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author:("Chen, zhikong")
1.  RECK Gene Polymorphism is Associated with Susceptibility and Prognosis of Wilms’ Tumor in Chinese Children 
Wilms’ tumor (WT) is the most common malignant renal tumor in children. Previous studies suggested the reversion-inducing, cysteine-rich protein with Kazal motifs (RECK) down-regulation might have a role in numerous human cancers. The current study was done to investigate the associations of RECK single-nucleotide polymorphisms (SNPs) with the WT susceptibility in Chinese children.
We analyzed 2 SNPs (rs10972727and rs11788747) in a total of 97 WT children and 194 healthy matched controls (1:2 ratio) by real-time PCR and PCR-RFLP genotyping analysis.
We found that the G allele of rs11788747 in the RECK gene was significantly associated with WT in Chinese children (OR=0.7, 95% CI: 0.45–0.99; P=0.042); as with another SNP rs10972727, however, no statistically significant difference was detected. Further analysis showed there was also a statistically significant difference in genotype frequencies between terminal tumor stage (P=0.026) and metastatic groups (P=0.002).
The present data indicate that there is a significant association between mutant G of rs11788747 in RECK and WT risk. G carriers with advanced tumor stage or with metastasis might have an increased risk of WT.
PMCID: PMC4496031  PMID: 26141647
Genes, Tumor Suppressor; Genes, Wilms Tumor; Nephrology; Polymorphism, Single Nucleotide
2.  Lipopolysaccharide-Induced Microglial Activation and Neuroprotection against Experimental Brain Injury Is Independent of Hematogenous TLR4 
Intraperitoneal injection of the Gram-negative bacterial endotoxin lipopolysaccharide (LPS) elicits a rapid innate immune response. While this systemic inflammatory response can be destructive, tolerable low doses of LPS render the brain transiently resistant to subsequent injuries. However, the mechanism by which microglia respond to LPS stimulation and participate in subsequent neuroprotection has not been documented. In this study, we first established a novel LPS treatment paradigm where mice were injected intraperitoneally with 1.0 mg/kg LPS for four consecutive days to globally activate CNS microglia. By using a reciprocal bone marrow transplantation procedure between wild-type and Toll-like receptor 4 (TLR4) mutant mice, we demonstrated that the presence of LPS receptor (TLR4) is not required on hematogenous immune cells but is required on cells that are not replaced by bone marrow transplantation, such as vascular endothelia and microglia, to transduce microglial activation and neuroprotection. Furthermore, we showed that activated microglia physically ensheathe cortical projection neurons, which have reduced axosomatic inhibitory synapses from the neuronal perikarya. In line with previous reports that inhibitory synapse reduction protects neurons from degeneration and injury, we show here that neuronal cell death and lesion volumes are significantly reduced in LPS-treated animals following experimental brain injury. Together, our results suggest that activated microglia participate in neuroprotection and that this neuroprotection is likely achieved through reduction of inhibitory axosomatic synapses. The therapeutic significance of these findings rests not only in identifying neuroprotective functions of microglia, but also in establishing the CNS location of TLR4 activation.
PMCID: PMC4461442  PMID: 22915113
3.  Phase II study of biomarker-guided neoadjuvant treatment strategy for IIIA-N2 non-small cell lung cancer based on epidermal growth factor receptor mutation status 
Neoadjuvant erlotinib and customized adjuvant therapy are appealing but controversial. The purpose of this study was to evaluate the role of biomarker-guided neoadjuvant treatment strategy in patients with IIIA-N2 non-small cell lung cancer (NSCLC) stratified by epidermal growth factor receptor (EGFR) mutation status.
Patients with resectable histologically documented stage IIIA-N2 NSCLC were assigned to a neoadjuvant erlotinib arm or a gemcitabine/carboplatin (GC) arm based on EGFR mutation status. The primary endpoint was response rate (RR). Secondary endpoints were progression-free survival (PFS) and overall survival (OS).
Twenty-four patients with IIIA-N2 NSCLC were enrolled in the trial from January 2008 until May 2011. The overall response rate was 41.7 % and the PFS and OS were 7.9 and 23.2 months, respectively, in overall population. The RR was 58.3 % (7/12) for the erlotinib arm with mutant EGFR and 25.0 % (3/12) for the GC arm with wild type EGFR (P = 0.18). Median PFS was 6.9 months versus 9.0 months, respectively (P = 0.071). Median OS was 14.5 months for the erlotinib arm and 28.1 months for the GC arm (P = 0.201). No unexpected toxicities were observed.
The primary endpoint was met and biomarker-guided neoadjuvant treatment strategy in patients with IIIA-N2 NSCLC is feasible. Erlotinib alone in neoadjuvant setting of EGFR mutant population showed an improved response but without survival benefits.
