Application and prospect of gene chip in genetic breeding of livestock and poultry.

Gene chip is a high-throughput technique for detecting specific DNA sequences by DNA or DNA-RNA complementary hybridization, among which SNP genotyping chips have been widely employed in the animal genetics and breeding, and have made great achievements in cattle (Bos taurus), pigs (Sus scrofa), sheep (Caprinae), chickens (Gallus gallus) and other livestock. However, genomic selection applied in production merely uses genomic information and cannot fully explain the molecular mechanism of complex traits genetics, which limits the accuracy of genomic selection. With the continuous progresses in epigenetic research, the development of commercial methylation chips and the application of the epigenome-wide association study (EWAS), DNA methylation has been extensively used to draw the causal connections between genetics and phenotypes. In the future, it is hopefully expected to develop methylation chips customized for livestock and poultry and explore methylation sites significantly related to economic traits of livestock and poultry through EWAS thereby extending the understanding of causal variation of complex traits. Combining methylation chips and SNP chips, we can capture the epigenomic and genomic information of livestock and poultry, interpret genetic variation more precisely, improve the accuracy of genome selection, and promote the fine evolution of molecular genetic breeding of livestock and poultry. In this review, we summarize the application of SNP chips and depict the prospects of the application of methylation chips in livestock and poultry. This review will provide valuable insights for further application of gene chips in farm animal breeding.

Correlation between serum trimethylamine-N-oxide concentration and protein energy wasting in patients on maintenance hemodialysis.

OBJECTIVES: Chronic kidney disease (CKD) is a serious health problem that is associated with several systemic changes, including protein energy wasting (PEW). However, the exact mechanism of PEW in CKD remains unclear. As one of the important intestinal flora metabolites and uremic toxins, trimethylamine-N-oxide (TMAO) is involved in CKD-associated mortality, which might play a role in the development of PEW in CKD patients especially in patients on maintenance hemodialysis (MHD). However, this possibility has not been investigated. METHODS: PEW was diagnosed in a group of CKD patients on MHD according to the criteria of the International Society of Renal Nutrition and Metabolism. Serum TMAO concentration was assessed by high-performance liquid chromatography and mass spectrometry. The association between TMAO concentration and PEW was assessed using linear regression and logistic analysis after adjustment for confounding factors, including basic characteristics, comorbidities, and laboratory findings. RESULTS: The circulating TMAO level was higher in the MHD patients than in control (healthy) individuals (5653.76 ± 2853.51 vs. 254.92 ± 197.88 ng/mL, p < 0.001). Further, after the MHD patients were screened for PEW, those with PEW were found to have significantly higher serum TMAO levels than those without PEW (6760.9 vs. 4016.1 ng/mL, p < 0.001). Further, the serum TMAO concentration exhibited a significant negative correlation with body mass index (BMI) and dietary protein intake. In the logistic regression analysis, after adjustment for confounding factors, the serum TMAO concentration was still significantly correlated with PEW occurrence. CONCLUSIONS: The circulating TMAO level is significantly correlated with the prevalence of PEW in MHD patients. TMAO might be a potential target in the prevention and treatment of PEW in CKD especially ESRD.

Testing Two Somatic Cell Count Cutoff Values for Bovine Subclinical Mastitis Detection Based on Milk Microbiota and Peripheral Blood Leukocyte Transcriptome Profile.

