Differentiation of Goat Meat Freshness Using Gas Chromatography with Ion Mobility Spectrometry.

To investigate the flavor changes in goat meat upon storage, the volatile components observed in goat meat after different storage periods were determined using gas chromatography-ion mobility spectrometry (GC-IMS). A total of 38 volatile organic compounds (VOCs) were determined from the goat meat samples, including alcohols, ketones, aldehydes, esters, hydrocarbons, ethers, and amine compounds. 1-Hexanol, 3-Hydroxy-2-butanone, and Ethyl Acetate were the main volatile substances in fresh goat meat, and they rapidly decreased with increasing storage time and can be used as biomarkers for identifying fresh meat. When combined with the contents of total volatile basic-nitrogen (TVB-N) and the total numbers of bacterial colonies observed in physical and chemical experiments, the characteristic volatile components of fresh, sub-fresh, and spoiled meat were determined by principal component analysis (PCA). This method will help with the detection of fraudulent production dates in goat meat sales.

CXCL1 stimulates decidual angiogenesis via the VEGF-A pathway during the first trimester of pregnancy.

Angiogenesis is critical to establishing a successful pregnancy. The chemokine (C-X-C motif) ligand 1 (CXCL1) is a small cytokine belonging to the CXC chemokine family that is an important chemokine involved in the processes of angiogenesis and arteriogenesis; however, little is known about its role in decidual angiogenesis. Effects of CXCL1 on cell proliferation and migration (propidium iodide staining and wound healing assays) of HUVEC cells were determined. The angiogenesis roles of CXCL1 in HUVEC-HTR8/SVneo co-culture system were detected by the tube formation assay. Signal transduction pathways in HUVEC cells in response to CXCL1 were determined by in-cell western analyses. In vivo, mice were injected with (1) PBS (Group A) or (2) CXCL1-neutralizing antibody (Group B) or (3) CXCL1-neutralizing antibody plus recombinant VEGF-A protein (Group C) from E1 to E5 and sacrificed at E6.5 of pregnancy. The decidual angiogenesis in mice was examined by immunohistochemistry of cluster designation 34 (CD34), and the expression levels of vascular endothelial growth factor-A (VEGF-A) in the decidual cells and vascular endothelial growth factor receptor 2 (VEGFR2) in decidual vascular endothelial cells were also tested. Exogenous recombinant human CXCL1 supported endothelial cell proliferation and migration, and this effect was blocked by CXCL1-neutralizing antibody or CXCR2 inhibitor SB265610. The tube formation of HUVEC-HTR8/SVneo co-culture system was significantly stimulated by CXCL1, but this effect was markedly abrogated once they were pretreated with CXCL1-neutralizing antibody or CXCR2 inhibitor SB265610. In addition, the level of vascular endothelial growth factor A (VEGF-A) expression in HUVEC cells was increased by CXCL1, and this level was suppressed by CXCL1-neutralizing antibody or CXCR2 inhibitor SB265610. In vivo, compared with Group A (n = 3), decidual angiogenesis was significantly reduced in Group B by CD34 immunostaining. But compared with Group B, decidual angiogenesis was significantly increased in Group C. In addition, the expression of VEGF-A and VEGFR2 was significantly increased after neutralizing of CXCL1 in Group B. In conclusions, CXCL1 may play essential roles in decidual angiogenesis during the first trimester, and this function may be mediated in part via altering VEGF-A expression.

Elevated expression of EP4 in human decidua is associated with delayed embryo expulsion during medical abortion by promoting decidual cell proliferation.

Purpose: Mifepristone in conjunction with misoprostol, is widely used in China as an effective medical abortifacient. However, a small proportion of women experience the unpleasant side effects of prolonged vaginal bleeding caused by delayed embryo expulsion. The aims of this study were to determine whether the expression levels of prostanoid receptors in human decidua are associated with delayed embryo expulsion in mifepristone-misoprostol induced an early medical abortion.Methods: Discharged decidua tissues were collected from females undergoing an artificial abortion (AA) (n = 28), females with early embryo expulsion during a medical abortion (EEMA) (n = 20) and delayed embryo expulsion in medical abortion (DEMA) (n = 30). The expression levels of prostanoid receptors in human decidua were assessed with immunohistochemistry and real-time PCR methods. Further, the RNAi method was used to silence prostanoid receptors 4 (EP4) in the primary decidual cells and human endometrial adenocarcinoma cell line Ishikawa cells in vitro and cell cycle analysis of these cells was performed.Results: All five prostanoid receptors (EP1-4, FP) were observed in human early pregnancy decidua. The protein and mRNA expression level of EP4 in the DEMA group were all significantly higher than that in the EEMA group. EP4 silence induced G1/S arrest of primary decidual cells and Ishikawa cells in vitro.Conclusions: Elevated expression level of EP4 in human decidua was significantly associated with delayed embryo expulsion in early medical abortion by promoting decidual cell proliferation. Detailed studies on the nature of roles EP4 plays in human decidua will help us to develop more effective prevention and noninvasive intervention approaches for delayed embryo expulsion during a medical abortion.

