Long-term outcomes and prognostic predictors of patients with Takayasu's arteritis along with pulmonary artery involvement.

OBJECTIVES: This study aimed to investigate the clinical characteristics, results, and prognostic predictors of patients with Takayasu's arteritis (TAK) along with pulmonary artery involvement (PAI). METHODS: A total of 806 patients with TAK admitted to the Fuwai Hospital were screened. Clinical symptoms, imaging features, and prognosis were analysed, and patients were categorised into those with and those without pulmonary hypertension (PH). Additionally, risk factors associated with cardiac death and repeated hospitalisation were explored. RESULTS: Among 806 patients with TAK, 142 patients with PAI were included, 90.8% (n=129) of whom had PH diagnosed by right heart catheterisation and 9.2% (n=13) of whom did not. The median follow-up time was 54 (range, 29-83) months. Sixteen patients died from right heart failure caused by PH. Patients with PH were significantly more likely to have worse outcomes than patients without PH (p=0.027). The multivariate Cox proportional regression hazard model showed that the 6-min walk distance (6MWD) and PH-targeted therapy were independent prognostic predictors of cardiac death and hospital readmissions. CONCLUSIONS: This study found that that a significant proportion of patients with TAK along with PAI had PH. Patients with PH had worse prognosis than those without. Further 6MWD and PH-targeted therapy were independent prognostic predictors of cardiac death or repeated hospitalization. In the future, multi-centre clinical studies are needed to further prospectively clarify this issue.

The Discovery and Biosynthesis of Nicotinic Myxochelins from an Archangium sp. SDU34.

Myxobacteria are a prolific source of structurally diverse natural products, and one of the best-studied myxobacterial products is the siderophore myxochelin. Herein, we report two new compounds, myxochelins N (1) and O (2), that are nicotinic paralogs of myxochelin A, from the terrestrial myxobacterium Archangium sp. SDU34; 2 is functionalized with a rare 2-oxazolidinone. A precursor-feeding experiment implied that the biosynthesis of 1 or 2 was due to altered substrate specificity of the loading module of MxcE, which likely accepts nicotinic acid and benzoic acid instead of more conventional 2,3-dihydroxybenzoic acid. We also employed a phylogenomic approach to map the evolutionary relationships of the myxochelin biosynthetic gene clusters (BGCs) in all the available myxobacterial genomes, to pave the way for the future discovery of potentially hidden myxochelin derivatives. Although the biological function of 1 and 2 is unclear yet, this work underpins that even extensively studied BGCs in myxobacteria can still produce new chemistry.

Discovery of a new autophagy inducer for A549 lung cancer cells.

Biological activities of a series of fluorescent compounds against human lung cancer cell line A549 were investigated. The results showed that (E)-1,3,3-trimethyl-2-(4-(piperidin-1-yl)styryl)-3H-indol-1-ium iodide (8) and (E)-2-(5,5-dimethyl-3-(4-(piperazin-1-yl)styryl)cyclohex-2-en-1-ylidene) malononitrile (11) could inhibit the growth of A549 cancer cells in a dose and time-dependent manner. Furthermore, compound 8 could trigger autophagy and apoptosis, but not obviously induce necrosis under the stimulatory condition. Therefore, 8 can be used as autophagy activator to investigate the regulatory mechanism of autophagy and may offer a new candidate for the treatment of lung cancer.

Exploring the N-terminal role of a heterologous protein in secreting out of Escherichia coli.

Escherichia coli is commonly used as a host for the extracellular production of proteins. However, its secretion capacity is often limited to a frustratingly low level compared with other expression hosts, because E. coli has a complex cell envelope with two layers. In previous report, we identified that the catalytic domain of a cellulase (Cel-CD) from Bacillus subtilis can be secreted into the medium from recombinant E. coli in large quantities without its native signal peptide. In this study, we proved the N-terminal sequence of the full length Cel-CD played a crucial role in transportation through both inner and outer membranes. By subcellular location analysis, we verified that the secretion was a two-step process via the SecB-dependent pathway through the inner membrane and an unknown pathway through the outer membrane. However, the N-terminal region of Cel-CD is polar and hydrophilic, which showed no similarities to other typical signal sequences. Random mutagenesis experiment suggested that the N-terminal sequence is a compromising result of transportation through inner and outer membranes. This is the first report that a "non-classical signal peptide" can guide recombinant proteins out of the cells from cytoplasm. Biotechnol. Bioeng. 2016;113: 2561-2567. © 2016 Wiley Periodicals, Inc.

