Genome-wide detection of multiple variants associated with teat number in French Yorkshire pigs.

BACKGROUND: Teat number is a vital reproductive trait in sows, crucial for providing immunity and nutrition to piglets during lactation. However, "missing heritability" in Single Nucleotide Polymorphism (SNP)-based Genome-Wide Association Studies (GWAS) has led to an increasing focus on structural variations in the genetic analysis of complex biological traits. RESULTS: In this study, we generated a comprehensive CNV map in a population of French Yorkshire pigs (n = 644) and identified 429 CNVRs. Notably, 44% (189 CNVRs) of these were detected for the first time. Subsequently, we conducted GWAS for teat number in the French Yorkshire pig population using both 80K chip and its imputed data, as well as a GWAS analysis based on CNV regions (CNVR). Interestingly, 80K chip GWAS identified two SNPs located on Sus scrofa chromosome 5 (SSC5) that were simultaneously associated with Total Teat Number (TTN), Left Teat Number (LTN), and Right Teat Number (RTN). The leading SNP (WU_10.2_5_76130558) explained 3.33%, 2.69%, and 2.67% of the phenotypic variance for TTN, LTN, and RTN, respectively. Moreover, through imputed GWAS, we successfully identified 30 genetic variants associated with TTN located within the 73.22 -73.30 Mb region on SSC5. The two SNPs identified in the 80K chip GWAS were also located in this region. In addition, CNVR-based GWAS revealed three significant CNVRs associated with TTN. Finally, through gene annotation, we pinpointed two candidate genes, TRIM66 and PRICKLE1, which are related to diverse processes such as breast cancer and abnormal vertebral development. CONCLUSIONS: Our research provides an in-depth analysis of the complex genetic structure underlying teat number, contributing to the genetic enhancement of sows with improved reproductive performance and, ultimately, bolstering the economic benefits of swine production enterprises.

Multi-omic characterization of allele-specific regulatory variation in hybrid pigs.

Hybrid mapping is a powerful approach to efficiently identify and characterize genes regulated through mechanisms in cis. In this study, using reciprocal crosses of the phenotypically divergent Duroc and Lulai pig breeds, we perform a comprehensive multi-omic characterization of regulatory variation across the brain, liver, muscle, and placenta through four developmental stages. We produce one of the largest multi-omic datasets in pigs to date, including 16 whole genome sequenced individuals, as well as 48 whole genome bisulfite sequencing, 168 ATAC-Seq and 168 RNA-Seq samples. We develop a read count-based method to reliably assess allele-specific methylation, chromatin accessibility, and RNA expression. We show that tissue specificity was much stronger than developmental stage specificity in all of DNA methylation, chromatin accessibility, and gene expression. We identify 573 genes showing allele specific expression, including those influenced by parent-of-origin as well as allele genotype effects. We integrate methylation, chromatin accessibility, and gene expression data to show that allele specific expression can be explained in great part by allele specific methylation and/or chromatin accessibility. This study provides a comprehensive characterization of regulatory variation across multiple tissues and developmental stages in pigs.

Partial mitochondrial genome of the Sanchiang Tree Toad Hyla sanchiangensis (Anura: Hylidae).

We reported the partial mitochondrial genome (mitogenome) for Hyla sanchiangensis (Anura: Hylidae), a arboreal frog and endemic in China. The length of partial mitogenome of H. sanchiangensis was 15,664 bp, and contained PCGs (COX1-3, ND1-6, ND4L, ATP6, ATP8 and CYTB), 2 ribosomal RNA genes, 22 transfer RNA genes, and 2 non-coding regions of a L-strand replication origin and a partial loop region. The overall base composition of the sequence is 29.91% A, 29.86% T, 14.58% G, and 25.65% C, with a total A + T content of 59.77%. The result of phylogenetic analysis showed that H. sanchiangensis formed a clade with other species belonging to the genus of Hyla. This mitogenome data could help in evolutionary biology and population genetics of the Hylid species.

