A2B5-positive oligodendrocyte precursor cell transplantation improves neurological deficits in rats following spinal cord contusion associated with changes in expression of factors involved in the Notch signaling pathway.

BACKGROUND: Oligodendrocyte precursor cells (OPCs) are myelinated glial cells of the central nervous system (CNS), able to regenerate oligodendrocytes and myelin. This study aimed to elucidate the effect of A2B5-positive (A2B5+) OPC transplantation in rats with spinal cord contusion (SCC) and to investigate changes in expression of various factors involved in the Notch signaling pathway after OPC transplantation. METHODS: OPCs were obtained from induced pluripotent stem cells (iPSCs) originating from mouse embryo fibroblasts (MEFs). After identification of iPSCs and iPSC-derived OPCs, A2B5+ OPCs were transplanted into the injured site of rats with SCC one week after SCC insult. Behavioral tests evaluated motor and sensory function 7 days after OPC transplantation. Real-time quantitative polymerase chain reaction (RT-qPCR) determined the expression of various cytokines related to the Notch signaling pathway after OPC transplantation. RESULTS: IPSC-derived OPCs were successfully generated from MEFs, as indicated by positive immunostaining of A2B5, PDGFα and NG2. Further differentiation of OPCs was identified by immunostaining of Olig2, Sox10, Nkx2.2, O4, MBP and GFAP. Importantly, myelin formation was significantly enhanced in the SCC+ OPC group and SCI-induced motor and sensory dysfunction was largely alleviated by A2B5+ OPC transplantation. Expression of factors involved in the Notch signaling pathway (Notch-1, Numb, SHARP1 and NEDD4) was significantly increased after OPC transplantation. CONCLUSIONS: A2B5+ OPC transplantation attenuates motor and sensory dysfunction in SCC rats by promoting myelin formation, which may be associated with change in expression of factors involved in the Notch signaling pathway.

Tid1-S regulates the mitochondrial localization of EGFR in non-small cell lung carcinoma.

The epidermal growth factor receptor (EGFR) is the major driver of non-small cell lung carcinoma (NSCLC). Mitochondrial accumulation of EGFR has been shown to promote metastasis in NSCLC, yet little is known about how the mitochondrial localization of EGFR is regulated. In this work, we show that Tid1 (also known as mitochondrial HSP40) is involved in the mitochondrial localization of EGFR, and that the DnaJ domain of Tid1-S is essential for the Tid1-S-mediated transportation of EGFR into mitochondria. Overexpression of Tid1-S increased the migration and invasion of NSCLC cells cultured in vitro. High levels of EGFR and Tid1-S were detected in the mitochondria of cancerous lesions from stage IV NSCLC patients, and high levels of mitochondrial Tid1-S/EGFR were correlated with lymph node metastasis and poor overall survival of NSCLC patients. We thus conclude that Tid1-S critically governs the mitochondrial localization of EGFR through the mtHSP70 transportation pathway, and that the mitochondrial accumulation of EGFR appears to promote metastasis in NSCLC.

Chondroitinase administration and pcDNA3.1-BDNF-BMSC transplantation promote motor functional recovery associated with NGF expression in spinal cord-transected rat.

STUDY DESIGN: We evaluated whether combination of chondroitinase (chABC) administration and brain-derived neurotrophic factor (BDNF)-mesenchymal stem cell (MSC) transplantation could provide an optimal effect for the treatment of spinal cord injury (SCI) subjected to complete transection. OBJECTIVES: Behavioral assessments and DBA tracing were used to evaluate the effects of combination of chABC administration and BDNF-MSC transplantation on axonal regeneration and functional improvement in SCT rats. SETTING: Sichuan, ChinaMethods:Bone mesenchymal stem cells (BMSCs) were cultured and overexpressed BDNF recombinant vector was constructed into MSCs, then transplanted into the impaired spinal cord, together with chABC administration. Finally, the cortical spinal tract regeneration was detected by DBA tracing at 4 weeks post operation, and the expression of nerve growth factor (NGF), BDNF, neurotrophic factor (NT)-3, NT-4, fibroblast growth factor (FGF-2)-2, B cell lymphoma 2 (BCL-2) assaciated X protein (BAX) and BCL-2 in the caudal cord tissues was assessed by reverse transcription-PCR. RESULTS: Animals received both BDNF-BMSC transplantation and chABC administration presented the best functional recovery and obvious axonal regeneration. Moreover, NGF expression was significantly higher than that in the other groups. CONCLUSION: Co-treated strategy could effectively promote motor functional recovery and axonal regeneration in SCT rats associated with NGF upregulation.

