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PTEN mutation occurs in a variety of aggressive cancers and is associated with poor patient outcomes. Recent studies have linked mutational loss of PTEN to reduced RAD51 expression and function, a key factor involved in the homologous recombination (HR) pathway. However, these studies remain controversial, as they fail to establish a definitive causal link to RAD51 expression that is PTEN-dependent, while other studies have not been able to recapitulate the relationship between the PTEN expression and the RAD51/HR function. Resolution of this apparent conundrum is essential due to the clinically-significant implication that PTEN-deficient tumors may be sensitive to poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) commonly used in the clinical management of BRCA -mutated and other HR-deficient (HRD) tumors., Methods: Primary Pten -deficient (and corresponding wild-type) mouse embryonic fibroblasts (MEFs) and astrocytes and PTEN -null human tumor cell lines and primary cells were assessed for RAD51 expression (via the Western blot analysis) and DNA damage repair analyses (via alkali comet and γH2AX foci assays). RAD51 foci analysis was used to measure HR-dependent DNA repair. Xrcc2 -deficient MEFs served as an HR-deficient control, while the stable knockdown of RAD51 ( shRAD51 ) served to control for the relative RAD51/HR-mediated repair and the phospho-53BP1 foci analysis served to confirm and measure non-homologous end joining (NHEJ) activity in PTEN-deficient and shRAD51 -expressing (HRD) lines. Cell proliferation studies were used to measure any potential added sensitivity of PTEN -null cells to the clinically-relevant PARPi, olaparib. RAD51 levels and DNA damage response signaling were assessed in PTEN-mutant brain tumor initiating cells (BTICs) derived from primary and recurrent glioblastoma multiforme (GBM) patients, while expression of RAD51 and its paralogs were examined as a function of the PTEN status in the RNA expression datasets isolated from primary GBM tumor specimens and BTICs., Results: Pten knockout primary murine cells display unaltered RAD51 expression, endogenous and DNA strand break-induced RAD51 foci and robust DNA repair activity. Defective HR was only observed in the cells lacking Xrcc2 . Likewise, human glioblastoma multiforme (GBM) cell lines with known PTEN deficiency (U87, PTEN -mutated; U251 and U373, PTEN -null) show apparent expression of RAD51 and display efficient DNA repair activity. Only GBM cells stably expressing shRNAs against RAD51 ( shRAD51 ) display dysfunctional DNA repair activity and reduced proliferative capacity, which is exacerbated by PARPi treatment. Furthermore, GBM patient-derived BTICs displayed robust RAD51 expression and intact DNA damage response signaling in spite of PTEN -inactivating mutations. RNA expression analysis of primary GBM tissue specimens and BTICs demonstrate stable levels of RAD51 and its paralogs ( RAD51B, RAD51C, RAD51D, XRCC2, XRCC3 , and DMC1 ), regardless of the PTEN mutational status., Conclusions: Our findings demonstrate definitively that PTEN loss does not alter the RAD51 expression, its paralogs, or the HR activity. Furthermore, deficiency in PTEN alone is not sufficient to impart enhanced sensitivity to PARPi associated with HRD. This study is the first to unequivocally demonstrate that PTEN deficiency is not linked to the RAD51 expression or the HR activity amongst primary neural and non-neural Pten -null cells, PTEN-deficient tumor cell lines, and primary PTEN -mutant GBM patient-derived tissue specimens and BTICs.

The most common cause of vision loss among the elderly is age‑related macular degeneration (AMD). The aim of the present study was to investigate the potential cytoprotective and anti‑inflammatory effects of carbon monoxide‑releasing molecules (CORMs), and their ability to activate the expression of nuclear factor erythroid 2‑related factor 2 (Nrf2)‑related genes in human retinal pigment epithelium (RPE) cells, as well as the inhibition of endothelial cell migration. It was first determined that CORM2 and CORM3 suppressed blue light‑induced cell damage. In addition, a decrease in the level of cleaved poly(ADP‑ribose) polymerase 1 protein and dissipation of mitochondrial membrane potential were considered to reflect the anti‑apoptotic activity of CORMs. Furthermore, CORM2 induced Nrf‑2 activation and the expression of the Nrf2‑related genes heme oxygenase‑1 and glutamate‑cysteine ligase. Pretreatment with CORM2 abolished the blue light‑induced increase in oxidative stress, suggesting that CORM2‑induced antioxidant activity was involved in the cytoprotection against blue light. It was also demonstrated that CORMs markedly suppressed tumor necrosis factor (TNF)α‑induced intercellular adhesion molecule‑1 expression. Moreover, it was further observed that CORMs exert their inhibitory effects through blocking nuclear factor‑κB/p65 nuclear translocation and IκBα degradation in TNFα‑treated RPE cells. It was observed that CORM2, but not CORM3, protected against oxidative stress‑induced cell damage. CORMs abolished vascular endothelial growth factor‑induced migration of endothelial cells. The findings of the present study demonstrated the cytoprotective, antioxidant and anti‑inflammatory effects of CORMs on RPE cells and anti‑angiogenic effects on endothelial cells, suggesting the potential clinical application of CORMs as anti‑AMD agents.

Oxidative stress induced by long-term cyclosporine A (CsA) administration is a major cause of chronic nephrotoxicity, which is characterized by tubular atrophy, tubular cell apoptosis, and interstitial fibrosis in the progression of organ transplantation. Although hydrogen-rich water (HRW) has been used to prevent various oxidative stress-related diseases, its underlying mechanisms remain unclear. This study investigated the effects of HRW on CsA-induced nephrotoxicity and its potential mechanisms. After administration of CsA (25 mg/kg/day), rats were treated with or without HRW (12 mL/kg) for 4 weeks. Renal function and vascular activity were investigated. Histological changes in kidney tissues were analyzed using Masson's trichrome and terminal deoxynucleotidyl transferase dUTP nick-end labeling stains. Oxidative stress markers and the activation of the Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway were also measured. We found that CsA increased the levels of reactive oxygen species (ROS) and malonaldehyde (MDA), but it reduced glutathione (GSH) and superoxide dismutase (SOD) levels. Such alterations induced vascular dysfunction, tubular atrophy, interstitial fibrosis, and tubular apoptosis. This was evident secondary to an increase in urinary protein, serum creatinine, and blood urea nitrogen, ultimately leading to renal dysfunction. Conversely, HRW decreased levels of ROS and MDA while increasing the activity of GSH and SOD. This was accompanied by an improvement in vascular and renal function. Moreover, HRW significantly decreased the level of Keap1 and increased the expression of Nrf2, NADPH dehydrogenase quinone 1, and heme oxygenase 1. In conclusion, HRW restored the balance of redox status, suppressed oxidative stress damage, and improved kidney function induced by CsA via activation of the Keap1/Nrf2 signaling pathway., (© 2020 Wiley Periodicals, Inc.)

Background: MYC transcriptional factors are members of the bHLH (basic helix-loop-helix) superfamily, and play important roles in plant growth and development. Recent studies have revealed that some MYCs are involved in the crosstalk between Jasmonic acid regulatory pathway and light signaling in Arabidopsis, but such kinds of studies are rare in wheat, especially in photo-thermo-sensitive genic male sterile (PTGMS) wheat line., Results: 27 non-redundant MYC gene copies, which belonged to 11 TaMYC genes, were identified in the whole genome of wheat (Chinese Spring). These gene copies were distributed on 13 different chromosomes, respectively. Based on the results of phylogenetic analysis, 27 TaMYC gene copies were clustered into group I, group III, and group IV. The identified TaMYC genes copies contained different numbers of light, stress, and hormone-responsive regulatory elements in their 1500 base pair promoter regions. Besides, we found that TaMYC3 was expressed highly in stem, TaMYC5 and TaMYC9 were expressed specially in glume, and the rest of TaMYC genes were expressed in all tissues (root, stem, leaf, pistil, stamen, and glume) of the PTGMS line BS366. Moreover, we found that TaMYC3, TaMYC7, TaMYC9, and TaMYC10 were highly sensitive to methyl jasmonate (MeJA), and other TaMYC genes responded at different levels. Furthermore, we confirmed the expression profiles of TaMYC family members under different light quality and plant hormone stimuli, and abiotic stresses. Finally, we predicted the wheat microRNAs that could interact with TaMYC family members, and built up a network to show their integrative relationships., Conclusions: This study analyzed the size and composition of the MYC gene family in wheat, and investigated stress-responsive and light quality induced expression profiles of each TaMYC gene in the PTGMS wheat line BS366. In conclusion, we obtained lots of important information of TaMYC family, and the results of this study was supposed to contribute novel insights and gene and microRNA resources for wheat breeding, especially for the improvement of PTGMS wheat lines.

