Long non-coding RNA SNHG17 may function as a competitive endogenous RNA in diffuse large B-cell lymphoma progression by sponging miR-34a-5p.

We investigated the functional mechanism of long non-coding small nucleolar host gene 17 (SNHG17) in diffuse large B-cell lymphoma (DLBCL). lncRNAs related to the prognosis of patients with DLBCL were screened to analyze long non-coding small nucleolar host gene 17 (SNHG17) expression in DLBCL and normal tissues, and a nomogram established for predicting DLBCL prognosis. SNHG17 expression in B-cell lymphoma cells was detected using qPCR. The effects of SNHG17 with/without doxorubicin on the proliferation and apoptosis of DoHH2 and Daudi were detected. The effects of combined SNHG17 and doxorubicin were analyzed. The regulatory function of SNHG17 in DLBCL was investigated using a mouse tumor xenotransplantation model. RNA sequencing was used to analyze the signaling pathways involved in SNHG17 knockdown in B-cell lymphoma cell lines. The target relationships among SNHG17, microRNA, and downstream mRNA biomolecules were detected. A higher SNHG17 level predicted a lower survival rate. SNHG17 was highly expressed in DLBCL patient tissues and cell lines. We established a prognostic model containing SNHG17 expression, which could effectively predict the overall survival rate of DLBCL patients. SNHG17 knockdown inhibited the proliferation and induced the apoptosis of B-cell lymphoma cells, and the combination of SNHG17 and doxorubicin had a synergistic effect. SNHG17, miR-34a-5p, and ZESTE gene enhancer homolog 2 (EZH2) had common hypothetical binding sites, and the luciferase reporter assay verified that miR-34a-5p was the direct target of SNHG17, and EZH2 was the direct target of miR-34a-5p. The carcinogenic function of SNHG17 in the proliferation and apoptosis of DLBCL cells was partially reversed by a miR-34a-5p inhibitor. SNHG17 increases EZH2 levels by inhibiting miR-34a-5p. Our findings indicate SNHG17 as critical for promoting DLBCL progression by regulating the EZH2 signaling pathway and sponging miR-34a-5p. These findings provide a new prognostic marker and therapeutic target for the prognosis and treatment of DLBCL.

Parvalbumin in the metabolic pathway of glutamate and γ-aminobutyric acid: Influence on expression of GAD65 and GAD67.

Parvalbumin-expressing neurons are a type of inhibitory intermediate neuron that play an important role in terminating seizures. The aim of the present study was to use lentiviral construction and packaging technology to overexpress and silence the parvalbumin gene in pheochromocytoma (PC12) cells, and to evaluate how parvalbumin influences the metabolic pathway involving glutamate and γ-aminobutyric acid (GABA). In this work, Immunofluorescence staining was used to verify the differentiation of PC12 cells into neurons after adding nerve growth factor (NGF). Western blotting and real-time quantitative polymerase chain reaction (qRT-PCR) were used to confirm lentivirus-mediated knockdown or overexpression of parvalbumin. Expression of parvalbumin, the 65-kDa GAD isoform (GAD65), and the 67-kDa GAD isoform (GAD67) in neuronal cells was examined at the mRNA and protein levels using qRT-PCR, western blotting and immunofluorescence staining, while intracellular glutamate and GABA levels were determined by high performance liquid chromatography (HPLC). We demonstrate that the expression of parvalbumin is associated with GAD65 and GAD67. Interestingly, overexpression of parvalbumin up-regulated GAD65 and GAD67, increased GABA concentration, and decreased glutamate concentration. Silencing of parvalbumin led to the opposite effects. Altogether, parvalbumin affected the expression of GAD65 and GAD67, thereby influencing the metabolic pathway involving glutamate and GABA.

Analysis and identification of m6A RNA methylation regulators in metastatic osteosarcoma.

