The protocol of CUT&Tag for metabolic tissue cells.

Cleavage under target and tagment (CUT&Tag) is a technology that utilizes the fusion protein of Tn5 transposase and protein A/G which can guide Tn5 enzyme to the antibody bound to target protein and cleave the chromatin regions adjacent to target protein. Chromatin libraries are then tagged and sequenced by the high-throughput sequencing to obtain chromatin information at specific sites or protein binding locations. CUT&Tag technology plays an important role in the research of DNA and protein interactions. It can be used to understand the modifications of histone and the bindings of transcription factors. Compared with the traditional chromatin immunoprecipitation-sequencing (ChIP-seq) technology, the CUT&Tag has the strengths of high signal-to-noise ratio, good repeatability, short experimental period, and low cell input. It shows great advantages in early embryonic development, stem cells, cancer, epigenetics and other research fields. In this article, we described the protocol of CUT&Tag for metabolic tissue cells (mouse primary islet cells), to provide an epigenetic method for studying metabolic cells.

Interaction Tolerance Detection Test for Understanding the Killing Efficacy of Directional Antibiotic Combinations.

Combination treatments are commonly prescribed for enhancing drug efficacy, as well as for preventing the evolution of resistance. The interaction between drugs is typically evaluated near the MIC, using growth rate as a measure of treatment efficacy. However, for infections in which the killing activity of the treatment is important, measurements far above the MIC are needed. In this regime, the killing rate often becomes weakly concentration dependent, and a different metric is needed to characterize drug interactions. We evaluate the interaction metric on killing using an easy visual assay, the interaction tolerance detection test (iTDtest), that estimates the survival of bacteria under antibiotic combinations. We identify antibiotic combinations that enable the eradication of tolerant bacteria. Furthermore, the visualization of the antibiotic interactions reveals directional drug interactions and enables predicting high-order combination outcomes, therefore facilitating the determination of optimal treatments. IMPORTANCE The killing efficacy of antibiotic combinations is rarely measured in the clinical setting. However, in cases where the treatment is required to kill the infecting organism and not merely arrest its growth, the information on the killing efficacy is important, especially when tolerant strains are implicated. Here, we report on an easy method for the determination of the killing efficacy of antibiotic combinations which enabled to reveal combinations effective against tolerant bacteria. The results could be generally used to guide antimicrobial therapy in life-threatening infections.

Noise Analysis of Monolayer Graphene Nanopores.

Graphene-based nanopore devices have shown tantalizing potential in single molecule detection for their monoatomic membrane thickness which is roughly equal to the gap between nucleobases. However, high noise level hampers applications of graphene nanopore sensors, especially at low frequencies. In this article, we report on a study of the contribution of suspended graphene area to noise level in full frequency band. Monolayer graphene films are transferred onto SiNx substrates preset with holes in varied diameters and formed self-supported films. After that, the films are perforated with smaller, nanoscale holes. Experimental studies indicate a dependency of low-frequency 1/f noise on the underlying SiNx geometry. The contribution of the suspended graphene area to capacitance which affects the noise level in the high frequency range reveals that the graphene free-standing film area influences noise level over a wide frequency region. In addition, the low-frequency noise demonstrates a weak dependency on salt concentration, in deviation from Hooge's relation. These findings and attendant analysis provide a systematic understanding of the noise characteristics and can serve as a guide to designing free-standing monolayer graphene nanopore devices.

[Terahertz Spectroscopic Study on the Property of Epoxy Resin Adhesive].