Notch pathway is activated in cell culture and mouse models of mutant SOD1-related familial amyotrophic lateral sclerosis, with suppression of its activation as an additional mechanism of neuroprotection for lithium and valproate.

Amyotrophic lateral sclerosis (ALS) is an idiopathic and lethal neurodegenerative disease that currently has no effective treatment. A recent study found that the Notch signaling pathway was up-regulated in a TAR DNA-binding protein-43 (TDP-43) Drosophila model of ALS. Notch signaling acts as a master regulator in the central nervous system. However, the mechanisms by which Notch participates in the pathogenesis of ALS have not been completely elucidated. Recent studies have shown that the mood stabilizers lithium and valproic acid (VPA) are able to regulate Notch signaling. Our study sought to confirm the relationship between the Notch pathway and ALS and whether the Notch pathway contributes to the neuroprotective effects of lithium and VPA in ALS. We found that the Notch pathway was activated in in vitro and in vivo models of ALS, and suppression of Notch activation with a Notch signaling inhibitor, N-[N-(3,5-difluorophenacetyl-L-alanyl)]-S-phenylglycine t-butyl ester (DAPT) and Notch1 siRNA significantly reduced neuronal apoptotic signaling, as evidenced by the up-regulation of Bcl-2 as well as the down-regulation of Bax and cytochrome c. We also found that lithium and VPA suppressed the Notch activation associated with the superoxide dismutase-1 (SOD1) mutation, and the combination of lithium and VPA produced a more robust effect than either agent alone. Our findings indicate that the Notch pathway plays a critical role in ALS, and the neuroprotective effects of lithium and VPA against mutant SOD1-mediated neuronal damage are at least partially dependent on their suppression of Notch activation.

Expression of dopamine transporter in the different cerebral regions of methamphetamine-dependent rats.

OBJECTIVES: To observe the expression of the dopamine transporter (DAT) in six cerebral regions of a methamphetamine (MA)-dependent rat, which were frontal cortex, nucleus accumbens septi, striatum, hippocampus, substantia nigra and ventral tegmental area. METHODS: The rats were administrated intraperitoneally with 10 mg/kg/day of MA for 10 days consecutively; the behaviour changes were measured via the conditioned place preference (CPP), and the scores of stereotyped behaviour (SB) were used to confirm animal addiction. Then, the animals were further injected with MA respectively for 1, 2, 4 and 8 weeks to establish different periods of MA-dependent models. The expressions of DAT and DAT messenger RNA in six cerebral regions were detected. RESULTS: The results of CPP and SB scores were significant different when comparing all four experimental groups with the control group (p < 0.05). Comparing between different experimental groups, the expression of DAT mainly decreased and had dynamic changes in the same regions (p < 0.05). Comparing the different regions with each other in the same experimental group, the expression of DAT also had significant difference in several regions p < 0.05). CONCLUSIONS: The expression of DAT mainly decreased and had different in the six cerebral regions at the same MA-dependent time period as well as at different time periods in the same cerebral region. It was speculated that DAT might play a crucial role in the mechanism of MA dependence.

Bortezomib enhances cancer cell death by blocking the autophagic flux through stimulating ERK phosphorylation.