Osteosarcoma (OS) is characterized by rapid growth and early metastasis. However, its mechanism remains unclear. N6-methyladenosine (m6A) modification and its regulatory factors play essential roles in most cancers, including OS. In this study, we screened out 21 m6A modifiers using the Therapeutically Applicable Research to Generate Effective Treatments (TARGET) database, followed by the identification of the critical m6A methylation modifiers. The results revealed that the expression levels of three m6A methylation regulators, namely RBM15, METTL3, and LRPPRC, were associated with the low survival rate of patients with OS. We further studied the independent prognostic factors by performing univariate and multivariate Cox analyses and found that metastasis was an independent prognostic factor for patients with OS. Furthermore, we found for the first time that RBM15 was specific for metastatic OS rather than non-metastatic OS. Moreover, the significant overexpression of RBM15 was validated in metastatic OS cell lines and in actual human clinical specimens. We also revealed that RBM15 promoted the invasion, migration, and metastasis of OS cells through loss-functional and gain-functional experiments and an animal metastatic model. In conclusion, RBM15 has a high correlation with OS metastasis formation and the decreased survival rate of patients with OS, and this may serve as a useful biomarker for predicting metastasis and prognosis of patients with OS.

[Corrigendum] Murine and Chinese cobra venom­derived nerve growth factor stimulate chondrogenic differentiation of BMSCs in vitro: A comparative study.

Subsequently to the publication of this paper, an interested reader drew to the authors' attention that Figs. 2 and 3, showing the results from experiments designed to assess the viability of bone­derived mesenchymal stem cells culture with or without nerve growth factors (NGFs) via fluorescein diacetate/propidium iodide or H&E staining respectively, contained apparently duplicated data panels within the figures. After having examined their original data, the authors have realized that these figures were inadvertently assembled incorrectly, and that there were also misassembled data panels in Fig. 6, which showed the secretion of types I and II collagens in bone­derived mesenchymal stem cells with or without NGFs. The corrected versions of Figs. 2, 3 and 6 are shown below and on the next page. Note that these errors did not significantly affect the results or the conclusions reported in this paper, and all the authors agree to this Corrigendum. Furthermore, the authors apologize to the readership for any inconvenience caused. [the original article was published in Molecular Medicine Reports 10.3892/mmr.2018.9307].

Prognostic Implications of the Complement Protein C1Q and Its Correlation with Immune Infiltrates in Osteosarcoma.

BACKGROUND: Osteosarcoma (OS) is the most widespread bone tumour among childhood cancers, and distant metastasis is the dominant factor in poor prognosis for patients with OS. Therefore, it is necessary to identify new prognostic biomarkers for identifying patients with aggressive disease. METHODS: Two OS datasets (GSE21257 and GSE33383) were downloaded from the Gene Expression Omnibus (GEO) and subsequently subjected to weighted gene co-expression network analysis (WGCNA) and differential gene expression analysis (DGE) to screen candidate genes. A prognostic model was constructed using OS data derived from the Therapeutically Applicable Research to Generate Effective Treatments (TARGET) program to further screen key genes and perform gene ontology (GO) analysis. The prognostic values of key genes were assessed using the Kaplan-Meier (KM) plotter. The GEO dataset was used for immune infiltration analysis and association analysis of key genes. In addition, quantitative real-time polymerase chain reaction (qRT-PCR) was employed to validate the expression levels of potentially crucial genes in OS cell lines. RESULTS: In the present study, we found 114 genes with a highly significant correlation in the module and 44 downregulated genes; 25 candidate genes overlapped in the two parts of the genes. Among these, three key genes, C1QA, C1QB, and C1QC, were the most significant hub genes, which had the highest node degrees, were clustered into one group, and implicated in most significant biological processes (regulation of immune effector process). Moreover, these three key genes were negatively associated with the prognosis of OS and positively associated with three immune cells (follicular helper T cells, memory B cells, and CD8 T cells). Additionally, compared to non-metastatic OS cell lines, the expression of three key genes was significantly downregulated in metastatic OS cell lines. CONCLUSION: Our results revealed that three key genes (C1QA, C1QB, and C1QC) were implicated in tumour immune infiltration and may be promising biomarkers for predicting metastasis and prognosis of patients with OS.

