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Glioblastoma is the most common glioma with high mortality and poor prognosis. Radiation resistance is one of the large challenges in the treatment of glioma. The study aimed to identify whether DNA polymerase ζ affects glioma cell radiosensitivity. The mRNA and protein levels of REV3L and REV7 were examined using quantitative real-time PCR and western blot. After REV3L and REV7 knockdown in a GBM cell line (A172), we assessed cell viability, colonies, apoptosis, and immune escape. The underlying mechanisms were evaluated using western blot and were confirmed using rescue experiments. The results showed that REV3L and REV7 levels were increased in glioma and related to poor survival. γ-ray treatment inhibited cell viability, survival fraction, and immune escape, and induced apoptosis of glioma cells from a GBM cell line, whereas knockdown of REV3L or REV7 enhanced these effects. Mechanically, silencing of REV3L or REV7 inactivated the PI3K/AKT/mTOR pathway. IGF-1 treatment abrogated the effects on cell viability, colonies, and apoptosis induced by REV3L or REV7 knocking down. Taken together, silencing of REV3L and REV7 inhibited radiation resistance via inactivating the PI3K/AKT/mTOR pathway, suggesting that targeting DNA polymerase ζ may be a new strategy to reduce the radiotherapy resistance of glioma.

In mammals, oxidative stress-induced apoptosis of granulosa cells is one of the major causes of follicular atresia, affecting ovarian physiological function. Protegrin-1 (PG-1) is an antimicrobial peptide with effective antimicrobial activity, immunomodulatory function, and porcine growth-promoting effects. PG-1 has been detected in porcine ovaries follicles. This study aimed to investigate the effect of PG-1 on oxidative stress-induced apoptosis of porcine ovarian granulosa cells and the underlying molecular mechanism. Granulosa cells were obtained from porcine follicles and treated with H2O2 to establish the oxidative stress model, and then treated with or without PG-1 (10 μg/mL). PG-1 significantly suppressed H2O2-induced apoptosis in granulosa cells after 24 h of treatment. Furthermore, these results revealed that PG-1 increased the mRNA and protein expression of anti-apoptotic B cell lymphoma/leukemia 2 (BCL2) and the BCL2/Bcl-2-associated X protein (BAX) ratio while decreasing the expression of pro-apoptotic BAX and active caspase-3. Using western blot analysis, it was found that PG-1 decreased the phosphorylation of RNA-like endoplasmic reticulum kinase (PERK) and the α-subunit of eukaryotic initiation factor 2 (eIF2α) as well as the protein expression level of CCAAT enhancer-binding protein homologous protein (CHOP), all of which were increased by H2O2. Moreover, inhibitors against PERK and phospho-eIF2ɑ both suppressed the H2O2-induced granulosa cells apoptosis and enhanced the anti-apoptosis effect of PG-1. Taken together, our findings demonstrated that PG-1 inhibited porcine ovarian granulosa cell apoptosis from oxidative stress via the PERK/eIF2α/CHOP signaling pathway in vitro, which suggests the novel regulatory function of the antimicrobial peptide in the ovary.

The heliostat field of tower solar thermal power station accounts for 40%–50% of the total cost, and influences the concentrating efficiency. Accordingly, it is necessary to optimize the layout of the heliostat field. Based on the optical efficiency model, an improved Gray Wolf Optimization (GWO) algorithm is proposed to optimize the field parameters of the heliostats, improve the convergence factor and weight updating formula, and effectively avoid the local optimal problem. Then SolarPILOT software is used to simulate the heliostat field distribution. In order to reduce the shadow and shielding efficiency loss, improve the land utilization rate and atmospheric attenuation efficiency, the heliostat field is initialized by radial staggered arrangement, which is easy to be optimized. By using the optical efficiency model, the program of heliostat field optimization algorithm is developed, and a Delingha tower power station is used to verify the algorithm. After the improved GWO algorithm optimizing the heliostat field, the optical or concentrating efficiency of the heliostat field is increased by 8.2% compared with the GWO algorithm. The improved GWO algorithm reduces the heliostat number by 3.4% compared with the Gray Wolf algorithm, and that reducing the cost of the heliostat field.

