Transcriptome profiling analysis of uterus during chicken laying periods.

The avian eggshell is formed in the uterus. Changes in uterine function may have a significant effect on eggshell quality. To identify the vital genes impacting uterine functional maintenance in the chicken, uteri in three different periods (22W, 31W, 51W) were selected for RNA sequencing and bioinformatics analysis. In our study, 520, 706 and 736 differentially expressed genes (DEGs) were respectively detected in the W31 vs W22 group, W51 vs W31 group and W51 vs W22 group. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated DEGs were enriched in the extracellular matrix, extracellular region part, extracellular region, extracellular matrix structural constituent, ECM receptor interaction, collagen-containing extracellular matrix and collagen trimer in the uterus (P < 0.05). Protein-protein interaction analysis revealed that FN1, LOX, THBS2, COL1A1, COL1A2, COL5A1, COL5A2, POSTN, MMP13, VANGL2, RAD54B, SPP1, SDC1, BTC, ANGPTL3 might be key candidate genes for uterine functional maintenance in chicken. This study discovered dominant genes and pathways which enhanced our knowledge of chicken uterine functional maintenance.

Grade follicles transcriptional profiling analysis in different laying stages in chicken.

During follicular development, a series of key events such as follicular recruitment and selection are crucially governed by strict complex regulation. However, its molecular mechanisms remain obscure. To identify the dominant genes controlling chicken follicular development, the small white follicle (SWF), the small yellow follicle (SYF), and the large yellow follicle (LYF) in different laying stages (W22, W31, W51) were collected for RNA sequencing and bioinformatics analysis. There were 1866, 1211, and 1515 differentially expressed genes (DEGs) between SWF and SYF in W22, W31, and W51, respectively. 4021, 2295, and 2902 DEGs were respectively identified between SYF and LYF in W22, W31, and W51. 5618, 4016, and 4809 DEGs were respectively identified between SWF and LYF in W22, W31, and W51. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis indicated that extracellular matrix, extracellular region, extracellular region part, ECM-receptor interaction, collagen extracellular matrix, and collagen trimer were significantly enriched (P < 0.05). Protein-protein interaction analysis revealed that COL4A2, COL1A2, COL4A1, COL5A2, COL12A1, ELN, ALB, and MMP10 might be key candidate genes for follicular development in chicken. The current study identified dominant genes and pathways contributing to our understanding of chicken follicular development.

Transcriptome landscapes of differentially expressed genes related to fat deposits in Nandan-Yao chicken.

Nandan-Yao chicken is a Chinese native chicken with lower fat deposition and better meat quality. Fat deposition is a quite complex and important economic trait. However, its molecular mechanism is still unknown in chickens. In the current study, Nandan-Yao chicken was divided into two groups based on the rate of abdominal fat at 120 days old, namely the high-fat group and low-fat group. The total RNAs were isolated and sequenced by RNA sequencing (RNA-seq). After quality control, we gained 1222, 902, 784, 624, and 736 differentially expressed genes (DEGs) in abdominal fat, back skin, liver, pectoral muscle, and leg muscle, respectively. Analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) showed that significantly enriched GO term and KEGG signaling pathway mainly involved cytosolic ribosome, growth development, PPAR signaling pathway, Wnt signaling pathway, and linoleic acid metabolism in abdominal fat, back skin, and liver. While in pectoral muscle and leg muscle, it is mainly enriched in phosphatidylinositol signaling system, adrenergic signaling in cardiomyocytes, cytosolic ribosome, and cytosolic part. Sixteen genes were differentially expressed in all five tissues. Among them, PLA2G4A and RPS4Y1 might be the key regulators for fat deposition in Nandan-Yao chicken. The protein-protein interaction (PPI) network analysis of DEGs showed that PCK1 was the most notable genes. The findings in the current study will help to understand the regulation mechanism of abdominal fat and intramuscular fat in Nandan-Yao chicken and provide a theoretical basis for Chinese local chicken breeding.

The effects of activin A on the migration of human breast cancer cells and neutrophils and their migratory interaction.