Seven new diterpenoids, euphorantones A-D (1, 3, 6, and 10), 8,12,13-epi-3,7,12-O-triacetyl-8-O-(2-methylbutanoyl)-ingol (9), 8,12,13-epi-3,12-O-diacetyl-7-O-benzoyl-8-methoxyingol (11), 2,3-epi-7,12-diacetate-8-benzoate-ingol (12), together with eighteen known compounds (2, 4-5, 7-8, and 13-25), were isolated from the aerial parts of Euphorbia antiquorum L.. The structures of new compounds 1, 3, 6, and 9-12 were elucidated by extensive spectroscopic analyses. The absolute configurations of new compounds were assigned using X-ray diffraction, Rh 2 (OCOCF 3 ) 4 -induced CD spectrum, and confirmed through comparison of the calculated and experimental 13 C NMR and electronic circular dichroism (ECD) data. Compounds 1-25 were evaluated for their inhibition of RANKL-induced osteoclastogenesis. Compound 1 showed the most potent inhibition of RANKL-induced osteoclastogenesis with IC 50 value of 0.3 μM. It inhibited NFAT transcript activity and osteoclast related genes TRAcP, CTSK, and NFATc1 expression., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Background: Hepatic radiation injury severely restricts irradiation treatment for liver carcinoma. The purpose of this study was to investigate the clinical application of gadolinium ethoxybenzyl diethylenetriamine pentaacetic acid (Gd-EOB-DTPA)-enhanced MRI (EOB-MRI) in the assessment of liver function after external radiation therapy and to determine the relationship between focal liver reaction (FLR) and liver function., Methods: A total of 47 patients with liver malignancies who underwent external beam radiation therapy were enrolled. EOB-MRI was performed on each patient at approximately one month post-radiotherapy. The hepatobiliary (HPB) phase images from EOB-MRI were fused with the planning CT images, and the isodose lines from the patients' treatment plans were overlaid onto the fused images. The correlation of the EOB-MR image intensity distribution with the isodose lines was studied. We also compared liver function in patients between pre-treatment and post-treatment., Results: Decreased uptake of Gd-EOB-DTPA, which was manifested by well-demarcated focal hypointensity of the liver parenchyma or FLR to high-dose radiation, was observed in the irradiated areas of 38 patients. The radiotherapy isodose line of decreased uptake area of Gd-EOB-DTPA was 30-46 Gy. The median corresponding dose curve of FLR was 34.4 Gy. Nine patients showed the absence of decreased uptake area of Gd-EOB-DTPA in the irradiated areas. Compared to the 38 patients with the presence of decreased uptake area of Gd-EOB-DTPA, 9 patients with the absence of decreased uptake area of Gd-EOB-DTPA showed significant higher levels of total bile acid, total bilirubin, direct bilirubin and alpha-fetoprotein (P < 0.05). There were no significant differences in alanine transaminase, aspartate aminotransferase, gamma-glutamyl transpeptidase or albumin levels between the two groups (P > 0.05)., Conclusions: Visible uptake of Gd-EOB-DTPA by the liver parenchyma was significantly associated with liver function parameters. EOB-MRI can be a valuable imaging biomarker for the assessment of liver parenchyma function outside of radiation area., (Copyright © 2019 First Affiliated Hospital, Zhejiang University School of Medicine in China. Published by Elsevier B.V. All rights reserved.)

To the best of our knowledge, this report presents, for the first time, the schematic of the possible chemical reaction for a one-pot synthesis of Zn 0.5 Cd 0.5 Se alloy quantum dots (QDs) in the presence of low/high oleylamine (OLA) contents. For high OLA contents, high-resolution transmission electron microscopy (HRTEM) results showed that the average size of Zn 0.5 Cd 0.5 Se increases significantly from 4 to 9 nm with an increasing OLA content from 4 to 10 mL. First, [Zn(OAc) 2 ]-OLA complex can be formed by a reaction between Zn(OAc) 2 and OLA. Then, Fourier transform infrared (FTIR) spectroscopy and X-ray diffraction (XRD) data confirmed that ZnO is formed by thermal decomposition of the [Zn(OAc) 2 ]-OLA complex. The results indicated that ZnO grew on the Zn 0.5 Cd 0.5 Se surface, thus increasing the particle size. For low OLA contents, HRTEM images were used to estimate the average sizes of the Zn 0.5 Cd 0.5 Se alloy QDs, which were approximately 8, 6, and 4 nm with OLA loadings of 0, 2, and 4 mL, respectively. We found that Zn(OAc) 2 and OLA could form a [Zn(OAc) 2 ]-OLA complex, which inhibited the growth of the Zn 0.5 Cd 0.5 Se alloy QDs, due to the decreasing reaction between Zn(oleic acid) 2 and Se 2- , which led to a decrease in particle size.

Aberrant aggregation of the protein tau is pathogenically involved in a number of neurodegenerative diseases, including Alzheimer's disease (AD). Although mouse models of tauopathy have provided a valuable resource for investigating the neurotoxic mechanisms of aggregated tau, it is becoming increasingly apparent that, due to interspecies differences in neurophysiology, the mouse brain is unsuitable for modeling the human condition. Advances in cell culture methods have made human neuronal cultures accessible for experimental use in vitro and have aided in the development of neurotherapeutics. However, despite the adaptation of human neuronal cell cultures, in vitro models of human tauopathy are not yet widely available. This protocol describes a cellular model of tau aggregation in which human neurons are transduced with lentiviral-derived vectors that code for pathogenically mutated tau fused to a yellow fluorescent protein (YFP) reporter. Transduced cultures produce tau aggregates that stain positively for thioflavin and display markers of neurotoxicity, such as decreased axonal length and increased lysosomal volume. This procedure may be a useful and cost-effective model for studying human tauopathies.

Objective: To explore the clinical significance of serum squamous cell carcinoma antigen (SCC-Ag) in early cervical squamous cell carcinoma. Methods: The clinicopathological data and follow-up information of 1435 patients with stage ⅠA2-ⅡA cervical squamous cell carcinoma were collected. The correlation between serum SCC-Ag level and clinicopathological feature and prognosis were analyzed. The best cut-off of serum SCC-Ag for predicting pelvic lymph node metastasis and survival of cervical squamous cell carcinoma patients were also identified. Results: The result of univariate analysis showed that The International Federation of Gynecology and Obstetrics (FIGO) staging, tumor size, depth of cervical stromal invasion, lymphovascular space involvement, pelvic lymph node metastasis, common iliac lymph node metastasis and para-aortic lymph node metastasis were significantly related with serum SCC-Ag level (all P <0.05). The result of multivariate logistic regression analysis showed that tumor size, depth of cervical stromal invasion, pelvic lymph node metastasis and common iliac lymph node metastasis were the independent risk factors of preoperative serum SCC-Ag>2.65 ng/ml (all P <0.001). Multivariate Cox regression analysis showed that lymphovascular space involvement, SCC-Ag>3.15 ng/ml, common iliac lymph node metastasis and tumor size >4 cm were the independent prognostic risk factors (all P <0.05). The univariate analysis showed that, the tumor size, FIGO stage, depth of cervical stromal invasion and SCC-Ag level were significantly related with the recurrence of 1 096 patients without postoperative high risk factors (all P <0.05). Multivariate logistic regression analysis showed that FIGO stage ( OR =1.671) and SCC-Ag>2.65 ng/ml ( OR =4.490) were the independent risk factors for recurrence (both P <0.05). The best cut off of SCC-Ag for predicting early postoperative cervical lymph node metastasis of cervical squamous cell carcinoma was 2.65 ng/ml, the sensitivity was 60.8%, the specificity was 71.8%. The best cut off of SCC-Ag for predicting prognosis of cervical squamous cell carcinoma was 3.15 ng/ml, the sensitivity was 53.5%, the specificity was 71.1%. Conclusions: Preoperative serum squamous cell carcinoma antigen is an independent prognostic risk factor of survival of patients with early cervical squamous cell carcinoma, and is significantly related with recurrence of patients without postoperative high-risk factors. It can be used as a reference factor for postoperative adjuvant radiotherapy.