The antitumor activity of an inhibitor of 26S proteasome bortezomib (Velcade) has been observed in various malignancies, including colon cancer, prostate cancer, breast cancer, and ovarian cancer. Bortezomib has been proposed to stimulate autophagy, but scientific observations did not always support this. Interactions between ERK activity and autophagy are complex and not completely clear. Autophagy proteins have recently been shown to regulate the functions of ERK, and ERK activation has been found to induce autophagy. On the other hand, sustained activation of ERK has also been shown to inhibit the maturation step of the autophagy process. In this study, we sought to identify the mechanism of autophagy regulation in cancer cells treated with bortezomib. Our results indicate that bortezomib blocked the autophagic flux without inhibiting the fusion of the autophagosome and lysosome. In ovarian cancer, as well as endometrial cancer and hepatocellular carcinoma cells, bortezomib inhibited protein degradation in lysosomes by suppressing cathepsins, which requires the participation of ERK phosphorylation, but not JNK or p38. Our findings that ERK phosphorylation reduced cathepsins further explain how ERK phosphorylation inhibits the autophagic flux. In conclusion, bortezomib may induce ERK phosphorylation to suppress cathepsin B and inhibit the catalytic process of autophagy in ovarian cancer and other solid tumors. The inhibition of cisplatin-induced autophagy by bortezomib can enhance chemotherapy efficacy in ovarian cancer. As we also found that bortezomib blocks the autophagic flux in other cancers, the synergistic cytotoxic effect of bortezomib by abolishing chemotherapy-related autophagy may help us develop strategies of combination therapies for multiple cancers.

The correlation between MR diffusion-weighted imaging and pathological grades on glioma.

OBJECTIVE: This work intends to quantitatively analyze on pathological grade of glioma using Magnetic Resonance (MR) diffusion weighted imaging (DWI) and exploring the relativity of pathological grade and Apparent Diffusion Coefficient (ADC) value of MR diffusion weighted imaging. PATIENTS AND METHODS: 40 patients with glioma accepted the MR diffusion weighted imaging to measure the ADC value of tumor with 3.0T MR machine before the surgery. Tumor samples were sent for pathologic diagnosis and tumor cell density measurement after the operation. The acquired data were analyzed statistically. RESULTS: The ADC values of low-grade (WHO I-II) glioma were higher than that of high-grade (WHO III-IV), but the cell density of low-grade glioma was apparently lower than that of high-grade glioma. The ADC values and density of tumor cells were negatively correlated with WHO malignant grades, while the density of cells of glioma was positively correlated with WHO malignant grades. CONCLUSIONS: MR diffusion weighted imaging is an objective and effective examination method.

Regulation of ovarian cancer progression by microRNA-187 through targeting Disabled homolog-2.

MicroRNAs (miRNAs) play important roles in tumorigenesis by regulating oncogenes and tumor-suppressor genes. In this study, miR-187 and miR-200a were found to be expressed at higher levels in ovarian cancers than in benign tumors. In patients with ovarian cancer, however, higher levels of miR-187 and miR-200a expression were paradoxically associated with better OS and recurrence-free survival. Further, multivariate analysis showed that miR-187 served as an independent prognostic factor for patients with ovarian cancer (n=176). Computational prediction and microarray results indicated that miR-187 directly targeted Disabled homolog-2 (Dab2), and luciferase reporter assays confirmed that the target site of miR-187 was located at the 3'-UTR of the Dab2 gene. Generally considered as a tumor-suppressor gene, Dab2 may actually promote tumor progression in advanced cancers through epithelial-to-mesenchymal transition (EMT). Ectopic expression of miR-187 in cancer cells promoted cell proliferation, but continued overexpression of miR-187 suppressed Dab2 and inhibited migration. Suppression of miR-187 upregulated Dab2, which, by inhibiting E-cadherin levels while stimulating vimentin and phospho-FAK levels, promoted EMT. Reduced ovarian cancer Dab2 histoscores correlated with high miR-187 levels and improved outcomes of patients. Collectively, these results demonstrate distinct dual roles of Dab2 in cell proliferation and tumor progression. In the initial steps of tumorigenesis, upregulated miR-187 suppresses Dab2, promoting cell proliferation. During the later stages, however, continued increased levels of miR-187 inhibits the Dab2-dependent EMT that is associated with tumor invasiveness, which is presumed to be the reason why cancers with high miR-187 levels were associated with better survivals.

The polymorphisms of C-reactive protein gene modify the association between central obesity and lung function in taiwan asthmatics.

