Design and experiment of high-field asymmetric waveform ion mobility spectrometry chip based on an integrated temperature control printed circuit board structure.

RATIONALE: During the detection of volatile organic compounds (VOC) using high-field asymmetric waveform ion mobility spectrometry (FAIMS), the ambient temperature significantly impacts the accuracy of planar FAIMS. To mitigate the influence of ambient temperature on detection accuracy and enhance resolution, a FAIMS system based on the inner impedance characteristics of a printed circuit board (PCB) was designed for temperature control. METHODS: This study, conducted under standard atmospheric pressure, aimed to assess the signal stability of a planar FAIMS instrument with and without temperature control, and the effect of temperature change on the detection ability of acetone, ethanol, and their mixture was studied using PCB self-heating. RESULTS: Experimental results demonstrated that the base noise in FAIMS with temperature control was 0.2 pA, whereas that in FAIMS without temperature control was 1.8 pA. Notably, with increasing temperature, the detection ability of FAIMS changes accordingly. The optimal relative detection ability of acetone was observed when the electrode plate was heated to 45°C under an electric field of 15 kV/cm. CONCLUSIONS: This study provides a novel approach to improve the resolving power of FAIMS systems and their signal-to-noise ratio. The utilization of a PCB-based temperature control proved effective in stabilizing FAIMS signal characteristics and optimizing detection capabilities, particularly for VOCs such as acetone. These findings have significant implications for improving the accuracy and resolving power of FAIMS systems in VOC detection applications.

Screening and Mutation of Saccharomyces cerevisiae UV-20 with a High Yield of Second Generation Bioethanol and High Tolerance of Temperature, Glucose and Ethanol.

A wild-type strain was isolated from slightly rotted pears after three rounds of enrichment culture, identified as Saccharomyces cerevisiae 3308, and evaluated for its fermentation capability of second generation bioethanol and tolerance of temperature, glucose and ethanol. S. cerevisiae 3308 was mutated by using the physical and chemical mutagenesis methods, ultraviolet (UV) and diethyl sulfate (DES), respectively. Positive mutated strains were mainly generated by the treatment of UV, but numerous negative mutations emerged under the treatment of DES. A positive mutated strain, UV-20, produced ethanol from 62.33 ± 1.34 to 122.22 ± 2.80 g/L at 30-45 °C, and had a maximum yield of ethanol at 37 °C. Furthermore, UV-20 produced 121.18 ± 2.51 g/L of second generation bioethanol at 37 °C. Simultaneously, UV-20 exhibited superior tolerance to 50% of glucose and 21% of ethanol. In a conclusion, all of these results indicated that UV-20 has a potential industrial application value.

Detection of Cyclic Dinucleotides by STING.

STING (stimulator of interferon genes) is an essential signaling adaptor protein mediating cytosolic DNA-induced innate immunity for both microbial invasion and self-DNA leakage. STING is also a direct receptor for cytosolic cyclic dinucleotides (CDNs), including the microbial secondary messengers c-di-GMP (3',3'-cyclic di-GMP), 3',3'cGAMP (3',3'-cyclic GMP-AMP), and mammalian endogenous 2',3'cGAMP (2',3'-cyclic GMP-AMP) synthesized by cGAS (cyclic GMP-AMP synthase). Upon CDN binding, STING undergoes a conformational change to enable signal transduction by phosphorylation and finally to active IRF3 (Interferon regulatory factor 3) for type I interferon production. Here, we describe some experimental procedures such as Isothermal Titration Calorimetry and luciferase reporter assays to study the CDNs binding and activity by STING proteins.

Physiological and metabolic differences between visceral and subcutaneous adipose tissues in Nile tilapia (Oreochromis niloticus).

Visceral adipose tissue (VAT) and subcutaneous adipose tissue (SCAT) have different structures and metabolic functions and play different roles in the regulation of the mammal endocrine system. However, little is known about morphology and physiological and metabolic functions between VAT and SCAT in fish. We compared the morphological, physiological, and biochemical characteristics of VAT and SCAT in Nile tilapia and measured their functions in energy intake flux, lipolytic ability, and gene expression patterns. SCAT contained more large adipocytes and nonadipocytes than VAT in Nile tilapia. VAT had higher lipid content and was the primary site for lipid deposition. Conversely, SCAT had higher hormone-induced lipolytic activity. Furthermore, SCAT had a higher percentage of monounsaturated and lower polyunsaturated fatty acids than VAT. SCAT had higher mitochondrial DNA, gene expression for fatty acid ß-oxidation, adipogenesis, and brown adipose tissue characteristics, but it also had a lower gene expression for inflammation and adipocyte differentiation than VAT. SCAT and VAT have different morphological structures, as well as physiological and metabolic functions in fish. VAT is the preferable lipid deposition tissue, whereas SCAT exhibits higher lipid catabolic activity than VAT. The physiological functions of SCAT in fish are commonly overlooked. The present study indicates that SCAT has specific metabolic characteristics that differ from VAT. The differences between VAT and SCAT should be considered in future metabolism studies using fish as models, either in biomedical or aquaculture studies.

