Network analysis of microRNAs, transcription factors, and target genes involved in axon regeneration.

Axon regeneration is crucial for recovery from neurological diseases. Numerous studies have identified several genes, microRNAs (miRNAs), and transcription factors (TFs) that influence axon regeneration. However, the regulatory networks involved have not been fully elucidated. In the present study, we analyzed a regulatory network of 51 miRNAs, 27 TFs, and 59 target genes, which is involved in axon regeneration. We identified 359 pairs of feed-forward loops (FFLs), seven important genes (Nap1l1, Arhgef12, Sema6d, Akt3, Trim2, Rab11fip2, and Rps6ka3), six important miRNAs (hsa-miR-204-5p, hsa-miR-124-3p, hsa-miR-26a-5p, hsa-miR-16-5p, hsa-miR-17-5p, and hsa-miR-15b-5p), and eight important TFs (Smada2, Fli1, Wt1, Sp6, Sp3, Smad4, Smad5, and Creb1), which appear to play an important role in axon regeneration. Functional enrichment analysis revealed that axon-associated genes are involved mainly in the regulation of cellular component organization, axonogenesis, and cell morphogenesis during neuronal differentiation. However, these findings need to be validated by further studies.

Network analysis identifies common genes associated with obesity in six obesity-related diseases.

Obesity has been reported to be associated with many diseases. However, common obesity-induced biological processes have not been evaluated across these diseases. We identified genes associated with obesity and obesity-related diseases, and used them to construct protein‒protein interaction networks. We also analyzed gene ontology (GO) in those genes overlapping between obesity and disease. Our work identifies gene modules common to obesity and obesity-related diseases, which can provide a basis for understanding the process of how obesity induces disease.

Identification of neuron-related genes for cell therapy of neurological disorders by network analysis.

Bone mesenchymal stem cells (BMSCs) differentiated into neurons have been widely proposed for use in cell therapy of many neurological disorders. It is therefore important to understand the molecular mechanisms underlying this differentiation. We screened differentially expressed genes between immature neural tissues and untreated BMSCs to identify the genes responsible for neuronal differentiation from BMSCs. GSE68243 gene microarray data of rat BMSCs and GSE18860 gene microarray data of rat neurons were received from the Gene Expression Omnibus database. Transcriptome Analysis Console software showed that 1248 genes were up-regulated and 1273 were down-regulated in neurons compared with BMSCs. Gene Ontology functional enrichment, protein-protein interaction networks, functional modules, and hub genes were analyzed using DAVID, STRING 10, BiNGO tool, and Network Analyzer software, revealing that nine hub genes, Nrcam, Sema3a, Mapk8, Dlg4, Slit1, Creb1, Ntrk2, Cntn2, and Pax6, may play a pivotal role in neuronal differentiation from BMSCs. Seven genes, Dcx, Nrcam, sema3a, Cntn2, Slit1, Ephb1, and Pax6, were shown to be hub nodes within the neuronal development network, while six genes, Fgf2, Tgfß1, Vegfa, Serpine1, Il6, and Stat1, appeared to play an important role in suppressing neuronal differentiation. However, additional studies are required to confirm these results.

Neuroprotective effects of Rhizoma Dioscoreae polysaccharides against neuronal apoptosis induced by in vitro hypoxia.

Rhizoma Dioscoreae polysaccharides (RDPS) are the primary active ingredient of Rhizoma Dioscoreae, which is a traditional Chinese medicine. RDPS have previously been shown to scavenge reactive oxygen species, and protect against D-galactose-induced mimetic aging. The present study aimed to investigate the neuroprotective effects of RDPS against hypoxia-induced neuronal cell apoptosis. Neuronal cells harvested from pregnant Sprague-Dawley rats were divided into groups, as follows: i) Normal control group; ii) hypoxia-induced apoptosis neuronal cell model; iii) 0.025 g/l RDPS-treated group; iv) 0.05 g/l RDPS-treated group; v) 0.1 g/l RDPS-treated group; and vi) 0.25 g/l RDPS treated group. Neuronal cell viability was investigated using an MTT assay, and neuronal cell apoptosis was analyzed using Annexin V-fluorescein isothiocyanate/propidium iodide double-staining, Hoechst 33342 fluorescent staining, Rhodamine 123 staining, polymerase chain reaction and immunocytochemical staining. The RDPS-treated neuronal cells exhibited improved viability, and decreased hypoxia-induced mitochondrial injury and apoptosis. In addition, the mRNA and protein expression levels of caspase-3 and B-cell lymphoma (Bcl)-2-associated X protein (Bax) were significantly downregulated, whereas the mRNA and protein expression levels of Bcl-2 were significantly upregulated, in the RDPS-treated hypoxic neurons, as compared with the apoptosis model (P<0.05). Furthermore, the ratio of Bcl-2 expression:Bax expression significantly increased following RDPS treatment, as compared with the apoptosis model (P<0.05). The results of the present study suggested that RDPS may attenuate hypoxia-induced neuronal cell apoptosis by altering the expression levels of key apoptosis-regulating proteins in hypoxic neurons.

[Effect of GABA(B) Receptor Signal to Corticosterone-induced Neuron Apoptosis].