High-sensitivity C-reactive protein (hs-CRP) concentrations and obesity are proposed to have a significant relationship with impairment of lung function, but little has been reported to date on the association between CRP gene and lung function. We studied the association of three tagSNPs (tag single nucleotide polymorphisms) of CRP gene and their interactions with central obesity on lung function. A total of 384 asthmatic adults and 384 controls who were 1:1 matched by sex and age were recruited for this study. Three tagSNPs polymorphisms for CRP rs1417938, rs1800947 and rs1205 were selected from HapMap data and genotyping by using TaqMan allelic discrimination assay. A questionnaire interview, body composition and pulmonary function tests were performed. CRP single nucleotide polymorphisms (SNPs) did not increase the risk of asthma, but CRP rs1205 CC genotype significantly decreased the predictive value of forced vital capacity (FVC) in the asthma group (adjusted mean change = -7.54%, 95% CI = -13.82 to -1.25%). Waist-to-hip ratio, not body mass index, also decreased the predictive value of FVC in asthmatics. The subjects with central obesity who carried CRP SNPs have a significant reduction effect in lung function. The current results suggest that central obesity may play a major role in lung function, and these effects were modified significantly by the polymorphisms for CRP gene.

Induction of production and secretion beta(1-->4) glucanase with Saccharomyces cerevesiae by replacing the MET10 gene with egl1 gene from Trichoderma reesei.

AIMS: To construct novel brewer's yeast strains with the ability to degrade beta-glucan and increase sulfite levels in beer brewing by genetic manipulation. METHODS AND RESULTS: The recombinant plasmid pA15ME containing P(met10)-egl1-T(met10) expression cassette was constructed. BamHI-linearized target plasmid pA15ME was transformed into the industrial brewer's yeast strain Z0103 to replace the MET10 locus through one-step gene replacement. The recombinants Z8, Z7 and Z3 with the ability to secrete active endo-beta-1,4-glucanase I into the culture medium were isolated by Congo red dyeing. The enzymatic activities of EG I of Z8, Z7 and Z3 were 3.3, 1.5, 1.3 U l(-1), and the hydrolysing degrees of beta-glucans in wort were increased 11.9%, 8.6% and 6.9%, respectively, than that of original strain Z0103. The MET10 gene deletions were confirmed by real-time PCR, and the sulfite levels of the culture mediums inoculated with Z8, Z7 and Z3 were increased 26%, 16% and 17%, respectively, compared to that of Z0103. CONCLUSIONS: The novel endoglucanase-producing brewer's yeast strains with inserted endoglucanase gene and deficient MET10 gene led to reduced content of barley beta-glucans, enhanced filterability and increased sulfur dioxide in fermenting wort. Thus, the cost for addition of microbial beta-glucanase enzyme and sulfite preparations in normal beer brewing processes could be reduced. SIGNIFICANCE AND IMPACT OF THE STUDY: These results suggested that genetic engineering approach is a powerful tool to construct the novel recombinant brewer's yeast strains with different properties to reduce the cost of beer brewing and improve the flavour of a beer, and the strains obtained have potential application value in beer brewing.

Prognostic significance of insulin-like growth factor II mRNA-binding protein 3 expression in gastric adenocarcinoma.

BACKGROUND: Insulin-like growth factor II mRNA-binding protein (IMP) 3 is expressed in embryonic tissues and multiple cancers. The aim was to establish the prognostic value of IMP-3 expression in gastric adenocarcinoma. METHODS: IMP-3 expression in resected gastric adenocarcinomas was analysed by immunohistochemistry. RESULTS: IMP-3 was expressed in 183 (58.1 per cent) of 315 tumours. Expression was associated with older age (P < 0.001), larger tumour size (P = 0.009), deep tumour invasion (P < 0.001) and lymph node metastasis (P < 0.001). IMP-3-positive tumours were associated with poorer 5-year survival than negative tumours at all stages (stage I, 82 versus 97 per cent; stage II, 55 versus 78 per cent; stage III and IV, 11 versus 25 per cent; P = 0.005, P = 0.033 and P = 0.036 respectively). Multivariable analysis identified IMP-3 (hazard ratio (HR) 1.93), depth of tumour invasion (HR 3.69, 9.77 and 10.69 for pathological tumour stage (pT) 2, pT3 and pT4 respectively versus pT1), and lymph node metastasis (HR 1.57, 3.29 and 3.40 for pathological node stage (pN) 1, pN2 and pN3 respectively versus pN0) as independent prognostic factors. CONCLUSION: IMP-3 expression correlates with the metastatic potential of gastric adenocarcinoma and is an independent prognostic factor.

