The present study was aimed to investigate how the induced pluripotent stem cells (iPSCs) and bone marrow mesenchymal stem cells (BMSCs) differentiate into neuron-like cells under the induction of hippocampal microenvironments and Reelin's regulation. iPSCs or BMSCs were co-cultured with WT (wild type) or genotypic hippocampal slice and cerebral homogenate supernatant, then the stem cells' differentiation under the induction of hippocampal environment was observed by using immunofluorescence technique. In the meantime, stem cells were co-cultured with hippocampal slice and cerebral conditioned medium of reeler (Reelin deletion) mouse respectively. The results showed that both adhesive iPSCs and BMSCs on WT hippocampal slice exhibited lamination of double "C" shape with high density on granular and pyramidal layers. The stem cells could differentiate into neuron-like cells with obvious polarization on WT hippocampal slice. In pyramidal cell layer, the differentiated neuron-like cells were oriented vertically with similar shapes of pyramidal cell in vivo, and the cells within molecule layer were arranged horizontally. In addition, adhesive iPSCs and BMSCs could differentiate into Nestin positive neural stem cells and NeuN positive neurons, respectively, under WT hippocampal microenvironment. On the other hand, under induction of hippocampal microenvironment of reeler mouse, iPSCs and BMSCs differentiation could also be seen, but their lamination was in disorder, and cell polarization was irregular. Moreover, differentiation and polarization of the iPSCs and BMSCs were delayed. These results suggest both iPSCs and BMSCs can differentiate into neuron-like cells under the induction of hippocampal microenvironments. Reelin is involved in the regulation of neuronal differentiation and cell polarization. Without Reelin, the cellular lamination and polarization appear irregular, and the stem cells' differentiation is delayed.
Animals ; Cell Adhesion Molecules, Neuronal ; metabolism ; Cell Differentiation ; Cells, Cultured ; Coculture Techniques ; Culture Media, Conditioned ; Extracellular Matrix Proteins ; metabolism ; Hematopoietic Stem Cells ; cytology ; Hippocampus ; Induced Pluripotent Stem Cells ; cytology ; Mice ; Mice, Inbred C57BL ; Nerve Tissue Proteins ; metabolism ; Neural Stem Cells ; cytology ; Neurons ; cytology ; Serine Endopeptidases ; metabolism

PURPOSE: Cell transplantation of myelin-producing exogenous cells is being extensively explored as a means of remyelinating axons in X-linked adrenoleukodystrophy. We determined whether 3,3',5-Triiodo-L-thyronine (T3) overexpresses the ABCD2 gene in the polysialylated (PSA) form of neural cell adhesion molecule (NCAM)-positive cells and promotes cell proliferation and favors oligodendrocyte lineage differentiation. MATERIALS AND METHODS: PSA-NCAM+ cells from newborn Sprague-Dawley rats were grown for five days on uncoated dishes in defined medium with or without supplementation of basic fibroblast growth factor (bFGF) and/or T3. Then, PSA-NCAM+ spheres were prepared in single cells and transferred to polyornithine/fibronectin-coated glass coverslips for five days to determine the fate of the cells according to the supplementation of these molecules. T3 responsiveness of ABCD2 was analyzed using real-time quantitative polymerase chain reaction, the growth and fate of cells were determined using 5-bromo-2-deoxyuridine incorporation and immunocytochemistry, respectively. RESULTS: Results demonstrated that T3 induces overexpression of the ABCD2 gene in PSA-NCAM+ cells, and can enhance PSA-NCAM+ cell growth in the presence of bFGF, favoring an oligodendrocyte fate. CONCLUSION: These results may provide new insights into investigation of PSA-NCAM+ cells for therapeutic application to X-linked adrenoleukodystrophy.
ATP-Binding Cassette Transporters/*metabolism ; Adrenoleukodystrophy/genetics/*therapy ; Animals ; Animals, Newborn ; Bromodeoxyuridine ; Cell Differentiation ; Fibroblast Growth Factor 2/pharmacology ; Fibronectins/metabolism ; Immunohistochemistry ; Neural Cell Adhesion Molecules/*genetics ; Rats ; Rats, Sprague-Dawley ; Real-Time Polymerase Chain Reaction ; Sialic Acids/metabolism ; Stem Cells ; Thyroid Hormones/*metabolism ; Triiodothyronine/pharmacology

OBJECTIVETo study the effects of astragalan (AG) on the expression of the neural cell adhesion molecule(NCAM) and c-fos of hippocampus CA1 region after the ischemic brain injury in rats.