Construction of cellulose-utilizing Escherichia coli based on a secretable cellulase.

BACKGROUND: The microbial conversion of plant biomass into value added products is an attractive option to address the impacts of petroleum dependency. The Gram-negative bacterium Escherichia coli is commonly used as host for the industrial production of various chemical products with a variety of sugars as carbon sources. However, this strain neither produces endogenous cellulose degradation enzymes nor secrets heterologous cellulases for its poor secretory capacity. Thus, a cellulolytic E. coli strain capable of growth on plant biomass would be the first step towards producing chemicals and fuels. We previously identified the catalytic domain of a cellulase (Cel-CD) and its N-terminal sequence (N20) that can serve as carriers for the efficient extracellular production of target enzymes. This finding suggested that cellulose-utilizing E. coli can be engineered with minimal heterologous enzymes. RESULTS: In this study, a ß-glucosidase (Tfu0937) was fused to Cel-CD and its N-terminal sequence respectively to obtain E. coli strains that were able to hydrolyze the cellulose. Recombinant strains were confirmed to use the amorphous cellulose as well as cellobiose as the sole carbon source for growth. Furthermore, both strains were engineered with poly (3-hydroxybutyrate) (PHB) synthesis pathway to demonstrate the production of biodegradable polyesters directly from cellulose materials without exogenously added cellulases. The yield of PHB reached 2.57-8.23 wt% content of cell dry weight directly from amorphous cellulose/cellobiose. Moreover, we found the Cel-CD and N20 secretion system can also be used for the extracellular production of other hydrolytic enzymes. CONCLUSIONS: This study suggested that a cellulose-utilizing E. coli was created based on a heterologous cellulase secretion system and can be used to produce biofuels and biochemicals directly from cellulose. This system also offers a platform for conversion of other abundant renewable biomass to biofuels and biorefinery products.

Identification of a heterologous cellulase and its N-terminus that can guide recombinant proteins out of Escherichia coli.

BACKGROUND: The Gram-negative bacterium Escherichia coli has been widely used as a cell factory for the production of proteins and specialty chemicals because it is the best characterized host with many available expression and regulation systems. However, recombinant proteins produced in Escherichia coli are generally intracellular and often found in the form of inclusion bodies. Extracellular production of proteins is advantageous compared with intracellular production because extracellular proteins can be purified more easily and can avoid protease attack, which results in higher product quality. In this study, we found a catalytic domain of a cellulase (Cel-CD) and its N-terminus can be employed as carriers for extracellular production of recombinant proteins. RESULTS: In this report, we identified the catalytic domain of a cellulase (Cel-CD) from Bacillus sp. that can be secreted into the medium from recombinant E. coli BL21 (DE3) in large quantities without its native signal peptide. By subcellular location analysis, we proved that the secretion was a two-step process and the N-terminal sequence of the full length Cel-CD played a crucial function in secretion. Both the Cel-CD and its N-terminal sequence can serve as carriers for efficient extracellular production of select target proteins. CONCLUSIONS: Fusion of heterologous proteins with N20 from Cel-CD can carry the target proteins out of the cells with a concentration from 101 to 691 mg/L in flask cultivation. The extracellular recombinant proteins with a relative high purity. The results suggested that this system has a potential application in plant biomass conversion and industrial production of enzymes and therapeutic proteins.

Evolution of a chimeric aspartate kinase for L-lysine production using a synthetic RNA device.