Trial registration NCT00600587
Electronic supplementary material
The online version of this article (doi:10.1186/s13045-015-0151-3) contains supplementary material, which is available to authorized users.
PMCID: PMC4455050  PMID: 25981169
IIIA-N2; Biomarker guided; EGFR mutation; Lung cancer; Neoadjuvant therapy
4.  Astrocyte response to IFN-γ limits IL-6-mediated microglia activation and progressive autoimmune encephalomyelitis 
Therapeutic modalities effective in patients with progressive forms of multiple sclerosis (MS) are limited. In a murine model of progressive MS, the sustained disability during the chronic phase of experimental autoimmune encephalomyelitis (EAE) correlated with elevated expression of interleukin (IL)-6, a cytokine with pleiotropic functions and therapeutic target for non-central nervous system (CNS) autoimmune disease. Sustained IL-6 expression in astrocytes restricted to areas of demyelination suggested that IL-6 plays a major role in disease progression during chronic EAE.
A progressive form of EAE was induced using transgenic mice expressing a dominant negative interferon-γ (IFN-γ) receptor alpha chain under control of human glial fibrillary acidic protein (GFAP) promoter (GFAPγR1Δ mice). The role of IL-6 in regulating progressive CNS autoimmunity was assessed by treating GFAPγR1Δ mice with anti-IL-6 neutralizing antibody during chronic EAE.
IL-6 neutralization restricted disease progression and decreased disability, myelin loss, and axonal damage without affecting astrogliosis. IL-6 blockade reduced CNS inflammation by limiting inflammatory cell proliferation; however, the relative frequencies of CNS leukocyte infiltrates, including the Th1, Th17, and Treg CD4 T cell subsets, were not altered. IL-6 blockade rather limited the activation and proliferation of microglia, which correlated with higher expression of Galectin-1, a regulator of microglia activation expressed by astrocytes.
These data demonstrate that astrocyte-derived IL-6 is a key mediator of progressive disease and support IL-6 blockade as a viable intervention strategy to combat progressive MS.
PMCID: PMC4410573  PMID: 25896970
Progressive multiple sclerosis; Experimental autoimmune encephalomyelitis; Astrocytes; Interferon-γ; Interleukin 6
5.  Molecular Diversity and Population Structure of a Worldwide Collection of Cultivated Tetraploid Alfalfa (Medicago sativa subsp. sativa L.) Germplasm as Revealed by Microsatellite Markers 
PLoS ONE  2015;10(4):e0124592.
Information on genetic diversity and population structure of a tetraploid alfalfa collection might be valuable in effective use of the genetic resources. A set of 336 worldwide genotypes of tetraploid alfalfa (Medicago sativa subsp. sativa L.) was genotyped using 85 genome-wide distributed SSR markers to reveal the genetic diversity and population structure in the alfalfa. Genetic diversity analysis identified a total of 1056 alleles across 85 marker loci. The average expected heterozygosity and polymorphism information content values were 0.677 and 0.638, respectively, showing high levels of genetic diversity in the cultivated tetraploid alfalfa germplasm. Comparison of genetic characteristics across chromosomes indicated regions of chromosomes 2 and 3 had the highest genetic diversity. A higher genetic diversity was detected in alfalfa landraces than that of wild materials and cultivars. Two populations were identified by the model-based population structure, principal coordinate and neighbor-joining analyses, corresponding to China and other parts of the world. However, lack of strictly correlation between clustering and geographic origins suggested extensive germplasm exchanges of alfalfa germplasm across diverse geographic regions. The quantitative analysis of the genetic diversity and population structure in this study could be useful for genetic and genomic analysis and utilization of the genetic variation in alfalfa breeding.
PMCID: PMC4406709  PMID: 25901573
6.  Aquaporins as diagnostic and therapeutic targets in cancer: How far we are? 
Aquaporins (AQPs) are a family of water channel proteins distributed in various human tissues, responsible for the transport of small solutes such as glycerol, even gas and ions. The expression of AQPs has been found in more than 20 human cancer types and is significantly correlated with the severity of histological tumors and prognosis of patients with cancer. More recent evidence showed that AQPs could also play a role in tumor-associated edema, tumor cell proliferation and migration, and tumor angiogenesis in solid and hematological tumors. Inhibitors of AQPs in tumor cells and microvessels have been suggested as new therapeutic strategies. The present review overviews AQPs structures, expression variation among normal tissues and tumors, AQPs functions and roles in the development of cancer with special focuses on lung, colorectal, liver, brain and breast cancers, and potential AQPs-target inhibitors. We call the special attention to consider AQPs important as diagnostic and therapeutic biomarkers. It may be a novel anticancer therapy by the AQPs inhibition.