Epoxy resin is an important adhesive applied in the manufacturing processes of fiber reinforced polymer (FRP) composites. Terahertz (THz) time-domain spectroscopy (TDS) technology is an effective supplementary method for nondestructive evaluation (NDE) of FRP composites. As one of the most important parameters for epoxy resin, different curing temperature can affect the properties of epoxy resin. In this paper, we carry out systematic investigations on THz transmission properties of epoxy resin cured respectively under room temperature and high temperature with THz TDS technology. At the same time, the authors extract the refractive indices and absorption coefficients of epoxy resin, and make comparisons d. As shown in the experiments, the epoxy resin samples cured under room temperature have no bubble, whereas there are some micro-bubbles in the samples cured under high temperature, which reduce the sample density. Hence, the refractive index and absorption coefficient of epoxy resin cured under room temperature are both greater than those cured under high temperature. The difference of refractive index of different samples cured under the same condition is not significant. In addition, the difference of absorption coefficient of different samples cured under room temperature is also slight. However, the difference of absorption coefficient of different samples cured under high temperature gradually increase within the frequency from 0.6 to 1.5 THz, which is mainly due to the heterogeneous distribution of the bubbles in the different samples cured under high temperature. Moreover, the absorption coefficient of epoxy resin prepared under both curing temperatures gradually increases with the frequency, and there is no obvious absorption peak. Finally, because of the existence of Fabry-Pérot interference, the power transmission ratio of thicker epoxy resin samples may be greater than thinner samples at the resonant frequency. This research is of great significance for the THz NDE of FRP composites.

[Experimental Study of PMI Foam Composite Properties in Terahertz].

Polymethacrylimide (PMI) foam composite has many excellent properties. Currently, PMI is heat-resistant foam, with the highest strength and stiffness. It is suitable as a high-performance sandwich structure core material. It can replace the honeycomb structure. It is widely used in aerospace, aviation, military, marine, automotive and high-speed trains, etc. But as new sandwich materials, PMI performance testing in the THz band is not yet visible. Based on the Terahertz (THz) time-domain spectroscopy technique, we conducted the transmission and reflection experiments, got the time domain waveforms and power density spectrum. And then we analyzed and compared the signals. The MATALB and Origin 8. 0 was used to calculate and obtain the transmittance (transfer function), absorptivity Coefficient, reflectance and the refractive index of the different thickness Degussa PMI (Model: Rohacell WF71), which were based on the application of the time-domain and frequency-domain analysis methods. We used the data to compared with the THz refractive index and absorption spectra of a domestic PMI, Baoding Meiwo Technology Development Co. , Ltd. (Model: SP1D80-P-30). The result shows that the impact of humidity on the measurement results is obvious. The refractive index of PMI is about 1. 05. The attenuation of power spectrum is due to the signal of the test platform is weak, the sample is thick and the internal scattering of PMI foam microstructure. This conclusion provides a theoretical basis for the THz band applications in the composite PMI. It also made a good groundwork for THz NDT (Non-Destructive Testing, NDT) technology in terms of PMI foam composites.

Aberrant hypermethylation and reduced expression of disabled-2 promote the development of lung cancers.

Disabled-2 (Dab2) is considered a tumor suppressor and is downregulated in cancers. We examined the promoter methylation status and expression levels of Dab2, and investigated their roles in the development of lung cancers. Methylation-specific PCR was employed to analyze the methylation status of Dab2 in 100 lung cancer tissues. The cytoplasmic and nuclear expression of the Dab2 protein was determined using western blot analysis. Demethylation treatment using 5-Aza-2-deoxycytidine (5-Aza-dC) was performed in three lung cancer cell lines. Dab2 expression was upregulated by Dab2 transfection or interrupted by Dab2 siRNA in lung cancer cells. Proliferative and invasive ability tests were performed with 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTS) and a Matrigel invasion assay, respectively. The methylation rate of Dab2 was significantly higher in lung cancer tissues compared to normal lung tissues. Dab2 methylation correlated with the reduced nuclear and cytoplasmic expression of Dab2, as well as the TNM stage and lymphatic metastasis of lung cancers. Treatment with 5-Aza-dC was able to eliminate the hypermethylation of Dab2, enhance Dab2 expression, and inhibit ß-catenin expression, and the proliferative and invasive ability of lung cancer cells. Upregulation of Dab2 expression reduced ß-catenin expression and proliferation and invasiveness of lung cancer cells. However, interruption of Dab2 expression induced the opposite results. Dab2 methylation is common in lung cancers, and is one of the most important factors responsible for the reduced expression of Dab2. Aberrant hypermethylation and reduced expression of Dab2 promote the development of lung cancers.

