The phytohormone abscisic acid (ABA) regulates growth, development, and response to abiotic stress in plants. The core ABA signaling has three key components: ABA receptors PYRABACTIN RESISTANCE 1 ...and PYR1-Like protein (PYR1/PYLs), type 2C protein phosphatases (PP2Cs) and SNF1- related protein Kinase 2s (SnRK2s). In the presence of ABA, PYR1/PYLs bind to and inhibit PP2Cs, leading to the release of SnRK2s from suppression, to activate downstream events such as ABA-responsive gene expression. To identify new components in the ABA signaling pathway, we performed a yeast two-hybrid screen for PYLs and SnRK2s interactors. We identified two proteins that interact with PYLs and SnRK2s: Ty pe One Protein Phosphatase 1 (TOPP1) and its regulatory protein At Inhibitor 2 (AtI-2). We show that TOPP1 and AtI-2 inhibit SnRK2.6 activity in vitro and that the inhibition is reversed by PYLs in the presence of ABA. Mutants topp1-1topp4-1 and ati-2 are hypersensitive to ABA during seed germination. ABA-responsive genes are highly up regulated in topp1-1topp4-1 and ati-2 mutants. Taken together, our results suggest that the TOPP1 and AtI-2 form a complex that negatively modulates ABA signaling by interacting with PYLs and SnRK2s
Sepsis is a major medical inflammatory response to bloodstream infection caused by
pathogens. Over 30 million infections occur due to sepsis in a year throughout the globe.
With over 6 million ...deaths, sepsis treatment demands timely diagnosis and medical
interventions to prevent large outspread of the infection. Rapid and sensitive detection of
infectious bacteria plays an important role in disease control. However, fast and accurate
diagnosis remains highly challenging for many hospitalized patients. Among various
bacterial diseases, Escherichia coli, (E. coli) a gram-negative bacterium, is the most
frequently and extensively researched and has become a global concern of food safety
problems. Traditional cell culture methods are considered as most accurate method for
bacterial diagnosis. But some microorganisms like Bartonella spp, Treponema pallidum, etc.
are difficult to grow on a culture plate, while some other virus species are unculturable, so
eventually the traditional cell culture may take several days to a week for the diagnostic
results. Hence, over several years, different novel methods have been developed, such as
optical methods, electrochemical methods, molecular detection by PCR etc. Even though
several techniques exist, concerns have been raised regarding their sensitivity, specificity etc.
In addition, various fluorescence-based methods, and Surface Plasmon resonance (SPR)-
based sensors have also been reported for bacterial detection. However, high cost, large
turnaround time, expensive instrumentation, and requirement of skilled personnel are some of
the disadvantages of these existing techniques.
FET biosensors have previously been used in different diagnosis areas. Nowadays, several
FET-based biosensors have been developed, which primarily concentrate on improving the
overall sensitivity of FET devices. However, the charge screening effect observed in a
physiological environment, and additional sample processing increase the complexity of the
sensor platform. To overcome this hurdle, an electrical double layer (EDL) gated FET
biosensor has been proposed and explored in the detection of a variety of biological entities.
Here, we proposed an EDL-FET sensor platform for the detection of Escherichia coli
O157:H7 strain in blood serum. A single-step surface functionalization strategy was used for
immobilizing receptor probes as ssDNA. The probes were designed and immobilized over a
disposable chip containing a pair of gold electrodes. A handheld reader which consists of an
enhancement mode N-channel MOSFETs was connected to the laptop through a USB port to
measure the sensor signals. Pulsed gate voltage was applied to the input electrode and which
modulated the EDL capacitance and potential change eventually resulting in the drain current
of the MOSFET. The probe was immobilized on the gate electrode and tested with different
concentrations of cDNA spiked in TE buffer. The probe-target binding was observed through
fluorescence change and electrically by measuring the change in drain current. In addition,
surface potential change was also examined by performing KPFM test. Similarly, the BioFET
detections were validated for E. coli O157 variants by spiking with short and long DNA
target sequences simultaneously in serum. The test results confirm selectivity by testing
various non-complementary sequences, which ensures the potential of this BioFET as a
point-of-care sensing device.