Aluminum-gallium oxide (AGO) thin films with wide bandgaps of greater than 5.0 eV were grown using pulsed laser deposition. As evidenced by X-ray photoelectron spectroscopy, X-ray diffraction, and transmission electron microscopy, the oxygen chamber pressure considerably affected the lattice deformation in the AGO materials. Under high oxygen pressure, the lattice deformation reduced the d-spacing of the AGO(-201) plane. In the measured transmittance spectra of the AGO films, this narrowing of the d-spacing in the main plane manifested as a high-energy shift of the absorption edge. The AGO films were then installed as the active layers in the metal-semiconductor-metal photodetectors (PDs). The lattice deformation was observed to enhance the photocurrent and reduce the dark current of the device. The responsivity was 20.7 times higher in the lattice-deformed AGO-based PD sample than that in the nondeformed sample. It appeared that the lattice deformation induced the separation of the piezopotential, improving the efficiency of the photogenerated carrier recombination and, consequently, shortening the decay time of the photodetector.

Introduction: Quantitative in vivo measurement of brain amyloid burden is important for both research and clinical purposes. However, the existence of multiple imaging tracers presents challenges to the interpretation of such measurements. This study presents a direct comparison of Pittsburgh compound B-based and florbetapir-based amyloid imaging in the same participants from two independent cohorts using a crossover design., Methods: Pittsburgh compound B and florbetapir amyloid PET imaging data from three different cohorts were analyzed using previously established pipelines to obtain global amyloid burden measurements. These measurements were converted to the Centiloid scale to allow fair comparison between the two tracers. The mean and inter-individual variability of the two tracers were compared using multivariate linear models both cross-sectionally and longitudinally., Results: Global amyloid burden measured using the two tracers were strongly correlated in both cohorts. However, higher variability was observed when florbetapir was used as the imaging tracer. The variability may be partially caused by white matter signal as partial volume correction reduces the variability and improves the correlations between the two tracers. Amyloid burden measured using both tracers was found to be in association with clinical and psychometric measurements. Longitudinal comparison of the two tracers was also performed in similar but separate cohorts whose baseline amyloid load was considered elevated (i.e., amyloid positive). No significant difference was detected in the average annualized rate of change measurements made with these two tracers., Discussion: Although the amyloid burden measurements were quite similar using these two tracers as expected, difference was observable even after conversion into the Centiloid scale. Further investigation is warranted to identify optimal strategies to harmonize amyloid imaging data acquired using different tracers.

The actinomycete genus Streptomyces is characterized by producing bioactive secondary metabolites, including antibiotics. In this study, chemical and biological investigations were carried out on Streptomyces strain 4205 isolated from the paddy soil, leading to the identification and characterization of 10 albocycline-type macrolides, among which 4 compounds were new, namely albocyclines A-D (1-4). The structures of 1-10 were identified according to the 1D- and 2D-NMR spectroscopic data. Furthermore, compounds 1-10 were evaluated for antimicrobial activity. Compounds 5-7 displayed antimicrobial activities against Candidaalbicans ATCC 90028 with the same MIC value of 10.0 mg/mL and the IC 50 values of 1.5, 1.0, and 1.0 mg/mL, respectively. Thus, the research on Streptomyces sp. is of vital significance for developing new antibiotic agents., (© 2019 Wiley-VHCA AG, Zurich, Switzerland.)

Progressive accumulation of aggregation-prone proteins, amyloid-β (Aβ) and hyperphosphorylated tau (p-tau), are the defining hallmarks of Alzheimer's disease (AD). The mechanisms by which Aβ and p-tau are transmitted throughout the diseased brain are not yet completely understood. Interest in exosome research has grown dramatically over the past few years, specifically due to their potential role as biomarkers for staging of neurodegenerative diseases, including AD. Despite their diagnostic utility, the pathogenic potential of exosomes has yet to be fully elucidated. In this study, we use a series of recombinant tau antibodies to characterize a new model of human tau in vivo. Exosome suspensions derived from neuronally-differentiated, human induced pluripotent stem cells that express the repeat domain of tau P301L and V337M mutations (NiPSCEs) were injected into the wild-type mouse brain and pathological changes were characterized by immunostaining at one- (1 m) and two-month (2 m) post-injection. We found that tau inclusions were present throughout the brain at 2 m post-injection, which were detectable using antibodies raised against full-length tau (K9JA) and misfolded tau (MC1). Furthermore, we found that phosphorylated tau immunoreactivity was elevated 1 m post-injection, which was surprisingly normalized after 2 m. Finally, we observed extensive degeneration of neuronal dendrites in both ipsilateral and contralateral hippocampi in NiPSCE treated mice. In summary, we demonstrate that exosomes are sufficient to cause long-distance propagation of tau pathology and neurodegeneration in vivo. These novel findings support an active role of exosomes in AD pathogenesis.

Extracellular vesicles (EVs) are a heterogeneous group of secreted particles consisting of microvesicles, which are released by budding of the cellular membrane, and exosomes, which are secreted through exocytosis from multivesicular bodies. EV cargo consists of a wide range of proteins and nucleic acids that can be transferred between cells. Importantly, EVs may be pathogenically involved in neurodegenerative diseases such as Alzheimer's disease (AD). While EVs derived from AD neurons have been found to be neurotoxic in vitro, little is known about the pathological consequences of AD EVs in vivo. Furthermore, although all known familial AD (fAD) mutations involve either amyloid-β protein precursor (AβPP) or the machinery that processes AβPP, hyperphosphorylation of the microtubule associated protein tau appears to play a critical role in fAD-associated neurodegeneration, and previous reports suggest EVs may propagate tau pathology in the AD brain. Therefore, we hypothesized that fAD EVs may have a mechanistic involvement in the development of fAD-associated tau pathology. To test this, we isolated EVs from iPSC-derived neuronal cultures generated from an fAD patient harboring a A246E mutation to presenilin-1 and stereotactically injected these EVs into the hippocampi of wild-type C57BL/6 mice. Five weeks after injection, mice were euthanized and pathology evaluated. Mice injected with fAD EVs displayed increased tau phosphorylation at multiple sites relative to PBS and non-disease control EV injected groups. Moreover, fAD EV injected hippocampi contained significantly more tau inclusions in the CA1 hippocampal neuronal field than controls. In total, these findings identify EVs as a potential mediator of fAD-associated tau dysregulation and warrant future studies to investigate the therapeutic potential of EV-targeted treatments for fAD.

There is increasing evidence that deep sequencing-based T cell repertoire can sever as a biomarker of immune response in cancer patients; however, the characteristics of T cell repertoire including diversity and similarity, as well as its prognostic significance in patients with cervical cancer (CC) remain unknown. In this study, we applied a high throughput T cell receptor (TCR) sequencing method to characterize the T cell repertoires of peripheral blood samples from 25 CC patients, 30 cervical intraepithelial neoplasia (CIN) patients and 20 healthy women for understanding the immune alterations during the cervix carcinogenesis. In addition, we also explored the signatures of TCR repertoires in the cervical tumor tissues and paired sentinel lymph nodes from 16 CC patients and their potential value in predicting the prognosis of patients. Our results revealed that the diversity of circulating TCR repertoire gradually decreased during the cervix carcinogenesis and progression, but the circulating TCR repertoires in CC patients were more similar to CIN patients than healthy women. Interestingly, several clonotypes uniquely detected in CC patients tended to share similar CDR3 motifs, which differed from those observed in CIN patients. In addition, the TCR repertoire diversity in sentinel lymphatic nodes from CC patients was higher than in tumor tissues. More importantly, less clonotypes in TCR repertoire of sentinel lymphatic node was associated with the poor prognosis of the patients. Overall, our findings suggested that TCR repertoire might be a potential indicator of immune monitoring and a biomarker for predicting the prognosis of CC patients. Although functional studies of T cell populations are clearly required, this study have expanded our understanding of T cell immunity during the development of CC and provided an experimental basis for further studies on its pathogenesis and immunotherapy.