BackgroundExosomes, internal proteins, lipids, and nucleic acids coated by phospholipid bilayer membranes, are one type of small extracellular vesicles, which can mediate cell-cell communication. In recent years, exosomes have gained considerable scientific interest due to their widely applied prospect in the diagnosis and therapeutics of human and animal diseases. In this study, we describe for the first time a feasible method designed to isolate and characterize exosomes from feline plasma, urine and adipose-derived mesenchymal stem cells.ResultsExosomes from feline plasma, urine and adipose-derived mesenchymal stem cells were successfully isolated by differential centrifugation. Quantification and sizing of exosomes were assessed by transmission electron microscopy, flow nano analysis and western blotting. Detected particles showed the normal size (30–100 nm) and morphology described for exosomes, as well as presence of the transmembrane protein (TSG101, CD9, CD63, and CD81) known as exosomal marker.ConclusionsThe results suggest that differential centrifugation is a feasible method for isolation of exosomes from different types of feline samples. Moreover, these exosomes can be used to further diagnosis and therapeutics in veterinary pre-clinical and clinical studies.

Background Mesenchymal stem cells (MSCs) have generated a great amount of interest in recent years as a novel therapeutic application for improving the quality of pet life and helping them free from painful conditions and diseases. It has now become critical to address the challenges related to the safety and efficacy of MSCs expanded in vitro. In this study, we establish a standardized process for manufacture of canine adipose-derived MSCs (AD-MSCs), including tissue sourcing, cell isolation and culture, cryopreservation, thawing and expansion, quality control and testing, and evaluate the safety and efficacy of those cells for clinical applications. Results After expansion, the viability of AD-MSCs manufactured under our standardized process was above 90 %. Expression of surface markers and differentiation potential was consistent with ISCT standards. Sterility, mycoplasma, and endotoxin tests were consistently negative. AD-MSCs presented normal karyotype, and did not form in vivo tumors. No adverse events were noted in the case treated with intravenously AD-MSCs. Conclusions Herein we demonstrated the establishment of a feasible bioprocess for manufacturing and banking canine AD-MSCs for veterinary clinical use.

Intrahepatic cholangiocarcinoma (ICC) is a malignant liver tumor derived from the epithelial cells of the second- or higher-order branch of the bile duct, with the features of insidious early symptoms, high degree of malignancy, rapid disease progression, lymph node metastasis, intrahepatic metastasis, and vascular invasion, and such patients tend to have poor prognosis. Early diagnosis can improve the radical resection rate and prognosis of ICC. At present, the preoperative diagnosis of ICC mainly relies on imaging technology, laboratory examination, and pathological examination. In addition, some new diagnostic methods have also been used in the diagnosis of ICC in recent years, but histopathological examination remains the only method for the diagnosis of ICC. This article reviews the latest research advances in the diagnosis of ICC.

Foot-and-mouth disease virus (FMDV) is one of the most important animal pathogens in the world. FMDV naturally infects swine, cattle, and other cloven-hoofed animals. FMD is not adequately controlled by vaccination. An alternative strategy is to develop swine that are genetically resistant to infection. Here, we generated FMDV-specific shRNA transgenic cells targeting either nonstructural protein 2B or polymerase 3D of FMDV. The shRNA-positive transgenic cells displayed significantly lower viral production than that of the control cells after infection with FMDV (P +, CD4+, CD8+, CD21+, and CD172+ cells, and the production of IFN-γ were analyzed, there were no significant differences observed between the TGCS and NS or Non-TGCS, suggesting that the FMDV resistance may be mainly derived from the RNAi-based antiviral pathway. Our work provides a foundation for a breeding approach to preventing infectious disease in swine.