Figure 1
ABSTRACT
Biliary Atresia (BA) is a severe liver disease that affects newborns. This disease may cause cholestasis and progressive hepatic failure or even death if not treated immediately. ELISA blood ...tests for metabolic screening are the current method of diagnosing BA. However, newborns need rapid BA detection for treatment in the future. The increased matrix metalloproteinase-7 (MMP7) cause BA-related liver fibrosis and newborns diagnosed with BA show higher MMP7 levels than those diagnosed with cholestasis. Several platforms have been developed for the fast detection of Biliary Atresia (BA). Here, we developed an electric-double-layer (EDL)-gated field-effect transistor (FET) platform for Biliary Atresia (BA) detection. The core idea of this sensor array is to use a uniquely designed DNA probe to capture MMP7 proteins and monitor the change in drain current caused by the target capturing. Different concentrations of MMP7 spiked and tested over a functionalized sensor. The simple and cost-effective fabrication and ease of use make this sensor a promising tool as a point care detection device.
Keywords: EDL, FET, Biliary atresia, MMP7, Biosensors
INTRODUCTION
Due to the symptomatic similarity to cholestatic liver diseases, it is challenging to diagnose a precise BA for newborns. False diagnosis may delay timely treatment (e.g., surgery) and worsens the prognosis. However, BA's underlying causes are still unclear. A level increase of intrahepatic matrix metalloproteinase-7 (MMP7) signifies BA-related liver fibrosis. BA cases found in newborns revealed higher MMP7 levels than cholestasis. MMP7 is a protease that contributes to tissue remodeling and breaks down extracellular matrix through a signal pathway. Based on the clinical results, MMP7 has emerged as a novel biomarker for BA diagnosis. ELISA was employed as the current method for detecting Biliary Atresia (BA). However, the procedure of ELISA has shortcomings, such as being time-consuming, high cost, and complicated operation. Hence, rapid and precise diagnostic methods need to be developed.
In this work, we propose to detect MMP7 proteins using an electric-double-layer (EDL)-gated field-effect transistor (FET). As shown in Figure 1, The aptamer of MMP7 was immobilized on sensor electrodes, allowing a binding with the protein. As the number of the captured analytes increases; the sensor signals, amplified by a FET, will respond to it. As such, we look forward to detecting MMP7 in serum samples collected from newborns and helping the clinical diagnostics by this EDL-gated FET biosensor.
MATERIALS AND METHODS
An extended gate chip was employed in the sensor array. The electrode surface was cleaned using oxygen plasma, following which, the probe was immobilized on it. The MMP7 aptamer is mixed with TCEP for 30 minutes at room temperature, TCEP was used as a reducing agent which helps in the formation of dithiol bonds (SS), making the attachment of the probe easier. The mixture solution is then dropped and cast over the sensor surface and immobilized at 24 ℃ for 24 hr. The sensor is washed with 0.01× PBS to remove unbound aptamers and was verified with fluorescent imaging.
ELECTRICAL MEASUREMENT
After the aptamer was immobilized over the sensor surface, a baseline measurement was carried out. The drain current was measured at different voltage biases (Vg=-2V and 3V, at Vd=3.5V). different concentration of MMP7 protein was spiked in 0.01x PBS and tested. A signal was read out every 1min, and 20 measurements were taken for each concentration of the target for 20 minutes.
RESULTS AND CONCLUSIONS
The platform established is to detect MMP7 protein precisely and further identify BA. Owning a characteristic of electrical amplification, the EDL-gated FET will be utilized to test MMP7 proteins with concentrations ranging from 10 ng/mL to 100 ng/mL, as shown in Figure 2. When testing proteins level at 100 ng/mL, the signal can increase by around 3.5 mA. The tests using EDL-FET were successfully validated for MMP7 detection in serum and 0.01x PBS and the results indicate that the proposed platform had a high sensitivity. The calibration curve (signal vs. analyte's concentration) will be established to quantify the unknown sample. Conclusively, the sensor developed in this study has the potential to be developed as a point-of-care testing device and is suitable for clinical diagnosis.