Nonspecific DNA Binding of cGAS N Terminus Promotes cGAS Activation.

The cytosolic DNA sensor cyclic GMP-AMP synthase (cGAS) mediates innate immune responses against invading pathogens, or against self-dsDNA, which causes autoimmune disorders. Upon nonspecific binding of cytosolic B-form DNA, cGAS synthesizes the second messenger 2'3'-cGAMP and triggers STING-dependent signaling to produce type I IFNs. The cGAS comprises less-conserved N-terminal residues and highly conserved nucleotidyltransferase/Mab21 domains. The function and structure of the well-conserved domains have been extensively studied, whereas the physiological function of the N-terminal domain of cGAS is largely uncharacterized. In this study we used a single-molecule technique combined with traditional biochemical and cellular assays to demonstrate that binding of nonspecific dsDNA by the N-terminal domain of cGAS promotes its activation. We have observed that the N terminus of human cGAS (hcGAS-N160) undergoes secondary structural change upon dsDNA binding in solution. Furthermore, we showed that the hcGAS-N160 helps full length hcGAS to expand the binding range on λDNA and facilitates its binding efficiency to dsDNA compared with hcGAS without the 160 N-terminal residues (hcGAS-d160). More importantly, hcGAS-N160 endows full length hcGAS relatively higher enzyme activity and stronger activation of STING/IRF3-mediated cytosolic DNA signaling. These findings strongly indicate that the N-terminal domain of cGAS plays an important role in enhancing its function.

Single Mutations Reshape the Structural Correlation Network of the DMXAA-Human STING Complex.

Subtle changes in protein sequences are able to alter ligand-protein interactions. Unraveling the mechanism of such phenomena is important for understanding ligand-protein interactions, including the DMXAA-STING interaction. DMXAA specifically binds to mouse STING instead of human STING. However, the S162A mutation and a newly discovered E260I mutation endow human STINGAQ with DMXAA sensitivity. Through molecular dynamics simulations, we revealed how these single mutations alter the DMXAA-STING interaction. Compared to mutated systems, structural correlations in the interaction of STINGAQ with DMXAA are stronger, and the correlations are cross-protomers in the dimeric protein. Analyses on correlation coefficients lead to the identification of two key interactions that mediate the strong cross-protomer correlation in the DMXAA-STINGAQ interaction network: DMXAA-267T-162S* and 238R-260E*. These two interactions are partially and totally interrupted by the S162A and E260I mutations, respectively. Moreover, a smaller number of water molecules are displaced upon DMXAA binding to STINGAQ than that on binding to its mutants, leading to a larger entropic penalty for the former. Considering the sensitivity of STINGAQ and two of its mutants to DMXAA, a strong structural correlation appears to discourage DMXAA-STING binding. Such an observation suggests that DMXAA derivatives, which are deprived of hydrogen-bond interaction with both 162S* and 267T, are potential agonists of human STING.

Effects of professional rehabilitation training on the recovery of neurological function in young stroke patients.

Young stroke patients have a strong desire to return to the society, but few studies have been conducted on their rehabilitation training items, intensity, and prognosis. We analyzed clinical data of young and middle-aged/older stroke patients hospitalized in the Department of Neurological Rehabilitation, China Rehabilitation Research Center, Capital Medical University, China from February 2014 to May 2015. Results demonstrated that hemorrhagic stroke (59.6%) was the primary stroke type found in the young group, while ischemic stroke (60.0%) was the main type detected in the middle-aged/older group. Compared with older stroke patients, education level and incidence of hyperhomocysteinemia were higher in younger stroke patients, whereas, incidences of hypertension, diabetes, and heart disease were lower. The average length of hospital stay was longer in the young group than in the middle-aged/older group. The main risk factors observed in the young stroke patients were hypertension, drinking, smoking, hyperlipidemia, hyperhomocysteinemia, diabetes, previous history of stroke, and heart disease. The most accepted rehabilitation program consisted of physiotherapy, occupational therapy, speech therapy, acupuncture and moxibustion. Average rehabilitation training time was 2.5 hours/day. Barthel Index and modified Rankin Scale scores were increased at discharge. Six months after discharge, the degree of occupational and economic satisfaction declined, and there were no changes in family life satisfaction. The degrees of other life satisfaction (such as friendship) improved. The degree of disability and functional status improved significantly in young stroke patients after professional rehabilitation, but the number of patients who returned to society within 6 months after stroke was still small.

Rat and human STINGs profile similarly towards anticancer/antiviral compounds.