OBJECTIVE: To investigate whether corticosterone results in neuron apoptosis through regulating γ-aminobutyric acid (GABA) receptor. METHODS: In vivo: the hyperglycemic rat model with applying chronic restraint stress to healthy male SD rats (3 months) was established, after paraffin embedding the brain was sliced, and the level of neuron apoptosis was tested by detecting active Caspase-3 with immune-histochemical staining and TUNEL. The level of corticosterone in serum was detected by using ELISA. In vitro: the level of active Caspase-3 in NG108-15 cells (neuroblastoma and glioma cell line) after treated with corticosterone (10(-7) mol/L) was detected with Western blot. In NG108-15 cells recombinanted with GABA(B2) receptor, after administrating separately with the GABA(B) agonist baclofen (100 µmol/L) and antagonist CGP35348 (100 µmol/L), the level of active Caspase-3 under the effect of corticosterone (10(-7) mol/L) was detected. RESULTS: Active Caspase-3 positive apoptotic cells and TUNEL-positive cells were detected in solitary nucleus of hyperglycemia rat induced by chronic restraint stress, and the level of serum corticosterone had recovered after an initial ascent. NG108-15 cells could express GABA(B1) receptor endogenously, and the expression of active Caspase-3 increased after corticosterone treatment (P < 0.05). In NG108-15 cells transfected with GABA(B2) receptor subunits, baclofen could reduce the effect of corticosterone- induced active Caspase-3 upexpression, while CGP35348 enhanced this effect (P < 0.05). CONCLUSION: Corticosterone may lead to abnormal neuron excitability and neuron apoptosis by means of inhibiting GABA receptor B.

[The distribution of caspase-3 positive apoptotic cells in 18 months old SD rat's brainstem compared with that in 3 months rat].

OBJECTIVE: To study whether there is apoptosis in brainstem neurons while aging by oberving the distribution of Caspase-3 positive apoptotic cells in in brainstem of young and old SD rats. METHODS: Healthy male SD rats were divided into 2 groups (3 and 18 month-old respectively), 3 rats each group. Brainstem specimens were treated followed the brainstem's common paraffin embedding, sectioning and HE staining procedures (sections were 6 µm in thickness). The sections were also determined by Caspase-3 immunostaining and TUNEL. The Caspase-3 positive cells on the rat stereotaxic atlas were drew, then composed the sections into a 3D model. RESULTS: Compared to 3 month-old rats, there were more Caspase-3 positive neurons in the brainstem and the positive neurons were distributed more extensively in 18 month-old rats spectially in nucleus of solitary tract and pontine reticular nuclei. CONCLUSION: More neurons suffer apoptotic changes in the brainstem while aging.

Orally administered DNA vaccine delivery by attenuated Salmonella typhimurium targeting fetal liver kinase 1 inhibits murine Lewis lung carcinoma growth and metastasis.

The vascular endothelial growth factor (VEGF) receptor 2 (VEGFR-2), also called fetal liver kinase 1 (FLK1) in mice and kinase insert domain receptor (KDR) in humans, is an endothelial cell specific receptor tyrosine kinase that mediates lung cancer angiogenesis. We hypothesized that an active immunotherapy approach targeting FLK1 may inhibit lung cancer growth and metastasis. To test this hypothesis, we evaluated whether immune responses to FLK1 could be elicited in mice by immunization with an orally administered DNA vaccine encoding the extracellular domain (ECD) of FLK1 (pcDNA3.1-FLK1(ECD)) carried by attenuated Salmonella typhimurium. We found that the vaccine was effective at protective antitumor immunity in Lewis lung carcinoma models in mice by breaking immune tolerance to FLK1 self-antigen. Both FLK1-specific humoral and cellular immune responses against endothelial cells can be induced in mice by immunization with pcDNA3.1-FLK1(ECD). Immunization with pcDNA3.1-FLK1(ECD) resulted in tumor suppression and prolonged survival in mice challenged with Lewis lung carcinomas cells. Experimental pulmonary metastases were strongly inhibited in pcDNA3.1-FLK1(ECD) immunized mice challenged with Lewis lung carcinoma cells. Thus, we conclude that the plasmid DNA vaccine encoding the extracellular domain of FLK1 could be an important component of FLK1 DNA vaccine to prevent lung carcinoma recurrence and metastasis after surgery.

[Staining improvement for Blastocystis hominis specimen].

Schaudinn solution was used to fix the Blastocystis hominis specimen and an improved Harris hematoxylin staining was applied to stain it. The method shows clearer internal structure of the parasite, simpler and less time-consuming than the traditional iron hematoxylin solution.

[Investigations on cordycepin production by solid culture of Cordyceps militaris].

An efficient method to produce cordycepin by solid culture using Cordyceps militaris was investigated in this study. Firstly, the changes of cordycepin during various growing periods of solid culture using 5 strains of C. militaris were detected, the best strain and optimal growing period for cordycepin production were determined. Then, by experiments of quadratic rotation-orthogonal combination design and orthogonal design, the medium composition and growth conditions for high yield of cordycepin were optimized. With the optimized method to produce cordycepin, the content of cordycepin in the medium was increased to 0.60%, which was nearly 2 times higher than the highest yield reported.

[Change rate in bone cells of mice].

OBJECTIVE: To study the anti-mutation action of acupuncture and moxibustion. METHODS: Mice were randomly divided into 6 groups, group 1 (normal control group), group 2 (positive control group), group 3 (prevention group I), group 4 (prevention group II ), group 5 (treatment group I) and group 6 (treatment group II). The mice in the group 2-6 were treated by cyclophosphamide (ip, 50 mg/kg body weight), and in the 3'-6 groups were given acupuncture at "Zusanli" (ST 36) and moxibustion at "Guanyuan" (CV 4). At the end of experiment, all the mice were decapitated and chromosome aberration rate and sister chromatid exchange rate of bone marrow cells were investigated. RESULTS: The chromosome aberration rate and the sister chromatid exchange rate of bone marrow cells in the positive control group increased significantly as compared with the normal control group, while they decreased significantly in the group 3, 4, 5, 6 as compared with the positive control group (P < 0.01). CONCLUSION: Acupuncture and moxibustion have anti-mutation action, inhibiting the increase of chromosome aberrations and sister chromatid exchange of bone marrow cells in mice induced by cyclophosphamide.