Figure 1
碩士
國立陽明大學
生命科學暨基因體科學研究所
96
Nitrate is not only a macronutrient but also acts as a signal molecule on regulating gene expression and root growth. Nitrate-induced rapid genes expression is referred as ...primary nitrate response. Little is known to be involved in nitrate signaling. In this study, signaling component genes from microarray data, two Calcineurin B-Like (CBL) and two F-box genes, were evaluated for their potential roles in nitrate signaling.
Previous study has shown that CBL-interacting protein kinase 8 (CIPK8) and CIPK23 was found to regulate primary nitrate response and nitrate-modulated primary root growth. Once signal perceived, CBLs directly activate CIPKs to trigger downstream response. Since CBL1 and CBL9 interact with CIPK8 and CIPK23, suggesting that these two CBLs may be involved in nitrate signaling. In this study, Q-PCR analysis showed that CBL1 and CBL9 were nitrate-inducible genes. The induction levels of primary nitrate response marker genes, nitrate transporters CHL1 and NRT2.1, were reduced in cbl1 and cbl9 mutants. However, the reduced expressions observed in single mutants were recovered in cbl1cbl9 double mutant. Yeast two-hybrid assay showed that CBL1 and CBL9 interacted with C-terminal region of CHL1. Xenopus oocyte uptake assay further revealed that CHL1 uptake activity was reduced while co-expressed with CBL1 and CHL1, implying that CBL1 might involved in modulation uptake ability or protein stability of CHL1. Furthermore, the cbl1cbl9, but not single mutants, displayed defect in primary root growth, indicating that CBL1 and CBL9 were redundant in regulating nitrate-mediated primary root growth. Taken together, this study showed that CBL1 and CBL9 played multiple roles in nitrate signaling.
Previous studies show that F-box genes participate in several plant signaling transduction such as hormone and light response. Although AFB3 and FBL6 were nitrate-inducible genes, the expressions of primary nitrate response genes and protein level of CHL1 were not significantly altered in afb3 and fbl6 mutants. Moreover, the primary root length of wild-type, afb3 and fbl6 were similar in nitrate medium. These results suggested that AFB3 and FBL6 might be involved in other unknown nitrate signaling pathway.
Background The angiotensin receptor–neprilysin inhibitor (LCZ696) has emerged as a promising pharmacological intervention against renin–angiotensin system inhibitor in reduced ejection fraction heart ...failure (HFrEF). Whether the therapeutic benefits may vary among heterogeneous HFrEF subgroups remains unknown. Methods and Results This study comprised a pooled 2‐center analysis including 1103 patients with symptomatic HFrEF with LCZ696 use and another 1103 independent HFrEF control cohort (with renin–angiotensin system inhibitor use) matched for age, sex, left ventricular ejection fraction, and comorbidity conditions. Three main distinct phenogroup clusterings were identified from unsupervised machine learning using 29 clinical variables: phenogroup 1 (youngest, relatively lower diabetes prevalence, highest glomerular filtration rate with largest left ventricular size and left ventricular wall stress); phenogroup 2 (oldest, lean, highest diabetes and vascular diseases prevalence, lowest highest glomerular filtration rate with smallest left ventricular size and mass), and phenogroup 3 (lowest clinical comorbidity with largest left ventricular mass and highest hypertrophy prevalence). During the median 1.74‐year follow‐up, phenogroup assignment provided improved prognostic discrimination beyond Meta‐Analysis Global Group in Chronic Heart Failure risk score risk score (all net reclassification index P <0.05) with overall good calibrations. While phenogroup 1 showed overall best clinical outcomes, phenogroup 2 demonstrated highest cardiovascular death and worst renal end point, with phenogroup 3 having the highest all‐cause death rate and HF hospitalization among groups, respectively. These findings were broadly consistent when compared with the renin–angiotensin system inhibitor control as reference group. Conclusions Phenomapping provided novel insights on unique characteristics and cardiac features among patients with HFrEF with sacubitril/valsartan treatment. These findings further showed potentiality in identifying potential sacubitril/valsartan responders and nonresponders with improved outcome discrimination among patients with HFrEF beyond clinical scoring.