Somatic cell count (SCC) is an important indicator of the health state of bovine udders. However, the exact cut-off value used for differentiating the cows with healthy quarters from the cows with subclinical mastitis remains controversial. Here, we collected composite milk (milk from four udder quarters) and peripheral blood samples from individual cows in two different dairy farms and used 16S rRNA gene sequencing combined with RNA-seq to explore the differences in the milk microbial composition and transcriptome of cows with three different SCC levels (LSCC: <100,000 cells/mL, MSCC: 100,000−200,000 cells/mL, HSCC: >200,000 cells/mL). Results showed that the milk microbial profiles and gene expression profiles of samples derived from cows in the MSCC group were indeed relatively easily discriminated from those from cows in the LSCC group. Discriminative analysis also uncovered some differentially abundant microbiota at the genus level, such as Bifidobacterium and Lachnospiraceae_AC2044_group, which were more abundant in milk samples from cows with SCC below 100,000 cells/mL. As for the transcriptome profiling, 79 differentially expressed genes (DEGs) were found to have the same direction of regulation in two sites, and functional analyses also showed that biological processes involved in inflammatory responses were more active in MSCC and HSCC cows. Overall, these results showed a similarity between the milk microbiota and gene expression profiles of MSCC and HSCC cows, which presented further evidence that 100,000 cells/ml is a more optimal cut-off value than 200,000 cells/mL for intramammary infection detection at the cow level.

An emerging role of radiation­induced exosomes in hepatocellular carcinoma progression and radioresistance (Review).

The incidence rates of hepatocellular carcinoma (HCC) worldwide are increasing, and the role of radiotherapy is currently under discussion. Radioresistance is one of the most important challenges in the therapy of HCC compared with other local advanced, recurrent and metastatic cancers. The mechanisms of radioresistance are complex and remain to be fully understood; however, extracellular vesicles have been investigated in recent studies. Exosomes, which are 40­ to 150­nm extracellular vesicles released by cancer cells, contain multiple pathogenic components, including proteins, nucleic acids and lipids, and play critical functions in cancer progression. Emerging data indicate a diagnosis potential for exosomes in HCC, since radiation­derived exosomes promote radioresistance. Radiation­based therapy alters the contents and components of exosomes, suggesting that exosomes and their components may serve as prognostic and predictive biomarkers to monitor radiation response. Therefore, understanding the roles and mechanisms of exosomes in HCC progression and radiation response during HCC therapy may increase our knowledge concerning the roles of exosomes in radioresistance, and may lead to novel approaches for HCC prognosis and treatment. The current review summarizes recent studies on exosome involvement in HCC and the molecular changes in exosome components during HCC progression. It also discusses the functions of exosomes in HCC therapy, and highlights the importance of exosomes in HCC progression and resistance for the development of novel therapies.

The Application of Peptides in Glioma: A Novel Tool for Therapy.

BACKGROUND: Glioma is the most aggressive and lethal tumor of the central nervous system. Due to the cellular heterogeneity, the invasiveness, and blood-brain barrier (BBB), current therapeutic approaches, such as chemotherapy and radiotherapy, are poorly to obtain great antitumor efficacy. However, peptides, a novel type of therapeutic agent, displayed excellent ability in the tumor, which becomes a new molecule for glioma treatment. METHODS: We review the current knowledge on peptides for the treatment of glioma through a PubMed-based literature search. RESULTS: In the treatment of glioma, peptides can be used as (i) decoration on the surface of the delivery system, facilitating the distribution and accumulation of the anti-tumor drug in target site; (ii) anti-tumor active molecules, inhibiting the growth of glioma and reducing solid tumor volume; (iii) immune-stimulating factor, and it activating immune cells in the tumor microenvironment or recruiting immune cells to the tumor for breaking out the immunosuppression by glioma cells. CONCLUSION: The application of peptides has revolutionized the treatment of glioma, which based on targeting, penetrating, anti-tumor activities and immunostimulatory. Moreover, better outcomes have been discovered in combining different kinds of peptides rather than a single one. Until now, more and more preclinical studies have been developed with multifarious peptides, which shows promising results in vitro or vivo with the model of glioma.

Quantitatively Recognizing Stimuli Intensity of Primary Taste Based on Surface Electromyography.