Low-dose Roundup induces developmental toxicity in bovine preimplantation embryos in vitro.

Roundup is a widely used glyphosate-based herbicide worldwide. Roundup residues can be detected in the organs and urine of animals. However, its toxicity on mammalian preimplantation embryos has not been well investigated. Here, we show Roundup impairs the development and quality of bovine preimplantation embryos in a dose-dependent manner. Exposure to the agricultural recommended doses of Roundup caused in vitro developmental arrest and quick death of bovine embryos. Furthermore, even a very low concentration (0.9 ppm) of Roundup was harmful to bovine preimplantation development. In addition, Roundup increases intracellular calcium levels and induces oxidative stress and apoptosis in bovine embryos. Even if the embryos developed to morphologically normal blastocysts when cultured with low concentrations of Roundup, abnormal intracellular calcium and oxidative stress could be detected inside the embryos and led to an increased incidence of apoptosis in the blastocysts. These data suggest Roundup residues from the agricultural application are potentially dangerous to mammalian preimplantation embryos.

Claudin 4 in pancreatic ß cells is involved in regulating the functional state of adult islets.

The functional state (FS) of adult pancreatic islets is regulated by a large array of regulatory molecules including numerous transcription factors. Whether any islet structural molecules play such a role has not been well understood. Here, multiple technologies including bioinformatics analyses were used to explore such molecules. The tight junction family molecule claudin 4 (Cldn4) was the highest enriched amongst over 140 structural genes analysed. Cldn4 expression was ~75-fold higher in adult islets than in exocrine tissues and was mostly up-regulated during functional maturation of developing islet cells. Cldn4 was progressively down-regulated in functionally compromised, dedifferentiating insulin-secreting ß cells and in db/db type 2 diabetic islets. Furthermore, the genetic deletion of Cldn4 impaired significantly the FS without apparently affecting pancreas morphology, islet architectural structure and cellular distribution, and secretion of enteroendocrine hormones. Thus, we suggest a previously unidentified role for Cldn4 in regulating the FS of islets, with implications in translational research for better diabetes therapies.

Association between the SNPs in trace element-related metabolic genes and the risk of gastric cancer: a case-control study in Xianyou of China.

This study aims to analyse the potential relationship between single-nucleotide polymorphism (SNPs) in trace element related metabolic genes GSTM3, GSTP1, GPX1 and NKG2D and the risk of gastric cancer. A case-control study was conducted in the hospital of Xianyou, Fujian, China. In this study, a total of 299 patients with histopathological diagnosis in gastric cancer and 295 healthy control subjects were involved. Association between the SNPs in trace element-related metabolic genes and gastric cancer risk was analysed using the unconditional logistic regression model. No relationship was found between the SNPs of GSTM3 and GPX1 genes and gastric cancer risk. However, the risk of gastric cancer is related to the SNPs of NKG2D gene (rs1049174). Patients who carry the rs1049174 GG genotype have a higher incidence of the gastric cancer and a multivariate odds ratio (OR) of 1.85 (95% confidence intervals (CI): 1.02-3.38). Through haplotype analysis, two haplotypes (i.e. A_rs1746123-T_rs10431294-G_rs1049174 and T_rs1746123-T_rs10431294-C_rs1049174), OR of 0.29 (95% CI: 0.15-0.56) and 0.33, (95% CI: 0.22-0.50), respectively, were found to have lower incidence of gastric cancer. Meanwhile, another two haplotypes (T_rs1746123-C_rs10431294-C_rs1049174 and T_rs1746123-T_rs10431294-G_rs1049174), OR of 1.81 (95% CI: 1.40-2.34) and 3.09 (95% CI: 2.30-4.16), respectively, were found to have a higher incidence of gastric cancer. Further, no influence of the haplotype on the risk of cardia gastric cancer was found. However, the haplotype T_rs1746123-T_rs10431294-C_rs1049174 had lower incidence of noncardia gastric cancer by 46%. Our data showed that polymorphisms of trace element-related metabolic genes are important in gastric cancer pathology.