Cyclic dinucleotides (CDNs) and antitumor/antiviral agents (DMXAA and CMA) trigger STING-dependent innate immunity activation. Accumulative evidences have showed that DMXAA and CMA selectively activate mouse, but not human STING signaling. The mechanism underlying this species selectivity remains poorly understood. In this report, we have shown that human and rat STINGs display more similar signaling profiles toward DMXAA and CMA than that of human and mouse STINGs, suggesting that rat is more suitable for preclinical testing of STING-targeted drugs. We have also determined the crystal structures of both apo rat STING and its complex with cyclic GMP-AMP with 2'5' and 3'5' phosphodiester linkage (2'3'-cGAMP), a human endogenous CDN. Structure-guided biochemical analysis also revealed the functional importance of the connecting loop (A140-N152) between membrane and cytosolic domains in STING activation. Taken together, these findings reveal that rat STING is more closely related to human STING in terms of substrate preference, serving as a foundation for the development of STING-targeted drugs.

The structural basis for the sensing and binding of cyclic di-GMP by STING.

STING (stimulator of interferon genes) is an essential signaling adaptor that mediates cytokine production in response to microbial invasion by directly sensing bacterial secondary messengers such as the cyclic dinucleotide bis-(3'-5')-cyclic dimeric GMP (c-di-GMP). STING's structure and its binding mechanism to cyclic dinucleotides were unknown. We report here the crystal structures of the STING cytoplasmic domain and its complex with c-di-GMP, thus providing the structural basis for understanding STING function.

Rheb1 is required for mTORC1 and myelination in postnatal brain development.

mTor kinase is involved in cell growth, proliferation, and differentiation. The roles of mTor activators, Rheb1 and Rheb2, have not been established in vivo. Here, we report that Rheb1, but not Rheb2, is critical for embryonic survival and mTORC1 signaling. Embryonic deletion of Rheb1 in neural progenitor cells abolishes mTORC1 signaling in developing brain and increases mTORC2 signaling. Remarkably, embryonic and early postnatal brain development appears grossly normal in these Rheb1f/f,Nes-cre mice with the notable exception of deficits of myelination. Conditional expression of Rheb1 transgene in neural progenitors increases mTORC1 activity and promotes myelination in the brain. In addition the Rheb1 transgene rescues mTORC1 signaling and hypomyelination in the Rheb1f/f,Nes-cre mice. Our study demonstrates that Rheb1 is essential for mTORC1 signaling and myelination in the brain, and suggests that mTORC1 signaling plays a role in selective cellular adaptations, rather than general cellular viability.

Antitumor effect of mSurvivinThr34→Ala in murine colon carcinoma when administered intravenously.

Colorectal cancer is one of the most common cancers. Survivin is strongly immunogenic in a fraction of colorectal cancer patients. The present study was designed to determine whether full-length mouse Survivin dominant-negative mutant SurvivinT34A has the antitumor activity in a murine colon carcinoma model. The complex of cationic liposome (DOTAP/Chol) to plasmid pORF9-mSurvivin T34A was administered intravenously in a mouse subcutaneous (S. C.) CT 26 tumor model. Apoptotic cells and anti-angiogenesis were evaluated by fluorescent in situ TUNEL assay and by immunohistochemistry with an antibody reactive to CD31, respectively. A 4 h 51Cr release assay was performed to determine Survivin-specific cytotoxicity. The adoptive transfer of CD8+ or CD4+ T-lymphocytes assay was to further explore the roles of immune cell subsets. We demonstrated the complex of cationic liposome (DOTAP/Chol) to plasmid pORF9--mSurvivin T34A when administered intravenously induced an efficient antitumor activity in a S. C. CT26 tumor model in mice. The main mechanism is involved in three aspects: triggering the apoptosis of tumor cells, inhibiting angiogenesis, and inducing Survivin-specific immune response. Our observations may have potential implications for the further exploration of the treatment of human colorectal cancer by intravenous delivery of dominant-negative mutant Survivin T34A.

Magnetic chitosan nanoparticles as a drug delivery system for targeting photodynamic therapy.

Photodynamic therapy (PDT) has become an increasingly recognized alternative to cancer treatment in clinic. However, PDT therapy agents, namely photosensitizer (PS), are limited in application as a result of prolonged cutaneous photosensitivity, poor water solubility and inadequate selectivity, which are encountered by numerous chemical therapies. Magnetic chitosan nanoparticles provide excellent biocompatibility, biodegradability, non-toxicity and water solubility without compromising their magnetic targeting. Nevertheless, no previous attempt has been reported to develop an in vivo magnetic drug delivery system with chitosan nanoparticles for magnetic resonance imaging (MRI) monitored targeting photodynamic therapy. In this study, magnetic targeting chitosan nanoparticles (MTCNPs) were prepared and tailored as a drug delivery system and imaging agents for PS, designated as PHPP. Results showed that PHPP-MTCNPs could be used in MRI monitored targeting PDT with excellent targeting and imaging ability. Non-toxicity and high photodynamic efficacy on SW480 carcinoma cells both in vitro and in vivo were achieved with this method at the level of 0-100 microM. Notably, localization of nanoparticles in skin and hepatic tissue was significantly less than in tumor tissue, therefore photosensitivity and hepatotoxicity can be attenuated.