Background: For advanced breast cancer with lymph node involvement, adjuvant radiotherapy (RT) with regional nodal irradiation (RNI) has been indicated to reduce cancer recurrence and mortality. ...However, an extensive RT volume is associated with normal organ exposure, which increases the toxicity and affects patient outcomes. Modern arc RT techniques can improve normal organ sparing compared with conventional techniques. The aim of this study was to explore the optimal technique for left-breast RT with RNI. Methods: We retrospectively reviewed patients receiving RT with RNI for left-breast cancer. We used modern arc RT techniques with either volumetric-modulated arc therapy (VMAT) or helical tomotherapy (HT) with a novel block technique, and compared differences in dosimetry parameters between the two groups. Subgroup analysis of RNI with or without internal mammary node (IMN) volume was also performed. Results: A total of 108 eligible patients were enrolled between 2017 and 2020, of whom 70 received VMAT and 38 received HT. The median RT dose was 55 Gy. No significant differences were found regarding the surgery, RT dose, number of fractions, target volume, and RNI volume between the VMAT and HT groups. VMAT reduced the heart mean dose more than HT (3.82 vs. 5.13 Gy, p < 0.001), as well as the cardiac parameters of V5–V20, whole-lung mean dose, lung parameters of V5–V20, and contralateral-breast and esophagus mean dose. In the subgroup analysis of RNI with IMNs, the advantage of VMAT persisted in protecting the heart, lung, contralateral breast, and esophagus. HT was beneficial for lowering the thyroid mean dose. For RNI without IMN, VMAT improved the low-dose exposure of the heart and lung, but HT was similar to VMAT in terms of heart, whole-lung, and contralateral-breast mean dose. Conclusions: For patients with left-breast cancer receiving adjuvant RT with RNI, VMAT reduced the exposure dose to the heart, lung, contralateral breast, and esophagus compared with HT. VMAT was superior to HT in terms of normal organ sparing in the patients who underwent RNI with IMN irradiation. Considering the reduction in normal organ exposure and potential toxicity, VMAT is the optimal technique for patients receiving RNI when deep inspiration breath-hold is not available.
Cellulose is a renewable feedstock for green industry. It is therefore important to develop a technique to construct a host with a high cellulolytic efficiency to digest cellulose. In this study, we ...developed a convenient host‐engineering technique to adjust the expression levels of heterologous genes in the host by promoter rearrangement and gene copy number adjustment. Using genes from different glycoside hydrolase (GH) families including GH2, GH3, GH5, GH6, GH7, and GH12 from Aspergillus niger, Trichoderma reesei, and Neocallimastix patriciarum, we constructed a cellulolytic Kluyveromyces marxianus with eight cellulase gene‐cassettes that produced a cellulase cocktail with a high cellulolytic efficiency, leading to a significant reduction in enzyme cost in a rice straw saccharification process. Our technique can be used to design a host that can efficiently convert biomass feedstock to biofuel.
Improving CBP by fine‐tuning cellulase cocktail in a cellulosic yeast via synthetic biology.
Physiological stress induces aversive memory formation and profoundly impacts animal behavior. In C. elegans, concurrent mitochondrial disruption induces aversion to the bacteria that the animal ...inherently prefers, offering an experimental paradigm for studying the neural basis of aversive memory. We find that, under mitochondrial stress, octopamine secreted from the RIC modulatory neuron targets the AIY interneuron through the SER-6 receptor to trigger learned bacterial aversion. RIC responds to systemic mitochondrial stress by increasing octopamine synthesis and acts in the formation of aversive memory. AIY integrates sensory information, acts downstream of RIC, and is important for the retrieval of aversive memory. Systemic mitochondrial dysfunction induces RIC responses to bacterial cues that parallel stress induction, suggesting that physiological stress activates latent communication between RIC and the sensory neurons. These findings provide insights into the circuit and neuromodulatory mechanisms underlying stress-induced aversive memory.