Introduction Targeted therapies are based on specific gene alterations. Various specimen types have been used to determine gene alterations, however, no systemic comparisons have yet been made. Herein, we assessed alterations in selected cancer‐associated genes across varying sample sites in lung cancer patients. Materials and Methods Targeted deep sequencing for 48 tumor‐related genes was applied to 153 samples from 55 lung cancer patients obtained from six sources: Formalin‐fixed paraffin‐embedded (FFPE) tumor tissues, pleural effusion supernatant (PES) and pleural effusion cell sediments (PEC), white blood cells (WBCs), oral epithelial cells (OECs), and plasma. Results Mutations were detected in 96% (53/55) of the patients and in 83% (40/48) of the selected genes. Each sample type exhibited a characteristic mutational pattern. As anticipated, TP53 was the most affected sequence (54.5% patients), however this was followed by NOTCH1 (36%, across all sample types). EGFR was altered in patient samples at a frequency of 32.7% and KRAS 10.9%. This high EGFR/ low KRAS frequency is in accordance with other TCGA cohorts of Asian origin but differs from the Caucasian population where KRAS is the more dominant mutation. Additionally, 66% (31/47) of PEC samples had copy number variants (CNVs) in at least one gene. Unlike the concurrent loss and gain in most genes, herein NOTCH1 loss was identified in 21% patients, with no gain observed. Based on the relative prevalence of mutations and CNVs, we divided lung cancer patients into SNV‐dominated, CNV‐dominated, and codominated groups. Conclusions Our results confirm previous reports that EGFR mutations are more prevalent than KRAS in Chinese lung cancer patients. NOTCH1 gene alterations are more common than previously reported and reveals a role of NOTCH1 modifications in tumor metastasis. Furthermore, genetic material from malignant pleural effusion cell sediments may be a noninvasive manner to identify CNV and participate in treatment decisions., Targeted deep sequencing for 48 tumor‐related genes was applied to 153 samples from 55 lung cancer patients obtained from six sources. NOTCH1 exhibited mutational changes in 70 % plasmas, much higher than in tumor tissue (20%), highlighting the extraordinary usefulness of NOTCH1 in liquid biopsy.

Long-term maintenance of chicken primordial germ cells (PGCs) in vitro has tremendous potential for transgenic chicken production. Feeder cells are essential for the establishment and culture of chicken PGCs in vitro. Buffalo rat liver (BRL) cells are the most commonly used feeder cells for PGCs culture; however, this feeder layers from other animal species usually cause immunogenic contaminations, compromising the potential of PGCs in applications. Therefore, we tested chicken source mensenchymal stem cell (MSCs) derived from bone marrow as feeder cells to further improve PGC culture conditions. MSCs derived from chicken bone marrow have a powerful capacity to proliferate and secrete cytokines. We found chicken primordial germ cells derived from circulating blood (cPGCs) and gonads (gPGCs) can be maintained and proliferated with MSCs feeder layer cells. PGCs co-cultured on MSCs feeder retained their pluripotency, expressed PGCs specific genes and stemness markers, and maintained undifferentiated state. Our study indicated that the xeno-free MSCs-feeders culture system is a good candidate for growth and expansion of PGCs as the stepping stone for transgenic chicken research.

Additional file 1. Western blot instructions and original images.

Objective. The aim of this study was to assess the efficacy of canine umbilical cord mesenchymal stem cells (UC-MSCs) on the treatment of knee osteoarthritis in dogs. Methods. Eight dogs were evenly assigned to two groups. The canine model of knee osteoarthritis was established by surgical manipulation of knee articular cartilage on these eight dogs. UC-MSCs were isolated from umbilical cord Wharton’s jelly by 0.1% type collagenase I and identified by immunofluorescence staining and adipogenic and osteogenic differentiation in vitro. A suspension of allogeneic UC-MSCs (1 × 106) and an equal amount of physiological saline was injected into the cavitas articularis in the treated and untreated control groups, respectively, on days 1 and 3 posttreatment. The structure of the canine knee joint was observed by magnetic resonance imaging (MRI), B-mode ultrasonography, and X-ray imaging at the 3rd, 7th, 14th, and 28th days after treatment. Concurrently, the levels of IL-6, IL-7, and TNF-α in the blood of the examined dogs were measured. Moreover, the recovery of cartilage and patella surface in the treated group and untreated group was compared using a scanning electron microscope (SEM) after a 35-day treatment. Results. Results revealed that the isolated cells were UC-MSCs, because they were positive for CD44 and negative for CD34 surface markers, and the cells were differentiated into adipocytes and osteoblasts. Imaging technology showed that as treatment time increased, the high signal in the MRI T2-weighted images decreased, the echo-free space in B ultrasonography images disappeared basically, and the continuous linear hypoechoic region at the trochlear sulcus thickened. On X-ray images, the serrate defect at the ventral cortex of the patella improved, and the low-density gap of the ventral patella and trochlear crest gradually increased in the treated group. On the contrary, the high signal in the MRI T2-weighted images and the echo-free space in B ultrasonography images still increased after a 14-day treatment in the untreated control group, and the linear hypoechoic region was discontinuous. On the X-ray images, there was no improvement in the serrate defect of the ventral cortex of the patella. Results for inflammatory factors showed that the blood levels of IL-6, IL-7, and TNF-α of the untreated control group were significantly higher than those of the treated group (P<0.05) 7–14 days posttreatment. The result of SEM showed that the cartilage neogenesis in the treated group had visible neonatal tissue and more irregular arrangement of new tissue fibers than that of the untreated control group. Furthermore, more vacuoles but without collagen fibers were observed in the cartilage of the untreated control group, and the thickness of the neogenetic cartilage in the treated group (65.13 ± 5.29, 65.30 ± 5.83) and the untreated control group (34.27 ± 5.42) showed a significant difference (P<0.01). Conclusion. Significantly higher improvement in cartilage neogenesis and recovery was observed in the treated group compared to the untreated control group. The joint fluid and the inflammatory response in the treated group decreased. Moreover, improved recovery in the neogenetic cartilage, damaged skin fascia, and muscle tissue around the joints was more significant in the treated group than in the untreated control group. In conclusion, canine UC-MSCs promote the repair of cartilage and patella injury in osteoarthritis, improve the healing of the surrounding tissues, and reduce the inflammatory response.