Sperm DNA fragmentation index, as measured by sperm chromatin dispersion, might not predict assisted reproductive outcome.

OBJECTIVE: Routine semen parameters have limited clinical diagnostic value for predicting male infertility. The aim of this study was to investigate the association between sperm DNA fragmentation index (DFI) and semen quality, and between DFI and clinical pregnancy rate of in vitro fertilization (IVF) and intracytoplasmic sperm injection (ICSI). METHODS AND MATERIALS: A total of 390 couples undergoing sperm fragmentation prior to receiving conventional IVF (n = 238) or ICSI (n = 152) were evaluated. RESULTS: We found that there were no significant differences in fertilization rate, good embryo rate, or pregnancy rate between high (≥30%) and low (<30%) DFI groups after IVF or ICSI. However, statistically different decreasing motility trends under higher DFI values in the IVF and ICSI groups were detected. Comparison of ROC curve of motility and DFI scores for achieved pregnancy revealed that the best DFI cut-off value was 20%. Also, no significant change was found when 20% DFI level was taken in IVF and ICSI outcomes. CONCLUSION: DFI scores did not provide independent information regarding fertilization, embryo quality, or pregnancy for infertile patients who received IVF or ICSI, but were consistent with semen analysis for infertile couples, regardless of IVF or ICSI outcome.

In vitro reprogramming of rat bmMSCs into pancreatic endocrine-like cells.

Islet transplantation provides curative treatments to patients with type 1 diabetes, but donor shortage restricts the broad use of this therapy. Thus, generation of alternative transplantable cell sources is intensively investigated worldwide. We previously showed that bone marrow-derived mesenchymal stem cells (bmMSCs) can be reprogrammed to pancreatic-like cells through simultaneously forced suppression of Rest/Nrsf (repressor element-1 silencing transcription factor/neuronal restrictive silencing factor) and Shh (sonic hedgehog) and activation of Pdx1 (pancreas and duodenal transcription factor 1). We here aimed to reprogram bmMSCs further along the developmental pathway towards the islet lineages by improving our previous strategy and by overexpression of Ngn3 (neurogenin 3) and NeuroD1 (neurogenic differentiation 1), critical regulators of the development of endocrine pancreas. We showed that compared to the previous protocol, the overexpression of only Pdx1 and Ngn3 reprogrammed bmMSCs into cells with more characteristics of islet endocrine lineages verified with bioinformatic analyses of our RNA-Seq datasets. These analyses indicated 2325 differentially expressed genes including those involved in the pancreas and islet development. We validated with qRT-PCR analysis selective genes identified from the RNA-Seq datasets. Thus, we reprogrammed bmMSCs into islet endocrine-like cells and advanced the endeavor to generate surrogate functional insulin-secreting cells.

Effect of anesthesia on cognitive status and MMP-2 expression in rats.

OBJECTIVE: To investigate the effect of anesthesia on the cognitive status damage and MMP-2 expression in rats. METHODS: A total of 120 healthy rats were selected and randomly divided into the control group, CF3-CH(OCH2F)-CF3 (Sevoflurane) group and CF3-CH2-O-CHF-CF3 group (Sevoflurane) (n=40). After training for 3 d by the Morris water maze, the control group were injected with fentanyl for analgesia, the CF3-CH(OCH2F)-CF3 group and the CF3-CH2-O-CHF-CF3 group were anesthesia with CF3-CH (OCH2F)-CF3 and CF3-CH2-O-CHF-CF3 on the basis of fentanyl, then rats in three groups underwent open surgery and suture conventional incision. Morris water maze was used to measure the rats' cognitive ability in three groups on the 1st d, 3rd d, 5th d and 7th d, and the brain tissue MMP-2 expression was detected. RESULTS: After 1 d/7 d of the surgery, Morris water maze performance and MMP-2 expression were not significantly different among three groups (P>0.05); After 3 d/5 d of the surgery, compared with the control group, the Morris water maze test result was significantly worsened, MMP-2 expression levels were significantly increased (P<0.05); After 3 d/5 d of the surgery, compared with the CF3-CH2-O-CHF-CF3 group, Morris water maze test result of CF3-CH(OCH2F)-CF3 group was significantly worsened, MMP-2 expression levels were significantly increased (P<0.05). CONCLUSIONS: Anesthesia can cause some injury on cognitive status, different anesthetic drugs may cause different injury, and the cognitive status injury is related to the MMP-2 expression.