A novel approach to quantitatively recognize the intensity of primary taste stimuli was explored based on surface electromyography (sEMG). We captured sEMG samples under stimuli of primary taste with different intensities and quantitatively recognized preprocessed samples with Support Vector Machine (SVM). The feasibility of quantitatively recognizing the intensity of Sour, Bitter, and Salty was verified. The sEMG signals were acquired under the stimuli of citric acid (aq), sucrose (aq), magnesium chloride (aq), sodium chloride (aq), and sodium glutamate (aq) with different concentrations, for five types of primary tastes: Sour, Sweet, Bitter, Salty, and Umami, whose order was fixed in this article. The acquired signals were processed with a method called Quadratic Variation Reduction to remove baseline wandering, and an adaptive notch to remove power frequency interference. After extracting 330 features for each sample, an SVM regressor with five-fold cross-validation was performed and the model reached R2 scores of 0.7277, 0.1963, 0.7450, 0.7642, and 0.5055 for five types of primary tastes, respectively, which manifested the feasibilities of the quantitative recognitions of Sour, Bitter, and Salty. To explore the facial responses to taste stimuli, we summarized and compared the muscle activities under stimuli of different taste types and taste intensities. To further simplify the model, we explored the impact of feature dimensionalities and optimized the feature combination for each taste in a channel-wise manner, and the feature dimensionality was reduced from 330 to 210, 120, 210, 260, 170 for five types of primary tastes, respectively. Lastly, we analyzed the model performance on multiple subjects and the relation between the model's performance and the number of experiment subjects. This study can provide references for further research and applications on taste stimuli recognition with sEMG.

Autophagy inhibition enhances anti-pituitary adenoma effect of tetrandrine.

Pituitary adenoma (PA) is a benign intracranial neoplasm originated from pituitary gland. Surgery is the first-line therapy for most of PAs, but lead to unsatisfactory prognosis in some cases. Tetrandrine (Tet) has anticancer effect on some cancers. However, growth inhibition effect on PA is unknown. To elucidate the inhibitory effect of Tet on the growth of PA and its potential mechanisms, we validated the in vitro and in vivo anti-PA effect of Tet and illustrated the cellular and molecular alterations by confocal microscopy observation, flow cytometry, and RNA interference. Tet inhibited PA cell growth in vitro and tumor progression in vivo. Tet induced autophagy and apoptosis in a dose-dependent manner. Low dosage (1.25 µM) of Tet induced PA cell autophagy by down-regulation of MAPK/STAT3 signal. While, higher dosage (5.0 µM) of Tet partially induced PA cell death through caspase-dependent apoptosis. Autophagy inhibitors enhanced Tet-induced caspase activity and apoptotic cell death. These findings demonstrated that Tet has anti-PA effect by inducing autophagy and apoptosis through MAPK/STAT3 signaling pathway attenuation and autophagy inhibition might enhance its anti-PA effect, indicating that Tet (or combined with autophagy inhibitor) is a potential therapeutic regimen for PAs.

A retrospective study of hypofractionated radiotherapy for keloids in 100 cases.

At present, the consensus on the best treatment for keloids is the combination of clinical and surgical therapies, if necessary, associated with adjuvant radiotherapy like brachytherapy. Whereas, the uniform scheme of radiotherapy in keloids is unclear. Here, we conducting a retrospective analysis to assess the efficacy and safety of a specific treatment regimen (20 Gy in 5 fractions) in keloid patients. We retrospectively analysed the medical records of keloid patients receiving auxiliary postoperative radiotherapy (PORT) treatment from 2009 to 2019. The patients were treated with the hypofractionation method of 20 Gy in 5 fractions. We compared the local control rate and complications, using the chi-square test and logistic regression analyses. After screening, we identified 100 keloid patients in this study, with a median follow-up of 59 months. In this study, the overall local control rate of keloid lesions was 84.8%. After multivariate analyses (primary keloid or not, family history, interval from surgery to irradiation and site), our research showed that primary keloid, site and interval from surgery to irradiation were significantly related to recurrence. Acute radiation injury and late radiation injury accounted for 3% (erythema) and 1% (skin sclerosis) of the total cases, respectively. Our results indicate that a postoperative hypofractionation with radiation dose of 20 Gy in 5 fractions may be effective, easy to accept and safe for keloid patients.