Tauopathies are neurodegenerative diseases characterized by tau protein pathology in the nervous system. EIF2AK3 (eukaryotic translation initiation factor 2 alpha kinase 3), also known as PERK (protein kinase R-like endoplasmic reticulum kinase), was identified by genome-wide association study as a genetic risk factor in several tauopathies. PERK is a key regulator of the Unfolded Protein Response (UPR), an intracellular signal transduction mechanism that protects cells from endoplasmic reticulum (ER) stress. PERK variants had previously been identified in Wolcott-Rallison Syndrome, a rare autosomal recessive metabolic disorder, and these variants completely abrogated the function of PERK's kinase domain or prevented PERK expression. In contrast, the PERK tauopathy risk variants were distinct from the Wolcott-Rallison variants and introduced missense alterations throughout the PERK protein. The function of PERK tauopathy variants and their effects on neurodegeneration are unknown. Here, we discovered that tauopathy-associated PERK alleles showed reduced signaling activity and increased PERK protein turnover compared to protective PERK alleles. We found that iPSC-derived neurons carrying PERK risk alleles were highly vulnerable to ER stress-induced injury with increased tau pathology. We found that chemical inhibition of PERK in human iPSC-derived neurons also increased neuronal cell death in response to ER stress. Our results indicate that tauopathy-associated PERK alleles are functional hypomorphs during the UPR. We propose that reduced PERK function leads to neurodegeneration by increasing neuronal vulnerability to ER stress-associated damage. In this view, therapies to enhance PERK signaling would benefit at-risk carriers of hypomorphic alleles.

Background: COI (CORONATINE INSENSITIVE), an F-box component of the Skp1-Cullin-F-box protein (SCF COI1 ) ubiquitin E3 ligase, plays important roles in the regulation of plant growth and development. Recent studies have shown that COIs are involved in pollen fertility. In this study, we identified and characterized COI genes in the wheat genome and analyzed expression patterns under abiotic stress., Results: A total of 18 COI candidate sequences for 8 members of COI gene family were isolated in wheat (Triticum aestivum L.). Phylogenetic and structural analyses showed that these COI genes could be divided into seven distinct subfamilies. The COI genes showed high expression in stamens and glumes. The qRT-PCR results revealed that wheat COIs were involved in several abiotic stress responses and anther/glume dehiscence in the photoperiod-temperature sensitive genic male sterile (PTGMS) wheat line BS366., Conclusions: The structural characteristics and expression patterns of the COI gene family in wheat as well as the stress-responsive and differential tissue-specific expression profiles of each TaCOI gene were examined in PTGMS wheat line BS366. In addition, we examined SA- and MeJA-induced gene expression in the wheat anther and glume to investigate the role of COI in the JA signaling pathway, involved in the regulation of abnormal anther dehiscence in the PTGMS wheat line. The results of this study contribute novel and detailed information about the TaCOI gene family in wheat and could be used as a benchmark for future studies of the molecular mechanisms of PTGMS in other crops.

Objective: To explore the influence of patatin-like phospholipase domain containing-3 (PNPLA3) wild type 148I/I and mutant type 148M/M on HepG2 cell proliferation and the relative mechanisms. Methods: HepG2 cell line stably overexpressing PNPLA3 148I/I, 148M/M and negative control (NC) were set up. Cell counting kit-8 (CCK8) assay was used to measure cell viability. Edu assay was used to determine the ability of cell proliferation. Western blot was used to detect the protein levels in the phosphatidylinositol 3-kinases (PI3K) pathway. Enzyme-linked immunosorbent assay (ELISA) was used to detect proliferation-related PNPLA3 metabolites[arachidonic acid (AA) and lysophosphatidic acid (LPA)]. Quantitative real-time PCR was used to detect the expression level of prostaglandin G/H synthase 2 (PTGS2) and proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) associated with PNPLA3. Results: The cell viability of overexpression of PNPLA3 148M/M group was about 1/3 times higher than that of overexpression of PNPLA3 148I/I group, and the difference was statistically significant[(98.02±1.29)% vs (71.51±2.89)%, P <0.001]. There was no significant difference between overexpression of PNPLA3 148M/M group and negative control group[(98.02±1.29)% vs (100±2.61)%, P =0.181]. The proliferative activity of overexpression of PNPLA3 148M/M group was about 1/3 times higher than that of overexpression of PNPLA3 148I/I group, and the difference was statistically significant(46.46±1.83 vs 35.96±2.65, P =0.001). There was no significant difference between overexpression of PNPLA3 148M/M group and negative control group(46.46±1.83 vs 46.64±7.33, P =0.965). The PGC1α mRNA expression, total PI3K, P Thr-308 AKT, P Ser2448 -mammalian target of rapamycin (P Ser2448 -mTOR) and PGC1α protein expression levels in the overexpression of PNPLA3 148M/M group were higher than those in the overexpression of PNPLA3 148I/I group, but there were no significant differences in AA and LPA levels, as well as PTGS2 mRNA expression levels. Conclusion: PNPLA3 148M/M cell proliferation was stronger than PNPLA3 148I/I.

Tauopathies are a class of neurodegenerative diseases, including Alzheimer's disease, frontotemporal dementia and progressive supranuclear palsy, which are associated with the pathological aggregation of tau protein into neurofibrillary tangles (NFT). Studies have characterized tau as a "prion-like" protein given its ability to form distinct, stable amyloid conformations capable of transcellular and multigenerational propagation in clonal fashion. It has been proposed that progression of tauopathy could be due to the prion-like propagation of tau, suggesting the possibility that end-stage pathologies, like NFT formation, may require an instigating event such as tau seeding. To investigate this, we applied a novel human induced pluripotent stem cell (hiPSC) system we have developed to serve as a human neuronal model. We introduced the tau repeat domain (tau-RD) with P301L and V337M (tau-RD-LM) mutations into hiPSC-derived neurons and observed expression of tau-RD at levels similar to total tau in postmortem AD brains. Tau aggregation occurred without the addition of recombinant tau fibrils. The conditioned media from tau-RD cultures contained tau-RD seeds, which were capable of inducing aggregate formation in homotypic mode in non-transduced recipient neuronal cultures. The resultant NFTs were thioflavin-positive, silver stain-positive, and assumed fibrillary appearance on transmission electron microscopy (TEM) with immunogold, which revealed paired helical filament 1 (PHF1)-positive NFTs, representing possible recruitment of endogenous tau in the aggregates. Functionally, expression of tau-RD caused neurotoxicity that manifested as axon retraction, synaptic density reduction, and enlargement of lysosomes. The results of our hiPSC study were reinforced by the observation that Tau-RD-LM is excreted in exosomes, which mediated the transfer of human tau to wild-type mouse neurons in vivo. Our hiPSC human neuronal system provides a model for further studies of tau aggregation and pathology as well as a means to study transcellular propagation and related neurodegenerative mechanisms., (Copyright © 2017 Elsevier Inc. All rights reserved.)

MicroRNAs (miRNAs) are endogenous small RNAs which play important negative regulatory roles at both the transcriptional and post-transcriptional levels in plants. Wheat is the most commonly cultivated plant species worldwide. In this study, RNA-seq analysis was used to examine the expression profiles of miRNA in the spikelets of photo-thermosenisitive genic male sterile (PTGMS) wheat line BS366 during male fertility transition. Through mapping on their corresponding precursors, 917-7,762 novel miRNAs were found in six libraries. Six novel miRNAs were selected for examination of their secondary structures and confirmation by stem-loop RT-PCR. In a differential expression analysis, 20, 22, and 58 known miRNAs exhibited significant differential expression between developmental stages 1 (secondary sporogenous cells had formed), 2 (all cells layers were present and mitosis had ceased), and 3 (meiotic division stage), respectively, of fertile and sterile plants. Some of these differential expressed miRNAs, such as tae-miR156, tae-miR164, tae-miR171, and tae-miR172, were shown to be associated with their targets. These targets were previously reported to be related to pollen development and/or male sterility, indicating that these miRNAs and their targets may be involved in the regulation of male fertility transition in the PTGMS wheat line BS366. Furthermore, target genes of miRNA cleavage sites were validated by degradome sequencing. In this study, a possible signal model for the miRNA-mediated signaling pathway during the process of male fertility transition in the PTGMS wheat line BS366 was developed. This study provides a new perspective for understanding the roles of miRNAs in male fertility in PTGMS lines of wheat.