Changes of the ocular refraction among freshmen in National Taiwan University between 1988 and 2005.

PURPOSE: The prevalence of myopia in Taiwan has been reported to be increasing in the sequential nationwide survey. The purpose of this study is to compare the change of ocular refraction among freshmen in 1988 and 2005 in National Taiwan University. METHODS: The refractive status of freshmen in National Taiwan University in 2005 was examined. The refractive status and corneal radius of each student were measured with autorefractometer. The data was compared with the data obtained in 1988. All the refractions of the right eye were chosen and myopia was defined as a mean spherical equivalent of -0.25 D or more. RESULTS: The mean refractive status of total 4686 freshmen was -4.25+/-2.74 D in 1988 (-4.12+/-2.72 D for males and -4.41+/-2.75 D for females). The prevalence of myopia was 91.3% (90.1% for males and 92.8% for females). The prevalence of high myopia (over -6.0 D) was 23.5% (22.2% for males and 25.1% for females). In 2005, the mean refractive status of total 3709 freshmen was -4.93+/-2.82 D (-4.93+/-2.83 D for males and -4.93+/-2.80 D for females). The prevalence of myopia was 95.9% (95.9% for males and 95.9% for females). The prevalence of high myopia was 38.4% (38.1% for males and 38.8% for females). CONCLUSIONS: The prevalence and severity of myopia in freshmen of National Taiwan University increased significantly in 2005 compared to 1988. The distribution of refractive status in different college changed also. These findings may be explained by the early onset of myopia.

24 Mb deletion of 6q22.1-->q23.2 in an infant with pulmonary atresia, ventricular septal defect, microcephaly, developmental delay and facial dysmorphism.

24 Mb deletion of 6q22.1-->q23.2 in an infant with pulmonary atresia, ventricular septal defect, microcephaly, developmental delay and facial dysmorphism.

High inguinal loupe-assisted varicocelectomy for subfertile men with varicococeles: technical feasibility, clinical outcomes and complications.

254 consecutive patients underwent high inguinal loupe-assisted varicocelectomy. All patients had at least a one year history of infertility with abnormal semen parameters and physical examination and/or color Doppler ultrasound proven varicocele. To facilitate the procedure, an x 3.0 loupe was used during spermatic cord dissection near or at level of internal inguinal ring. Semen analysis and physical examination were performed at 3 monthly intervals. No intra-operative complications occurred. The most common post-operative complications were transient scrotal pain and stitch reaction, occurring in 12% and 4% of men, respectively. Only one permanent and two transient hydroceles were observed. Recurrent or persistent varicocele was identified by physical examination and color Doppler in 5 varicocelectomies (1.4%), and by color Doppler only in 6 varicocelectomies (1.7%). Sperm motility increased from 30 +/- 8% to 46 +/- 20%, and sperm concentration. (10(6)/cc) increased from 24 +/- 18 to 41 +/- 28. The one-year pregnancy rate was 37%. High inguinal loupe-assisted varicocelectomy is a safe, simple, and effective treatment for varicocele.

Paclitaxel (Taxol) upregulates expression of functional interleukin-6 in human ovarian cancer cells through multiple signaling pathways.