Electrospun PLGA/PCL/OCP nanofiber membranes promote osteogenic differentiation of mesenchymal stem cells (MSCs).

Nanofibers as niche-biomimetic scaffolds hold promise in guided bone regeneration (GBR). Here we fabricated poly (lactic-co-glycolic acid) (PLGA)/poly(caprolactone) (PCL)-doped octacalcium phosphate (OCP) nanofiber membranes via electrospinning and investigated the osteogenic behavior of marrow mesenchymal stem cells (MSCs) on the membranes. By adjusting different ratio of OCP to PLGA/PCL, three hybrid stents including PLGA/PCL, PLGA/PCL/2 wt%OCP, PLGA/PCL/4wt%OCP were successfully prepared. The PLGA/PCL/OCP membranes showed a decrease in fiber diameter compared with PLGA/PCL, leading to enhanced mechanical strength. In-vitro studies showed that PLGA/PCL/OCP membranes better supported cell adhesion, spreading and proliferation than PLGA/PCL. The incorporation of OCP via electrospinning also endowed the membranes with osteoinductive capacity, as evidenced by activation of ALP activity, increased gene expression of bone specific markers (such as Runx2, ALP, Col 1a1, OPN, OCN, BMP2), and mineral nodules formation compared to PLGA/PCL. Comparatively, PLGA/PCL/4wt%OCP showed better mechanical and biological performance than PLGA/PCL/2 wt%OCP, demonstrating the role of OCP in nanofiber membranes. Thus, the electrospun PLGA/PCL/OCP nanofiber membranes can be potentially developed as a promising hybrid stent for GBR.

Osteogenic Potential of Electrospun Poly(3-hydroxybutyrate-co-4-hydroxybutyrate)/ Poly(ethylene glycol) Nanofiber Membranes.

Nanofibers as niche-biomimetic scaffolds exhibit potential in bone tissue engineering (BTE). Here, poly(3-hydroxybutyrate-co-4-hydroxybutyrate) co-polymer (P34HB)/poly(ethylene glycol) (PEG) nanofiber membranes with a high hydrophilicity and mechanical properties were fabricated by introducing PEG to P34HB via electrospinning. The P34HB/PEG nanofibrous scaffolds were investigated for their potential in the osteogenic differentiation and mineralization of bone marrow mesenchymal stem cells (BMSCs). By adjusting the ratio of PEG to P34HB, three scaffolds, including P34HB, P34HB/10 wt%PEG, and P34HB/30 wt%PEG, were successfully fabricated. The composite P34HB/PEG nanofiber membranes showed an enhanced hydrophilicity, a decreased fiber size, and an increased mechanical strength compared with those of P34HB. In-vitro studies showed that the P34HB/PEG membranes better supported cell adhesion, spreading, and proliferation than those of P34HB. The incorporation of PEG into the P34HB scaffold also promoted the osteoinduction capacity, as evidenced by activation of the alkaline phosphatase activity (ALP) activity, increased gene expression of bone specific markers (such as RUNX2, ALP, Col1a1, OPN, OCN, and BMP2), and mineral nodules formation. Comparatively, P34HB/10 wt%PEG showed a higher hydrophilicity and mechanical properties, as well as a better biological performance than the other membranes. Thus, the electrospun P34HB/PEG nanofiber membranes may be potentially developed as regenerative materials for BTE applications.

Comparative profiling of chondrogenic differentiation of mesenchymal stem cells (MSCs) driven by two different growth factors.