Activin A belongs to the superfamily of transforming growth factor beta (TGFß) and is a critical regulatory cytokine in breast cancer and inflammation. However, the role of activin A in migration of breast cancer cells and immune cells was not well characterized. Here, a microfluidic device was used to examine the effect of activin A on the migration of human breast cancer cell line MDA-MB-231 cells and human blood neutrophils as well as their migratory interaction. We found that activin A promoted the basal migration but impaired epidermal growth factor (EGF)-induced migration of breast cancer cells. By contrast, activin A reduced neutrophil chemotaxis and transendothelial migration to N-Formyl-Met-Leu-Phe (fMLP). Finally, activin A promoted neutrophil chemotaxis to the supernatant from breast cancer cell culture. Collectively, our study revealed the different roles of activin A in regulating the migration of breast cancer cells and neutrophils and their migratory interaction. These findings suggested the potential of activin A as a therapeutic target for inflammation and breast cancers.

The extremely high level expression of human serum albumin in the milk of transgenic mice.

The recombinant production of human serum albumin has been challenging due to the low unit price and huge amount needed, for the commercial production of rhSA at an economically feasible level, It will be well worth the effort to exploit new method for the extremely high level expression of rhSA. To this end, here a hybrid gene locus strategy was employed, a 37 Kb mWAP-hSA hybrid gene locus was constructed and used as mammary gland specific expression vector, in which the 3 Kb genomic coding sequence in the 24 Kb mouse whey acidic protein (mWAP) gene locus was substituted by the 16 Kb genomic coding sequence of human serum albumin (hSA), exactly from the start codon to the end codon. Corresponding transgenic mice were generated and rhSA was secreted into the milk at an extremely high level of 11.9 g/L. Our transgenic mice carrying the mWAP-hSA hybrid gene locus represent a model system for the cost-effective production of human serum albumin.

Plk1-mediated phosphorylation of UAP56 regulates the stability of UAP56.

Polo-like kinase 1 (Plk1) is a conserved serine/threonine protein kinase that plays pivotal roles during the cell cycle and cell proliferation. Although a number of important targets have been identified, the mechanism of Plk1-regulated pathways and the bulk of the Plk1 interactome are largely unknown. Here, we demonstrate that Plk1 interacts with the DExH/D RNA helicase, UAP56. The protein levels of UAP56 and Plk1 are inversely correlated during the cell cycle. We also show that Plk1 phosphorylates UAP56 in vitro and in vivo and that Plk1-dependent phosphorylation of UAP56 triggers ubiquitination and degradation of UAP56 through proteasomes. This result suggests that Plk1-mediated phosphorylation of UAP56 regulates the stability of UAP56. Our results will be helpful in further understanding mRNA metabolism, cell cycle progression, and the link between mRNA metabolism and cellular function.

Mitochondrial DNA polymorphisms are associated with the longevity in the Guangxi Bama population of China.

Human longevity is an interesting and complicated subject, with many associated variations, geographic and genetic, including some known mitochondrial variations. The population of the Bama County of Guangxi Province of China is well known for its longevity and serves as a good model for studying a potential molecular mechanism. In this study, a full sequence analysis of mitochondrial DNA (mtDNA) has been done in ten Bama centenarians using direct sequencing. Polymorphisms of the displacement loop (D-loop) region of mtDNA and several serum parameters were analyzed for a total of 313 Bama individuals with ages between 10 and 110 years. The results showed that there were seven mitochondrial variations, A73G, A263G, A2076G, A8860G, G11719A, C14766T, and A15326G, and four haplogroups, M(*), F1, D* and D(4) in 10 Bama centenarians. In the D-loop region of mtDNA, the mt146T occurred at a significantly lower frequency in those is the older age group (90-110 years) than in the middle (80-89 years) and in the younger (10-79 years) groups (P < 0.05). The mt146T also had lower systolic blood pressure and serum markers such as total cholesterol, triglyceride and low density lipoprotein than did mt146C in the older age group (P < 0.05). No significant differences were observed between the mt146C and the mt146T individuals in the middle and the younger groups (P > 0.05). The mt5178C/A polymorphisms did not show any significant differences among the three age-groups (P > 0.05), but different nationalities in the Bama County did show a significant difference in the mt5178C/A polymorphisms (P < 0.05). These results suggest that the mt146T/C polymorphisms in Guangxi Bama individuals may partly account for the Bama longevity whereas the mt5178C/A polymorphisms are strongly associated with the nationalities in the Guangxi Bama population.

