM 50 Hz, intensity 1 mA, needle-retained time 20 min, twice per week; for high cholesterol (CHO) type, AM 100 Hz, intensity 1 mA, needle-retained time 30 min, thrice per week; for high low-density-lipoprotein (LDL-C) type, the same parameters as the high CHO type except the tolerable and comfortable intensity; for the mixing type, corresponding methods were alternatively used. The patients in the Xuezhikang group received Xuezhikang capsule orally, 2 capsules each time and twice daily, for total 11 weeks. RESULTS: The total effective rates of the Fenglong group and the Xuezhi-kang group were 83.0% and 85.9%, respectively, with no significant difference between the two groups (P > 0.05), and there was no significant differences in the function of regulating blood lipids between the two groups (all P > 0.05). After one month follow-up survey, the total CHO, TG and LDL-C decreased and high-density-lipoprotein (HDL-C) increased, of which there was a significant difference in TG reduction (P < 0.05). There were no relapses in both groups. CONCLUSION: EA at Fenglong (ST 40) can effectively regulate blood lipids with a better after-effect, which can be applied as a safe and effective method to replace medication for regulating blood lipids.