PMCID: PMC4382841  PMID: 25886458
AQPs; Cancer; Water channel; Inhibition; Biomarkers
7.  Loss of plexin-B3 in hepatocellular carcinoma 
Plexins are the primary receptors of semaphorins, and participate in the majority of intracellular pathways triggered by semaphorins, including the regulation of cell adhesion and the motility of numerous cell types. Recently, several studies have reported that plexins can significantly affect different aspects of cancer cell biology, and the aberrant expression of plexins has been observed in a wide variety of tumor types. However, the expression and role of plexin-B3 in hepatocellular carcinoma (HCC) is yet to be investigated. In the present study, plexin-B3 expression was measured in 14 paired HCC samples and the corresponding adjacent non-cancerous tissue by quantitative polymerase chain reaction and western blot analysis. The results indicated that the mRNA and protein expression levels of plexin-B3 were downregulated in HCC samples when compared with the corresponding adjacent non-cancerous tissue. In order to elucidate the correlation between clinicopathological data and the expression of plexin-B3 in patients with HCC, 84 HCC archived specimens were analyzed by immunohistochemistry (IHC). The IHC results revealed that the protein expression level of plexin-B3 was lower in the HCC samples compared with the corresponding adjacent non-cancerous tissue, and plexin-B3 underexpression was correlated with the patient gender and tumor size. In conclusion, these results indicated that loss of plexin-B3 in HCC may be of predictive value for the occurrence and progression of HCC. Thus, plexin-B3 may be a promising biomarker for the diagnosis and treatment of tumors in the future.
PMCID: PMC4353781  PMID: 25780417
plexin-B3; hepatocellular carcinoma
8.  Chronic Deep Cerebellar Stimulation Promotes Long-Term Potentiation, Microstructural Plasticity, and Reorganization of Perilesional Cortical Representation in a Rodent Model 
The Journal of Neuroscience  2014;34(27):9040-9050.
Control over postinjury CNS plasticity is a major frontier of science that, if conquered, would open new avenues for treatment of neurological disorders. Here we investigate the functional, physiological, and structural changes in the cerebral cortex associated with chronic deep brain stimulation of the cerebellar output, a treatment approach that has been shown to improve postischemia motor recovery in a rodent model of cortical infarcts. Long–Evans rats were pretrained on the pasta-matrix retrieval task, followed by induction of focal cortical ischemia and implantation of a macroelectrode in the contralesional lateral cerebellar nucleus. Animals were assigned to one of three treatment groups pseudorandomly to balance severity of poststroke motor deficits: REGULAR stimulation, BURST stimulation, or SHAM. Treatment initiated 2 weeks post surgery and continued for 5 weeks. At the end, animals were randomly selected for perilesional intracortical microstimulation mapping and tissue sampling for Western blot analysis or contributed tissue for 3D electron microscopy.
Evidence of enhanced cortical plasticity with therapeutically effective stimulation is shown, marked by greater perilesional reorganization in stimulation- treated animals versus SHAM. BURST stimulation was significantly effective for promoting distal forepaw cortical representation. Stimulation-treated animals showed a twofold increase in synaptic density compared with SHAM. In addition, treated animals demonstrated increased expression of synaptic markers of long-term potentiation and plasticity, including synaptophysin, NMDAR1, CaMKII, and PSD95. These findings provide a critical foundation of how deep cerebellar stimulation may guide plastic reparative reorganization after nonprogressive brain injury and indicate strong translational potential.
PMCID: PMC4078081  PMID: 24990924
cerebello-thalamo-cortical; deep brain stimulation; focal lesion; plasticity; reorganization
9.  Cuprizone does not induce CNS demyelination in nonhuman primates 
Cognitive decline is a common symptom in multiple sclerosis patients, with profound effects on the quality of life. A nonhuman primate model of multiple sclerosis would be best suited to test the effects of demyelination on complex cognitive functions such as learning and reasoning. Cuprizone has been shown to reliably induce brain demyelination in mice. To establish a nonhuman primate model of multiple sclerosis, young adult cynomolgus monkeys were administered cuprizone per os as a dietary supplement. The subjects received increasing cuprizone doses (0.3–3% of diet) for up to 18 weeks. Magnetic resonance imaging and immunohistological analyses did not reveal demyelination in these monkeys.