In vitro reprogramming of rat bone marrow-derived mesenchymal stem cells into insulin-producing cells by genetically manipulating negative and positive regulators.

Islet cell replacement therapy represents the most promising approach for the cure of type 1 diabetes if autoimmunity to ß cells is under control. However, this potential is limited by a shortage of pancreas donors. To address the donor shortage problem, we determined whether bone marrow-derived mesenchymal stem cells (bmMSCs) can be directly reprogrammed to islet lineages by simultaneously forced suppression and over-expression of key regulator genes that play critical roles during pancreas development. Here, we report that rat bmMSCs were converted in vitro into insulin-producing cells by suppressing two-repressor genes repressor element-1 silencing transcription factor/neuronal restrictive silencing factor (Rest/Nrsf) and sonic hedgehog (Shh) and by over-expressing pancreas and duodenal transcription factor 1 (Pdx1). The reprogrammed bmMSCs expressed both genes and proteins specific for islet cells. These converted cells were capable of releasing insulin in a glucose-responsive manner. Our study suggests that bmMSCs may ultimately be reprogrammed to functional insulin-secreting cells.

[Relationship between neuronal restricted silencing factor and induced differentiation from rat mesenchymal stem cells to neurons].

OBJECTIVE: To analyze the change of the neuronal restricted silencing factor (NRSF) gene as well as the NRSF regulation genes in beta-mercaptoethanol induction of the marrow mesenchymal stem cells (MSCs) to neurons, and to discuss the function of NRSF in neural induction of the MSCs and the mechanism of the differentiation from MSCs to neurons. METHOD: We used beta-mercaptoethanol, serum-free DMEM, and dimethyl sulfoxide to induce rat MSCs to differentiate to neurons, and then analyzed the changes of the expressions of NRSF gene and NRSF-regulated genes through real-time PCR. RESULTS: The rat MSCs were successfully induced to differentiate into neuron-like cells. The induced neuron marker, neuron-specific enolase, was positive. Real-time PCR showed that the expression of NRSF gene remarkably declined. The expressions of neurotrophic tyrosine kinase receptor, type 3, synaptosomal-associated protein 25, L1 cell adhesion molecular,neuronal pentraxin receptor in the NRSF-regulated genes also increased at varied extents. CONCLUSIONS: The differentiation from MSCs to neurons is relevant with the decline of NRSF expression and the increase of the expressions of NRSF-regulated genes. The NRSF may be the key gene during the differentiation from MSCs to neurons.

[Construction and identification of the lentiviral vector of repressor element-1/neuron-restrictive silencer element double-stranded RNA].

OBJECTIVE: To construct a lentiviral vector of repressor element-1/neuron-restrictive silencer element (RE-1/NRSE) double-stranded RNA (dsRNA). METHODS: The RE-1/NRSE cDNA containing both sense and antisense oligo DNA fragments of the targeting sequence was synthesized and cloned into the pGC-LV vector. The obtained lentiviral vector containing RE-1/NRSE dsRNA was confirmed by PCR and sequencing. A total of 293T cells were cotransfected with lentiviral vector of L-smNRSE/RE-1, pHelper 1.0, and pHelper 2.0. The titer of virus was measured based on the expression level of green fluorescent protein. The transfection efficiency of green fluorescent protein into rat mesenchymal stem cells was calculated. RESULTS: PCR and DNA sequencing demonstrated that the constructed lentivirus vector of L-smNRSE/RE-1 produced RE-1/NRSE dsRNA.The titer of the concentrated virus was 4x108 TU/m1. The virus was stably transfected into rat mesenchymal stem cells, and the infection efficiency reached 100% when the multiplicity of infection was 80. CONCLUSION: The lentivirus vector of RE-1/NRSE dsRNA is successfully constructed.