This study aimed to investigate the mechanism of nerve growth factor (NGF) from cobra venom and human transforming growth factor-ß1 (TGF-ß1) on the chondrogenic induction of mesenchymal stem cells (MSCs). NGF and TGF-ß1 were used to induce chondrogenesis of MSCs from rabbits for 7 days. Total RNA was extracted for mRNA sequencing. Differentially expressed genes (DEGs), gene ontology (GO), KEGG pathway enrichment, and PPI network analysis were conducted to screen the specific signalling pathways and target genes. Quantitative real-time polymerase chain reaction (qRT-PCR) was performed to further confirm the relative target genes. The results showed that NGF could significantly promote the expression of hyaline cartilage specific genes (collagen type II alpha 1 chain, COL2A1) compared with TGF-ß1. PI3K-AKT signalling pathway is commonly involved in the chondrogenesis of MSCs induced by NGF and TGF-ß1. However, the expression levels of the genes in the PI3K-AKT signalling pathway were significantly higher in NGF group than that in the TGF-ß1 group. In the process of chondrogenesis of MSCs induced by NGF and TGF-ß1, integrin (ITGAs) were the targeted hub genes to activate the PI3K-AKT signalling pathway. NGF could activate more proliferation and differentiation genes in the process of chondrogenesis of MSCs than TGF-ß1. TGF-ß1 promoted angiogenesis by targeting the thrombospondin (THBS1) and THBS2 which might contribute to the osteophyte formation. PI3K-AKT was the crucial signalling pathway for chondrogenic differentiation. NGF could activate the PI3K-AKT signalling pathway to a higher level, and NGF had more specificity for promoting expression of specific genes of chondrocyte compared with TGF-ß1. SIGNIFICANCE OF THE STUDY: In our study, we compared two different growth factors in promoting cartilage differentiation of MSCs and found some similarities and differences. We revealed that both NGF and TGF-ß1 could activate the PI3K-AKT signalling pathway (the expression of it in NGF was higher) by targeting the ITGAs in the process of chondrogenesis from MSCs. However, NGF could activate more proliferation and differentiation genes in the process of chondrogenesis of MSCs, whereas TGF-ß1 caused osteophyte formation by activating THBS1 and THBS2. These might be the reason why NGF could promote cartilage differentiation more specifically.

Murine and Chinese cobra venom­derived nerve growth factor stimulate chondrogenic differentiation of BMSCs in vitro: A comparative study.

Mesenchymal stem cell (MSC)­based therapy has been commonly used in cartilage reconstruction, due to its self­renewing ability and multi­differentiation potential. Nerve growth factor (NGF) from cobra venom has been reported to regulate chondrogenesis of bone­derived MSCs (BMSCs) and chondrocyte metabolism. Therefore, the present study aimed to determine whether other sources of NGF behave in the same manner as NGF from natural venom. The present study compared the effects of NGF from two sources, the commercially purchased recombinant murine ß­NGF (mNGF) and cobra venom­derived NGF (cvNGF), on chondrogenesis of BMSCs by performing hematoxylin and eosin and fluorescein diacetate/propidium iodide staining, DNA and glycosaminoglycan quantization and reverse transcription­quantitative polymerase chain reaction to investigate cell morphology, viability, proliferation, glycosaminoglycan synthesis and cartilage­specific gene expression. The results demonstrated that cvNGF significantly accelerated cell proliferation and upregulated the expression of cartilage­specific genes, including aggrecan, SRY­box 9 and collagen type II α1 chain. Conversely, cvNGF reduced the expression levels of collagen type I α1 chain (a fibrocartilage marker), runt­related transcription factor 2 and enolase 2 compared with in the mNGF and control groups. In addition, Chinese cobra venom, which is the main resource of cvNGF, is abundant and inexpensive, thus greatly decreasing the cost. In conclusion, the present study demonstrated that cvNGF may be considered a potential growth factor for inducing chondrogenic differentiation of BMSCs.

Collagen, agarose, alginate, and Matrigel hydrogels as cell substrates for culture of chondrocytes in vitro: A comparative study.

