Serotonin neurons derived from rhesus monkey embryonic stem cells: similarities to CNS serotonin neurons
| Autor: | Hung-Chih Kuo, Don P. Wolf, Arubala P. Reddy, Francis K.Y. Pau, Nick Z. Lu, Cynthia L. Bethea, Nurgul C. Salli, Ugur Salli |
|---|---|
| Rok vydání: | 2004 |
| Předmět: |
Central Nervous System
Receptors Steroid Serotonin medicine.medical_specialty Cellular differentiation Cell Culture Techniques Fluorescent Antibody Technique Embryoid body Serotonergic Cell Line Developmental Neuroscience Internal medicine medicine Animals Cell Lineage RNA Messenger Neurons biology Reverse Transcriptase Polymerase Chain Reaction Gene Expression Profiling Stem Cells Cell Differentiation Tryptophan hydroxylase Antigens Differentiation Macaca mulatta Molecular biology Extracellular Matrix Endocrinology medicine.anatomical_structure Neurology biology.protein Neuron NeuN Stem cell |
| Zdroj: | Experimental Neurology. 188:351-364 |
| ISSN: | 0014-4886 |
| Popis: | We sought an in vitro primate model for serotonin neurons. Rhesus monkey embryonic stem (ES) cell colonies were isolated and differentiated into embryoid bodies (EBs), then transferred to serum-free medium with 1% insulin-transferrin-selenium for 7 days to induce neural precursor cell (NPC) formation. NPCs were cultured in medium with 1% N-2 neural supplement and human fibroblast growth factor 2 (FGF2, 10 ng/ml) for 7 days to stimulate cell proliferation. Lastly, NPCs were dispersed into single cells and cultured without FGF2 for another 7 days to obtain terminal differentiation. Terminal cells were characterized for neuronal and serotonergic markers. Over 95% of the NPCs were immunopositive for nestin and Musashi1. Terminally differentiated cells appeared in both small and large morphologies. Most (>95%) of the mature cells (both small and large) were immunopositive for neuron-specific nuclear protein (NeuN), synaptophysin, microtubule-associated protein (MAP2C), Tau-1, neurofilament 160 (NF-160), beta-tubulin (TujIII), tryptophan hydroxylase (TPH), serotonin, the serotonin reuptake transporter (SERT), estrogen receptor-beta (ERbeta), and progestin receptor (PR), but not estrogen receptor-alpha (ERalpha). Less than 2-3% of cells were positive for tyrosine hydroxylase (TH). Reverse transcriptase polymerase chain reaction (RT-PCR) detected mRNA transcripts for TPH-1, TPH-2, SERT, 5-HT1A-autoreceptor, ERbeta, and PR in the differentiated population. A low level of expression of ERalpha mRNA was also detected. Quantitative RT-PCR indicated that the relative abundance of TPH-2 mRNA was greater than TPH-1 mRNA. Serotonin as measured by ELISA increased 3-fold in the mature stage compared to the selection and expansion stages. In summary, a remarkably high percentage of cells derived from monkey ES cells exhibited neuronal plus serotonergic markers as well as nuclear steroid receptors similar to primate CNS serotonin neurons, suggesting that these cells may serve as a useful primate model for serotonergic neurons. |
| Databáze: | OpenAIRE |
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