Introduction: Childhood asthma is globally one of the most common respiratory disorders and accounts for more school absences and more hospitalizations than any other chronic illness. The worldwide economic burden of this disease exceeds those of HIV/AIDS and tuberculosis combined. Proper intervention and effective management is of paramount importance for the control and prognosis of paediatric asthma. Unfortunately, the rate of uncontrolled and partially controlled paediatric asthma in China is >90%. This study will use a new management model to investigate the status of asthma control and the adherence of patients to a medication protocol., Methods: This prospective, multicentre, observational study will be conducted at 15 hospitals on children (n=800) diagnosed with asthma. Each patient will be assigned to either the nearest community hospital or Shanghai Children's Medical Center, whichever is closer to the patient's home, according to the decision of parents. Participants were divided into two groups: tertiary care hospital (Shanghai Children's Medical Center) follow-up group and community hospital follow-up group. The primary outcome will be the difference in the proportion of controlled, partially controlled and uncontrolled asthma among the two groups. Secondary outcomes will be the differences in adherence rate, lung function, exacerbations, growth development, total asthma-related unscheduled visits, days absent from school and loss of working days for the patient's caregiver. Data will be analysed on an intention-to-treat and a per-protocol basis., Ethics and Dissemination: Ethics approval was obtained from the Institutional Review Board of Shanghai Children's Medical Center affiliated with Shanghai Jiao Tong University of Medicine. We plan to publish the results of this study in a peer-reviewed journal article., Competing Interests: Competing interests: None declared., (© Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.)

Objective: To study the difference between intensity-modulated radiation therapy (IMRT) and three dimensional conformal radiation therapy (3D-CRT) for pelvic radiation of post-operative treatment with gynecologic malignant tumor. Methods: A prospective investigation study was conducted on 183 patients of post-operative patients with whole pelvic radiation therapy of cervical cancer or endometrial cancer in Zhejiang Cancer Hospital [IMRT group ( n =85) and 3D-CRT group ( n =98)] from Oct. 2015 to Oct. 2016. The two groups received same dose (45 Gy in 25 fractions). Comparison of two groups with radiation dosimetry:the score according to the Radiation Therapy Oncology Group (RTOG) acute radiation injury grading standards before and after radiotherapy reaction, the score from functional assessment of cancer therapy scale-cervix (FACT-Cx) scale and expanded prostate cancer index composite for clinical practice (EPIC-CP) scale were also analyzed. Results: (1) There were no significant effect with age, culture level, family economic condition and ratio of radiochemotherapy between two groups (all P >0.05). (2) Dosimetric comparison for IMRT vs 3D-CRT: the average dose of planning target volume (PTV) decreased (46.1±0.4) vs (46.4±0.5) Gy, V (45) dose percentage increased (95.2±1.0) % vs (93.3±2.0) %, intestinal bag dose of V (4)0 decreased (24.4±6.8) % vs (36.5±15.9) %, rectal V (40) dose percentage decreased (73.9±12.3) % vs (85.4±8.4) %, and lower rectal V (45) dose percentage (32.8±13.4) % vs (71.5±13.7) %, bladder V (40) dose percentage decreased (55.5±13.0) % vs (84.4±13.0) %. Bone marrow V (20) lower: (67.9±5.4) % vs (79.5±6.6) %, V (1)0 lower: (82.1±6.0) % vs (86.3±6.6) %; there were significant differences (all P <0.05). There was no significant difference between the dose of V (45) in the intestinal pouch and bladder ( P >0.05). (3) Acute radiation injury classification for IMRT vs 3D-CRT: big or small intestine: Ⅱ-Ⅲ reaction [13% (11/85) vs 24% (24/98); χ(2)=3.925, P =0.048], there was significant difference. Bladder: Ⅲ reaction [19% (16/85) vs 26% (25/98); χ(2)=1.171, P =0.279], there was no significant difference. Radiochemotherapy of bone marrow suppression: Ⅲ-Ⅳ reaction (14/20), the incidence rate [26% (14/54) vs 31% (20/65); χ(2)=0.339, P =0.562], the difference was not statistically significant. (4) Quality of life scale by FACT-Cx scale in IMRT vs 3D-CRT: there were no significant difference before radiotherapy (82±16 vs 85±16; t =1.279, P =0.203), while there was significant difference after radiotherapy (76±14 vs 71±18; t =-2.160, P=0.032). EPIC-CP scale score: before radiotherapy they were (16±7 vs 15±6; t =-0.174, P =0.862),but after radiotherapy (18±7 vs 22±7; t =3.158, P =0.002), there was significant difference between them. Before and after radiotherapy, the increased EPIC-CP scale of the IMRT group vs 3D-CRT group were 3±4 and 6±4, the 3D-CRT group was significantly higher, the difference was statistically significant ( t =5.500, P =0.000). Conclusion: IMRT has shown that there are a significant benefit for the post-operative patients with cervical cancer and endometrial cancer compared to 3D-CRT.

Objective: To investigate the effect of exendin-4, a glucagon-like peptide-1 (GLP-1) receptor agonist, on reducing lipid deposition and improving insulin resistance in skeletal muscle and the underlying mechanisms in high-fat diet (HFD)-induced obese mice. Methods: Twelve male C57BL/6J mice were challenged with HFD for 12 weeks to induce obesity and then randomly divided into two groups: exendin-4 group (intraperitoneal injection of 24 nmol·kg -1 ·d -1 exendin-4 for 4 weeks) and HFD group (intraperitoneal injection of normal saline for 4 weeks), with 6 mice in each group. Additional 6 mice were also selected as control group. Body weight, fasting blood glucose were recorded. Serum triglyceride (TG), total cholesterol (TC), insulin and skeletal muscle triglyceride levels were measured by enzyme-linked immunosobent assay (ELISA). Oil red O staining was used for morphologic changes of frozen sections from skeletal muscle. The protein levels of lipid metabolic pathway mediated by AMP-activated protein kinase (AMPK) and insulin signailing pathway were determined by Western blot. Results: Compared with mice in HFD group, exendin-4 significantly decreased body weight[(37.68±1.80) vs (46.03±5.00) g, P <0.025], fasting blood glucose[(5.40±0.33) vs (7.65±1.92) mmol/L, P <0.025], serum TG[(37.78±7.14) vs (80.76±34.22) mg/dl, P <0.025], TC[(180.13±18.75) vs (217.57±22.52) mg/dl, P <0.025], insulin[(0.58±0.01) vs (1.67±1.23) ng/ml, P <0.025]and skeletal muscle TG levels[(9.84±1.08) vs (19.35±7.44) mg/g, P <0.025]of obese mice. Oil red O staining revealed that exendin-4 alleviated the accumulation of larger lipid droplets in skeletal muscle. The protein expressions of lipolysis and lipid oxidation mediated by AMPK and insulin signailing pathway were up-regulated, and the protein expressions of lipogenesis mediated by AMPK were down-regulated after intervention of exendin-4. Conclusion: Exendin-4 reduces lipid deposition and insulin resistance in skeletal muscle of HFD-induced obese mice via activating AMPK and up-regulating insulin signailing pathway.