Aspartate kinase (AK) is a key enzyme involved in catalyzing the first step of the aspartate-derived amino acid biosynthesis, including L-lysine and L-threonine, which is regulated by the end-metabolites through feedback inhibition. In order to accumulate the end-metabolites in the host, the feedback inhibition of AK has to be released. In this study, a chimeric aspartate kinase, which is composed of the N-terminal catalytic region from Bacillus subtilis AKII and the C-terminal region from Thermus thermophilus, was evolved through random mutagenesis and then screened using a high-throughput synthetic RNA device which comprises of an L-lysine-sensing riboswitch and a selection module. Of three evolved aspartate kinases, the best mutant BT3 showed 160 % increased in vitro activity compared to the wild-type enzyme. Recombinant Escherichia coli harboring BT3 produced 674 mg/L L-lysine in batch cultivation, similar to that produced by the strain harboring the typical commercial widely used feedback resistant aspartate kinase AKC (fbr) from E. coli. The results suggested that this strategy can be extended for screening of other key enzymes involved in lysine biosynthesis pathways.

A highly sensitive ratiometric fluorescent probe for detecting HSO3-/SO32- and viscosity change based on FRET/TICT mechanism.

BACKGROUND: Sulfur dioxide (SO2) is a significant gas signaling molecule in organisms, and viscosity is a crucial parameter of the cellular microenvironment. They are both involved in regulating many physiological processes in the human body. However, abnormalities in SO2 and viscosity levels are associated with various diseases, such as cardiovascular disease, lung cancer, respiratory diseases, neurological disorders, diabetes and Alzheimer's disease. Hence, it is essential to explore novel and efficient fluorescent probes for simultaneously monitoring SO2 and viscosity in organisms. RESULTS: We selected quinolinium salt with good stability, high fluorescence intensity, good solubility and low cytotoxicity as the fluorophore and developed a highly sensitive ratiometric probe QQD to identify SO2 and viscosity changes based on Förster resonance energy transfer/twisted intramolecular charge transfer (FRET/TICT) mechanism. Excitingly, compared with other probes for SO2 detection, QQD not only identified HSO3-/SO32- with a large Stokes shift (218 nm), low detection limit (1.87 µM), good selectivity, high energy transfer efficiency (92 %) and wide recognition range (1.87-200 µM), but also identified viscosity with a 26-fold fluorescence enhancement and good linearity. Crucially, QQD was applied to detect HSO3-/SO32- and viscosity in actual water and food samples. In addition, QQD had low toxicity and good photostability for imaging HSO3-/SO32- and viscosity in cells. These results confirmed the feasibility and reliability of QQD for HSO3-/SO32- and viscosity imaging and environmental detection. SIGNIFICANCE: We reported a unique ratiometric probe QQD for detecting HSO3-/SO32- and viscosity based on the quinolinium skeleton. In addition to detecting HSO3-/SO32- and viscosity change in actual water and food samples, QQD could also monitor the variations of HSO3-/SO32- and viscosity in cells, which provided an experimental basis for further exploration of the role of SO2 derivatives and viscosity in biological systems.

Association between changes in the retina with major depressive disorder and sleep quality.

BACKGROUND: Major depressive disorder (MDD) is linked to degenerative and inflammatory processes in the brain. The retina, as well as the brain, originates from the ectoderm. The changes in the retina that occur in concert with MDD and sleep quality are not very clear. This study aimed to detect changes in the retinal nerve fibre layer (RNFL) and macular thickness in Chinese patients in correlation with MDD and sleep quality. METHODS: Thirty-six MDD patients and 187 age- and sex-matched healthy controls were enrolled in this study. All subjects were scanned by spectral-domain optical coherence tomography (OCT) and completed the Stroop Colour-Word Test (SCWT), Pittsburgh Sleep Quality Index, and Patient Health Questionnaire-9. RESULTS: The RNFL and macular thickness were significantly thinner and disc-area enlargement of the right eye and cup-volume enlargement of both eyes were present in MDD patients. Linear regression analysis revealed that the RNFL and macular thickness in patients with MDD were correlated with sleep quality and the severity of depression. SCWT scores were higher following expansion of the cup volume of the right eye (all p < 0.01) and in correlation with larger disc areas. CONCLUSIONS: Serious degrees of depression and insomnia in MDD patients may reflect obvious atrophy of the inner and outer rings of the right macula and enlargement of the optic disc in both eyes. These retinal structural changes might influence the visual cognition of patients with MDD.

Co-delivery of Docetaxel and Resveratrol by liposomes synergistically boosts antitumor efficiency against prostate cancer.