Adipose tissue houses different subtypes of stem cells.

Adipose tissue stromal fraction (ASF) contains multipotent cells capable of differentiation towards several lineages and may be used for the treatment of various degenerative diseases. However, the multipotent cells within ASF have not been fully characterized. In this study we have attempted to characterize stem cells in the ASF obtained through serial dilution. Five single-cell clones were studied. It was found that the single-cell clones exhibited slight but significant differences in proliferative capacity and differentiation potential. We conclude that ASF houses different subtypes of stem cells.

Whole-transcriptome RNA sequencing reveals the global molecular responses and circRNA/lncRNA-miRNA-mRNA ceRNA regulatory network in chicken fat deposition.

Fat deposition is a vital factor affecting the economics of poultry production. Numerous studies on fat deposition have been done. However, the molecular regulatory mechanism is still unclear. In the present study, the whole-transcriptome RNA sequencing in abdominal fat, back skin, and liver both high- and low-abdominal fat groups was used to uncover the competitive endogenous RNA (ceRNA) regulation network related to chicken fat deposition. The results showed that differentially expressed (DE) genes in abdominal fat, back skin, liver were 1207(784 mRNAs, 330 lncRNAs, 41 circRNAs, 52 miRNAs), 860 (607 mRNAs, 166 lncRNAs, 26 circRNAs, 61 miRNAs), and 923 (501 mRNAs, 262 lncRNAs, 15 circRNAs, 145 miRNAs), respectively. The ceRNA regulatory network analysis indicated that the fatty acid metabolic process, monocarboxylic acid metabolic process, carboxylic acid metabolic process, glycerolipid metabolism, fatty acid metabolism, and peroxisome proliferator-activated receptor (PPAR) signaling pathway took part in chicken fat deposition. Meanwhile, we scan the important genes, FADS2, HSD17B12, ELOVL5, AKR1E2, DGKQ, GPAM, PLIN2, which were regulated by gga-miR-460b-5p, gga-miR-199-5p, gga-miR-7470-3p, gga-miR-6595-5p, gga-miR-101-2-5p. While these miRNAs were competitive combined by lncRNAs including MSTRG.18043, MSTRG.7738, MSTRG.21310, MSTRG.19577, and circRNAs including novel_circ_PTPN2, novel_circ_CTNNA1, novel_circ_PTPRD. This finding provides new insights into the regulatory mechanism of mRNA, miRNA, lncRNA, and circRNA in chicken fat deposition.

Viability and proliferation potential of adipose-derived stem cells following labeling with a positron-emitting radiotracer.