PMCID: PMC4338961  PMID: 25750925
10.  Green tea extracts reduce adipogenesis by decreasing expression of transcription factors C/EBPα and PPARγ 
Objectives: This study is to determine if green tea (Camellia sinensis) extracts (GTE) affects adipogenesis and further investigate the related molecular mechanisms. Methods: Patients with metabolic syndrome were recruited in this study. Of them, 70 patients received GTE and 64 received water to serve as the control group. The human serum adiponectin, visfatin, and leptin concentrations were determined by enzyme-linked immunosorbent assay. Adipogenesis of 3T3-L1 preadipocytes was induced with reagents and then the cells were treated with GTE. The lipids were stained with Oil Red O for analysis of adipogenesis of 3T3-L1 preadipocytes. The 3T3-L1 preadipocytes were treated with increasing concentrations (0.2-0.5%, w/v) of GTE for 2 days and the cell viability was determined by MTT assay. Reverse transcription real-time PCR and immunoblotting assays were performed to determine RNA and protein levels of relative molecules. Results: GTE increases the serum concentrations of adiponectin but decreases visfatin levels in patients received GTE. The leptin concentrations in serum were not significantly affected. The GTE reduces the adipogenesis-induced lipid accumulation in 3T3-L1 preadipocytes. GTE decreases the mRNA and protein expression of adipogenic transcription factors C/EBPα and PPARγ in 3T3-L1 cells. Expression levels of the adipocyte-specific genes encoding adipocyte protein 2, lipoprotein lipase, and glucose transporter 4 were also decreased by GTE. Furthermore, it was found that GTE reduces phosphorylation of Akt during adipocyte differentiation. Conclusions: GTE reduces adipogenesis by decreasing expression of transcription factors C/EBPα and PPARγ by reduction of phosphorylation of Akt during adipocyte differentiation.
PMCID: PMC4307434  PMID: 25663987
Green tea extracts; adipogenesis; transcription factor; C/EBPα; PPARγ
11.  IL-4 confers resistance to IL-27-mediated suppression on CD4+ T cells by impairing STAT1 signaling 
Th2 cells play a critical role in the pathogenesis of allergic asthma. Established Th2 cells have been shown to resist reprogramming into Th1 cells. The inherent stability of Th2 cells poses a significant barrier to treating allergic diseases.
We sought to understand the mechanisms by which CD4+ T cells from asthmatic patients resist the IL-27-mediated inhibition.
We isolated and cultured CD4+ T cells from both healthy individuals and allergic asthmatic patients in order to test whether IL-27 can inhibit IL-4 production by the cultured CD4+ T cells using ELISA. Culturing conditions that resulted in resistance to IL-27 were determined using both murine and human CD4+ T cell culture systems. STAT1 phosphorylation was analyzed by Western blot and flow cytometry. Suppressor of cytokine signaling (Socs) mRNA expression was measured by quantitative PCR. The small interfering RNA method was used to knockdown the expression of Socs3 mRNA.
Main Results
We demonstrated that CD4+ T cells from asthmatic patients resisted the suppression of IL-4 production mediated by IL-27. We observed that repeated exposure to Th2-inducing conditions rendered healthy human CD4+ T cells resistant to IL-27-mediated inhibition. Using an in vitro murine culture system, we further demonstrated that repeated or higher doses of IL-4 stimulation, but not IL-2 stimulation, upregulated Socs3 mRNA expression and impaired IL-27-induced STAT1 phosphorylation. The Knockdown of Socs3 mRNA expression restored IL-27-induced STAT1 phosphorylation and IL-27-mediated inhibition of IL-4-production.
Our findings demonstrate that differentiated Th2 cells can resist IL-27-induced reprogramming toward Th1 cells by downregulating STAT1 phosphorylation and likely explain why the CD4+ T cells of asthmatic patients are resistant to IL-27-mediated inhibition.
PMCID: PMC3788709  PMID: 23958647
asthma; Th2 stability; IL-4; STAT1 signaling; SOCS3; IL-27
ACS chemical biology  2014;9(4):1003-1014.
NRAS is the second most frequently mutated gene in melanoma. Previous reports have demonstrated the sensitivity of cancer cell lines carrying KRAS mutations to apoptosis initiated by inhibition of protein kinase C delta (PKCδ). Here, we report that PKCδ inhibition is cytotoxic in melanomas with primary NRAS mutations. Novel small-molecule inhibitors of PKCδ were designed as chimeric hybrids of two naturally-occurring PKCδ inhibitors, staurosporine and rottlerin. The specific hypothesis interrogated and validated is that combining two domains of two naturally-occurring PKCδ inhibitors into a chimeric or hybrid structure retains biochemical and biological activity, and improves PKCδ isozyme selectivity. We have devised a potentially general synthetic protocol to make these chimeric species using Molander trifluorborate coupling chemistry. Inhibition of PKCδ, by siRNA or small molecule inhibitors, suppressed the growth of multiple melanoma cell lines carrying NRAS mutations, mediated via caspase-dependent apoptosis. Following PKCδ inhibition, the stress-responsive JNK pathway was activated, leading to the activation of H2AX. Consistent with recent reports on the apoptotic role of phospho-H2AX, knockdown of H2AX prior to PKCδ inhibition mitigated the induction of caspase-dependent apoptosis. Furthermore, PKCδ inhibition effectively induced cytotoxicity in BRAF-mutant melanoma cell lines that had evolved resistance to a BRAF inhibitor, suggesting the potential clinical application of targeting PKCδ in patients who have relapsed following treatment with BRAF inhibitors. Taken together, the present work demonstrates that inhibition of PKCδ by novel small molecule inhibitors causes caspase-dependent apoptosis mediated via the JNK-H2AX pathway in melanomas with NRAS mutations or BRAF inhibitor-resistance.