Obesity-associated insulin resistance is a major risk factor for most metabolic diseases, including dyslipidemia and type 2 diabetes. Acanthopanax senticosus (Rupr. et Maxim.) Harms (Goka) root has been used in traditional Chinese medicine for treatment of diabetes and other conditions; however, little is known about the effects of Goka fruit (GF). Goka fruit is rich in anthocyanin, which has beneficial effects on obesity and insulin resistance via activation of adenosine monophosphate-activated protein kinase (AMPK). We hypothesized that GF can improve obesity-associated insulin resistance. The aim of the present study was to investigate whether GF improves insulin resistance in high-fat diet (HFD)-induced obese mice. High-fat diet mice treated with GF (500 and 1000 mg/kg) for 12 weeks showed an improved glucose tolerance and insulin sensitivity, as well as reduced plasma insulin and liver lipid accumulation. Moreover, GF administration to HFD mice resulted in down-regulation of fatty acid synthase expression and up-regulation of cholesterol 7-alpha-hydroxylase expression in the liver. Notably, AMPK phosphorylation in the liver increased after GF administration. In summary, GF supplementation improved obesity-associated insulin resistance and hepatic lipid accumulation through modulation of AMPK activity and lipid metabolism-associated gene expression., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Objective: To investigate the effect of silent mating type information regulation 2 homolog 1 (SIRT1) deficiency on function of brown adipose tissue (BAT) in high-fat diet (HFD)-induced obese mice., Methods: Male SIRT1 deficient heterozygous (SIRT1(+ /-)) mice and their wild-type (WT) littermates were challenged with a HFD diet for 16 weeks to induce obesity model.Energy metabolic cages were used to measure oxygen consumption and heat production, and cold tolerance test was to evaluate the adaptive thermogenic function.With brown fat collected after the diet intervention, determination techniques were adopted included HE staining for morphologic changes, immunohistochemical staining and Western blotting for uncoupling protein 1 (UCP1) expression, quantitative real-time PCR for relative content of mitochondria DNA (mtDNA)., Results: Compared to WT controls, SIRT1(+ /-) mice displayed significant decreases in both oxygen consumption and heat production[(2 681±297) vs (3 017±313) ml·kg(-1)·h(-1,) (19.05±2.40) vs (21.15±2.49) kcal·kg(-1)·h(-1,) both P<0.05)], as well as an impairment in maintaining their body temperature during the cold challenge.HE staining revealed the accumulation of larger lipid droplets in BAT of SIRT1(+ /-) mice, and both immunohistochemical staining and Western blotting indicated an obvious reduction in expression of UCP1 (P<0.05). Quantitative real-time PCR showed a significant decrease in the relative mtDNA content in BAT of SIRT1(+ /-) mice (0.38±0.10 vs 1.00±0.40, P<0.05)., Conclusion: SIRT1 deficiency promotes BAT dysfunction, meaning that whitening in obese mice.

Objective: To explore the influence of patatin-like phospholipase domain-containing protein 3 (PNPLA3) on palmitic acid (PA)-induced hepatocyte apoptosis and its mechanism., Methods: Human hepatocarcinoma cell line HepG2 cells were transfected with PNPLA3(WT)-pcDNA3.1 (PNPLA3(WT) group) and PNPLA3(I148M)-pcDNA3.1 (PNPLA3(I148M) group) plasmids respectively to overexpress wild type or mutant type PNPLA3, and cells transfected with empty vector pcDNA3.1 (NC group) were set as control group.After 24 h PA incubation, Oil red staining was used to determine lipid deposition, and terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling assay (TUNEL) was used to measure apoptosis.Western blot was used to detect the protein level of endoplasmic reticulum (ER) stress and associated apoptosis.Enzyme-linked immunosorbent assay (ELISA) was used to test lysolecithin (LPC) levels in the cellular supernatant., Results: After 24 h PA incubation, there was no significant difference in lipid deposition among three groups.Compared to NC group, the cell apoptosis rates of PNPLA3(WT) and PNPLA3(I148M) groups were increased by 2 times and 3 times respectively.The levels of ER stress PRKR-like endoplasmic reticulum kinase (PERK) pathway associated proteins, immunoglobulin-binding protein (BIP), p-PERK, p-eIF2α, and ER stress associated apoptosis pathway proteins, CCAAT/enhancer binding homologous protein (CHOP), p53 upregulated modulator of apoptosis (PUMA), Bax, caspase-3 were higher, and were more significant in PNPLA3(I148M) group.The LPC level in the supernatant of PNPLA3(WT) and PNPLA3(I148M) groups were about 5 times and 1.5 times of NC group respectively after PA incubation., Conclusion: PNPLA3 may be involved in palmitic acid-induced apoptosis mediated by hepatocyte ER stress through regulating LPC metabolism.

Introduction: Breast cancer is a commonly diagnosed cancer, in which most patients' metastases (about 75%) occurred in 5 years after the initial diagnosis, especially in 3 years. Recrudescence exceeding 20 years is rarely reported in the past several decades.Case information: A 68-year-old female patient presented with breast cancer in which 3 focal increased 18F-fluorodeoxyglucose uptake in the right supraclavicular lymph node, the mediastinum and sternum were found on positron emission tomography and computed tomography. Then we learned that the patient had suffered from breast cancer and been given a right-sided mastectomy 24 years ago. Histopathology from the mediastinum revealed metastatic, moderately differentiated breast adenomatous cell carcinoma., Conclusion: We report this late recurrence of breast cancer 24 years following mastectomy, suggesting that possible recurrence of this disease with a 24-year latency period should be taken into consideration.

Human papillomavirus (HPV) are firmly established as the principal causative agent for cervical carcinoma. Current vaccines may provide some protection for women from cervical carcinoma linked to HPV genotype 16 and 18. This may be the best vaccine for Western women, but the geographical variation in HPV distributions may not make it the most appropriate vaccine for China or Asia. This study provided an observational, retrospective, hospital-based cross-sectional study on the distribution of HPV genotypes among 5410 women with invasive cervical cancer (ICC) or cervical intraepithelial neoplasia (CIN). Overall, the positive rates of the four HPV types included in current prophylactic vaccines were counted, the two high-risk types (HPV-16 and -18) covered by current vaccines represented 66.9% of women with squamous cancer, 55.0% with adenocarcinoma, 64.9% with adenosquamous carcinoma and 77.4% of other type ICC, as well as 59.5% of CIN III, 45.0% of CIN II and 38.1% of CIN I cases. As expected, two low-risk types (HPV-6 and -11) included in the quadrivalent vaccine did not show good coverage data. Particularly worth mentioning is the fact that the addition of HPV-52 and -58 to the vaccine cocktail would increase cancer protection in our population, potentially preventing up to beyond 16% of squamous/adenosquamous carcinoma and other type of cervical cancers, and 7.75% of adenocarcinomas. It might also potentially reduce the rate of CIN III by a further 28.6% and CIN II and I by a third. This study established the baseline for surveillance in Zhejiang Province, and provides data for further vaccine designs: a quadrivalent HPV vaccine covering HPV-16/-58/-18/-52, would be more welcome in our region in the forthcoming year compared to the currently available vaccine.

The aim of the study was to analyze the expressions of vascular endothelial growth factor (VEGF), transforming growth factor β1 (TGF-β1) and endostatin in non-small cell lung caner (NSCLC), and to explore their correlations with NSCLC. 80 NSCLC patients during January 2013 to June 2014 were selected. The expression levels of VEGF, TGF-β1, and endostatin before surgeries were detected, and compared with 40 healthy individuals and 40 patients with benign pulmonary diseases. Serum VEGF, TGF-β1, and endostatin levels were (573.6 ± 25.4) pg/mL, (36.2 ± 10.5) ng/mL, and (20.3 ± 7.8) ng/mL, respectively, in NSCLC group, which were obviously higher than those in healthy individuals and patients with benign pulmonary diseases, and the difference was statistically significant (p < 0.05); there was no statistical difference of the VEGF, TGF-β1, and endostatin levels between the healthy individuals and patients with benign pulmonary diseases (p > 0.05), or among patients with different physiological characteristics such as gender, age, and smoking history in NSCLC group (p > 0.05). Serum VEGF, TGF-β1, and endostatin levels also showed no statistical difference among patients with different pathological characteristics such as histological types, with or without lymphatic metastasis (p > 0.05). However, the three indicators were significantly different among patients with different TNM stages, with or without distant metastasis and different cell differentiation degrees (p < 0.05). Serum VEGF and TGF-β1 were positively related (r = 0.479, p < 0.05), endostatin was negatively related to both VEGF and TGF-β1 (r = -0.392, -0.354, p < 0.05 in both comparisons). The expression levels of VEGF, TGF-β1, and endostatin significantly contributed to the poor cell differentiation in NSCLC. They had important effects on the occurrence, development, and metastasis of NSCLC, which could be applied as the indicators to predict the malignancy of NSCLC.

Predisposition to sporadic Alzheimer's disease (SAD) involves interactions between a person's unique combination of genetic variants and the environment. The molecular effect of these variants may be subtle and difficult to analyze with standard in vitro or in vivo models. Here we used hIPSCs to examine genetic variation in the SORL1 gene and possible contributions to SAD-related phenotypes in human neurons. We found that human neurons carrying SORL1 variants associated with an increased SAD risk show a reduced response to treatment with BDNF, at the level of both SORL1 expression and APP processing. shRNA knockdown of SORL1 demonstrates that the differences in BDNF-induced APP processing between genotypes are dependent on SORL1 expression. We propose that the variation in SORL1 expression induction by BDNF is modulated by common genetic variants and can explain how genetic variation in this one locus can contribute to an individual's risk of developing SAD., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Small cell lung cancer (SCLC) represents a group of highly malignant tumors that give rise to early and widespread metastases at the time of diagnosis. The preferential metastatic sites are the brain, liver, adrenal glands, bone, and bone marrow. However, metastases of the gastrointestinal system, especially the stomach, are rare; most cases of stomach metastasis are asymptomatic and, as a result, are usually only discovered at autopsy. We report a case of gastric metastasis originating from SCLC. The patient was a 66-year-old man admitted to our hospital due to abdominal pain. He underwent gastroscopy, with the pathological report of the tissue biopsy proving it to be a small cell cancer. Immunohistochemistry was positive for CD56, synaptophysin, and pan-cytokeratin. These results confirmed the diagnosis of gastric metastasis of a neuroendocrine small cell carcinoma from the lung.