METHODSOne hundred male Wistar rats (180-220 g) were divided into ten groups randomly, they were sham operated group (SOG, n = 10), three model group(MG-ld, 3d, 7d, n = 10), as well as three low and high dose astragalan treatment groups (L/H-AGTG-1d, 3d, 7d, n = 10), respectively. And then, middle cerebral artery of MG and AGTG were intercepted by operation inducing brain injured. Their cerebral blood vessel were reperfused on 1, 2, 3 d, respectively, after the L/H-AGTG were treated with the AG (5 mg/kg and 15 mg/kg, ip). After neurologic impairment(NIP) was scored, animals were decapitated to take out hippocampus for counting apoptosis , determining the expression of the NCAM and c-fos by immunohistochemistry method and RT-PCR semiquantitative analysis, respectively.

RESULTSThe NIP scores and apoptotic cell of the L-AGTG and H-AGTG were significantly lower than MG (P < 0.05 or P < 0.01). The expression of NCAM and c-fos were significantly higher than the MG (P < 0.05 or P < 0.01).

CONCLUSIONAstragalan could improve significantly neural function of ischemia brain injury in rats,the mechanism concerned probably with blocking or reversing apoptosis of hippocampus by promoting the expression of the NCAM and c-fos of hippocampus CA1 region.

Animals ; Apoptosis ; drug effects ; Astragalus membranaceus ; chemistry ; Brain Ischemia ; metabolism ; pathology ; CA1 Region, Hippocampal ; cytology ; Kaempferols ; pharmacology ; Male ; Neural Cell Adhesion Molecules ; metabolism ; Neurons ; drug effects ; Neuroprotective Agents ; pharmacology ; Polysaccharides ; pharmacology ; Rats ; Rats, Wistar

OBJECTIVETo observe the effects of flutamide (Flu) on the development of testicular germocytes in SD rats, and to establish a rat model for further researches on the maldevelopment of cryptorchidism gonocytes (Go).

METHODSPregnant SD rats were subcutaneously injected with Flu from gestational day (GD) 12 to 21 to establish a model of cryptorchidism. The testes of the newborns were harvested on postnatal day (PD) 1, 10, 20 and 80 for observation of their morphological and histological changes by HE staining and detection of the expression of neural cell adhesion molecules (NCAM) by immunohistochemistry and RT-PCR.

RESULTSFlu induced 43.9% (29/66) of cryptorchidism in the exposed rats. Significant differences were found in the testicular weight and organ coefficient between the Flu and the control groups on PD 20 and 80. Gos remained in the center of seminiferous tubules of the Flu-induced testis on PD 10, and in the center of seminiferous tubules in the cryptorchids' testicular tissues on PD 20 and 80. Immunohistochemistry showed the expression of NCAM on the membrane of the remaining Gos, and RT-PCR revealed significantly up-regulated expression of NCAM mRNA in the Flu-induced testes on PD 10 and 20 as compared with the controls (P < 0.05).

CONCLUSIONA rat model of Flu-induced cryptorchidism with remaining Gos was successfully established, which could be used to study the mechanism and management of the maldevelopment of cryptorchidism gonocytes.

Animals ; Cryptorchidism ; chemically induced ; pathology ; Female ; Flutamide ; adverse effects ; Male ; Neural Cell Adhesion Molecules ; metabolism ; Pregnancy ; Rats ; Rats, Sprague-Dawley ; Testis ; drug effects ; metabolism ; pathology

