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5-Hydroxytryptamine (5-HT) is an inhibitory neurotransmitter widely distributed in mammalian tissues, exerting its effects through binding to various receptors. It plays a crucial role in the proliferation of granulosa cells (GCs) and the development of follicles in female animals, however, its effect on porcine follicle development is not clear. The aim of this study is to investigate the expression of 5-HT and its receptors in various parts of the pig ovary, as well as the effect of 5-HT on porcine follicular development by using ELISA, quantitative real-time PCR (qPCR) and EdU assays. Firstly, we examined the levels of 5-HT and its receptors in porcine ovaries, follicles, and GCs. The findings revealed that the expression of different 5-HT receptors varied among follicles of different sizes. To investigate the relationship between 5-HT and its receptors, we exposed the GCs to 5-HT and found a decrease in 5-HT receptor expression compared to the control group. Subsequently, the treatment of GCs with 0.5 μM, 5 μM, and 50 μM 5-HT showed an increase in the expression of cell cycle-related genes, and EdU results indicated cell proliferation after the 0.5 μM 5-HT treatment. Additionally, the expression of genes involved in E2 synthesis was examined after the treatment of granulosa cells with 0.5 μM 5-HT. The results showed that CYP19A1 and HSP17β1 expression was decreased. These results suggest that 5-HT might affect the development of porcine follicle by promoting the proliferation of GCs and inhibiting the synthesis of estrogen. This provides a new finding for exploring the effect of 5-HT on follicular development, and lays a foundation for further research on the mechanism of 5-HT in follicles.

Objective: To explore the effects and mechanisms of chidamide on the osteogenic differentiation of bone marrow mesenchymal stromal cells (MSC) from myelodysplastic syndromes (MDS)., Methods: MSC were isolated and cultured from bone marrow of MDS patients and healthy donors. CCK-8 assay was used to detect the effects of chidamide on the proliferation of MSC. The effects of chidamide on the activity of histone deacetylase (HDAC) in MSC was measured by a fluorescence assay kit and Western blot. Alkaline phosphatase (ALP) activity was detected on day 3 and calcium nodule formation was observed by Alizarin Red staining on day 21 after osteogenic differentiation. The expression of early and late osteogenic genes was detected on day 7 and day 21, respectively. RT-PCR and Western blot were used to detect the effects of chidamide on mRNA and protein expression of RUNX2 which is the key transcription factor during osteogenesis., Results: As the concentration of chidamide increased, the proliferation of MSC was inhibited. However, at a low concentration (1 μmol/L), chidamide had no significant inhibitory effect on MSC proliferation but significantly inhibited HDAC activity. In MSC from both MDS patients and healthy donors, chidamide (1 μmol/L) significantly increased ALP activity, calcium nodule formation, thereby mRNA expression of osteogenic genes, and restored the reduced osteogenic differentiation ability of MDS-MSC compared to normal MSC. Mechanistic studies showed that the osteogenic-promoting effect of chidamide may be related to the upregulation of RUNX2 ., Conclusion: Chidamide can inhibit HDAC activity in MSC, upregulate the expression of the osteogenic transcription factor RUNX2 , and promote the osteogenic differentiation of MDS-MSC.

Objective: To investigate the inhibitory effect of decitabine (DAC) in various dosages on the proliferention of MDS-RAEB cell line MDS-L and its mechanism., Methods: LC-MS/MS method was used to test the blood DAC concentration of 2 groups of MDS patients being treated with DAC 20 and 15 mg/m 2 ×5 d. In according to the various blood DAC concentration levels, the MDS-L cells were treated with different DAC dosages for 24, 48, 72 and 96 h, respectively. The CCK-8 method was applied to determine the cell proliferation, the flow cytometry was used to analyze the cell cycle and cell apoptosis changes, the P15 INK4B DNA methylation status was measured by methylation specific PCR using EZ DNA Methylation-Gold Kit., Results: The blood DAC concentration of MDS patients treated with DAC 20 mg/m 2 ×5 d was 174.08±80.15(84.7-311) ng/ml, which was significantly higher than 89.87±32.94(43.2-165)ng/ml for the group treated with 15 mg/m 2 ×5 d (P=0.014). DAC could notably inhibit the proliferation of MDS-L cells, and the effect was in dose- and- time-dependent manner(r=0.786). However, when DAC concentration was ≥0.1 µg/ml, the proliferation inhibition rates were not significantly different between various dosages. After DAC treatment, MDS-L cells in G 1 phase increased notably, while cells in S phase decreased significantly. Also, the P15 INK4B DNA methylation status of MDS-L cells decreased after being treated with DAC for 96 h, but the difference was not significant between various dosages., Conclusion: DAC can significantly suppress MDS-L cell proliferation, block MDS-L cells in G 1 phase and induce the apoptosis at low concentration (0.1-0.2 µg/ml).