The complete mitochondrial genome of Quasipaa exilispinosa (Anura: Dicroglossidae).

The complete mitochondrial genome (mitogenome) of Quasipaa exilispinosa (Anura: Dicroglossidae) we sequenced can provide critical information for mitogenome evolution. This mitogenome with 17,046 bp long contains 13 protein-coding genes (PCGs), 22 transfer RNA genes, two ribosomal RNA genes, and one non-coding regions. The overall AT content of 58.6%. The data using Bayesian phylogenetic inference supports the close relationship with Q. spinosa from the genus Quasipaa. Our results will be helpful for detail study on phylogenetic relationships of the related taxa.

miR-590-5P inhibits the progression of tongue squamous cell carcinoma by targeting FasL.

OBJECTIVE: This study evaluated microRNA-590-5P (miR-590-5P), which functions as an anti-onco-miRNA in TSCC by downregulating FasL expression. METHODS: In this study, immunohistochemistry was used to detect FasL protein expression in 30 OSCC samples and 8 normal oral mucosa tissue samples. Target Scan was used to predict miRNAs that target FasL. Luciferase reporter assays were used to confirm the effects of miRNA on FasL. Subsequently, the SCC3 tongue cancer cell line was transfected with a miR-590-5P mimic or miR-590-5P inhibitor. qPCR and Western blots were used to detect the expression levels of miR-590-5P and FasL. SCC3 cell viability, apoptosis and growth were assayed by MTT assays, colony formation assays, and a xenograft model. RESULTS: FasL expression was significantly higher in OSCC tissue samples than in normal oral mucosa tissue samples. miR-590-5P could downregulate the expression of FasL in vitro via direct binding to its 3'-untranslated region (3'-UTR). Overexpression of miR-590-5P inhibited the proliferation of SCC3 cells. Moreover, miR-590-5P increased the sensitivity of SCC3 cells to the chemotherapeutic agent cisplatin (DDP) and led to a significant decrease in colony formation ability. The xenograft experiment confirmed that miR-590-5P can suppress the development of TSCC. CONCLUSIONS: These results suggest that miR-590-5P targets FasL to inhibit the development of tongue cancer and that miR-590-5P may be a novel therapeutic target for TSCC.

Ghrelin is a possible new predictor associated with executive function in patients with type 2 diabetes mellitus.

AIMS/INTRODUCTION: The aim of the present research was to study the ghrelin level, executive function and their possible association in patients with type 2 diabetes mellitus. MATERIALS AND METHODS: A total of 370 people were recruited between March 2015 and March 2016 in this study. Among them, 212 participants were patients with type 2 diabetes mellitus and 158 participants were included as the control group. Their blood sample was analyzed for the level of ghrelin and other clinical indexes. Cognitive function was measured by the Montreal Cognitive Assessment, and executive function was evaluated by the Wisconsin Card Sorting Test. RESULTS: In the type 2 diabetes mellitus group, age, years of education, duration of diabetes, fasting blood glucose, glycated hemoglobin, hypertension and waist-to-hip ratio were correlated with total Montreal Cognitive Assessment scores. No association was found between ghrelin level and total Montreal Cognitive Assessment score in patients with type 2 diabetes mellitus. However, ghrelin was found to be a significant predictor for executive function impairment measured by the Wisconsin Card Sorting Test in patients with type 2 diabetes mellitus. CONCLUSIONS: The level of serum ghrelin might be a biomarker of executive function and become a strong predictor of executive function impairment in patients with type 2 diabetes mellitus. Ghrelin might have a potential protective effect against cognitive function impairment in type 2 diabetes patients.

The clinical significance of CDK1 expression in oral squamous cell carcinoma.

