Single-cell transcriptome analysis of the germ cells and somatic cells during mitotic quiescence stage in goats.

The mitotic quiescence of prospermatogonia is the event known to occur during genesis of the male germline and is tied to the development of the spermatogenic lineage. The regulatory mechanisms and the functional importance of this process have been demonstrated in mice; however, regulation of this process in human and domestic animal is still largely unknown. In this study, we employed single-cell RNA sequencing to identify transcriptional signatures of prospermatogonia and major somatic cell types in testes of goats at E85, E105, and E125. We identified both common and specific Gene Ontology categories, transcription factor regulatory networks, and cell-cell interactions in cell types from goat testis. We also analyzed the transcriptional dynamic changes in prospermatogonia, Sertoli cells, Leydig cells, and interstitial cells. Our datasets provide a useful resource for the study of domestic animal germline development.

Hypoxia Treatment of Adipose Mesenchymal Stem Cells Promotes the Growth of Dermal Papilla Cells via HIF-1α and ERK1/2 Signaling Pathways.

Dermal papilla cells (DPCs) cultured in vitro induce hair follicle formation. Using a hypoxic microenvironment to culture adipose mesenchymal stem cells (ADSCs) can promote hair follicle growth. However, the exact molecular mechanisms underlying this process remain unclear. In this study, ADSCs and DPCs from Arbas Cashmere goats were used. A hypoxic microenvironment promoted the proliferation of ADSCs and increased the pluripotency of ADSCs. The growth factors vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and platelet-derived growth factor (PDGF) were upregulated in ADSCs in the hypoxia-conditioned medium (Hypo-cm). Hypo-cm also enhanced the ability of DPCs to induce hair follicle formation. Inhibitors of the ERK1/2 signaling pathway caused the expressions of growth factors that increased in hypoxic microenvironments to decrease; moreover, hypoxia-inducible factor-1α (HIF-1α) increased the expression levels of VEGF, bFGF, and PDGF and inhibited the expression of bone morphogenic protein 7 (BMP7). In conclusion, these findings improve the theoretical basis for the development of gene therapy drugs for the treatment of alopecia areata and hair thinning.

Neural, adipocyte and hepatic differentiation potential of primary and secondary hair follicle stem cells isolated from Arbas Cashmere goats.

BACKGROUND: Arbas Cashmere goats are excellent domestic breeds with high yields of wool and cashmere. Their wool and cashmere can bring huge benefits to the livestock industry. Our studies intend to more fully understand the biological characteristics of hair follicle stem cells (HFSCs) in order to further explore the mechanisms of wool and cashmere regular regeneration. And they have been increasingly considered as promising multipotent cells in regenerative medicine because of their capacity to self-renew and differentiate. However, many aspects of the specific growth characteristics and differentiation ability of HFSCs remain unknown. This study aimed to further explore the growth characteristics and pluripotency of primary hair follicle stem cells (PHFSCs) and secondary hair follicle stem cells (SHFCs). RESULTS: We obtained PHFSCs and SHFSCs from Arbas Cashmere goats using combined isolation and purification methods. The proliferation and vitality of the two types of HFSCs, as well as the growth patterns, were examined. HFSC-specific markers and genes related to pluripotency, were subsequently identified. The PHFSCs and SHFSCs of Arbas Cashmere goat have a typical cobblestone morphology. Moreover, the PHFSCs and SHFSCs express HFSC surface markers, including CD34, K14, K15, K19 and LGR5. We also identified pluripotency-associated gene expression, including SOX2, OCT4 and SOX9, in PHFSCs and SHFSCs. Finally, PHFSCs and SHFSCs displayed multipotent abilities. PHFSCs and SHFSCs can be directed to differentiate into adipocyte-like, neural-like, and hepatocyte-like cells. CONCLUSIONS: In conclusion, this study confirmed that the biological characteristics and differentiation potential of PHFSCs and SHFSCs from Arbas Cashmere goats. These findings broaden and refine our knowledge of types and characteristics of adult stem cells.