Umbilical cord blood mononuclear cells (UCBMC) transplantation may improve hypoxia-induced brain injury in neonatal rats, but the mechanism is unclear. This study examines whether UCBMC promote neural stem cell (NSC) proliferation via the Sonic hedgehog (Shh) signaling pathway. The rats underwent left carotid ligation followed by hypoxic stress. UCBMC were transplanted 24h after hypoxia ischemia (HI), and immunohistochemistry, immmunoblotting, and morphology analyses were performed at different time points after transplantation. Increased numbers of NSCs were observed in the subventrical zone (SVZ) of the HI+UCBMC group, but these increases were attenuated by cyclopamine treatment. There were significant increases in Shh and Gli1 protein levels after transplantation in the HI group treated with UCBMC compared to HI rats treated with phosphate-buffered solution (PBS). Significantly more Gli1(+)DAPI(+) cells were observed in the SVZ of the HI+UCBMC group compared to the HI+PBS and N+UCBMC groups, but few Gli1(+)DAPI(+) cells were found in the SVZ of the HI+cyclopamine+UCBMC group. The HI+UCBMC group had significantly less neuronal loss in the cortex and CA1 sector of the hippocampus compared to the HI+PBS group, but more neuron loss was observed in the HI+cyclopamine+UCBMC group compared to HI+UCBMC. These results indicate that UCBMC may promote NSC proliferation and alleviate brain injury in HI neonatal rats via Shh signaling., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Epidemiological and experimental studies have correlated hyperhomocysteinemia to a range of neurodegenerative conditions, including Alzheimer's disease, stroke, and Parkinson's disease. Although homocysteine-induced apoptosis in neurons has been extensively studied, little information is available regarding the effect of homocysteine on microglia. In this report, we demonstrated that homocysteine promoted proliferation and up-regulated the expression of CD11b (a marker of microglial activation). Consistent with our in vitro results, a significant increase in the number of CD11b-positive microglia was also observed in brain sections of mice with hyperhomocysteinemia. Homocysteine promoted the activity of NAD(P)H oxidases, resulting in the generation of reactive oxygen species. Up-regulation of NAD(P)H oxidase activity by homocysteine appears to be due to its ability to induce the phosphorylation of p47phox through the p38 MAPK pathway. Furthermore, inhibition of reactive oxygen species significantly blocked cellular proliferation and activation in microglia. Since microglial proliferation and activation play an important role in the development of several neurodegenerative disorders, our results reveal a novel role of homocysteine in the pathogenesis of neurodegenerative diseases., (Copyright © 2008 Elsevier Inc. All rights reserved.)

Elevated homocysteine levels are defined as hyperhomocysteinemia (HHcy), a disorder that is associated with cardiovascular and neurodegenerative diseases as well as with hepatic fibrosis. Recent studies have shown that HHcy promotes hepatic injury by increasing oxidative stress. Although homocysteine induces cell cycle arrest in a variety of different cell types, it is not known whether HHcy has a definitive role in hepatocyte proliferation during liver regeneration. In this report, we investigated the effect of homocysteine on liver regeneration. Our results demonstrated that mice with HHcy exhibited an impairment in liver regeneration after partial hepatectomy, as measured by immunohistochemical staining of proliferation cell nuclear antigen and bromodeoxyuridine incorporation. Impaired proliferation was also correlated with reduced cyclin D1 induction and elevated expression levels of both p53 and p21Cip1. In addition, the phosphorylation of Akt, which plays an essential role in normal regeneration responses, was attenuated during the early phases of liver regeneration in HHcy mice. Our results also indicated that the cAMP/protein kinase A pathway mediated the inhibitory effect of homocysteine on liver regeneration. These findings provide evidence that impairment of liver regeneration by HHcy may result in delayed recovery from liver injury induced by homocysteine itself.