Combination therapy is frequently used in cancer treatments. Delivery of combined anticancer agents loaded in a nanocarrier would be a promising option for combination therapy. Here, we designed PEGylated nano-liposomes for co-delivery Docetaxel (Doc) and Resveratrol (Res) to evaluate antitumor efficiency of the combined drugs in prostate cancer. The average diameter of the liposomes was 99.67 nm with a spheral-like shape. Drug release studies showed that both drugs could synchronously leak from the liposomes in a sustained release behavior. Cellular uptake results demonstrated that liposomes could effectively deliver more cargos into cells than other formulations. Moreover, co-loaded liposomes with Doc/Res in a molar ratio of 1:2 exhibited significantly higher cytotoxicity than a mixed solution containing both drugs on cancer cells. In the study of caspase 3, we found that the combination of Doc and Res could significantly increase the activity of caspase 3 enzyme compared with Doc alone. Animal studies revealed that co-encapsulated Doc/Res in liposomes predominantly inhibited tumor growth in PC3 bearing Balb/c nude mice, as evidenced by a change in cell proliferation and apoptosis parameters. Importantly, little toxicities and prolonged survival time were observed in mice treated with liposome-loaded Doc/Res than control group exposed to liposome-free Doc/Res. These results provided evidence that loading of Doc/Res in a nano-liposome is an efficient delivery formulation for synergistic treating prostate cancer.

Direct and indirect parameter imaging methods for dynamic PET.

The method of reconstructing parametric images from dynamic positron emission tomography (PET) data with the linear Patlak model has been widely used in scientific research and clinical practice. Whether for direct or indirect image reconstruction, researchers have deeply investigated the associated methods and effects. Among the existing methods, the traditional maximum likelihood expectation maximization (MLEM) reconstruction algorithm is fast but produces a substantial amount of noise. If the parameter images obtained by the MLEM algorithm are postfiltered, a large amount of image edge information is lost. Additionally, although the kernel method has a better noise reduction effect, its calculation costs are very high due to the complexity of the algorithm. Therefore, to obtain parametric images with a high signal-to-noise ratio (SNR) and good retention of detailed information, here, we use guided kernel means (GKM) and dynamic PET image information to conduct guided filtering and perform parametric image reconstruction. We apply this method to direct and indirect reconstruction, and through computer simulations, we show that our proposed method has higher identifiability and a greater SNR than conventional direct and indirect reconstruction methods. We also show that our method produces better images with direct than with indirect reconstruction.

Technical Note: A preliminary study of dual-tracer PET image reconstruction guided by FDG and/or MR kernels.

PURPOSE: Clinically, single radiotracer positron emission tomography (PET) imaging is a commonly used examination method; however, since each radioactive tracer reflects the information of only one kind of cell, it easily causes false negatives or false positives in disease diagnosis. Therefore, reasonably combining two or more radiotracers is recommended to improve the accuracy of diagnosis and the sensitivity and specificity of the disease when conditions permit. METHODS: This paper proposes incorporating 18 F-fluorodeoxyglucose (FDG) as a higher-quality PET image to guide the reconstruction of other lower-count 11 C-methionine (MET) PET datasets to compensate for the lower image quality by a popular kernel algorithm. Specifically, the FDG prior is needed to extract kernel features, and these features were used to build a kernel matrix using a k-nearest-neighbor (kNN) search for MET image reconstruction. We created a 2-D brain phantom to validate the proposed method by simulating sinogram data containing Poisson random noise and quantitatively compared the performance of the proposed FDG-guided kernelized expectation maximization (KEM) method with the performance of Gaussian and non-local means (NLM) smoothed maximum likelihood expectation maximization (MLEM), MR-guided KEM, and multi-guided-S KEM algorithms. Mismatch experiments between FDG/MR and MET data were also carried out to investigate the outcomes of possible clinical situations. RESULTS: In the simulation study, the proposed method outperformed the other algorithms by at least 3.11% in the signal-to-noise ratio (SNR) and 0.68% in the contrast recovery coefficient (CRC), and it reduced the mean absolute error (MAE) by 8.07%. Regarding the tumor in the reconstructed image, the proposed method contained more pathological information. Furthermore, the proposed method was still superior to the MR-guided KEM method in the mismatch experiments. CONCLUSIONS: The proposed FDG-guided KEM algorithm can effectively utilize and compensate for the tissue metabolism information obtained from dual-tracer PET to maximize the advantages of PET imaging.