PURPOSE: Adipose-derived stem cells (ASCs) have promising potential in regenerative medicine and cell therapy. Our objective is to examine the biological function of the labeled stem cells following labeling with a readily available positron emission tomography (PET) tracer, (18)F-fluoro-2-deoxy-D: -glucose (FDG). In this work we characterize labeling efficiency through assessment of FDG uptake and retention by the ASCs and the effect of FDG on cell viability, proliferation, transdifferentiation, and cell function in vitro using rat ASCs. METHODS: Samples of 10(5) ASCs (from visceral fat tissue) were labeled with concentrations of FDG (1-55 Bq/cell) in 0.75 ml culture medium. Label uptake and retention, as a function of labeling time, FDG concentration, and efflux period were measured to determine optimum cell labeling conditions. Cell viability, proliferation, DNA structure damage, cell differentiation, and other cell functions were examined. Non-labeled ASC samples were used as a control for all experimental groups. Labeled ASCs were injected via tail vein in several healthy rats and initial cell biodistribution was assessed. RESULTS: Our results showed that FDG uptake and retention by the stem cells did not depend on FDG concentration but on labeling and efflux periods and glucose content of the labeling and efflux media. Cell viability, transdifferentiation, and cell function were not greatly affected. DNA damage due to FDG radioactivity was acute, but reversible; cells managed to repair the damage and continue with cell cycles. Over all, FDG (up to 25 Bq/cell) did not impose severe cytotoxicity in rat ASCs. Initial biodistribution of the FDG-labeled ASCs was 80% + retention in the lungs. In the delayed whole-body images (2-3 h postinjection) there was some activity distribution resembling typical FDG uptake patterns. CONCLUSION: For in vivo cell tracking studies with PET tracers, the parameter of interest is the amount of radiotracer that is present in the cells being labeled and consequent biological effects. From our study we developed a labeling protocol for labeling ASCs with a readily available PET tracer, FDG. Our results indicate that ASCs can be safely labeled with FDG concentration up to 25 Bq/cell, without compromising their biological function. A labeling period of 90 min in glucose-free medium and efflux of 60 min in complete media resulted in optimum label retention, i.e., 60% + by the stem cells. The initial biodistribution of the implanted FDG-labeled stem cells can be monitored using microPET imaging.

Malignant transformation of 293 cells induced by ectopic expression of human Nanog.

Tumor development has long been known to resemble abnormal embryogenesis. The ESC self-renewal gene NANOG is purportedly expressed in some epithelial cancer cells and solid tumors, but a casual role in tumor development has remained unclear. In order to more comprehensively elucidate the relationship between human Nanog and tumorigenesis, the hNanog was ectopically expressed in the 293 cell line to investigate its potential for malignant transformation of cells both in vitro and in vivo. Here we provide compelling evidence that the overexpression of hNanog resulted in increased cell proliferation, anchor-independent growth in soft agar, and formation of tumors after subcutaneous injection of athymic nude mice. Pathologic analysis revealed that these tumors were poorly differentiated. In analysis of the underlying molecular mechanism, two proteins, FAK and Ezrin, were identified to be upregulated in the hNanog expressing 293 cells. Our results demonstrate that hNanog is a potent human oncogene and has the ability to induce cellular transformation of human cells.

Transcriptome Analysis Identifies Candidate Genes and Signaling Pathways Associated With Feed Efficiency in Xiayan Chicken.

Feed efficiency is an important economic factor in poultry production, and the rate of feed efficiency is generally evaluated using residual feed intake (RFI). The molecular regulatory mechanisms of RFI remain unknown. Therefore, the objective of this study was to identify candidate genes and signaling pathways related to RFI using RNA-sequencing for low RFI (LRFI) and high RFI (HRFI) in the Xiayan chicken, a native chicken of the Guangxi province. Chickens were divided into four groups based on FE and sex: LRFI and HRFI for males and females, respectively. We identified a total of 1,015 and 742 differentially expressed genes associated with RFI in males and females, respectively. The 32 and 7 Gene Ontology (GO) enrichment terms, respectively, identified in males and females chiefly involved carbohydrate, amino acid, and energy metabolism. Additionally, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis identified 11 and 5 significantly enriched signaling pathways, including those for nutrient metabolism, insulin signaling, and MAPK signaling, respectively. Protein-protein interaction (PPI) network analysis showed that the pathways involving CAT, ACSL1, ECI2, ABCD2, ACOX1, PCK1, HSPA2, and HSP90AA1 may have an effect on feed efficiency, and these genes are mainly involved in the biological processes of fat metabolism and heat stress. Gene set enrichment analysis indicated that the increased expression of genes in LRFI chickens was related to intestinal microvilli structure and function, and to the fat metabolism process in males. In females, the highly expressed set of genes in the LRFI group was primarily associated with nervous system and cell development. Our findings provide further insight into RFI regulation mechanisms in chickens.