Multiple potential roles of thymosin ß4 in the growth and development of hair follicles.

The hair follicle (HF) is an important mini-organ of the skin, composed of many types of cells. Dermal papilla cells are important signalling components that guide the proliferation, upward migration and differentiation of HF stem cell progenitor cells to form other types of HF cells. Thymosin ß4 (Tß4), a major actin-sequestering protein, is involved in various cellular responses and has recently been shown to play key roles in HF growth and development. Endogenous Tß4 can activate the mouse HF cycle transition and affect HF growth and development by promoting the migration and differentiation of HF stem cells and their progeny. In addition, exogenous Tß4 increases the rate of hair growth in mice and promotes cashmere production by increasing the number of secondary HFs (hair follicles) in cashmere goats. However, the molecular mechanisms through which Tß4 promotes HF growth and development have rarely been reported. Herein, we review the functions and mechanisms of Tß4 in HF growth and development and describe the endogenous and exogenous actions of Tß4 in HFs to provide insights into the roles of Tß4 in HF growth and development.

MiR-455-5p monitors myotube morphogenesis by targeting mylip.

As a posttranscriptional regulatory factor, microRNA (miRNA) plays an important role in the formation of myotubes. However, little is known about the mechanism of miRNA regulating myotube morphogenesis. Here, we aimed to characterize the function of miR-455-5p in myotube morphogenesis by inducing differentiation in C2C12 myoblasts containing murine Mylip fragments with the miR-455-5p target sequence. We found that miR-455-5p overexpression promoted the differentiation and hypertrophy of myotubes, while miR-455-5p inhibition led to the failure of myotube differentiation and formation of short myotubes. Furthermore, we demonstrated that miR-455-5p directly targeted the Mylip 3'-untranslated region, which plays a key role in monitoring myotube morphogenesis. Interestingly, the expression and function of Mylip were opposite to those of miR-455-5p during myogenesis. Our data uncovered novel miR-455-5p targets and established a functional link between Mylip and myotube morphogenesis. Understanding the involvement of Mylip in myotube morphogenesis provides insight into the function of the gene regulatory network.

Identifying new compounds with potential pharmaceutical and physiological activity in Areca catechu and Areca triandra via a non-targeted metabolomic approach.

INTRODUCTION: The fruits of Areca catechu, also called areca nuts, are widely used as popular masticatory and traditional herbal medicine in Asia. Besides arecoline and related alkaloids, limited information is available about further primary and secondary metabolites and their potential biological activities. OBJECTIVE: Here we aimed to further enhance our knowledge on phytochemical profiles of A. catechu and Areca triandra fruits. We intended to comprehensively identify metabolites in A. catechu and A. triandra fruits. METHODOLOGY: Metabolites were identified by ultra-performance liquid chromatography triple-quadrupole tandem mass spectrometry (UPLC-MS/MS). The occurrence of 12 selected bioactive compounds in 4 different developmental stages of A. catechu and A. triandra was quantified by LC-MS/MS. RESULTS: A total of 791 metabolites was identified. Of these, 115 metabolites could successfully be mapped to 44 Kyoto Encyclopedia of Genes and Genomes metabolic pathways, and 154 metabolites occurred at significantly different levels in A. catechu compared to A. triandra. Several components with known biological activities were identified for the first time in A. catechu and A. triandra. The abundance of many of these new components was similar in A. catechu and A. triandra, but significantly different between the pericarp and the seeds of A. catechu fruits. CONCLUSIONS: Metabolic profiles indicate that fruits of the Areca species compared here have similar primary and secondary metabolites. Our findings provide new insights into A. catechu and A. triandra as valuable sources for traditional medicine and they pave the way for further studies to potentially improve the underlying pharmaceutical and physiological effects.

HOXC10 promotes proliferation and attenuates lipid accumulation of sheep bone marrow mesenchymal stem cells.