Larger workpiece's size visual detection needs to combine motion control and image mosaic to ensure accuracy. Usually the complex flat parts have many test items therefore require tedious path planning which affect detection efficiency and accuracy. This paper researched and developed a intelligent path generation method using image interaction. The method is based on DXF drawing file analysis, combined with smart interaction, image restructure and sample point automatic positioning to achieve plane size detection automatic processing and path planning. Application results shown that this method surely reduced difficulty of complex workpiece's detection process planning and increased detection system's efficiency and accuracy. Furthermore, its easily extended application to 3D or other measure system also improved intelligent level.

Highlights • We generated transgenic pig expressing PRRSV-specific siRNA. • Stability of siRNA expression was proved in two generations. • Type I interferon was not elicited by the expression of siRNA in vivo. • We proved that transgenic pigs showed substantially decreased virus load in serum after PRRSV infection., Porcine reproductive and respiratory syndrome (PRRS) is an economically devastating viral disease causing heavy losses to the swine industry worldwide. Many studies have shown that transient delivery of small interfering RNA (siRNA) or adenovirus-mediated RNA interfere (RNAi) could potentially inhibit porcine reproductive and respiratory syndrome virus (PRRSV) replication in vivo and in vitro. Here, we applied RNAi to produce transgenic (TG) pigs that constitutively expressed PRRSV-specific siRNA derived from small hairpin RNA (shRNA). First, we evaluated siRNA expression in the founding and F1 generation pigs and confirmed stable transmission. Then, we detected the expression of IFN-β and protein kinase R (PKR) and found no difference among TG, non-transgenic (NTG), and wild-type pigs. Lastly, the F1 generation pigs, including TG and NTG piglets, were challenged with 3 × 104.5 TCID50 of JXA1, a highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV). Our results showed that the in vivo siRNA expression substantially reduced the serum HP-PRRSV titers and increased survival time by 3 days when TG pigs were compared with the NTG controls. These data suggested that RNAi-based genetic modification might be used to breed viral-resistant livestock with stable siRNA expression with no complications of siRNA toxicity.