Aim: To investigate the sphingosine 1-phosphate (S1P) receptor expression profile in human esophageal cancer cells and the effects of S1P5 on proliferation and migration of human esophageal cancer cells., Methods: S1P receptor expression profile in human esophageal squamous cell carcinoma cell line Eca109 was detected by semi-quantitative reverse transcription polymerase chain reaction. Eca109 cells were stably transfected with S1P5-EGFP or control-EGFP constructs. The relation between the responses of cell proliferation and migration to S1P and S1P5 expression was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide and migration assay, respectively., Results: Both normal human esophageal mucosal epithelium and Eca109 cells expressed S1P1, S1P2, S1P3 and S1P5, respectively. Esophageal mucosal epithelium expressed S1P5 at a higher level than Eca109 cell line. S1P5 over-expressing Eca109 cells displayed spindle cell morphology with elongated and extended filopodia-like projections. The proliferation response of S1P5-transfected Eca109 cells was lower than that of control vector-transfected cells with or without S1P stimulation (P < 0.05 or 0.01). S1P significantly inhibited the migration of S1P5-transfected Eca109 cells (P < 0.001). However, without S1P in transwell lower chamber, the number of migrated S1P5-transfected Eca109 cells was greater than that of control vector-transfected Eca109 cells (P < 0.001)., Conclusion: S1P binding to S1P5 inhibits the proliferation and migration of S1P5-transfected Eca109 cells. Esophageal cancer cells may down-regulate the expression of S1P5 to escape the inhibitory effect.

Homocysteine is an intermediate in sulfur amino acid metabolism, which takes place mainly in the liver. Recent studies have shown that hyperhomocysteinemia in patients and murine models develop hepatic fibrosis. To define mechanisms underlying homocysteine-induced hepatic fibrosis, the effect of homocysteine on hepatic stellate cell (HSC) proliferation was examined. In the present study, homocysteine promoted proliferation in myofibroblastic HSCs. Homocysteine elicited a transient formation of reactive oxygen species (ROS). The initial ROS activated extracellular signal-regulated kinase and p38 mitogen-activated protein kinase, which were involved in the activation of NAD(P)H oxidases and the generation of more ROS. The activation of NAD(P)H oxidases resulted from upregulation of the expression of p22(phox) and the phosphorylation of p47(phox). The ROS derived from NAD(P)H oxidases activated the PI3K/Akt pathway, thus promoting cellular proliferation in HSCs. These findings provide a mechanistic explanation for the development and progression of hepatic fibrosis in hyperhomocysteinemia.

Objective: To explore a new specific therapy of nasopharyngeal carcinoma (NPC) using an anti-nasopharyngeal carcinoma (NPC) monoclonal antibody BAC5 conjugate with Chinese cobra (CT) and iodine-131(131I)., Methods: BAC5 was labeled with 131I by chloramine-T method, CT was labeled with 125I using iodogen method, and BAC5 and 125I-CT were conjugated by SPDP method. The inhibitory effect of the conjugate on cultured human NPC CNE2 cells was observed using MTT assay., Results: The IC50 of 125I-CT-BAC5 conjugate was 9.17x10(-8) mol/L, and that of 131I-BAC5 was 5.83x10(8) Bq/L, and their combined administration showed obvious inhibitory effect on the NPC cells., Conclusion: Both 125I-CT-BAC5 and 131I-BAC5 have obvious inhibition effects against NPC cells, but their combined use shows stronger effects.