Homeodomain-containing gene C10 (HOXC10), known to regulate cell differentiation and proliferation, is a key negative regulator in the browning of white adipose tissue in mice. Sheep is an important farm animal that provides meat for human consumption, with fat content being an important meat quality determinant; however, there is no report about the role of HOXC10 in sheep adipocytes or adipogenesis. In this study, we investigated the effect of HOXC10 on proliferation and adipogenic differentiation in sheep bone marrow mesenchymal stem cells (sBMSCs). In sBMSCs, HOXC10 overexpression promoted cell proliferation and upregulated the expression of p-PI3K, p-AKT, p-p70S6K, p-MEK, and p-ERK, whereas HOXC10 knockdown was associated with the opposite effects. These results suggested that HOXC10 may promote cell proliferation by activating the MEK/ERK and PI3K/AKT/mTOR/p70S6K signaling pathways. In addition, we found that HOXC10 expression was negatively associated with lipid accumulation in adipogenic-differentiated sBMSCs. HOXC10 overexpression in sBMSCs significantly decreased lipid droplet accumulation and suppressed the expression of adipogenic-specific genes, including ACC, LPL, PPARG, and FABP4, while HOXC10 knockdown was associated with the opposite effects. Furthermore, our study suggested a new regulatory mechanism of the effect of HOXC10 on lipid accumulation and metabolism; HOXC10 may negatively regulate lipid accumulation in adipogenic-differentiated sBMSCs, at least in part, by suppressing LPL expression. Overall, our research not only contributes to a better understanding of the mechanism of lipid accumulation and metabolism in sheep, but also shed light on meat quality control in the future.

Dlgap1 negatively regulates browning of white fat cells through effects on cell proliferation and apoptosis.

BACKGROUND: Obesity is a metabolic imbalance characterized by excessive deposition of white fat. The browning of white fat can effectively treat obesity and related diseases. Although Dlgap1 (Discs, Large (Drosophila) Homolog-Associated Protein 1) is suspected to have an effect on this process, no empirical evidence is available. METHODS: To understand the role of Dlgap1, we cultured white and brown fat cells, then performed overexpression and knockout experiments. RESULTS: We found that Dlgap1 overexpression in brown adipocytes inhibits brown-fat-related gene expression, promotes white-fat-related genes, while also increasing brown-adipocyte proliferation and apoptosis. However, the gene overexpression has no effect on brown adipocyte maturation. Knocking out Dlgap1 in white fat cells promotes the expression and inhibition of brown-fat-related and white-fat-related genes, respectively. Additionally, the knockout inhibits white fat cell proliferation and apoptosis, while also promoting their maturation. CONCLUSIONS: Dlgap1 negatively regulates the browning of white adipocytes by influencing cell proliferation and apoptosis.

Thymosin ß4 Identified by Transcriptomic Analysis from HF Anagen to Telogen Promotes Proliferation of SHF-DPCs in Albas Cashmere Goat.

Increasing cashmere yield is one of the important goals of cashmere goat breeding. To achieve this goal, we screened the key genes that can improve cashmere performance. In this study, we used the RNA raw datasets of the skin and dermal papilla cells of secondary hair follicle (SHF-DPCs) samples of hair follicle (HF) anagen and telogen of Albas cashmere goats and identified a set of significant differentially expressed genes (DEGs). To explore potential associations between gene sets and SHF growth features and to identify candidate genes, we detected functional enrichment and constructed protein-protein interaction (PPI) networks. Through comprehensive analysis, we selected Thymosin ß4 (Tß4), Rho GTPase activating protein 6 (ARHGAP6), ADAM metallopeptidase with thrombospondin type 1 motif 15, (ADAMTS15), Chordin (CHRD), and SPARC (Osteonectin), cwcv and kazal-like domains proteoglycan 1 (SPOCK1) as candidate genes. Gene set enrichment analysis (GSEA) for these genes revealed Tß4 and ARHGAP6 have a close association with the growth and development of SHF-DPCs. However, the expression of Tß4 in the anagen was higher than that in the telogen, so we finally chose Tß4 as the ultimate research object. Overexpressing Tß4 promoted and silencing Tß4 inhibited the proliferation of SHF-DPCs. These findings suggest that Tß4 can promote the growth and development of SHF-DPCs and indicate that this molecule may be a valuable target for increasing cashmere production.