Mesenchymal stem cells (MSCs) are the most promising seed cells for cell therapy. Comparing the biological and transcriptome gene characteristics of MSCs from different sources provides an important basis for the screening of clinically used cells. The main purpose of this experiment was to establish methods for the isolation and culture of MSCs from five different canine sources, including adipose tissue, bone marrow, umbilical cord, amniotic membrane, and placenta, and compare biological and transcriptome characteristics of MSCs, in order to provide a basis for the clinical application of canine MSCs. MSCs were isolated from Chinese pastoral dogs, and the following experiments were performed: (1) the third, sixth, and ninth generations of cells were counted, respectively, and a growth curve was plotted to calculate the MSC population doubling time, (2) the expression of CD34 and CD44 surface markers was studied by immunofluorescence, (3) the third generation of cells were used for osteogenetic and adipogenic differentiation experiments, and (4) MSC transcriptome profiles were performed using RNA sequencing. All of the five types of MSCs showed fibroblast-like adherent growth. The cell surface expressed CD44 instead of CD34, the third-generation MSCs had the highest proliferative activity. The average population doubling time of adipose mesenchymal stem cells (AD-MSCs), placenta mesenchymal stem cells (P-MSCs), bone marrow mesenchymal stem cells (BM-MSCs), umbilical cord mesenchymal stem cells (UC-MSCs), and amniotic mesenchymal stem cells (AM-MSCs) were 15.8 h, 21.2 h, 26.2 h, 35 h, and 41.9 h, respectively. All five types of MSCs could be induced to differentiate into adipocytes and osteoblasts in vitro, with lipid droplets appearing after 8 days and bone formation occurring 5 days after AD-MSC induction. However, the multilineage differentiation for the remaining of MSCs was longer compared to that of the AD-MSCs. The MSC transcriptome profiles showed that AD-MSC and BM-MSCs had the highest homology, while P-MSCs were significantly different compared to the other four types of MSCs. All the isolated MSCs had the main biological characteristics of MSCs. AD-MSCs had the shortest time for proliferation, adipogenesis, and osteogenic differentiation.

At present, the variation of gas content resulted from the surface coalbed methane (CBM) development has not been reported at home. It is introduced that adopting the CBM-SIM software and combining with the key oil and gas special research task, the productivity prediction and reservoir simulation analysis of the impact and variation of the CBM in the surface CBM development process are conducted respectively in the Sihe north block and Chengzhuang block of the Jincheng mining area. Combined with the measured results of the gas content in the inspect well on the drainage effect after 5 years on the surface CBM development in the Sihe north block, the variation rules of the CBM in the surface coalbed methane development process are discussed and researched.

Porcine reproductive and respiratory syndrome (PRRS) is one of the most devastating diseases for the pig industry. Our goal was to identify microRNAs involved in the host immune response to PRRS. We generated microRNA expression profiles of lung tissues from Tongcheng or Landrace pigs infected with a highly pathogenic PRRS virus (PRRSV) at 3, 5, 7 dpi (day post infection) and control individuals from these two breeds. Our data showed that 278 known and 294 novel microRNAs were expressed in these combined microRNA transcriptomes. Compared with control individuals, almost half of the known microRNAs (116 in Tongcheng and 153 in Landrace) showed significantly differential expression (DEmiRNAs) at least once. The numbers of down-regulated DEmiRNAs were larger than the corresponding number of up-regulated DEmiRNAs in both breeds. Interestingly, miR-2320-5p, which was predicted to bind to conserved sequences of the PRRSV genome, was down-regulated significantly at 3 dpi after PRRSV infection in both breeds. In addition, PRRSV infection induced a significant increase of microRNA editing level in both breeds. Our results provide novel insight into the role of microRNA in response to PRRSV infection in vivo, which will aid the research for developing novel therapies against PRRSV.

An improved particle swarm optimization algorithm with opposition mutation (OMPSO) is presented and applied to choose satisfied parameter of grinding process. The proposed OMPSO employs opposition-based learning algorithms, which can accelerate the learning and searching process in soft computing. The mutation threshold of OMPSO is adapted to the evolution information of the global best, which is very useful to keep the global search ability and fast convergence of the optimization algorithm. The OMPSO has the same tuning parameters as standard particle swarm optimization algorithm (PSO) and is easily implemented in practice. At last, OMPSO is applied to several benchmark problems. Results of numerical examples indicate that the proposed algorithm is an effective method for grinding process optimization problem.

The robust output feedback control problem is studied for a class of Markovian jump time-delay systems with uncertain switching probabilities. Sufficent conditions on the stochastic stability with prescribed Hinfin tracking performance are proposed using a slack weighting matrix method. The conditions and the optimal design of the controller are formulated as a set of direct linear matrix inequalities(LMIs). Further more, the tolerable bound of the perturbation of the mode transition rates is also presented.

Based on Takagi-Sugeno (T-S) fuzzy models, the energy-to-peak filtering problem is studied for a class of nonlinear stochastic neutral systems with delays. Sufficient conditions on the stochastic stability with prescribed L 2 -L ? noise attenuation level are proposed. The conditions and the optimal design of the controller are formulated as a set of direct linear matrix inequalities (LMIs). The effectiveness of the proposed method is illustrated by a simulation example.