Differential proteomics analysis of proteins from human diabetic and age-related cataractous lenses.

BACKGROUND: To investigate the differential lens proteomics between diabetic cataract, age-related cataract, and natural subjects. MATERIALS AND METHODS: Two-dimensional electrophoresis (2-DE), mass spectrometry (MS), and enzyme-linked immunosorbent assay (ELISA) were employed. Total soluble proteins in lenses of type I diabetic cataract, age-related cataract (nondiabetic) patients, and normal control were extracted and subjected to 2-DE. The differential protein spots were recovered, digested with trypsin, and further applied to MALDI-TOF-MS. ELISA analysis was used to determine the levels of differential proteins in lenses of three groups. RESULTS: 2-DE analysis reflected that lens proteins of normal control, diabetic, and age-related cataract subjects were in the section of pH 5-9 and the relative molecular weights were 14-97 kDa, while relative molecular weight of more abundant crystallines was localized at 20-31 kDa. five differential protein spots were detected and identified using MALDI-TOF-MS, including beta-crystallin A3, alpha-crystallin B chain, chain A of crystal structure of truncated human beta-B1-crystallin, beta-crystallin B1, and an interesting unnamed protein product highly similar to alpha-crystallin B chain, respectively. ELISA analysis revealed that lenses of diabetic cataract patients should contain significantly more concentrations of beta-crystallin A3, alpha-crystallin B chain, and beta-crystallin B1 than those of age-related cataract patients and normal control. CONCLUSION: This study clearly reflected the differential proteins of diabetic cataract, age-related cataract lenses compared with natural subjects, and it is helpful for the further research on the principles and mechanisms of different types of cataract.

[Effects of electroacupuncture assistant general anesthesia on postoperative cognitive dysfunction of aged patients].

OBJECTIVE: To observe the effects of electroacupuncture (EA) assistant general anesthesia on postoperative cognitive dysfunction (POCD) of aged patients. METHODS: One hundred and twenty senile American Society of Anesthesiology (ASA) grade I - III patients (more than 65 years old) with non-cardiac surgery were randomly assigned to two groups, Group A and Group B, 60 cases in each group. Patients in Group A received general anesthesia, while those in Group B received EA assistant general anesthesia. Patients in Group B received EA at Baihui (DU20), Hegu (LI4), Neiguan (PC6), Zusanli (ST36) 30 min before anesthesia induction to the end of operation. Anesthesia was induced by etomidate 0.2 - 0.3 mg/kg, fentanyl 3-5 microg/kg, cisatracurium 0.15 mg/kg in the two groups. Anesthesia maintenance was provided by sevoflurane and continuous pumping of remifentanil. Tramadol 50 mg was given 30 min before ending the operation. The scores of mini-mental state examination (MMSE), nausea and vomiting were recorded in the two groups one day before anesthesia, the 2nd, 4th, and 6th day after operation. The occurrence of postoperative cognitive dysfunction (POCD) on day 2, 4, and 6 was compared. RESULTS: The occurrence of POCD on day 2 and 4 was obviously lower in Group B than in Group A at the same time period (40.0% vs 66.7%, 13.3% vs 43.3%), showing statistical difference (P < 0.05). There was no significant difference in the pre-anesthesia scores of MMSE between the two groups. The scores of MMSE on the 2nd and the 4th day were higher in Group B than in Group A (21.3 +/- 3.9 vs 18.3 +/- 3.8, 26.4 +/- 2.9 vs 22.9 +/- 3.9, P < 0.05). Compared with one day before anesthesia in the same group, the scores of MMSE were significantly different on the 2nd and the 4th day (Group A: 18.3 +/- 3.8, 22.9 +/- 3.9 vs 27.9 +/- 2.1; Group B: 21.3 +/- 3.9 vs 27.5 +/- 2.5; P < 0.05). The occurrence of post-aesthesia nausea and vomiting was lower in Group B (23.3%, 14/60) than in Group A (46.7%, 28/60) with statistical difference (P < 0.05). CONCLUSION: EA assistant general anesthesia could reduce the occurrence of POCD in aged patients.