Paclitaxel (Taxol) is an antineoplastic agent that specifically targets microtubules and arrests cells at the G2/M phase of the cell cycle. In addition to mitotic arrest, the activation of c-Jun N-terminal kinase (JNK) signaling pathway has been demonstrated to be involved in the process leading to apoptosis. In an attempt to explore what genes are transcriptionally regulated by the activated JNK signaling pathway upon paclitaxel treatment, we used cDNA microarrays to analyse the changes of gene expression in human ovarian cancer cells that were treated with paclitaxel and/or the JNK inhibitor SP600125. Among 20 genes that were specifically regulated by the paclitaxel-activated JNK pathway, interleukin (IL)-6 was shown to elicit function through the JAK-STAT signaling pathway in an autocrine and/or paracrine fashion. Subsequently, we identified that 87.5% of eight tested ovarian cancer lines secreted detectable levels of IL-6, which could be further upregulated 2-3.2 fold by 1 microM paclitaxel. Dissection on regulatory pathways for IL-6 indicated that (i) when ovarian cancer cells were treated with paclitaxel at low but clinically achievable concentrations (exemplified by 1 microM in this study), the JNK signaling pathway was the major stimulator of IL-6 gene regulation and (ii) at suprapharmacologically high concentrations (exemplified by 50 microM), paclitaxel exerted lipopolysaccharide-like effects, most likely through the Toll-like receptor 4 signaling pathway. Collectively, these results suggest that paclitaxel upregulates functional IL-6 expression in human ovarian cancer cells through multiple signaling pathways.

Gene-gene synergistic effect on atopic asthma: tumour necrosis factor-alpha-308 and lymphotoxin-alpha-NcoI in Taiwan's children.

BACKGROUND: Asthma is now known to be an inflammatory response caused by the release of inflammatory mediators and cytokines. Tumour necrosis factor (TNF) is a potent cytokine in the inflammation response of the airway, and the polymorphisms of TNF genes have been associated with asthma. OBJECTIVE: This study investigated two variants, TNF-alpha-308*2 and lymphotoxin (LT)-alpha-NcoI*1, which may predispose individuals to asthma and atopy pathogenesis. METHODS: PCR-based assays were performed to determine LT-alpha-NcoI*1 and TNF-alpha-308*2 genotypes among our subjects, with 128 atopic asthmatics and 51 non-atopic asthmatics, 55 atopic controls, and 78 non-atopic controls in this genetic case-control study. RESULTS: The TNF-alpha-308*2 polymorphism increased in subjects with atopic asthma vs. non-atopic controls after adjusting for age distribution (adjusted odds ratios, AOR=2.73, 95% confidence interval, CI=1.16-6.64), but was not associated with non-atopic asthma (AOR=2.40, 95% CI=0.81-7.09). LT-alpha-NcoI*1 did not show an independent association with either atopic asthma or any one phenotype of specific IgE. The synergistic effect between these two genes was conducted, and the interaction between TNF-alpha-308*2 and LT-alpha-NcoI*1 polymorphisms was seen for atopic asthma (OR=2.59, 95% CI=1.10-6.10) when compared with all controls. CONCLUSION: We have concluded that TNF-alpha-308 may be a risk factor for atopic asthma, whereas the LT-alpha-NcoI polymorphism may modify risk to atopic asthma with TNF-alpha-308.

Detection of Divergent Hepatitis C Virus Envelope Sequences.

The nucleotide sequences of the putative envelope region (E1) and the junction between the E1 and envelope 2/nonstructural 1 (E2/NS1) region of the hepatitis C virus (HCV) genome are divergent among different genotypes. To characterize them, we introduced a set of nested primers that are conserved among four different genotypes (types I-IV) of HCV for polymerase chain reaction (PCR) amplification. The amplified products include the variable full-length E1 region, and the 5' end of the E2/NS1 region, the so-called hypervariable region-1 (HVR-1). Of 53 patients with histologically confirmed chronic liver disease and HCV viremia, type II virus was the most dominant strain as detected by the PCR genotyping method and the envelope region could be amplified in more than half of them irrespective of their genotypes. The specificity was confirmed by subsequent nucleotide sequence analysis. The positivity of envelope region PCR was not correlated with histologic diagnosis and hepatitis activities in these patients. Our results suggest that the nested primers can amplify the variable E1 and hypervariable 5' end of E2/NS1 of the HCV genome with moderate efficiency, and thus will be useful in future studies of HCV infections. Copyright 1994 S. Karger AG, Basel