Autologous chondrocyte implantation (ACI) has emerged as a new approach to cartilage repair through the use of harvested chondrocytes. But the expansion of the chondrocytes from the donor tissue in vitro is restricted by limited cell numbers and dedifferentiation of chondrocytes. In this study, we used four types of hydrogels including agarose, alginate, Matrigel, and collagen type I hydrogels to serve as cell substrates and investigated the effect on proliferation and phenotype maintenance of chondrocytes. As a substrate for monolayer culture, collagen facilitated cell expansion and effectively suppressed the dedifferentiation of chondrocytes, as evidenced by fluorescein diacetate/propidium iodide (FDA/PI), hematoxylin-eosin staining (HE), Safranin O, immunofluorescenceassay, biochemistry analysis, and quantitative real-time polymerase chain reaction (qRT-PCR). Compared with that in agarose gels, alginate, and Matrigel, collagen accelerated cell proliferation and enhanced the expression of cartilage specific genes such as ACAN, SOX9, and COLII more markedly. Furthermore, significantly lower expression of COL I (an indicator of dedifferentiation) and COL X (the chondrocyte hypertrophy marker) was present in collagen group than in other groups. This indicated that collagen substrate can better support chondrocyte growth and maintain cell phenotype, due to that it might serve as a cartilage-like ECM to provide adhesive site for chondrocytes. In summary, collagen hydrogel is a promising cell substrate for chondrocytes culture for ACI.

Promoter DNA methylation analysis reveals a novel diagnostic CpG-based biomarker and RAB25 hypermethylation in clear cell renel cell carcinoma.

Clear-cell renal cell carcinoma (ccRCC) is a common aggressive urinary malignant tumor that cannot be easily diagnosed at an early stage. The DNA methylation occurs within promoter before precancerous lesion plays a pivotal role that could help us in diagnosing and understanding ccRCC. In this study, based on a whole-genome promoter DNA methylation profiling, we used shrunken centroids classifier method to identify a CpG-based biomarker that is capable of differentiating between ccRCC tumor and adjacent tissues. The biomarker was validated in 19 ccRCCs and three public datasets. We found that both CYP4B1 and RAB25 are downregulated with promoter hypermethylation and CA9 is upregulated with promoter hypomethylation, and we validated their mRNA differential expressions in 19 ccRCCs and 10 GEO datasets. We further confirmed that hypermethylated RAB25 is inversely correlated with its mRNA level. Log-rank test showed that ccRCC patients with low levels of CA9 promoter methylation had a higher survival rate. This reveals clinically a potential biomarker for use in early detection for ccRCC, and provides a better understanding of carcinogenesis.

Nerve growth factor from Chinese cobra venom stimulates chondrogenic differentiation of mesenchymal stem cells.

Growth factors such as transforming growth factor beta1 (TGF-ß1), have critical roles in the regulation of the chondrogenic differentiation of mesenchymal stem cells (MSCs), which promote cartilage repair. However, the clinical applications of the traditional growth factors are limited by their high cost, functional heterogeneity and unpredictable effects, such as cyst formation. It may be advantageous for cartilage regeneration to identify a low-cost substitute with greater chondral specificity and easy accessibility. As a neuropeptide, nerve growth factor (NGF) was involved in cartilage metabolism and NGF is hypothesized to mediate the chondrogenic differentiation of MSCs. We isolated NGF from Chinese cobra venom using a three-step procedure that we had improved upon from previous studies, and investigated the chondrogenic potential of NGF on bone marrow MSCs (BMSCs) both in vitro and in vivo. The results showed that NGF greatly upregulated the expression of cartilage-specific markers. When applied to cartilage repair for 4, 8 and 12 weeks, NGF-treated BMSCs have greater therapeutic effect than untreated BMSCs. Although inferior to TGF-ß1 regarding its chondrogenic potential, NGF showed considerably lower expression of collagen type I, which is a fibrocartilage marker, and RUNX2, which is critical for terminal chondrocyte differentiation than TGF-ß1, indicating its chondral specificity. Interestingly, NGF rarely induced BMSCs to differentiate into a neuronal phenotype, which may be due to the presence of other chondrogenic supplements. Furthermore, the underlying mechanism revealed that NGF-mediated chondrogenesis may be associated with the activation of PI3K/AKT and MAPK/ERK signaling pathways via the specific receptor of NGF, TrkA. In addition, NGF is easily accessed because of the abundance and low price of cobra venom, as well as the simplified methods for separation and purification. This study was the first to demonstrate the chondrogenic potential of NGF, which may provide a reference for cartilage regeneration in the clinic.