Prevalence of human papillomavirus infection in women in the Autonomous Region of Inner Mongolia: A population-based study of a Chinese ethnic minority.

Human papillomavirus (HPV) is one of the most common sexually transmitted infectious pathogens. Persistent infection has been linked to cancer development, in particular to cervical cancer. This study aims to investigate the epidemiology of HPV infection in women in Inner Mongolia of China and to dissect the disparities between the Han and Mongolian ethnic populations. Cervical cell samples from 5655 women (17-68 years old) were collected during routine gynecologic examination. HPV infection was established using the HPV GenoArray kit detecting 21 HPV genotypes. The overall HPV prevalence was 14.5%. HPV16 (5.0%), HPV58 (2.2%), and HPV52 (1.5%) are the most common genotypes. Of the 21 genotypes investigated, high-risk HPV genotypes dominate in all age groups. HPV16 and HPV58 are the most common genotypes in patients with cervical lesions. HPV prevalence among Han women is 11.5% and the most common genotypes are HPV16 (4%) and HPV58 (2.1%). HPV prevalence is significantly higher in Mongolian women (32.6%), with the most common genotypes being HPV16 (10.7%), HPV31 (7.1%), and HPV52 (4.3%). The multiple infection rate in Mongolian participants (14.9%) is also higher than that of Han participants (4.3%). Urbanization, the number of sex partners, and PAP history appear as risk factors for HPV infection in Han, but not in Mongolian participants. HPV infection is highly prevalent in women in Inner Mongolia, China. HPV16 remains the most common genotype in this area. However, there are clear ethnical disparities in respect to the HPV epidemiology between the Han and Mongolian population.

Whole-genome resequencing of Ujumqin sheep to investigate the determinants of the multi-vertebral trait.

The Ujumqin sheep is one of the most profitable breeds in China, with unique multi-vertebral characteristics. We performed high-throughput genome resequencing of five multi-vertebral and three non-multi-vertebral sheep in an Ujumqin population. We identified the genomic regions that correlated with the germplasm characteristics to establish the cause of the "multi-vertebral" phenotype in this breed. Sequencing generated a total of 314 952 000 000 bp of raw data. The alignment rate of all the samples was between 98.53% and 99.11%, and the mean depth of coverage relative to the reference genome was between 11.58× and 14.92×. After comparing the differences between the two groups, we identified 21 homozygous single nucleotide polymorphisms (SNPs) in the mutant exons of 14 genes. Nineteen loci of 10 genes contained nonsynonymous mutations, while two loci contained synonymous mutations. Resequencing revealed homozygous mutations comprised of 44 indels located within exons of 19 genes. These indels included 37 frameshift mutations, 6 non-frameshift mutations, and 1 stopgain single nucleotide variation (SNV). Finally, comparisons of genotypic variations revealed 17 genes with homozygous mutations in their coding regions, 5 of which have previously been associated with vertebral development and the remaining 12 genes were newly identified in this study.

N-Glycosylation is required for FDNC5 stabilization and irisin secretion.

Irisin, a myokine derived from the extracellular domain of FNDC5, has been shown to mediate thermogenesis of white adipose tissue. Biochemical data have shown that N-glycosylation of FNDC5 is unlikely to affect ligand or receptor activation of irisin. The N-glycosylation of FNDC5 remains poorly understood. In the present study, we analysed N-glycosylation sites of FNDC5 and found that two potential N-glycosylation sites (Asn36 and Asn81) could indeed be occupied by N-glycan. Furthermore we showed that the lack of N-glycosylation decreases the secretion of irisin, which is relevant to the instability of FNDC5 and the deficiency of cleavage of the signal peptide. We also found that the expression level of N-glycosylated FNDC5 was elevated after myoblast differentiation. These findings show that the secretion of irisin is modulated by N-glycosylation, which in turn enhances our understanding of the secretion of glycosylated irisin.