Targeting c-kit inhibits gefitinib resistant NSCLC cell growth and invasion through attenuations of stemness, EMT and acquired resistance.

EGFR tyrosine kinase inhibitors (TKIs) are the first-line drugs for NSCLC. But, the acquired resistance limited their efficacy, so that the patients deteriorate eventually. Therefore, it is necessary to clarify the mechanism of the acquired resistance and overcome it for effective NSCLC therapy. In this experimental study, a stable gefitinib resistant lung adenocarcinoma cell line (PC9/GR) infected with shRNA-c-kit-homo-1386 were established; c-kit siRNA and c-kit inhibitors were used to block c-kit signaling; the acquired resistance of PC9/GR cells and the effects of c-kit siRNA and c-kit inhibitors on the growth and invasion of PC9/GR cells were investigated with CCK-8 assay, colony formation and cell invasion assays in vitro; the tumor growth inhibition effects of c-kit inhibitors on PC9/GR cell generated tumors were tested in vivo; the mechanisms involved in the acquired resistance reverse, growth and invasion inhibition effects of c-kit siRNA and c-kit inhibitors on PC9/GR cells were evaluated with qRT-PCR, Western blot and immunohistochemistry staining. The proliferation, colony formation, and invasion of PC9/GR cells were decreased by c-kit siRNA and inhibitors in vitro significantly; c-kit inhibitors suppressed the tumor growth of PC9/GR cell generated tumors in vivo. In the stable shRNA-c-kit transfected PC9/GR cells, the protein expressions of c-kit signaling and stemness phenotype related proteins, including ALDH1A1, Oct4, Sox2 and ABCG2 were decreased, and EMT phenotype related protein expressions including vimentin, N-cadherin, and Slug, were downregulated and with upregulation of E-cadherin; c-kit inhibitors reduced stemness phenotype related protein expressions, downregulated EMT phenotype related protein expressions including vimentin, N-cadherin, and Slug, with upregulation of E-cadherin, and the stemness related protein expressions of c-kit, ALDH1A1, ABCG2 and EMT-related proteins of vimentin and slug were decreased in the imatinib treated tumor tissues. The findings of this study indicated that c-kit signaling mediated the acquired gefitinib resistance, cell growth, invasion, stemness and EMT phenotype of PC9/GR cells. Targeting c-kit signaling with c-kit siRNA and small molecule c-kit inhibitors might overcome the acquired gefitinib resistance, and inhibit PC9/GR cell growth in vitro and in vivo.

Glioblastoma Stem Cell-Derived Exosomes Enhance Stemness and Tumorigenicity of Glioma Cells by Transferring Notch1 Protein.

Exosomes contain plenty of bioactive information, playing an important role in intercellular communication by transfer their bioactive molecular contents to recipient cells. Glioblastoma stem cells (GSCs) and non-GSC glioma cells coexist in GBM microenvironment; GSC-released exosomes contain intracellular signaling molecules, which may affect the biological phenotypes of recipient cells. However, whether GSC exosomes could affect the biological phenotype of non-GSC glioma cells has not yet been defined. To explore whether GSC exosomes could reprogramme non-GSC glioma cells into GSCs and its possible mechanism involved, non-GSC glioma cells were treated with GSCs released exosomes; the potential mechanisms of action were studied with RNA interference, Notch inhibitors and Western blot analysis. The proliferation, neurosphere formation, invasive capacities, and tumorigenicity of non-GSC glioma cells were increased significantly after GSC exosome treatment; Notch1 signaling pathway was activated in GSCs; Notch1 protein was highly enriched in GSC exosomes; Notch1 signaling pathway and stemness-related protein expressions were increased in GSC exosome treated non-GSC glioma cells and these cell generated tumor tissues; Notch1 protein expression in GSCs and their exosomes, and the neurosphere formation of GSCs were decreased by Notch1 RNA interference; Notch1 signaling pathway protein and stemness protein expressions were decreased in GSC exosome treated non-GSC glioma cells by Notch1 RNA interference and Notch inhibitors. The findings in this study indicated that GSC exosomes act as information carriers, mediated non-GSC glioma cell dedifferentiation into GSCs by delivering Notch1 protein through Notch1 signaling activation, and enhanced stemness and tumorigenicity of non-GSC glioma cells.