OBJECTIVES: To evaluate the clinical significance of cyclin-dependent kinase 1 (CDK1) in 77 oral squamous cell carcinomas (OSCC) using immunohistochemical methods. STUDY DESIGN: Immunohistochemical expression of CDK1 was compared with various clinicopathological features in 77 OSCC and 60 controlled epithelia adjacent to the tumours. In addition, correlation of CDK1 expression and prognostic and the 5-year accumulative survival rate of OSCC were investigated. RESULTS: The CDK1 protein was expressed in 52 cases of 77 tumor tissues (67.5%), compared with 21 cases of 60 controlled (35.0%). The expression of CDK1 was significantly correlated with the histological grade of OSCC (P<0.05). The CDK1 protein was over-expressed in recurrent tumors or in those with lymph node metastasis. Statistical analysis showed a significant reduction in the 5-year accumulative survival rate in CDK1 positive cases compared with CDK1 negative cases (P<0.05). Namely, the CDK1 positive patients had poor prognosis. CONCLUSIONS: The expression of CDK1 might serve as malignant degree and prognostic markers for the survival of OSCC.

[Experimental study of canine bone marrow mesenchymal stem cells combined with calcium phosphate cement for repair of mandibular bone defects in Beagle dogs].

PURPOSE: To evaluate the effects of bone marrow mesenchymal stem cells (BMSCs) combined with calcium phosphate cement (CPC) scaffold for repair of mandibular defect in Beagle dogs. METHODS: BMSCs were isolated from Beagle dogs and cultured in DMEM plus 10% FBS. The induction effect was determined using alizarin red staining or alkaline phosphate staining at 14-day of culture. BMSCs were added to the CPC scaffold for animal experiments. In vivo, three critical size bone defects were surgically created in each side of the mandible. The bone defects were repaired with BMSCs-CPC (scaffolds with composite seeding cells), CPC (scaffold alone) or no materials (blank group). Two dogs were sacrificed at 4-week and 8-week after operation. Gross observation, X-ray imaging, histologic and histometric analyses were performed to evaluate the level of bone formation. RESULTS: Newly formed bones were detected within all defect sites after operation. The BMSCs-CPC group and CPC group showed increased bone formation compared with the blank group. The BMSCs-CPC group exhibited more bone formation and degradation of the material than the CPC group. The percentage of new bone in the BMSCs-CPC and CPC treated group were significantly higher than that in the control group (P<0.05), while the percentage of new bone in the BMSCs-CPC sites was higher than that in the CPC sites (P<0.01); the percentage of residual material in the BMSCs-CPC sites was lower than that in the CPC sites (P<0.01) 4 weeks and 8 weeks after operation. CONCLUSIONS: Using the theory of tissue engineering, BMSCs composite CPC compound is an effective method in promoting new bone regeneration, which has a positive influence on the bone space preservation.

Effects of FasL expression in oral squamous cell cancer.

PURPOSE: To probe the role of FasL in cell apoptosis in oral squamous cell carcinomas (OSCCs). METHODS: The expression of Fas/FasL was assessed in 10 cases of normal oral epithelium, 38 cases of OSCC and tumor infiltrating lymphocytes (TIL), and 11 cases of metastatic lymph nodes by immunohistochemistry. Apoptosis of tumor cells and TIL was detected by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay (TUNEL). FasL-induction of T cell apoptosis was tested by co-culture assay in vitro with SCC-9 and Jurkat T cells. RESULTS: The 10 cases of normal oral epithelium all demonstrated extensive expression of Fas, the positive rate being largely down-regulated in OSCC (21/38) (P<0.05) compared to the normal (10/10). At the same time, the positive rate of FasL significantly increased in OSCC (P<0.05) especially those with lymph node metastasis (P<0.05). The positive rates of Fas in well and middle differentiated OSCC were higher than those in poor differentiated OSCC (P<0.05). The AI of tumor cells in Fas-positive OSCC was remarkably higher than that in Fas-negative OSCC (P<0.01), with a positive correlation between Fas expression and cell differentiation as well as apoptosis (r=0.68, P<0.01). The AI of tumor cells in FasL positive OSCC was remarkably lower than that in control while the AI of TIL was higher than in FasL negative OSCC (P< 0.05). The AI of tumor cells reversely correlated with that of TIL (r = -0. 72, P<0.05). It was found that SCC-9 cells expressing functional FasL could induce apoptosis of Jurkat cells as demonstrated by co-culture assays. As a conclusion, it is evident that OSCC cells expressing FasL can induce apoptosis in Fas-expressing T cells. CONCLUSIONS: In progression of OSCC, expression of the Fas/FasL changes significantly. The results suggest that FasL is a mediator of immune privilege in OSCC and may serve as an marker for predicting malignant change in oral tissues.