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•Systemic mitochondrial stress induces aversive associative memory•Distinct neurons regulate the formation and retrieval of aversive memory•Octopamine modulates the avoidance circuit via the SER-6 receptor•Functional circuit reorganization underlies stress-induced aversive memory
Liao et al. identify a neuronal circuit that mediates aversive learning under mitochondrial stress in C. elegans. Stress induces functional and behavioral reorganization through the neuromodulator octopamine that targets key neurons in the avoidance circuit.
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•Nanofiltration achieves SF up to 10 for 120 g/L ZnCl2 and 30 g/L glucose.•Retention of glucose reaches 95% in a solution of 120 g/L ZnCl2 by nanofiltration.•The rejection of ZnCl2 ...depends on Donnan exclusion and dielectric exclusion.•Desalinated bagasse hydrolysis sugar solution achieved 82% yield of bio-ethanol.•The availability of the membranes makes the industrial process scale-up easily.
The effects of pH, transmembrane pressure, and ZnCl2 concentration on the purification of glucose from solution through nanofiltration were investigated. The solutions used contained ZnCl2 and glucose in concentrations as high as 180 and 80 g/L, respectively. A high concentration of glucose was discovered in the retentate as the product, and ZnCl2 was filtered out into the permeate; the highest separation factor achieved was 10. The derivative Hagen–Poiseuille equation was applied to model the experimental data and calculate the effective thickness and pore radius of the membranes used. Moreover, an integrated multiple-nanofiltration system was designed to process a real bagasse hydrolysis solution. The fermentability of the purified sugar solution was confirmed, with an 82% total alcohol yield obtained. The multiple-nanofiltration system is both economically and environmentally sustainable for the production of bioethanol through chemical hydrolysis of biomass.
A strengthened chain of survival benefits patient outcomes after out-of-hospital cardiac arrest (OHCA).22OHCA, out-of-hospital cardiac arrest; TFD, Taipei Fire Department; CPR, cardiopulmonary ...resuscitation; PAD, public-access defibrillation; EMS, emergency medical service; GRA, Global Resuscitation Alliance; QA, quality assurance; BLS, basic life support; ALS, advanced life support; AED, automated external defibrillator; OR, odds ratio; CI, confidence interval; STHD, survival to hospital discharge; CPC, cerebral performance category; ROSC, return of spontaneous circulation; aOR, adjusted odds ratio; PEA, pulseless electrical activity; AHA, American Heart Association; TOR, termination of resuscitation. Over the past decade, the Taipei Fire Department (TFD) has continuously implemented system-wide initiatives on this issue.We hypothesised that for adult, non-trauma OHCA patients, the bundle of these system-wide initiatives are associated with better outcomes.
We conducted a registry-based, retrospective study to examine the association between consecutive system-level initiatives and OHCA survival on a two-yearly basis using trend analysis and multivariable logistic regression. The primary outcome was survival to hospital discharge (STHD) and favourable neurological status.
We analysed 18,076 cases from 2008 to 2017. The numbers of two-yearly cases of OHCA with resuscitation attempts from 2008 to 2017 were 3,576, 3,456, 3,822, 3,811, and 3,411. There was a significant trend of improved STHD (Two-fold) and favourable neurological outcome (Six-fold) over the past decade. Similar trends were observed in the shockable and non-shockable groups. Considering the first 2 years as baseline, the odds of STHD and favourable neurological status in the end of the initiatives increased significantly after adjusting for universally recognised predictors for OHCA survival.
For non-trauma adult OHCA in Taipei, continuous, multifaceted system-wide initiatives on the community chain of survival were associated with improved odds of STHD and favourable neurologic outcomes.
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