Genome-Wide Association Study Using Whole-Genome Sequencing Identifies a Genomic Region on Chromosome 6 Associated With Comb Traits in Nandan-Yao Chicken.

Comb traits have potential economic value in the breeding of indigenous chickens in China. Identifying and understanding relevant molecular markers for comb traits can be beneficial for genetic improvement. The purpose of this study was to utilize genome-wide association studies (GWAS) to detect promising loci and candidate genes related to comb traits, namely, comb thickness (CT), comb weight (CW), comb height, comb length (CL), and comb area. Genome-wide single-nucleotide polymorphisms (SNPs) and small insertions/deletions (INDELs) in 300 Nandan-Yao chickens were detected using whole-genome sequencing. In total, we identified 134 SNPs and 25 INDELs that were strongly associated with the five comb traits. A remarkable region spanning from 29.6 to 31.4 Mb on chromosome 6 was found to be significantly associated with comb traits in both SNP- and INDEL-based GWAS. In this region, two lead SNPs (6:30,354,876 for CW and CT and 6:30,264,318 for CL) and one lead INDEL (a deletion from 30,376,404 to 30,376,405 bp for CL and CT) were identified. Additionally, two genes were identified as potential candidates for comb development. The nearby gene fibroblast growth factor receptor 2 (FGFR2)-associated with epithelial cell migration and proliferation-and the gene cytochrome b5 reductase 2 (CYB5R2)-identified on chromosome 5 from INDEL-based GWAS-are significantly correlated with collagen maturation. The findings of this study could provide promising genes and biomarkers to accelerate genetic improvement of comb development based on molecular marker-assisted breeding in Nandan-Yao chickens.

Transcriptome analysis reveals key genes and pathways associated with egg production in Nandan-Yao domestic chicken.

BACKGROUND: Egg production is a very important economic trait in chicken breeding, but its molecular mechanism is unclear until now. Nandan-Yao chicken (Gallus gallus domesticus) is a native breed in Guangxi province, China, which is famous for good meet quality, but with low egg production. METHODS: To explore the molecular regulation related to egg production, high egg production (HEP) and low egg production (LEP) were divided according to the total egg number at 55 weeks, and the concentration of serum sex hormones was tested to evaluate the physiological function of ovary and uterus. RNA sequencing (RNA-Seq) was used to explore the transcriptome from the ovary and uterus of Nandan-Yao chicken. RESULTS: The levels of serum sex hormone showed that concentrations of estradiol (E2), follicle-stimulating hormone (FSH), and luteotropic hormone (LH) were significantly higher in HEP than those in LEP (P < 0.01), while the concentration of testosterone (T) was significantly lower in HEP (P < 0.01). RNA-Seq analysis identified 901 and 2763 differentially expressed genes (DEGs) in ovary and uterus, respectively. Enrichment analysis showed that DEGs were significantly involved in the regulation of tight junction in the ovary (P < 0.05), while in uterus, DEGs were mainly enriched in the phagosome, ECM-receptor interaction, cell adhesion molecules (CAMs), focal adhesion, cardiac muscle contraction, cytokine-cytokine receptor interaction, and the regulation of MAPK signaling pathway (P < 0.05). Protein network interaction and function analyses revealed that FN1, FGF7, SOX2 identified from the ovary, and UQCRH, COX5A, FN1 from the uterus might be key candidate genes for egg production in Nandan-Yao chicken. CONCLUSIONS: Our study provided key candidate genes and pathways involved in the egg-laying process of Nandan-Yao chicken and could help to further understand the molecular mechanisms of chicken reproduction.

Metagenomic analysis reveals linkages between cecal microbiota and feed efficiency in Xiayan chickens.