Protective effects of total flavones of rhododendra against global cerebral ischemia reperfusion injury.

The purpose of this research was to demonstrate the protective effects and possible mechanisms of total flavones of rhododendra (TFR) against global cerebral ischemia reperfusion injury in rats. Global cerebral ischemia/reperfusion injury was caused by four vessel occlusion (bilateral vertebral arteries and bilateral carotid arteries, 4-VO). The electroencephalographic (EEG) changes were recorded. The EEG, brain water content, levels of malondialdehyde (MDA) and lactate dehydrogenase (LDH) activity in plasma, aggregation of platelets induced by ADP, and the resting and CaCl(2)-induced increase of free intracellular calcium concentration ([Ca(2+)](i)), were also evaluated. TFR dramatically elevated EEG amplitude, reduced the brain water content and the resting cytoplasmic free calcium concentration, inhibited the increase of [Ca(2+)](i) induced by CaCl(2) and had an inhibitory effect on platelet aggregation. The LDH activity and the MDA content in plasma were also decreased. These results indicate that TFR has protective effects against cerebral injury in rats, which might be associated with its antioxidant properties, antiplatelet effects and possible inhibition of Cal(2+) influx to reduce [Ca(2+)](i).

Enhancement of immune responses to hepatitis B vaccine in immunodepressed mice by CpG oligodeoxynucleotide.

AIM: To investigate enhancement of immune response to hepatitis B vaccine in immunosuppressed mice by CpG oligodeoxynucleotide (ODN). METHODS: Immunodepressed C57BL/6 mice caused by cytoxan were given an injection of either hepatitis B vaccine alone or hepatitis B vaccine and CpG ODN into the left tibialis anterior muscle, then the mice were given another shot after 2 weeks using the same formulation. Blood was collected at 5 weeks after immunization and anti-hepatitis B surface antigen IgG and IL-12 levels were measured by ELISA. Spleens of immunized mice were observed under microscope. RESULTS: There was a two-fold increase in anti-hepatitis B surface antigen IgG level when hepatitis B vaccine was mixed with CpG ODN to immunize immunodepressed mice. There was a significant increase in IL-12 level when hepatitis B vaccine was mixed with CpG ODN. Under optical microscope, there were fewer lymphocytes in immunodepressed mice than in normal control group. The proliferation level of spleen lymphocytes in CpG ODN group was elevated compared with that of normal control group and nuclei of lymphocytes became larger. CONCLUSION: CpG ODN can enhance the immune response to hepatitis B vaccine in immunodepressed mice.

CpG ODN enhances immunization effects of hepatitis B vaccine in aged mice.

Oligodeoxynucleotides (ODN) containing unmethylated CpG dinucleotides in contexts of unique sequence (CpG motifs) is active as adjuvant in induction of cellular and humoral immune responses in young mice. To date, there are only limited reports about effect of CpG ODN on immune responses against hepatitis B (HB) infection in aged mice. Our studies demonstrated there were significant increases in secreting of total anti-HB IgG, IgG1 and IgG2a, as well as of IL-12 and IFN-gamma, when CpG ODNs were injected together with hepatitis B antigen in aged mice. Moreover, CpG ODN could stimulate proliferation of spleen lymphocytes in a dose-dependent manner. Taken together, the results we obtained indicate that the adding of CpG ODN into the vaccine antigen might be useful in development of more effective vaccination for inducing anti-HB virus responses in the elderly.