Zoledronic acid inhibits the growth of cancer stem cell derived from cervical cancer cell by attenuating their stemness phenotype and inducing apoptosis and cell cycle arrest through the Erk1/2 and Akt pathways.

BACKGROUND: Zoledronic acid is the most potent osteoclast inhibitor and is widely used for advanced cancer patients with bone metastasis, but its role on cancer stem cells (CSCs) remains unclear. In the present study, we aimed to identify the stemness phenotypic characteristics of CSCs derived from cervical cancer cells and explore the anti-cancer efficiency of zoledronic acid on these cells, as well as the possible molecular mechanisms. METHODS: Stemness phenotypic identification of cervical cancer cells derived CSCs was performed via sphere formation efficiency (SFE), tumorigenesis, immunofluorescence staining, Transwell assay, and western blot. Anti-cancer efficiency of zoledronic acid on these cells (including proliferation, stemness phenotype, apoptosis, and cell cycle) was carried out through MTT assay, SFE, transwell, DAPI staining, flow cytometry, immunofluorescence, TUNEL staining, and western blot, both in vitro and in vivo. RESULTS: Enhanced self-renewal ability, including SFE and tumorigenesis, was verified in cervical cancer cells derived CSCs compared to parental cervical cancer cells. Specifically, the expression of ALDH1, Sox2, CD49f, Nanog, and Oct4 was significantly up-regulated in cervical cancer cells derived CSCs. Furthermore, enhanced migratory ability was observed in these cells along with up-regulated N-cadherin and Vimentin and down-regulated E-cadherin. Zoledronic acid inhibited cervical cancer cells derived CSCs proliferation in vitro and in vivo. The stemness phenotype of these CSCs including tumor sphere formation, migration, as well as the expression of the aforementioned associated markers was also suppressed. In addition, zoledronic acid significantly induced apoptosis and cell cycle arrest of cervical cancer cells derived CSCs in a dose-dependent manner. Mechanistically, the expression of phosphorylated Erk1/2 and Akt was significantly increased in cervical cancer cells derived CSCs compared to parental cervical cancer cells. Zoledronic acid inhibited phosphorylated Erk1/2 and Akt in cervical cancer cells derived CSCs. IGF-1, a potent stimulator for Erk1/2 and PI3K/Akt, attenuated the aforementioned anti-cancer effect of zoledronic acid. CONCLUSIONS: Zoledronic acid inhibited the growth of cervical cancer cells derived CSCs through attenuating their stemness phenotype, inducing apoptosis, and arresting cell cycle. The suppression of phosphorylated Erk1/2 and Akt was involved in this process.

Species-dependent neuropathology in transgenic SOD1 pigs.

Mutations in the human copper/zinc superoxide dismutase 1 (hSOD1) gene cause familial amyotrophic lateral sclerosis (ALS). It remains unknown whether large animal models of ALS mimic more pathological events seen in ALS patients via novel mechanisms. Here, we report the generation of transgenic pigs expressing mutant G93A hSOD1 and showing hind limb motor defects, which are germline transmissible, and motor neuron degeneration in dose- and age-dependent manners. Importantly, in the early disease stage, mutant hSOD1 did not form cytoplasmic inclusions, but showed nuclear accumulation and ubiquitinated nuclear aggregates, as seen in some ALS patient brains, but not in transgenic ALS mouse models. Our findings revealed that SOD1 binds PCBP1, a nuclear poly(rC) binding protein, in pig brain, but not in mouse brain, suggesting that the SOD1-PCBP1 interaction accounts for nuclear SOD1 accumulation and that species-specific targets are key to ALS pathology in large mammals and in humans.