Construction of FGF21 knockout mouse models by the CRISPR/Cas9 system.

Fibroblast growth factors (FGFs) are multifunctional signal molecules between cells, regulating the various physiological functions of the organism. FGF21 is a regulatory factor of the FGF family and has been postulated to play important roles in hair follicle development and hair follicle growth cycle. To evaluate the roles of FGF21, we had established a FGF21 knockout mouse model, using the CRISPR/Cas9 technology. We had constructed a FGF21 targeting vector and microinjected it with Cas9 mRNA and gRNA into fertilized ova of FVB mice. The gRNA was designed to target the exon 1 of the endogenous mouse FGF21 gene. Three lines of Fgf21 -/- mice were obtained from these experiments, and confirmed to harbor Fgf21 -/- genotypes and null expression phenotype, using DNA sequencing, qRT-PCR and Western blotting. FGF21 mRNA and FGF21 protein were not detected in tissues of these Fgf21 -/- mice. Depilation and histochemistry analyses showed that the Fgf21 -/- mice had lower body weight, slower hair regrowth and poorer hair quantities and smaller hair follicles diameters, as compared to WT mice. The Fgf21 -/- mice reported here could provide a useful genetic model for future studies of FGF21 functions in hair follicle development and hair follicle growth cycle.

Identification and characterization of an endoplasmic reticulum localization motif.

Sorting motifs are involved in the transport of diverse proteins. In the present study, we identified a hydrophobic peptide (WRPWRNFWWSIRVPWRRN) that was able to target enhanced green fluorescent protein- or DsRed2-enriched vesicular-like sub-compartments of the endoplasmic reticulum (ER). Analysis of mutation constructs revealed that the sequence WRPWRNFWW was responsible for the ER-targeting activity, and the arginine residue of the peptide is a critical determinant of ER localization. Results from co-immunoprecipitation, glutathione S-transferase pull-down, liquid chromatography-tandem mass spectrometry, and western blotting analyses demonstrated that this motif could bind with the γ2-COP subcomplex of coat protein complex I (COPI), which is involved in the retrieval and transport of ER-resident proteins from the Golgi apparatus to the ER. Overall, we report a new hydrophobic peptide that possesses an arginine-based ER localization motif, which can help elucidate the mechanisms of ER sorting mediated by COPI.

Role of thymosin beta 4 in hair growth.

Although thymosin beta 4 (Tß4) is known to play a role in hair growth, its mechanism of action is unclear. We examined the levels of key genes in a Tß4 epidermal-specific over-expressing mouse model and Tß4 global knockout mouse model to explore how Tß4 affects hair growth. By depilation and histological examination of the skin, we confirmed the effect of Tß4 on hair growth, the number of hair shafts and hair follicle (HF) structure. The mRNA and protein expression of several genes involved in hair growth were detected by real-time PCR and western blotting, respectively. Changes in the expression of ß-catenin and Lef-1, the two key molecules in the Wnt signaling pathway, were similar to the changes observed in Tß4 expression. We also found that compared to the control mice, the mRNA and protein expression of MMP-2 and VEGF were increased in the Tß4 over-expressing mice, while the level of E-cadherin (E-cad) remained the same. Further, in the Tß4 global knockout mice, the mRNA and protein levels of MMP-2 and VEGF decreased dramatically and the level of E-cad was stable. Based on the above results, we believe that Tß4 may regulate the levels of VEGF and MMP-2 via the Wnt/ß-catenin/Lef-1 signaling pathway to influence the growth of blood vessels around HFs and to activate cell migration. Tß4 may have potential for the treatment of hair growth problems in adults, and its effects should be further confirmed in future studies.