MRI-aided kernel PET image reconstruction method based on texture features.

We investigate the reconstruction of low-count positron emission tomography (PET) projection, which is an important, but challenging, task. Using the texture feature extraction method of radiomics, i.e. the gray-level co-occurrence matrix (GLCM), texture features can be extracted from magnetic resonance imaging images with high-spatial resolution. In this work, we propose a kernel reconstruction method combining autocorrelation texture features derived from the GLCM. The new kernel function includes the correlations of both the intensity and texture features from the prior image. By regarding the GLCM as a discrete approximation of a probability density function, the asymptotically gray-level-invariant autocorrelation texture feature is generated, which can maintain the accuracy of texture features extracted from small image regions by reducing the number of quantized image gray levels. A computer simulation shows that the proposed method can effectively reduce the noise in the reconstructed image compared to the maximum likelihood expectation maximum method and improve the image quality and tumor region accuracy compared to the original kernel method for low-count PET reconstruction. A simulation study on clinical patient images also shows that the proposed method can improve the whole image quality and that the reconstruction of a high-uptake lesion is more accurate than that achieved by the original kernel method.

Association of retinal nerve fiber abnormalities with serum CNTF and cognitive functions in schizophrenia patients.

BACKGROUND: Recent studies have reported reductions in retinal nerve fiber layers (RNFL) in schizophrenia. Ciliary neurotrophic factor (CNTF) has shown protective effects on both the neurogenesis and retina. This study aimed at investigating retinal abnormalities and establishing their correlation with serum CNTF and cognitive impairments in schizophrenic Chinese patients. METHODS: In total, 221 patients diagnosed with schizophrenia and 149 healthy controls were enrolled. Serum CNTF and clinical features of patients were investigated. Cognitive functions were evaluated with Repeatable Battery for the Assessment of Neuropsychology Status (RBANS). RNFL thickness and macular thickness (MT) of both eyes were measured with optical coherence tomography (OCT). T-tests and analysis of covariance were used to compare the variables between the patient and control groups, while multiple linear regression analysis was performed to determine the associations of RNFL thickness, CNTF and cognitive impairments. RESULTS: RNFL was found thinner in patients than in healthy controls (right: 88.18 ± 25.84 µm vs.102.13 ± 14.32 µm, p = 0.001; left: 92.84 ± 13.54 µm vs.103.71 ± 11.94 µm, p < 0.001). CNTF was lower in the schizophrenia group (1755.45 ± 375.73 pg/ml vs. 1909.99 ± 368.08 pg/ml, p = 0.001). Decline in RNFL thickness was found correlated with course of illness and serum CNTF in patients (all p < 0.05). Similarly, cognitive functions such as immediate memory and visuospatial functions were also found correlated with decline in RNFL thickness. CONCLUSION: Decline in RNFL thickness was associated with cognitive impairments of schizophrenia and CNFT serum concentration. The possibility of reduction in RNFL thickness as a biomarker for schizophrenia needs to be further examined.

Design and performance of SIAT aPET: a uniform high-resolution small animal PET scanner using dual-ended readout detectors.

In this work, a small animal PET scanner named SIAT aPET was developed using dual-ended readout depth encoding detectors to simultaneously achieve high spatial resolution and high sensitivity. The scanner consists of four detector rings with 12 detector modules per ring; the ring diameter is 111 mm and the axial field of view (FOV) is 105.6 mm. The images are reconstructed using an ordered subset expectation maximization (OSEM) algorithm. The spatial resolution of the scanner was measured by using a 22Na point source at the center axial FOV with different radial offsets. The sensitivity of the scanner was measured at center axis of the scanner with different axial positions. The count rate performance of the system was evaluated by scanning mouse-sized and rat-sized phantoms. An ultra-micro hot-rods phantom and two mice injected with 18F-NaF and 18F-FDG were scanned on the scanner. An average depth of interaction (DOI) resolution of 1.96 mm, energy resolution of 19.1% and timing resolution of 1.20 ns were obtained for the detector. Average spatial resolutions of 0.82 mm and 1.16 mm were obtained up to a distance of 30 mm radially from the center of the FOV when reconstructing a point source in 1% and 10% warm backgrounds, respectively, using OSEM reconstruction with 16 subsets and 10 iterations. Sensitivities of 16.0% and 11.9% were achieved at center of the scanner for energy windows of 250-750 keV and 350-750 keV respectively. Peak noise equivalent count rates (NECRs) of 324 kcps and 144 kcps were obtained at an activity of 26.4 MBq for the mouse-sized and rat-sized phantoms. Rods of 1.0 mm diameter can be visually resolved from the image of the ultra-micro hot-rods phantom. The capability of the scanner was demonstrated by high quality in-vivo mouse images.