PMCID: PMC4160068  PMID: 24506253
13.  Microglial displacement of inhibitory synapses provides neuroprotection in the adult brain 
Nature Communications  2014;5:4486.
Microglia actively survey the brain microenvironment and play essential roles in sculpting synaptic connections during brain development. While microglial functions in the adult brain are less clear, activated microglia can closely appose neuronal cell bodies and displace axosomatic presynaptic terminals. Microglia-mediated stripping of presynaptic terminals is considered neuroprotective, but the cellular and molecular mechanisms are poorly defined. Using 3D electron microscopy, we demonstrate that activated microglia displace inhibitory presynaptic terminals from cortical neurons in adult mice. Electrophysiological recordings further establish that the reduction in inhibitory GABAergic synapses increased synchronized firing of cortical neurons in γ-frequency band. Increased neuronal activity results in the calcium-mediated activation of CaM kinase IV, phosphorylation of CREB, increased expression of antiapoptotic and neurotrophic molecules and reduced apoptosis of cortical neurons following injury. These results indicate that activated microglia can protect the adult brain by migrating to inhibitory synapses and displacing them from cortical neurons.
Microglia play essential roles in sculpting synaptic connections during brain development but their role in the adult brain is less clear. Here the authors show that activated microglia can prophylactically protect the adult rodent brain from injury by migrating to and displacing inhibitory synapses from cortical neurons.
PMCID: PMC4109015  PMID: 25047355
14.  Activation of the germ-cell potential of human bone marrow-derived cells by a chemical carcinogen 
Scientific Reports  2014;4:5564.
Embryonic/germ cell traits are common in malignant tumors and are thought to be involved in malignant tumor behaviors. The reasons why tumors show strong embryonic/germline traits (displaced germ cells or gametogenic programming reactivation) are controversial. Here, we show that a chemical carcinogen, 3-methyl-cholanthrene (3-MCA), can trigger the germ-cell potential of human bone marrow-derived cells (hBMDCs). 3-MCA promoted the generation of germ cell-like cells from induced hBMDCs that had undergone malignant transformation, whereas similar results were not observed in the parallel hBMDC culture at the same time point. The malignant transformed hBMDCs spontaneously and more efficiently generated into germ cell-like cells even at the single-cell level. The germ cell-like cells from induced hBMDCs were similar to natural germ cells in many aspects, including morphology, gene expression, proliferation, migration, further development, and teratocarcinoma formation. Therefore, our results demonstrate that a chemical carcinogen can reactivate the germline phenotypes of human somatic tissue-derived cells, which might provide a novel idea to tumor biology and therapy.
PMCID: PMC4083294  PMID: 24998261
15.  Gene Expression Profiling Reveals Large Regulatory Switches between Succeeding Stipe Stages in Volvariella volvacea 
PLoS ONE  2014;9(5):e97789.
The edible mushroom Volvariella volvacea is an important crop in Southeast Asia and is predominantly harvested in the egg stage. One of the main factors that negatively affect its yield and value is the rapid transition from the egg to the elongation stage, which has a decreased commodity value and shelf life. To improve our understanding of the changes during stipe development and the transition from egg to elongation stage in particular, we analyzed gene transcription in stipe tissue of V. volvacea using 3′-tag based digital expression profiling. Stipe development turned out to be fairly complex with high numbers of expressed genes, and regulation of stage differences is mediated mainly by changes in expression levels of genes, rather than on/off modulation. Most explicit is the strong up-regulation of cell division from button to egg, and the very strong down-regulation hereof from egg to elongation, that continues in the maturation stage. Button and egg share cell division as means of growth, followed by a major developmental shift towards rapid stipe elongation based on cell extension as demonstrated by inactivation of cell division throughout elongation and maturation. Examination of regulatory genes up-regulated from egg to elongation identified three potential high upstream regulators for this switch. The new insights in stipe dynamics, together with a series of new target genes, will provide a sound base for further studies on the developmental mechanisms of mushroom stipes and the switch from egg to elongation in V. volvacea in particular.
PMCID: PMC4035324  PMID: 24867220
16.  A nontoxic and low-cost hydrothermal route for synthesis of hierarchical Cu2ZnSnS4 particles 
Nanoscale Research Letters  2014;9(1):208.