The cecal microbiota plays a critical role in energy harvest and nutrient digestion, influencing intestinal health and the performance of chickens. Feed efficiency (FE) is essential for improving economic efficiency and saving social resources in chicken production and may be affected by the cecal microbiota. Therefore, to investigate the composition and functional capacity of cecum microbes related to FE in Xiayan chicken, an indigenous breed in Guangxi province, metagenome sequencing was performed on chicken cecal contents. 173 male and 167 female chickens were divided into high and low FE groups according to the residual feed intake. The cecal microbial genome was extracted and sequenced. The results showed that the genera Bacteroides, Prevotella, and Alistipes were the 3 most abundant in each cecal microbiome. The linear discriminant analysis effect size revealed 6 potential biomarkers in male and 14 in female chickens. Notably, the relative abundance of Lactobacillus in the high FE group was higher than that of the low FE group both in the male and female chickens, and the species Limosilactobacillus oris has a higher score in the high FE group of male chickens. In contrast, some potentially pathogenic microorganisms such as Campylobacter avium in females and Helicobacter pullorum in males were enriched in the low FE group. Predictive functional analysis showed that the high FE group in male chickens had a greater ability of xenobiotics biodegradation and metabolism and signaling molecules and interaction. In addition, the host sex was found to exert effects on the cecal microbial composition and function associated with FE. These results increased our understanding of the cecal microbial composition and identified many potential biomarkers related to FE, which may be used to improve the FE of the chickens.

Analysis of Population Structure and Differentially Selected Regions in Guangxi Native Breeds by Restriction Site Associated with DNA Sequencing.

Guangxi indigenous chicken breeds play a very important role in promoting the high-quality development of the broiler industry in China. However, studies on genomic information of Guangxi indigenous chicken to date remain poorly explored. To decipher the population genetic structure and differentially selected regions (DSRs) in Guangxi indigenous chickens, we dug into numerous SNPs from seven Guangxi native chickens (GX) by employing the restriction site associated with DNA sequencing (RAD-seq) technology. Another three breeds, Cobb, White Leghorn, and Chahua (CH) chicken, were used as a control. After quality control, a total of 185,117 autosomal SNPs were kept for further analysis. The results showed a significant difference in population structure, and the control breeds were distinctly separate from the Guangxi native breeds, which was also strongly supported by the phylogenetic tree. Distribution of FST indicated that there were three SNPs with big genetic differentiation (FST value all reach to 0. 9427) in GX vs. CH group, which were located on chr1-96,859,720,chr4-86,139,601, and chr12-8,128,322, respectively. Besides, we identified 717 DSRs associated with 882 genes in GX vs. Cobb group, 769 DSRs with 476 genes in GX vs. Leghorn group, and 556 DSRs with 779 genes in GX vs. CH group. GO enrichment showed that there were two significant terms, namely GPI-linked ephrin receptor activity and BMP receptor binding, which were enriched in GX vs. Leghorn group. In conclusion, this study suggests that Guangxi native chickens have a great differentiation with Cobb and Leghorn. Our findings would be beneficial to fully evaluate the genomic information on Guangxi native chicken and facilitate the application of these resources in chicken breeding.

Adipose-derived stem cells are an effective cell candidate for treatment of heart failure: an MR imaging study of rat hearts.

This study assessed the potential therapeutic efficacy of adipose-derived stem cells (ASCs) on infarcted hearts. Myocardial infarction was induced in rat hearts by occlusion of the left anterior descending artery (LAD). One week after LAD occlusion, the rats were divided into three groups and subjected to transplantation of ASCs or transplantation of cell culture medium (CCM) or remained untreated. During a 1-mo recovery period, magnetic resonance imaging showed that the ASC-treated hearts had a significantly greater left ventricular (LV) ejection fraction and LV wall thickening than did the CCM-treated and untreated hearts. The capillary density in infarct border zone was significantly higher in the ASC-treated hearts than in the CCM-treated and untreated hearts. However, only 0.5% of the ASCs recovered from the ASC-treated hearts were stained positive for cardiac-specific fibril proteins. It was also found that ASCs under a normal culture condition secreted three cardiac protective growth factors: vascular endothelial growth factor, hepatocyte growth factor, and insulin-like growth factor-1. Results of this study suggest that ASCs were able to improve cardiac function of infarcted rat hearts. Paracrine effect may be the mechanism underlying the improved cardiac function and increased capillary density.