Baicalin inhibits biofilm formation, attenuates the quorum sensing-controlled virulence and enhances Pseudomonas aeruginosa clearance in a mouse peritoneal implant infection model.

The quorum sensing (QS) circuit plays a role in the precise regulation of genes controlling virulence factors and biofilm formation in Pseudomonas aeruginosa. QS-controlled biofilm formation by Pseudomonas aeruginosa in clinical settings has remained controversial due to emerging drug resistance; therefore, screening diverse compounds for anti-biofilm or anti-QS activities is important. This study demonstrates the ability of sub-minimum inhibitory concentrations (sub-MICs) of baicalin, an active natural compound extracted from the traditional Chinese medicinal Scutellaria baicalensis, to inhibit the formation of Pseudomonas aeruginosa biofilms and enhance the bactericidal effects of various conventional antibiotics in vitro. In addition, baicalin exerted dose-dependent inhibitory effects on virulence phenotypes (LasA protease, LasB elastase, pyocyanin, rhamnolipid, motilities and exotoxin A) regulated by QS in Pseudomonas aeruginosa. Moreover, the expression levels of QS-regulatory genes, including lasI, lasR, rhlI, rhlR, pqsR and pqsA, were repressed after sub-MIC baicalin treatment, resulting in significant decreases in the QS signaling molecules 3-oxo-C12-HSL and C4-HSL, confirming the ability of baicalin-mediated QS inhibition to alter gene and protein expression. In vivo experiments indicated that baicalin treatment reduces Pseudomonas aeruginosa pathogenicity in Caenorhabditis elegans. Greater worm survival in the baicalin-treated group manifested as an increase in the LT50 from 24 to 96 h. In a mouse peritoneal implant infection model, baicalin treatment enhanced the clearance of Pseudomonas aeruginosa from the implants of mice infected with Pseudomonas aeruginosa compared with the control group. Moreover, the combination of baicalin and antibiotics significantly reduced the numbers of colony-forming units in the implants to a significantly greater degree than antibiotic treatment alone. Pathological and histological analyses revealed mitigation of the inflammatory response and reduced cell infiltration in the peritoneal tissue surrounding the implants after baicalin treatment. Measurement of the cytokine levels in the peritoneal lavage fluid of mice in the baicalin treatment group revealed a decrease in IL-4, an increase in interferon γ (IFN-γ), and a reversed IFN-γ/IL-4 ratio compared with the control group, indicating that baicalin treatment activated the Th1-induced immune response to expedite bacterial load clearance. Based on these results, baicalin might be a potent QS inhibitor and anti-biofilm agent for combating Pseudomonas aeruginosa biofilm-related infections.

Effect of NGF From Venom of Chinese Cobra (Naja Atra)on Chondrocytes Proliferation and Metabolism In Vitro.

Autologous chondrocyte implantation (ACI) is promising strategy for cartilage repair. However, chondrocyte phenotype is easily lost when expanded in vitro which defined as "dedifferentiation." To ensure successful therapy, an effective pro-chondrogenic agent is necessary to overcome the obstacle of limited cell numbers in the restoration process, and dedifferentiation is a prerequisite. In the present study, we investigated the chondro-protective effect of NGF from Chinese cobra venom on human chondrocytes by determination of its specific effect on cell viability, proliferation, morphology, GAG production, and cartilage specific gene expression. The results suggested that NGF showed no cytotoxicity to chondrocytes below the concentration of 16 µg/mL. DNA and glycosaminoglycan (GAG) content were, respectively, improved in NGF groups comparing to the control (P < 0.05). NGF up-regulate the gene expression of ACAN, SOX9, and COL2A1 while down-regulate the expression level of COL1A1 (P < 0.05). Moreover, the results of viability assay, hematoxylin-eosin, safranin O, and immunohistochemical staining also suggested better performances in NGF groups. NGF of 6 µg/mL shown lower cytotoxicity on chondrocytes, more glycosaminoglycans (GAGs) synthesis and up-regulated chondrogenic gene expression. This study may provide a basis for the development of a novel agent for the treatment of articular cartilage defects. J. Cell. Biochem. 118: 4308-4316, 2017. © 2017 Wiley Periodicals, Inc.