We explore a facile and nontoxic hydrothermal route for synthesis of a Cu2ZnSnS4 nanocrystalline material by using l-cysteine as the sulfur source and ethylenediaminetetraacetic acid (EDTA) as the complexing agent. The effects of the amount of EDTA, the mole ratio of the three metal ions, and the hydrothermal temperature and time on the phase composition of the obtained product have been systematically investigated. The addition of EDTA and an excessive dose of ZnCl2 in the hydrothermal reaction system favor the generation of kesterite Cu2ZnSnS4. Pure kesterite Cu2ZnSnS4 has been synthesized at 180°C for 12 h from the reaction system containing 2 mmol of EDTA at 2:2:1 of Cu/Zn/Sn. It is confirmed by Raman spectroscopy that those binary and ternary phases are absent in the kesterite Cu2ZnSnS4 product. The kesterite Cu2ZnSnS4 material synthesized by the hydrothermal process consists of flower-like particles with 250 to 400 nm in size. It is revealed that the flower-like particles are assembled from single-crystal Cu2ZnSnS4 nanoflakes with ca. 20 nm in size. The band gap of the Cu2ZnSnS4 nanocrystalline material is estimated to be 1.55 eV. The films fabricated from the hierarchical Cu2ZnSnS4 particles exhibit fast photocurrent responses under intermittent visible-light irradiation, implying that they show potentials for use in solar cells and photocatalysis.
PMCID: PMC4013540  PMID: 24855463
Cu2ZnSnS4; Nanocrystalline material; Hierarchical particles; Hydrothermal process; Photoelectrochemical property
17.  Semi-automated method for estimating lesion volumes 
Journal of neuroscience methods  2012;213(1):76-83.
Accurately measuring the volume of tissue damage in experimental lesion models is crucial to adequately control for the extent and location of the lesion, variables that can dramatically bias the outcome of preclinical studies. Many of the current commonly used techniques for this assessment, such as measuring the lesion volume with primitive software macros and plotting the lesion location manually using atlases, are time-consuming and offer limited precision. Here we present an easy to use semi-automated computational method for determining lesion volume and location, designed to increase precision and reduce the manual labor required. We compared this novel method to currently used methods and demonstrate that this tool is comparable or superior to current techniques in terms of precision and has distinct advantages with respect to user interface, labor intensiveness and quality of data presentation.
PMCID: PMC3570743  PMID: 23261655
lesion volume; stroke; traumatic brain injury; cell death; lesion estimation; computational method
18.  Protein kinase C-delta inactivation inhibits the proliferation and survival of cancer stem cells in culture and in vivo 
BMC Cancer  2014;14:90.
A subpopulation of tumor cells with distinct stem-like properties (cancer stem-like cells, CSCs) may be responsible for tumor initiation, invasive growth, and possibly dissemination to distant organ sites. CSCs exhibit a spectrum of biological, biochemical, and molecular features that are consistent with a stem-like phenotype, including growth as non-adherent spheres (clonogenic potential), ability to form a new tumor in xenograft assays, unlimited self-renewal, and the capacity for multipotency and lineage-specific differentiation. PKCδ is a novel class serine/threonine kinase of the PKC family, and functions in a number of cellular activities including cell proliferation, survival or apoptosis. PKCδ has previously been validated as a synthetic lethal target in cancer cells of multiple types with aberrant activation of Ras signaling, using both genetic (shRNA and dominant-negative PKCδ mutants) and small molecule inhibitors. In contrast, PKCδ is not required for the proliferation or survival of normal cells, suggesting the potential tumor-specificity of a PKCδ-targeted approach.
shRNA knockdown was used validate PKCδ as a target in primary cancer stem cell lines and stem-like cells derived from human tumor cell lines, including breast, pancreatic, prostate and melanoma tumor cells. Novel and potent small molecule PKCδ inhibitors were employed in assays monitoring apoptosis, proliferation and clonogenic capacity of these cancer stem-like populations. Significant differences among data sets were determined using two-tailed Student’s t tests or ANOVA.
We demonstrate that CSC-like populations derived from multiple types of human primary tumors, from human cancer cell lines, and from transformed human cells, require PKCδ activity and are susceptible to agents which deplete PKCδ protein or activity. Inhibition of PKCδ by specific genetic strategies (shRNA) or by novel small molecule inhibitors is growth inhibitory and cytotoxic to multiple types of human CSCs in culture. PKCδ inhibition efficiently prevents tumor sphere outgrowth from tumor cell cultures, with exposure times as short as six hours. Small-molecule PKCδ inhibitors also inhibit human CSC growth in vivo in a mouse xenograft model.
These findings suggest that the novel PKC isozyme PKCδ may represent a new molecular target for cancer stem cell populations.
PMCID: PMC3927586  PMID: 24528676
Protein Kinase C isozymes; Synthetic lethal interaction; Cancer-initiating cell; Xenograft tumor model
19.  Effect of sericin on diabetic hippocampal growth hormone/insulin-like growth factor 1 axis 
Neural Regeneration Research  2013;8(19):1756-1764.