A GPU-accelerated fully 3D OSEM image reconstruction for a high-resolution small animal PET scanner using dual-ended readout detectors.

In this work, a GPU-accelerated fully 3D ordered-subset expectation maximization (OSEM) image reconstruction with point spread function (PSF) modeling was developed for a small animal PET scanner with a long axial field of view (FOV). Dual-ended readout detectors that provided high depth of interaction (DOI) resolution were used for the small animal PET scanner to simultaneously achieve uniform high spatial resolution and high sensitivity. First, we developed a novel sinogram generation method, in which the dimension of the sinogram was determined first and then an event was assigned to a few neighboring sinogram elements by using weights that are inversely proportional to the distance from the measured line of response (LOR) to the LOR of the sinogram elements. System geometric symmetry, precomputation of LOR-driven ray-tracing and texture memory were applied to accelerate the GPU-based reconstruction. We developed a spatially variant PSF model where the PSF parameters were obtained by using point source images measured at 18 positions in the FOV and a spatial invariant PSF model where the PSF parameters were obtained by using only one image measured at the center FOV. The performance of the image reconstruction method was evaluated by using simulated phantom data as well as phantom and in-vivo mouse data acquired on the scanner. The results showed that the proposed reconstruction method provided better spatial resolution, a higher contrast recovery coefficient and lower noise than the OSEM reconstruction and was more than 1000 times faster than the CPU-based reconstruction. The spatially variant PSF model did not result in any spatial resolution improvement compared to the spatial invariant PSF model, and thus, the latter that is much easier to implement in image reconstruction and can be used in a small animal PET scanner using detectors with very high DOI resolution. A whole body 18F-FDG mouse image with high resolution and a high contrast to noise ratio was obtained by using the proposed reconstruction method.

Discovery of new fluorescent thiazole-pyrazoline derivatives as autophagy inducers by inhibiting mTOR activity in A549 human lung cancer cells.

A series of fluorescent thiazole-pyrazoline derivatives was synthesized and their structures were characterized by 1H NMR, 13C NMR, and HRMS. Biological evaluation demonstrated that these compounds could effectively inhibit the growth of human non-small cell lung cancer (NSCLC) A549 cells in a dose- and time-dependent manner in vitro and inhibit tumor growth in vivo. The structure-activity relationship (SAR) of the compounds was analyzed. Further mechanism research revealed they could induce autophagy and cell cycle arrest while had no influence on cell necrosis. Compound 5e inhibited the activity of mTOR via FKBP12, which could be reversed by 3BDO, an mTOR activator and autophagy inhibitor. Compound 5e inhibited growth, promoted autophagy of A549 cells in vivo. Moreover, compound 5e showed good selectivity with no influence on normal vascular endothelial cell growth and the normal chick embryo chorioallantoic membrane (CAM) capillary formation. Therefore, our research provides potential lead compounds for the development of new anticancer drugs against human lung cancer.

Pemetrexed exerts anticancer effects by inducing G0/G1-phase cell cycle arrest and activating the NOXA/Mcl-1 axis in human esophageal squamous cell carcinoma cells.