Characterization and potential function of a novel pre-implantation embryo-specific RING finger protein: TRIML1.

Members of the super-class of zinc finger proteins are key regulators in early embryogenesis. Utilizing in silico mining of EST Databases for pre-implantation Embryo-Specific Zinc Finger Protein Genes, we characterized a novel zygotic mouse gene-tripartite motif family-like 1 (TRIML1), which expresses in embryo before implantation. Knocking down of TRIML1 resulted in the fewer cell number of blastocysts and failture to give rise to neonates after embryo transfer. The binding partner of TRIML1, Ubiquitin-specific protease 5 (USP5), was identified by yeast two-hybrid screening assay. The interaction was confirmed by GST pull-down and coimmunoprecipitation analysis. The role of TRIML1 in ubiquitin pathway during the development stage of mouse blastocyst was further discussed.

Superparamagnetic iron oxide does not affect the viability and function of adipose-derived stem cells, and superparamagnetic iron oxide-enhanced magnetic resonance imaging identifies viable cells.

The objectives of this study were (1) to determine whether superparamagnetic iron oxide (SPIO) affects viability, transdifferentiation potential and cell-factor secretion of adipose-derived stem cells (ASCs); and (2) to determine whether SPIO-enhanced magnetic resonance (MR) imaging highlights living stem cells. Rat ASCs were incubated in SPIO-containing cell culture medium for 2 days. The SPIO-treated ASCs were then subjected to adipogenic, osteogenic and myogenic transdifferentiation. Expression of vascular endothelial growth factor, hepatocyte growth factor and insulin-like growth factor 1 by the SPIO-treated ASCs was measured using reverse transcription polymerase chain reaction. Cell viability was assessed using trypan blue stain. For in vivo experiments, SPIO-labeled ASCs were injected into 10 rat hearts. The hearts were monitored using MRI. We found that survival rate of the ASCs cultured in the SPIO-containing medium was very high (97-99%). The SPIO-treated ASCs continued to express specific markers for the three types of transdifferentiation. Expression of the cell factors by the ASCs was not affected by SPIO. Signal voids on MR images were associated with the living SPIO-labeled ASCs in the rat hearts. We conclude that SPIO does not affect viability, transdifferentiation potential or cell-factor secretion of ASCs. MRI mainly highlights living SPIO-labeled stem cells.

Isolation, characterization, and SREBP1 functional analysis of mammary epithelial cell in buffalo.

Compared to cow milk, buffalo milk contains more protein, fat, and vitamin. Buffalo milk is an ideal food in human life. Sterol regulatory element-binding protein 1 (SREBP1), an important transcription factor, regulates the expression and activity of enzyme and protein involved in milk fat synthesis to influence on the synthesis and secretion of triglyceride in mammary epithelial cells. In the present study, we successfully isolated buffalo mammary epithelial cell by using enzymatic digestion, and then described the growth characteristics and expression characteristics of mammary epithelial cells. Moreover, we cloned the SREBP1 gene from total RNA isolated from milk fat globule and analyzed the function of the SREBP1 gene. After infected with shRNA-SREBP1 lentiviral particle and treated with fatty acid, the expression trend of ACACA, FABP3, FAS, SCD, ERK1, ERK2, PPARy, and Insigl genes was consistent with the expression trend of SREBP1 gene. These results suggested that SREBP1 gene is a central transcription factor in regulating milk fat synthesis and SREBP1 gene may act on ERK1/ERK2 signaling pathway to regulate the expression of PPARy gene. The current study will provide a theoretical basis for further reveal the molecular mechanism of milk fat synthesis in buffalo mammary epithelial cells. PRACTICAL APPLICATIONS: This study aim to separate and analysis characterization of mammary epithelial cell in buffalo. Compared to cow milk, buffalo milk contains more protein, fat, and vitamin. Buffalo milk is an ideal food in human life. This study will provide a theoretical basis for further research on the molecular mechanism of milk fat synthesis in buffalo mammary epithelial cells.