Previous studies have shown that sericin extracted from silk cocoon significantly reduces blood glucose levels and protects the nervous system against diabetes mellitus. In this study, a rat type 2 diabetes mellitus model was established by intraperitoneal injection of 25 mg/kg streptozotocin for 3 successive days, following which the rats were treated with sericin for 35 days. After treatment, the blood glucose levels of the diabetic rats decreased significantly, the growth hormone level in serum and its expression in the hippocampus decreased significantly, while the insulin-like growth factor-1 level in serum and insulin-like growth factor-1 and growth hormone receptor expression in the hippocampus increased significantly. The experimental findings indicate that sericin improves disorders of the growth hormone/insulin-like growth factor 1 axis to alleviate hippocampal damage in diabetic rats.
PMCID: PMC4145953  PMID: 25206472
neural regeneration; traditional Chinese medicine; sericin; type 2 diabetes mellitus; hippocampus; growth hormone; insulin-like growth factor 1; growth hormone receptor; growth hormone/insulin-like growth factor 1 axis; streptozotocin; blood glucose; western blot assay; reverse transcription-PCR; grants-supported paper; neuroregeneration
20.  Downregulation of Beclin1 and Impairment of Autophagy in a Small Population of Colorectal Cancer 
Digestive Diseases and Sciences  2013;58(10):2887-2894.
Autophagy is a highly conserved mechanism for degradation and recycling of long-lived proteins and damaged organelle to maintain cell homeostasis. Deregulation of autophagy has been associated with tumorigenesis. Beclin 1 is an essential autophagy protein and its upregulation has been observed in most colorectal cancer tissues. However, there is a small population of colorectal cancers with downregulation of Beclin 1.
The purpose of this study was to investigate the role autophagy plays in colorectal cancers with downregulaion of Beclin 1.
LC3 protein, an autophagosome marker, was assessed by ICH and WB in colorectal cancers tissues. An anti-tumor effect of Beclin 1 was examined by introducing exogenous Beclin 1 in vitro. Colony formation assay, growth curves and mouse xenograft were analysed.
Our results showed that LC3 was suppressed in the colorectal cancers (9.86 %) with downregulation of Beclin 1. Moreover, overexpression of Beclin 1 inhibited colorectal cancer cell growth and enhanced the rapamycin-induced antitumor effect in vitro.
Downregulation of Beclin 1 and autophagy inhibition play an important role in a part of colorectal cancers. Activating autophagy or overexperssion of Beclin 1 may be an effective treatment for some colorectal cancers. Detection of expression profile of Beclin 1 in colorectal cancers could be a strategy for new diagnostic and therapeutic methods.
Electronic supplementary material
The online version of this article (doi:10.1007/s10620-013-2732-8) contains supplementary material, which is available to authorized users.
PMCID: PMC3781302  PMID: 23812859
Colorectal cancer; Beclin 1; Autophagy; Rapamycin
21.  Sericin protects against diabetes-induced injuries in sciatic nerve and related nerve cells★ 
Neural Regeneration Research  2013;8(6):506-513.
Sericin from discarded silkworm cocoons of silk reeling has been used in different fields, such as cosmetology, skin care, nutrition, and oncology. The present study established a rat model of type 2 diabetes by consecutive intraperitoneal injections of low-dose (25 mg/kg) streptozotocin. After intragastrical perfusion of sericin for 35 days, blood glucose levels significantly declined, and the expression of neurofilament protein in the sciatic nerve and nerve growth factor in L4–6 spinal ganglion and anterior horn cells significantly increased. However, the expression of neuropeptide Y in spinal ganglion and anterior horn cells significantly decreased in model rats. These findings indicate that sericin protected the sciatic nerve and related nerve cells against injury in a rat type 2 diabetic model by upregulating the expression of neurofilament protein in the sciatic nerve and nerve growth factor in spinal ganglion and anterior horn cells, and downregulating the expression of neuropeptide Y in spinal ganglion and anterior horn cells.
PMCID: PMC4146054  PMID: 25206693
neural regeneration; traditional Chinese medicine; peripheral nerve injury; diabetes mellitus; sericin; sciatic nerve; spinal ganglion cells; anterior horn cells; nerve cells; neurofilament protein; nerve growth factor; neuropeptide Y; streptozotocin; photographs-containing paper; neuroregeneration
22.  Long-term effects of a combination of D-penicillamine and zinc salts in the treatment of Wilson’s disease in children 
The aim of this study was to investigate the effectiveness of a high-dose zinc sulfate and low-dose D-penicillamine combination in the treatment of pediatric Wilson’s disease (WD). A retropective chart review of 65 patients with WD was conducted. These patients received D-penicillamine (8–10 mg/kg/day) and zinc sulfate as the primary treatment. The pediatric dose of elemental zinc is 68–85 mg/day until 6 years of age, 85–136 mg/day until 8 years of age, 136–170 mg/day until 10 years of age and then 170 mg/day, in 3 divided doses 1 h before meals. After clinical and biochemical improvement or stabilization, zinc sulfate alone was administered as the maintenance therapy. Under treatment, the majority of patients (89.2%) had a favourable outcome and 3 patients succumbed due to poor therapy compliance. No penicillamine-induced neurological deterioration was noted and side-effects were observed in <11% of patients over the entire follow-up period. Benefical results on the liver and neurological symptoms were reported following extremely long-term treatment with a combination of low-dose D-penicillamine and high-dose zinc sulfate. Therefore, this regimen is an effective and safe treatment for children with WD.