Esophageal squamous cell carcinoma (ESCC) is a dominant histological subtype of esophageal cancer with notably high incidence and mortality rates. Pemetrexed is a clinical antifolate therapeutic agent with anticancer properties. The present study aimed to understand whether pemetrexed is able to exert anticancer effects on ESCC cells, and to determine the underlying molecular mechanism. ESCC cells were treated with pemetrexed and cell survival was assessed with MTT assays. The cell cycle and apoptosis were evaluated using flow cytometry analysis, and proteins were detected using western blotting. It was demonstrated that pemetrexed inhibited cell survival and induced G0/G1 cell cycle arrest and apoptosis in human ESCC cells. Furthermore, the results demonstrated that the phorbol-12-myristate-13-acetate-induced protein 1/induced myeloid leukemia cell differentiation protein Mcl-1 axis is involved in intrinsic apoptosis induced by pemetrexed. The protein expression of endoplasmic reticulum stress markers inositol-requiring enzyme 1α, binding immunoglobulin protein and CCAAT-enhancer-binding protein homologous protein were upregulated following treatment with pemetrexed. These results suggest that pemetrexed may induce an endoplasmic reticulum stress response while activating intrinsic apoptosis. The present study provided important mechanistic insights into potential cancer treatments involving pemetrexed and enhanced the understanding of human ESCC.

Downregulation of MUTYH contributes to cisplatin­resistance of esophageal squamous cell carcinoma cells by promoting Twist­mediated EMT.

Acquired resistance to cisplatin (CDDP) in esophageal squamous cell carcinoma (ESCC) remains a major challenge in cancer therapy. Although progress has been made in identifying the mechanisms responsible for resistance to CDDP, the underlying mechanisms of resistance in ESCC are still not entirely understood. In the present study, a CDDP­resistant ESCC cell line EC109/CDDP was established by culturing parental EC109 cells in increasing concentrations of CDDP, and it was demonstrated that MutY homolog (MUTYH), a critical base excision repair gene, was significantly downregulated in the resistant EC109/CDDP cells compared with that noted in the parental cells. Ectopic expression of MUTYH by transient transfection of pcDNA3.1­MUTYH plasmid significantly enhanced the CDDP­mediated inhibitory effect on resistant cell proliferation and induction of apoptosis, while silencing of MUTYH by transiently transfecting MUTYH­targeted siRNA in parental cells led to decreased sensitivity to CDDP as demonstrated by MTT assay, suggesting the crucial involvement of MUTYH in CDDP resistance. Further experiments demonstrated that the CDDP­resistant cells went through epithelial­mesenchymal transition (EMT) driven by its master regulator Twist, and MUTYH overexpression significantly reduced the Twist expression level and reversed the phenotype of EMT as detected by western blot analysis and RT­qPCR assays, suggesting that downregulation of MUTYH contributed to the Twist­mediated EMT. Moreover, it was observed that the effect of MUTYH on Twist was also associated with its degradation in addition to transcription. MUTYH acted as a positive regulator of reactive oxygen species (ROS) that showed a low level in resistant cells via flow cytometry assay, as demonstrated by increased ROS production in response to MUTYH overexpression. Reduced ROS by using N­acetylcysteine led to a decrease in proteasome activity and sequentially inhibited the degradation of Twist. In conclusion, the present data demonstrated that EMT activation mediated by MUTYH downregulation, by both enhancing Twist transcription and blocking its degradation, is one of the mechanisms for acquisition of CDDP resistance in ESCC.

Secretory production of antimicrobial peptides in Escherichia coli using the catalytic domain of a cellulase as fusion partner.

Antimicrobial peptides (AMPs) are small molecules which serve as essential components of the innate immune system in various organisms. AMPs possess a broad spectrum of antimicrobial activities. However, the scaled production of such peptides in Escherichia coli faces many difficulties because of their small size and toxicity to the host. Here, we described a new fusion strategy to extracellularly produce significant amounts of these antimicrobial peptides in recombinant E. coli at significant amount. Employing the catalytic domain of a cellulase (Cel-CD) from Bacillus subtilis KSM-64 as the fusion partner, five recombinant antimicrobial peptides were confirmed to accumulate in the culture medium at concentrations ranging from 184 mg/L to 297 mg/L. The radical diffusion experiment demonstrated that the released model antimicrobial peptide, bombinin, had antibacterial activities against both E. coli and Staphylococcus aureus. This strategy will be suitable for the production of antimicrobial peptides and other toxicity proteins.