Although the pluripotent and proliferative capacity of embryonic germ (EG) cells is thought to be equivalent to that of embryonic stem (ES) cells, there has been far less attention focused on the potential use of EG cells for applications in developing novel strategies of tissue transplantation in the treatment of degenerative diseases. In this study, EG cells were derived from primordial germ cells (PGCs) of genital ridges of 4-day-old chicken embryos. These cells satisfied the criteria previously used for defining chicken EG cells by using the expression of markers characteristic to ES cells. When injected subcutaneously, chicken EG cells could form teratomas that enable differentiation into a wide range of tissue types of all three primary cell lineages including neural cells, cartilage, forming bone, adipocytes, blood vessels, smooth muscle, and secretory epithelia in the recipients. Furthermore, cells in embryoid bodies (EBs) expressed lineage-specific markers of three germ layers and could be induced to differentiate into more advanced stages of various committed cell types, including dopamine and cholinergic neurons, astrocytes, oligodendrocytes, adipocytes, and hepatocytes, which were demonstrated by immunocytochemical staining or RT-PCR analysis. These findings support the multilineage differentiation capability of chicken pluripotent EG cells, thus confirming the presumption that chicken embryos may be used as a potential model for better understanding the mechanisms of tissue-specific differentiation and regeneration that will help to devise strategies based on the transplantation of stem cell-derived tissues for restoring function to damaged or diseased tissues.
The effect of ginsenosides on proliferation of chicken primordial germ cells (PGCs) was evaluated and involvement of nuclear factor (NF)-kappaB in the signaling pathway was investigated. PGCs were isolated from the genital ridge of 3.5-4 day embryos and cultured in Medium 199 supplemented with 5% FCS and 10 ng/ml LIF. PGCs subcultured on chicken embryonic fibroblast feeder were challenged with ginsenosides alone or in combination with PKC inhibitor H(7) or activator phorbol 12-myristate 13-acetate (PMA) for 24h. Moreover, the translocation of NF-kappaB and degradation level of IkappaBalpha were investigated by Western blot analysis. Results show that PGCs were identified by periodic acid-Schiff, alkaline phosphatase histochemistry as well as c-kit, SSEA-1 and Oct-4 immunocytochemistry. Treatment with ginsenosides at 1-100 microg/ml significantly increased the number and area of PGC colonies in a dose-dependent manner. However, this proliferating effect was obviously attenuated by combined treatment of H(7) (10(-7)-10(-5)M). Similarly, PKC staining of PGC colonies was more intensive after ginsenosides treatment compared with the control group. In addition, treatment with ginsenosides at 1-10 microg/ml stimulated the translocation of NF-kappaB (p65). However, the NF-kappaB translocation and the degradation of IkappaBalpha were significantly blocked by combined treatment with 10(-6)M H(7). These results indicated that ginsenosides promote proliferation of chicken PGCs through activation of PKC-involved NF-kappaB signaling pathway.
Many studies demonstrated that chicken primordial germ cells (PGCs) could maintain undifferentiated state on mouse embryonic fibroblast feeders supplemented with growth factors and cytokines. However, the xenosupport systems may run risk of cross-transfer of animal pathogens from the other animal feeder, matrix to the PGCs, then influencing later transgenic technology. In this study, chicken PGCs were identified by alkaline phosphatase, stage-specific embryonic antigen-1 and Oct-4 immunocytochemical stainings. Three different homologous somatic cell feeder layers (chicken embryonic fibroblast feeder layer, CEF; embryonic skeletal myoblast feeder layer; follicular granulosa cell feeder layer) were used to support growth and proliferation of PGCs to find a better supporting culture system. In addition, the effects of fetal calf serum (FCS), leukemia inhibitory factor (LIF) and the combination of insulin, transferring and selenite (ITS) on PGC proliferation were compared. Results showed that CEF was the best supporter for PGC growth and proliferation, which was verified by 5-bromo-2'-deoxyuridine incorporation stain. FCS alone or in combination with LIF could significantly promote PGC proliferation in the presence of CEF in ITS medium. This study will contribute to providing a safer supporting system for chicken PGC amplification in vitro, and may be applied in transgenic chicken production and transplantation therapy.
Three polymorphisms were identified in the sixth intron of the CAST gene by PCR-RFLP using enzymes Msp I, Hinf I and Rsa I in 45 Jinpi F2 pigs. However, only three genotypes AACCEE, BBDDFF, and ABCDEF were detected in those pigs and the genotype frequencies were 0.1778, 0.2222 and 0.6000, respectively. Longissimus dorsi muscles were moved for paraffin serial sections and stained with hematoxylin-eosin or myosin heavy chain (MHC) by immunohistochemistry respectively. Fiber cross-sectional area, fiber density, fiber diameter, the rate of MHC type I fiber and the carcass characteristics were recorded. Correlation analysis showed that the skeletal muscle fiber area and the eye muscle area of the BBDDFF individual were significantly higher than those of the ABCDEF individual (P < 0.05).
neutrophils are crucial in the process of gout flare and remission. The signal transduction pathway of chemokine plays a vital role in the chemotaxis and activation of neutrophils. CXCR2 gene knocked out can avoid the acute neutrophilic inflammation stimulated by monosodium urate (MSu) crystals in mice. To investigate the relationship among CXCR1 rs2234671, CXCR2 rs1126579, and rs2230054 polymorphisms with gout arthritis flare in the Chinese han male population, a case-control study was carried out in 412 gout patients and 508 gout-free individuals. TaqMan probes fluorescence real-time polymerase chain reaction (PCR) was used to genotype CXCR1 rs2234671, CXCR2 rs1126579, and rs2230054 SnPs. There was a clear link between CXCR2 rs2230054 T included genotypic and T allelic frequencies and gout cases (c 2 = 9.286, p = 0.002 by genotype, c 2 = 8.639, p = 0.003 by allele), while no significant differences were observed between the gouty arthritis group and the control group in CXCR1 rs2234671 and CXCR2 rs1126579 genotypic and allelic frequencies. Multivariate logistic regression analysis showed that the T genotype included in rs2230054 can decrease the risk of gouty arthritis (adjusted OR = 0.47; 95% Ci: 0.31-0.74) compared with the CC genotype. Our study might suggest that rs2230054 in CXCR2 is associated with susceptibility to gout in Chinese males.
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