PMCID: PMC3628594  PMID: 23599735
Wilson’s disease; D-penicillamine; zinc sulfate; child
23.  Protein Kinase C-delta Inactivation Inhibits Cellular Proliferation and Decreases Survival in Human Neuroendocrine Tumors 
Endocrine-related cancer  2011;18(6):759-771.
The concept of targeting cancer therapeutics towards specific mutations or abnormalities in tumor cells which are not found in normal tissues has the potential advantages of high selectivity for the tumor and correspondingly low secondary toxicities. Many human malignancies display activating mutations in the Ras family of signal-transducing genes or over-activity of p21Ras-signaling pathways. Carcinoid and other neuroendocrine tumors similarly have been demonstrated to have activation of Ras signaling directly by mutations in Ras, indirectly by loss of Ras-regulatory proteins, or via constitutive activation of upstream or downstream effector pathways of Ras, such as growth factor receptors or PI3-Kinase and Raf/MAP kinases. We previously reported that aberrant activation of Ras signaling sensitizes cells to apoptosis when the activity of the PKCδ isozyme is suppressed, and that PKCδ suppression is not toxic to cells with normal levels of p21Ras signaling. We demonstrate here that inhibition of PKCδ by a number of independent means, including genetic mechanisms (shRNA) or small molecule inhibitors, is able to efficiently and selectively repress the growth of human neuroendocrine cell lines derived from bronchopulmonary, foregut or hindgut tumors. PKCδ inhibition in these tumors also efficiently induced apoptosis. Exposure to small-molecule inhibitors of PKCδ over a period of 24 hr is sufficient to significantly suppress cell growth and clonogenic capacity of these tumor cell lines.
Neuroendocrine tumors are typically refractory to conventional therapeutic approaches. This Ras-targeted therapeutic approach, mediated through PKCδ suppression, which selectively takes advantage of the very oncogenic mutations which contribute to the malignancy of the tumor, may hold potential as a novel therapeutic modality.
PMCID: PMC3527126  PMID: 21990324
carcinoid; Ras; apoptosis; cancer
24.  Sericin can reduce hippocampal neuronal apoptosis by activating the Akt signal transduction pathway in a rat model of diabetes mellitus☆ 
Neural Regeneration Research  2012;7(3):197-201.
In the present study, a rat model of type 2 diabetes mellitus was established by continuous peritoneal injection of streptozotocin. Following intragastric perfusion of sericin for 35 days, blood glucose levels significantly reduced, neuronal apoptosis in the hippocampal CA1 region decreased, hippocampal phosphorylated Akt and nuclear factor kappa B expression were enhanced, but Bcl-xL/Bcl-2 associated death promoter expression decreased. Results demonstrated that sericin can reduce hippocampal neuronal apoptosis in a rat model of diabetes mellitus by regulating abnormal changes in the Akt signal transduction pathway.
PMCID: PMC4353114  PMID: 25767499
sericin; type 2 diabetes mellitus; hippocampus; apoptosis; Akt signal transduction pathway; neural regeneration
25.  Role of aquaporin 5 in antigen-induced airway inflammation and mucous hyperproduction in mice 
Airway inflammation and mucus hyperproduction play the central role in the development of asthma, although the mechanisms remain unclear. The aquaporin (AQP)-5 may be involved in the process due to its contribution to the volume of liquid secreted from the airways. The present study firstly found the overexpression of AQP5 in the airway epithelium and submucosal glands of asthmatics. Furthermore, we aimed at evaluating the role of AQP5 in airway inflammation and mucous hyperproductions during chronic allergic responses to house dust mite (HDM). Bronchoalveolar lavage levels of interleukin (IL)-2, IL-4, IL-10, interferon-γ and Mucin 5AC (MUC5AC), and number of peribronchial and perivascular cells were measured in AQP5 wild-type and AQP5 knockout (KO) mice. We found that HDM induced airway inflammation, lung Th2 cell accumulation and mucin hypersecretion in C57BL/6 mice rather than AQP5 KO mice. Expression of MUC5AC and MUC5B proteins and genes in the lung tissue was significantly lower in AQP5 KO mice. Thus, our results implicate involvement of AQP5 in the development of airway inflammation and mucous hyperproduction during chronic asthma.
PMCID: PMC4373335  PMID: 20550619
aquaporins; mucin; house dust mite; chronic allergy; inflammation

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