The mammalian target of rapamycin signaling pathway regulates myocyte enhancer factor-2C phosphorylation levels through integrin-linked kinase in goat skeletal muscle satellite cells.

Mammalian target of rapamycin (mTOR) signaling pathway plays a key role in muscle development and is involved in multiple intracellular signaling pathways. Myocyte enhancer factor-2 (MEF2) regulates muscle cell proliferation and differentiation. However, how the mTOR signaling pathway regulates MEF2 activity remains unclear. We isolated goat skeletal muscle satellite cells (gSSCs) as model cells to explore mTOR signaling pathway regulation of MEF2C. We inhibited mTOR activity in gSSCs with PP242 and found that MEF2C phosphorylation was decreased and that muscle creatine kinase (MCK) expression was suppressed. Subsequently, we detected integrin-linked kinase (ILK) using MEF2C coimmunoprecipitation; ILK and MEF2C were colocalized in the gSSCs. We found that inhibiting mTOR activity increased ILK phosphorylation levels and that inhibiting ILK activity with Cpd 22 and knocking down ILK with small interfering RNA increased MEF2C phosphorylation and MCK expression. In the presence of Cpd 22, mTOR activity inhibition did not affect MEF2C phosphorylation. Moreover, ILK dephosphorylated MEF2C in vitro. These results suggest that the mTOR signaling pathway regulates MEF2C positively and regulates ILK negatively and that ILK regulates MEF2C negatively. It appears that the mTOR signaling pathway regulates MEF2C through ILK, further regulating the expression of muscle-related genes in gSSCs.

[Bone marrow mesenchymal stem cells to repair the reproductive system of male azoospermia rats].

OBJECTIVE: To study the ability of bone marrow mesenchymal stem cells (BMSCs) to repair the internal environment of the testis in male azoospermia rats. METHODS: We established azoospermia models in 22 six-week-old male SD rats by intraperitoneal injection of busulfan at 20 mg per kg body weight. We transplanted allogeneic rat BMSCs (rBMSCs) into the testicular seminiferous tubules of the model rats and, 30 days after transplantation, observed the composition and structure of the seminiferous tubular cells by HE staining and detected the expressions of CD44, CD106, and c-kit in the rBMSCs by immunohistochemistry. RESULTS: The number of epididymal sperm was significantly reduced in the model rats as compared with the normal controls (P < 0.01). CD44 and CD106, but not c-kit, were expressed in the isolated rBMSCs. At 30 days after transplantation of rBMSCs, lots of new cells were observed in the seminiferous tubules, some expressing CD106 and some expressing the germ cell surface marker c-kit. CONCLUSION: BMSCs can transdifferentiate into germ cells and repair the damaged seminiferous tubules of sterile rats.

Transcriptome sequencing reveals differences between anagen and telogen secondary hair follicle-derived dermal papilla cells of the Cashmere goat (Capra hircus).

Dermal papilla is considered the control center of hair follicle growth and hair cycle. The secondary hair follicle (producing cashmere) growth cycle of the Cashmere goat (Capra hircus) is circannual, and each growth phase can be easily distinguished by its long duration. To identify gene expression patterns and differences of the dermal papilla cell (DPC) between the anagen and telogen phases, we established two DPC lines: ana-DPCs (DPCs derived from the anagen secondary hair follicle) and tel-DPCs (DPCs derived from the telogen secondary hair follicle). Compared with the ana-DPCs, the tel-DPCs lost the capacity to form cell aggregates and showed lower cell proliferation rate. Transcriptome sequencing revealed that 825 genes were differentially expressed by at least threefold between the two DPC lines. These genes were significantly enriched in cell cycle control, cell division, and chromosome partitioning from the Eukaryotic Orthologous Groups of proteins (KOG) database and in cell cycle, cell adhesion molecules, cytokine-cytokine receptor interaction, and p53 signaling pathway from the Kyoto Encyclopedia of Gene and Genomes (KEGG) database. Enrichment analyses revealed that in the middle of the telogen the DPCs of secondary hair follicles (SHFs) seemed on the one hand to promote the degeneration of SHFs and cessation of cashmere growth, while on the other hand to resist self-apoptosis and prepare for the regeneration or revivification of fully functional dermal papillae. These findings provide a better understanding of hair follicle growth and will be useful for identification of novel molecules associated with the control of hair growth cycle.