Combined hepatocellular carcinoma and cholangiocarcinoma (combined HCC-CC) is a rare subtype of primary liver cancer. We investigated the histopathologic features of transitional or intermediate areas in 21 combined HCC-CCs and immunophenotypes using different hepatic progenitor cell markers (CK7, CK19, c-kit, NCAM, and EpCAM). Major histologic findings of transitional or intermediate areas of 21 combined HCC-CCs included strands/trabeculae of small, uniform, oval-shaped cells with scant cytoplasm and hyperchromatic nuclei embedded within an abundant stroma, small cells with an antler-like anastomosing pattern, and solid nests of intermediate hepatocyte-like cells surrounded by small cells in periphery, in order of frequency. The intermediate area of one tumor was composed predominantly of spindle cells arranged in short fascicles. Immunophenotype of tumor cells with intermediate morphology suggested a progenitor cell origin for this tumor. Clinical findings of combined HCC-CC showed a closer resemblance with those of HCC than those of CC. In univariate analysis, tumor size, TNM stage, and serum alpha-fetoprotein levels showed a significant association with poor patient survival. Serum alpha-fetoprotein level was an independent prognostic indicator in multivariate analysis. In conclusion, an awareness of the clinicopathologic features, specifically the various morphologic features of intermediate areas in this tumor, is essential for prevention of potential misdiagnosis as another tumor.
Adult ; Aged ; Antigens, Neoplasm/metabolism ; Carcinoma, Hepatocellular/*pathology ; Cell Adhesion Molecules/metabolism ; Cholangiocarcinoma/*pathology ; Female ; Humans ; Immunophenotyping ; Keratin-19/metabolism ; Keratin-7/metabolism ; Liver Neoplasms/*pathology ; Male ; Middle Aged ; Neural Cell Adhesion Molecules/metabolism ; Prognosis ; Proto-Oncogene Proteins c-kit/metabolism ; alpha-Fetoproteins/analysis

OBJECTIVEThis study explored the effects of levetiracetam (LEV) on the expression of nerve cell adhesion molecule (NCAM) and growth-associated protein 43 (GAP-43) mRNA in the hippocampus of rats with epilepsy induced by lithium-pilocarpine (Li-PILO) in order to provide a basis for investigating the antiepileptic mechanism of LEV and its doseresponse.

METHODSForty-eight Wistar rats were randomly divided into a normal control, a Li-PILO model and two LEV treatment groups (LEV: 150 and 300 mg/kg) (n=12 each). The LEV treatment groups received LEV by intragastric administration 6 hrs after status epilepticus (once daily for 2 two weeks). The expressions of NCAM and GAP-43 mRNA in the hippocampus was determined by real-time PCR.

RESULTSThe expression of NCAM and GAP-43 mRNA in the Li-PILO model group was significantly higher than in the normal control group (P<0.05). LEV treatment of 150 and 300 mg/kg significantly decreased the expression of NCAM and GAP-43 mRNA compared with the Li-PILO model group (P<0.05). The LEV treatment group at the dose of 300 mg/kg showed significantly lower expression of NCAM and GAP-43 mRNA than the 150 mg/kg LEV treatment group (P<0.05).

CONCLUSIONSLi-PILO can up-regulate the expressions of NCAM and GAP-43 mRNA in the hippocampus of rats with epilepsy. LEV can inhibit the expression of NCAM and GAP-43 mRNA and the effect is associated with the dose of LEV.

Animals ; Anticonvulsants ; therapeutic use ; Epilepsy ; drug therapy ; metabolism ; GAP-43 Protein ; genetics ; Hippocampus ; metabolism ; Male ; Neural Cell Adhesion Molecules ; genetics ; Piracetam ; analogs & derivatives ; pharmacology ; therapeutic use ; RNA, Messenger ; analysis ; Rats ; Rats, Wistar

Altitude ; Animals ; CA1 Region, Hippocampal ; metabolism ; physiology ; Male ; Neural Cell Adhesion Molecules ; metabolism ; Rats ; Rats, Sprague-Dawley ; Time Factors

OBJECTIVETo explore the influences of the restoration of neural adhesion molecule NECL1 on the morphology, migration, and invasion of NECL1-deficient glioma cell lines.

METHODSScratch and Transwell assays were used to observe the cell migration and invasion, the activities of extracellular metalloproteinases were measured, and the cell morphology was observed. Astrocytes marker glial fibrillary acidic protein was detected by Western blot after the restoration of NECL1 in glioma U251 cell line.