Importance: Although considerable effort has been expended developing drug candidates for Alzheimer disease, none have yet succeeded owing to the lack of efficacy or to safety concerns. One potential shortcoming of current approaches to Alzheimer disease drug discovery and development is that they rely primarily on transformed cell lines and animal models that substantially overexpress wild-type or mutant proteins. It is possible that drug development failures thus far are caused in part by the limits of these approaches, which do not accurately reveal how drug candidates will behave in naive human neuronal cells., Objective: To analyze purified neurons derived from human induced pluripotent stem cells from patients carrying 3 different presenilin 1 (PS1) mutations and nondemented control individuals in the absence of any overexpression. We tested the efficacy of γ-secretase inhibitor and γ-secretase modulator (GSM) in neurons derived from both normal control and 3 PS1 mutations (A246E, H163R, and M146L)., Design, Setting, and Participants: Adult human skin biopsies were obtained from volunteers at the Alzheimer Disease Research Center, University of California, San Diego. Cell cultures were treated with γ-secretase inhibitor or GSM. Comparisons of total β-amyloid (Aβ) and Aβ peptides 38, 40, and 42 in the media were made between vehicle- vs drug-treated cultures., Main Outcomes and Measures: Soluble Aβ levels in the media were measured by enzyme-linked immunosorbent assay., Results: As predicted, mutant PS1 neurons exhibited an elevated Aβ42:Aβ40 ratio (P < .05) at the basal state as compared with the nondemented control neurons. Treatment with a potent non-nonsteroidal anti-inflammatory druglike GSM revealed a new biomarker signature that differs from all previous cell types and animals tested. This new signature was the same in both the mutant and control neurons and consisted of a reduction in Aβ42, Aβ40, and Aβ38 and in the Aβ42:Aβ40 ratio, with no change in the total Aβ levels., Conclusions and Relevance: This biomarker discrepancy is likely due to overexpression of amyloid precursor protein in the transformed cellular models. Our results suggest that biomarker signatures obtained with such models are misleading and that human neurons derived from human induced pluripotent stem cells provide a unique signature that will more accurately reflect drug response in human patients and in cerebrospinal fluid biomarker changes observed during GSM treatment.

It has been proposed that genetic factors contribute to the susceptibility of non-small cell lung cancer (NSCLC). The programmed cell death 6 interacting protein (PDCD6IP) encodes for a protein that has been known to bind to the products of the PDCD6 gene, a required protein in apoptosis. The aim of this study is to investigate the relationship between PDCD6IP insertion/deletion (I/D) polymorphism (rs28381975) and NSCLC risk in a Chinese population. A population-based case-control study was conducted in 449 NSCLC patients and 512 cancer-free controls. The genotype of the PDCD6IP gene was determined by using a polymerase chain reaction assay. The promoter activity was analyzed by luciferase reporter assay in A549 and H1299 cells. Statistically significant difference was observed when the patients and controls were compared according to ID + II versus DD (OR = 1.72, 95 % CI 1.29-2.31, P < 0.01). The I allele was significantly associated with NSCLC risk (OR = 1.41, 95 % CI 1.18-1.69, P < 0.01). Compared to TNM stage I + II, PDCD6IP I/D polymorphism significantly increased advanced NSCLC risk (OR = 2.06, 95 % CI 1.30-3.26, P < 0.01). Promoter reporter structures carrying the I allele displayed significantly higher promoter activity than the D allele in A549 and H1299 cells (P = 0.001). The results from this study suggested that PDCD6IP I/D polymorphism was potentially related to NSCLC susceptibility in Chinese Han population.

Objective: To determine the clinicopathologic factors associated with survival in small cell neuroendocrine cervical cancer (SCNEC) patients., Materials and Methods: The study was approved by the ethics committee of the hospital. The records of 64 SCNEC patients from 9,474 Chinese patients with cervical cancer at the Zhejiang Cancer Hospital were reviewed. Kaplan-Meier and Cox regression methods were used for analyses., Results: Of 64 patients, 47 had Stages I-IIA, 12 had Stages IIB-IVA, and five had Stage IV-B disease. A total of 81.25% underwent surgery, 89.1% received chemotherapy, 62.5% received radiation, 34.4% received neoadjuvant chemotherapy (NACT), and 34.4% received concurrent chemoradiation (CCRT). The median follow-up for surviving patients was 35.7 months (range: 0.5-160), and 29 (50%) of the 58 patients with Stages I-III had either disease recurrence or progression. The median time to first relapse was 10.5 months (range: 0-88.2). The five-year overall survival of patients in Stages I-IIA and IIB-IVB disease was 54.4% and 9.8%, respectively (p = 0.001). Women with early-stage (Stages IIBIIA) disease had median survival rates of 94 months compared with 21.4 months in the advanced-stage (Stages IIB-IVB) group. In univariate analysis, advanced-stage (p = 0.001), without radical surgery (p = 0.002) and deep stromal invasion (DSI) (p = 0.000) were considered poor prognostic factors. In a multivariable analysis, tumor size > four cm (p = 0.048), postoperative radiation (p = 0.038) for early-stage patients and the FIGO stage (p = 0.040) of disease in the overall population remained as independent prognostic factor of survival., Conclusion: The FIGO stage was found to be an independent prognostic factor of SCNEC. In addition, tumor size > four cm and DSI was associated with poor survival. Postoperative radiation for early-stage patients may not improve survival. The role of primary and postoperative NACT or CCRT is unclear. Clinical trials are needed.

Presenilin 1 (PS1) is the catalytic core of γ-secretase, which cleaves type 1 transmembrane proteins, including the amyloid precursor protein (APP). PS1 also has γ-secretase-independent functions, and dominant PS1 missense mutations are the most common cause of familial Alzheimer's disease (FAD). Whether PS1 FAD mutations are gain- or loss-of-function remains controversial, primarily because most studies have relied on overexpression in mouse and/or nonneuronal systems. We used isogenic euploid human induced pluripotent stem cell lines to generate and study an allelic series of PS1 mutations, including heterozygous null mutations and homozygous and heterozygous FAD PS1 mutations. Rigorous analysis of this allelic series in differentiated, purified neurons allowed us to resolve this controversy and to conclude that FAD PS1 mutations, expressed at normal levels in the appropriate cell type, impair γ-secretase activity but do not disrupt γ-secretase-independent functions of PS1. Thus, FAD PS1 mutations do not act as simple loss of PS1 function but instead dominantly gain an activity toxic to some, but not all, PS1 functions., (Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.)

The recent discovery of a simple method for making induced pluripotent stem cells (iPSC) from human somatic cells was a major scientific advancement that opened the way for many promising new developments in the study of developmental and degenerative diseases. iPSC have already been used to model many different types of neurological diseases, including autism, schizophrenia, Alzheimer's disease and Parkinson's disease. Because of their pluripotent property, iPSC offer the possibility of modeling human development in vitro. Their differentiation seems to follow the developmental timeline and obeys environmental cues. Clinically relevant phenotypes of neurodegenerative pathologies have also been observed using iPSC derived human neuronal cultures. Options for treatment are still some way off. Although some early research in mouse models has been encouraging, major obstacles remain for neural stem cell (NSC) transplantation therapy. However, iPSC now offer the prospect of an unlimited amount of human neurons or astrocytes for drug testing. The aim of this review is to summarize the recent progress in modeling neural development and neurological diseases using iPSC and to describe their applications for aging research and personalized medicine., (Copyright © 2013 Elsevier B.V. All rights reserved.)