Analysis of the stem cell characteristics of adult stem cells from Arbas white Cashmere goat.

Studies have shown that multipotent adult stem cells possess differentiation characteristics similar to embryonic stem cells and pluripotent stem cells. We aimed to explore these similarities further by examining the expression of the pluripotency and stemness biomarkers, AKP, IL-6, Nanog, Oct-4, Rex-1, Sox-2 and TERT, as well as the triploblastic biomarkers, Sox-1, Myod1 and Gata-6 in adipose-derived stem cells (ADSCs), bone marrow stem cells (BMSCs) and muscle-derived satellite cells (MDSCs). These were isolated from adult Arbas white Cashmere goats and cultured in vitro. Immunocytochemistry, reverse transcription quantitative PCR and Western blotting were used to analyze the protein and mRNA expression of the markers. To investigate the ability of ADSCs, BMSCs and MDSCs to differentiate and cause tumors in vivo they were injected into immunodeficient mice (NOD-SCID). All results were compared to those for mouse embryonic stem cells (mESCs). Immunocytochemistry showed that AKP, IL-6, Nanog, Oct-4, Rex-1 and TERT were expressed in ADSCs, BMSCs and MDSCs, whereas Sox-2 was not. In ADSCs, the expression of IL-6 mRNA was relatively high, followed by Nanog and Oct-4, while Rex-1 and TERT expression were the lowest (P<0.01). In BMSCs, the expression of Rex-1 was relatively high, followed by IL-6, while Oct-4, Nanog and TERT were comparatively low (P<0.01). In MDSCs, the expression of IL-6, Nanog and Oct-4 were relatively high, while TERT was comparatively low (P<0.01). However, no expression of Sox-2 mRNA was detected in any of the three cell lines. The expression of Sox-1, Myod1 and Gata-6 was observed to different degrees in all three cell lines (P<0.01); the expression pattern in MDSCs was different from that in ADSCs and BMSCs. Western blotting indicated that no expression of Sox-2 and Rex-1 protein occurred in ADSCs, BMSCs and MDSCs, while the other five proteins were all expressed to different degrees (P<0.01); the expression pattern was consistent with the mRNA results. In contrast to the mESCs, no teratoma tissue or triploblastic differentiation appendages were formed in the immunodeficient mice after injection of ADSCs, BMSCs and MDSCs. Our results suggest that the three adult goat stem cell types are non-oncogenic and have stemness characteristics similar to embryonic stem cells. Of these, MDSCs were found to exhibit the most ESC-like properties and would make the best candidates for clinical application.

Recombinant Goat VEGF164 Increases Hair Growth by Painting Process on the Skin of Shaved Mouse.

To detect goat vascular endothelial growth factor (VEGF)-mediated regrowth of hair, full-length VEGF164 cDNA was cloned from Inner Mongolia cashmere goat (Capra hircus) into the pET-his prokaryotic expression vector, and the recombinant plasmid was transferred into E. coli BL21 cells. The expression of recombinant 6×his-gVEGF164 protein was induced by 0.5 mM isopropyl thio-ß-D-galactoside at 32°C. Recombinant goat VEGF164 (rgVEGF164) was purified and identi ed by western blot using monoclonal anti-his and anti-VEGF antibodies. The rgVEGF164 was smeared onto the dorsal area of a shaved mouse, and we noted that hair regrowth in this area was faster than in the control group. Thus, rgVEGF164 increases hair growth in mice.