RESULTSIn NECL1-deficient U251 glioma cell lines, migration and invasion were inhibited. The U251 cells was differentiated potentially to astrocytes, and glial fibrillary acidic protein was up-regulated after the restoration of the NECL1 expression.

CONCLUSIONAs a potential tumor repressor, the neural adhesion molecule NECL1 can inhibit the migration and invasion of glioma cell and induces its differentiation.

Brain Neoplasms ; metabolism ; pathology ; Cell Differentiation ; Cell Line, Tumor ; Cell Movement ; Glioma ; metabolism ; pathology ; Humans ; Neoplasm Invasiveness ; Neural Cell Adhesion Molecules ; metabolism

In the post-genomic era, various computational methods that predict protein-protein interactions at the genome level are available; however, each method has its own advantages and disadvantages, resulting in false predictions. Here we developed a unique integrated approach to identify interacting partner(s) of Semaphorin 5A (SEMA5A), beginning with seven proteins sharing similar ligand interacting residues as putative binding partners. The methods include Dwyer and Root-Bernstein/Dillon theories of protein evolution, hydropathic complementarity of protein structure, pattern of protein functions among molecules, information on domain-domain interactions, co-expression of genes and protein evolution. Among the set of seven proteins selected as putative SEMA5A interacting partners, we found the functions of Plexin B3 and Neuropilin-2 to be associated with SEMA5A. We modeled the semaphorin domain structure of Plexin B3 and found that it shares similarity with SEMA5A. Moreover, a virtual expression database search and RT-PCR analysis showed co-expression of SEMA5A and Plexin B3 and these proteins were found to have co-evolved. In addition, we confirmed the interaction of SEMA5A with Plexin B3 in co-immunoprecipitation studies. Overall, these studies demonstrate that an integrated method of prediction can be used at the genome level for discovering many unknown protein binding partners with known ligand binding domains.
Binding Sites ; genetics ; Cell Line, Tumor ; Cluster Analysis ; Computational Biology ; methods ; Databases, Protein ; Gene Expression Profiling ; Humans ; Hydrophobic and Hydrophilic Interactions ; Immunoprecipitation ; Membrane Proteins ; chemistry ; genetics ; metabolism ; Models, Molecular ; Nerve Tissue Proteins ; chemistry ; genetics ; metabolism ; Neural Cell Adhesion Molecules ; chemistry ; genetics ; metabolism ; Protein Binding ; Protein Interaction Mapping ; methods ; Protein Structure, Tertiary ; Reverse Transcriptase Polymerase Chain Reaction

It has been shown that neural cell adhesion molecule (NCAM)-induced neuronal differentiation is extracellular signal-regulated kinase (ERK)-dependent. However, an involvement of the mitogen activated protein kinase (MAPK) kinase (MEK), an upstream kinase of ERK, has not been directly demonstrated in this process. Therefore, we investigated whether the MEK1 plays a critical role in the NCAM-induced neuronal differentiation of hippocampal neural progenitor cells (NPCs). NPCs were transiently transfected with expression plasmids encoding activated or dominant negative (DN) forms of MEK1. The expression of DN MEK1 inhibited neuronal phenotype acquisition and soluble NCAM rescued the defect in the neuronal phenotype acquisition in DN-MEK1-transfected cells, suggesting that NCAM might contribute to the neuronal differentiation via distinct, parallel pathways including the MEK pathway. In cells expressing wild type MEK1 or constitutively active MEK1 on the other hand, the percentage of cells positive for beta-tubulin type III (Tuj1), a marker for early postmitotic neurons, was higher than seen in vector-transfected cells. These results suggest that the activation of MEK1 is required for obtaining neuronal phenotype in NPCs.
Transfection ; Stem Cells/cytology/drug effects/*metabolism ; Solubility ; Rats ; *Phenotype ; Neurons/cytology/drug effects/*metabolism ; Neural Cell Adhesion Molecules/*pharmacology ; Mutation/genetics ; MAP Kinase Kinase 1/genetics/*metabolism ; Hippocampus/cytology/drug effects/*metabolism ; Gene Expression Regulation ; Cells, Cultured ; Cell Differentiation ; Animals