Serum Macroelement and Microelement Concentrations in Patients with Polycystic Ovary Syndrome: a Cross-Sectional Study.

Polycystic ovarian syndrome (PCOS) is one of the most common endocrine diseases. However, its pathogenesis is unclear. We aim to explore the potential relationships between serum macroelements/microelements and PCOS. A total of 1137 women were involved in the current study. PCOS was defined according to ESHRE/ASRM, and complete blood samples were collected. Serum macroelements (calcium and magnesium) and microelements (copper, zinc, and iron) were assayed through atomic absorption spectrophotometry. PCOS patients had significantly higher copper concentrations than patients without PCOS (P < 0.001). By contrast, PCOS patients had lower serum calcium levels than patients without PCOS (P < 0.001). No significant differences were observed in the levels of serum zinc, magnesium, and iron between PCOS and non-PCOS patients. PCOS patients with acne had higher magnesium levels than those without acne (P = 0.020), and PCOS patients with hirsutism had lower magnesium levels than those without hirsutism (P = 0.037). High serum copper and low calcium levels may be correlated with PCOS. Magnesium concentrations are correlated with acne and hirsutism in PCOS patients. These results provide clues to explore the mechanism of PCOS and guidance for element treatments in PCOS patients.

Integrated Analysis Reveals together miR-182, miR-200c and miR-221 Can Help in the Diagnosis of Prostate Cancer.

Research has shown that microRNAs are promising biomarkers that can be used to promote a more accurate diagnosis of cancer. In this study, we developed an integrated multi-step selection process to analyze available high-throughput datasets to obtain information on microRNAs as cancer biomarkers. Applying this approach to the microRNA expression profiles of prostate cancer and the datasets in The Cancer Genome Atlas Data Portal, we identified miRNA-182, miRNA-200c and miRNA-221 as possible biomarkers for prostate cancer. The associations between the expressions of these three microRNAs with clinical parameters as well as their diagnostic capability were studied. Several online databases were used to predict the target genes of these three microRNAs, and the results were confirmed by significant statistical correlations. Comparing with the other 18 types of cancers listed in The Cancer Genome Atlas Data Portal, we found that the combination of both miRNA-182 and miRNA-200c being up-regulated and miRNA-221 being down-regulated only happens in prostate cancer. This provides a unique biological characteristic for prostate cancer that can potentially be used for diagnosis based on tissue testing. In addition, our study also revealed that these three microRNAs are associated with the pathological status of prostate cancer.

[Research for the production of recombinant human epidermal growth factor using Samia Cynthia Ricini pupae bioreactor].

The protein production system using a baculovirus Antheraea pernyi nucleopolyhedrovirus (AnpeNPV) as a gene expression vector and its host insect as a natural bioreactor was successful established and its excellent performance in the protein production has been demonstrated. In this paper, the system is used to produce recombinant human epidermal growth factor (rhEGF), which have been widely used in medical and cosmetic treatment. A recombinant AnpehEGF virus has been constructed by replacing the viral polyhedrin gene with the rhEGF gene, and then injected it to Samia cynthia ricini pupae. Amplification and expression of rhEGF gene in the pupae was clearly detected by PCR, Western blot and ELISA analyses. These analyses have also revealed that rhEGF in the pupae was significantly increased at 6 days post-infection, and reached maximum level at the 12th day. The concentrations of rhEGF were 19.77, 24.90, 618.59 and 1 952.46 ng/g pupae at 3, 6, 9 and 12 days post-infection, respectively. However, the rhEGF concentration reduced at later stage (days 15). The rhEGF in the pupae could be purified using ammonium sulfate precipitation and Ni-NTA agrose affinity chromatography. Results demonstrate that Samia cynthia ricini pupae can be used as a bioreactor to produce rhEGF and, if successfully improved, will be a novel method of rhEGF production with lower cost and more efficient.