Temporal correlation of the memory deficit with Alzheimer-like lesions induced by activation of glycogen synthase kinase-3.

We have reported that activation of glycogen synthase kinase-3 (GSK-3) by ventricle injection of wortmannin (WT) and GF-109203X (GFX) induces Alzheimer-like memory deficit in rats [Liu et al., J. Neurochem. 87 (2003), 1333]. To further explore the factors responsible for the memory loss, we studied here the temporal alterations of GSK-3, tau phosphorylation, beta-amyloid (Abeta), and acetylcholine (ACh) after injection of WT/GFX, and analyzed their correlation with the memory loss. We observed that the severe memory deficit occurred at 24 and 48 h, and simultaneously, GSK-3 activation, tau hyperphosphorylation at Thr231, Ser396, and Ser404 and decline of ACh in hippocampus were detected, and these changes were mostly recovered at 72 and 96 h after the injection of WT/GFX. Remarkable increase of Abeta and intracellular accumulation of argentophilic substances were detected at 72 h. Pearson analysis showed that the memory deficit was correlated with GSK-3 activation, tau hyperphosphorylation, and decline of ACh but not with Abeta overproduction. Our data provide direct evidence demonstrating that activation of GSK-3 by WT/GFX may cause memory deficit through tau hyperphosphorylation and suppression of ACh in hippocampus.

Correlation between choline signal intensity and acetylcholine level in different brain regions of rat.

Acetylcholine is an important excitatory neurotransmitter, which plays a crucial role in synaptic transmission. The level of acetylcholine is decreased in the early stages of Alzheimer disease (AD), the most common neurodegenerative disease. Therefore, measurement of acetylcholine in the brain may help the clinical diagnosis of AD. However, the methods used till now to detect the brain acetylcholine level are invasive, which are neither recommended nor acceptable in the clinic. Acetylcholine is synthesized from choline-containing compounds (Cho), the latter can be estimated by noninvasive proton magnetic resonance spectroscopy ((1)H MRS). To explore whether the Cho signal intensity could be used to represent the acetylcholine level in the brain, we employed (1)H MRS to detect the Cho signal, and simultaneously, we also used microdialysis and high-performance liquid chromatography (HPLC) to measure the level of acetylcholine in hippocampus, striatum, frontal cortex, and somatosensory barrel field (S1BF cortex) of rats, respectively. The results showed that the correlations between Cho signal intensity and acetylcholine level in hippocampus, striatum, frontal cortex, and S1BF cortex were, respectively, 0.823 (p = 0.044), 0.851 (p = 0.032), 0.817 (p = 0.047), and 0.822 (p = 0.045). The F-values of the regression model were, respectively, 8.404 (p = 0.044), 10.47 (p = 0.032), 8.000 (p = 0.047), and 8.326 (p = 0.045). And the derived regression equations were y = 0.67x + 1.363 (hippocampus), y = 5.398x + 6.684 (striatum), y = 0.656x + 0.564 (frontal cortex), and y = 0.394x + 1.127 (S1BF cortex), respectively (y means acetylcholine, and x means Cho). These data suggest that the Cho signal intensity observed by (1)H MRS may be used as an indicator of acetylcholine level in different brain regions of the rats.

[Etiologic analysis of 1137 inpatients of chronic renal failure].

OBJECTIVE: To investigate the main causes of chronic renal failure (CRF) and the differences in how subpopulation is affected by these causes, and to determine the morbidity of anemia and hypertension, and the relation between age and gender in CRF patients. METHODS: We chose 1137 inpatients diagnosed as CRF in the Department of Nephropathy of 2nd Xiangya Hospital of Center South University from January 1996 to March 2002. We collected the information on name, gender, age, admission number, diagnosis, blood pressure, blood routine, renal function, and creatinine clearance rate. RESULTS: Chronic glomerulonephitied accounted for 56.4% as the leading cause, followed by hypertensive nephropathy (14.3%), obstructive nephropathy (7.2%), and diabetic nephropath (6.7%). Gender influenced the incidence of some diseases. Most patients were companied with anemia (90.7%) and hypertension (72.6%). CONCLUSION: The main causes of CRF include chronic glomerulonephitis, hypertensive nephropathy, obstructive nephropathy, and diabetic nephropath.