Background: Posterior pedicle screw device is widely used in treatment of thoracolumbar burst fractures. As the clinical operation is not based upon quantitative data of adjustments, the results are not optimal. At present, no study has assessed the associations between the device adjustments and the restoration of stiffness. We investigated the biomechanical effects that adjustments of a pedicle screw device had on the burst fracture, and explored an optimal adjustment., Methods: Burst fractures were produced at L1 vertebra in 24 fresh calf spines (T12-L3). The specimens were divided into four groups at random. Pedicle screw devices were attached to T13 and L2. Four device adjustments, consisting of distraction and extension, were applied. Adjustment 1 was pure 6° extension, adjustment 2 was pure 5 mm distraction, adjustment 3 was 6° extension followed by 5 mm distraction, and adjustment 4 was 5 mm distraction followed by 6° extension. The effect of each adjustment on the stiffness restoration, anatomical reduction, and neural decompression for the burst fractures was analyzed and evaluated., Results: Pure extension (Group 1) produced the closest segment height and the least restoration of the canal to the intact. Pure distraction (Group 2) restored stiffness most, but with only 60% stiffness of the intact value, and lost the segmental angle most to the intact. The combination of extension-distraction (Group 3 and Group 4) produced the maximum reduction of the anatomy and restoration of the canal in the burst fracture, and the least stiffness restoration. The sequence of extension and distraction did not affect stiffness restoration, anatomical reduction, and neural decompression., Conclusions: The device adjustments affected stiffness restoration, anatomical reduction, and neural decompression. The combined extension-distraction adjustment may be the most suitable considering the anatomical reduction and neural decompression, but the stiffness decreased the most; it should be considered to reconstruct L1 vertebra.

The aim of the study was to investigate the effect of recombinant adeno-associated virus (AAV) with morphogenetic protein 4/7 fusion gene on the ossification of rabbit bone marrow stromal cells (BMSCs). The genes BMP-4 and BMP-7 were obtained through one-step reverse transcriptase polymerase chain reaction from human placental cells. The BMP-4/7 fusion gene was generated through recombination. The rabbit BMSCs were transfected with the recombinant AAV vectors carrying AAV-BMP-4/7 with various multiplicity of infection (MOI) values. Expression of fusion gene BMP-4/7 protein was determined by the ELISA method. The ossification of cells was evaluated by observing alkaline phosphatase (ALP) and osteocalcin (OC) activity after transfection for 7 and 14 days. We successfully constructed the recombinant AAV with the BMP-4/7 fusion gene. A 29-20 kDa protein was shown by SDS-PAGE electrophoresis following transfection with AAV-BMP-4/7. The fusion protein BMP-4/7 in BMSCs transferred by AAV showed a positive correlation with various MOI values. There was significantly higher ALP and OC activity in the AAV-BMP-4/7 transfection groups than in the AAV-EGFP groups (t(ALP)=896.88, P<0.001, t(OC)=543.24, P<0.01). The fusion gene BMP-4/7 is capable of enhancing the expression of BMPs and possesses significant ossification activity through AAV.

An unresolved issue about many neurodegenerative diseases is why neurons are particularly sensitive to defects in ubiquitous cellular processes. One example is Niemann Pick type C1, caused by defects in cholesterol trafficking in all cells, but where neurons are preferentially damaged. Understanding this selective failure is limited by the difficulty in obtaining live human neurons from affected patients. To solve this problem, we generated neurons with decreased function of NPC1 from human embryonic stem cells and used them to test the hypothesis that defective cholesterol handling leads to enhanced pathological phenotypes in neurons. We found that human NPC1 neurons have strong spontaneous activation of autophagy, and, contrary to previous reports in patient fibroblasts, a block of autophagic progression leading to defective mitochondrial clearance. Mitochondrial fragmentation is an exceptionally severe phenotype in NPC1 neurons compared with fibroblasts, causing abnormal accumulation of mitochondrial proteins. Contrary to expectation, these abnormal phenotypes were rescued by treatment with the autophagy inhibitor 3-methyladenine and by treatment with the potential therapeutic cyclodextrin, which mobilizes cholesterol from the lysosomal compartment. Our findings suggest that neurons are especially sensitive to lysosomal cholesterol accumulation because of autophagy disruption and accumulation of fragmented mitochondria, thus defining a new route to effective drug development for NPC1 disease.

Our understanding of Alzheimer's disease pathogenesis is currently limited by difficulties in obtaining live neurons from patients and the inability to model the sporadic form of the disease. It may be possible to overcome these challenges by reprogramming primary cells from patients into induced pluripotent stem cells (iPSCs). Here we reprogrammed primary fibroblasts from two patients with familial Alzheimer's disease, both caused by a duplication of the amyloid-β precursor protein gene (APP; termed APP(Dp)), two with sporadic Alzheimer's disease (termed sAD1, sAD2) and two non-demented control individuals into iPSC lines. Neurons from differentiated cultures were purified with fluorescence-activated cell sorting and characterized. Purified cultures contained more than 90% neurons, clustered with fetal brain messenger RNA samples by microarray criteria, and could form functional synaptic contacts. Virtually all cells exhibited normal electrophysiological activity. Relative to controls, iPSC-derived, purified neurons from the two APP(Dp) patients and patient sAD2 exhibited significantly higher levels of the pathological markers amyloid-β(1-40), phospho-tau(Thr 231) and active glycogen synthase kinase-3β (aGSK-3β). Neurons from APP(Dp) and sAD2 patients also accumulated large RAB5-positive early endosomes compared to controls. Treatment of purified neurons with β-secretase inhibitors, but not γ-secretase inhibitors, caused significant reductions in phospho-Tau(Thr 231) and aGSK-3β levels. These results suggest a direct relationship between APP proteolytic processing, but not amyloid-β, in GSK-3β activation and tau phosphorylation in human neurons. Additionally, we observed that neurons with the genome of one sAD patient exhibited the phenotypes seen in familial Alzheimer's disease samples. More generally, we demonstrate that iPSC technology can be used to observe phenotypes relevant to Alzheimer's disease, even though it can take decades for overt disease to manifest in patients.

In previous studies, we have demonstrated that spinal grafting of human or rat fetal spinal neural precursors leads to amelioration of spasticity and improvement in ambulatory function in rats with spinal ischemic injury. In the current study, we characterize the survival and maturation of three different human embryonic stem (ES) cell line-derived neural precursors (hNPCs) once grafted into ischemia-injured lumbar spinal cord in rats or in naive immunosuppressed minipigs. Proliferating HUES-2, HUES-7, or HUES-9 colonies were induced to form embryoid bodies. During the nestin-positive stage, the rosettes were removed and CD184(+)/CD271(-)/CD44(-)/CD24(+) population of ES-hNPCs FAC-sorted and expanded. Male Sprague-Dawley rats with spinal ischemic injury or naive immunosuppressed Gottingen-Minnesota minipigs received 10 bilateral injections of ES-NPCs into the L2-L5 gray matter. After cell grafting, animals survived for 2 weeks to 4.5 months, and the presence of grafted cells was confirmed after staining spinal cord sections with a combination of human-specific (hNUMA, HO14, hNSE, hSYN) or nonspecific (DCX, MAP2, CHAT, GFAP, APC) antibodies. In the majority of grafted animals, hNUMA-positive grafted cells were identified. At 2-4 weeks after grafting, double-labeled hNUMA/DCX-immunoreactive neurons were seen with extensive DCX(+) processes. At survival intervals of 4-8 weeks, hNSE(+) neurons and expression of hSYN was identified. Some hSYN-positive terminals formed putative synapses with the host neurons. Quantitative analysis of hNUMA(+) cells at 2 months after grafting showed comparable cell survival for all three cell lines. In the presence of low-level immunosuppression, no grafted cell survival was seen at 4.5 months after grafting. Spinal grafting of proliferating pluripotent HUES-7 cells led to consistent teratoma formation at 2-6 weeks after cell transplantation. These data show that ES-derived, FAC-sorted NPCs can represent an effective source of human NPCs to be used in CNS cell replacement therapies.

Background: Quercetin has been shown to induce apoptosis in a number of cancer cell lines, but a quercetin-loaded nanoliposomal formulation with enhanced antitumor activity in C6 glioma cells and its effect on cancer cell death has not been well studied. The aim of this study was to examine if quercetin-loaded liposomes (QUE-NL) has enhanced cytotoxic effects and if such effects involve type III programmed cell death in C6 glioma cells., Methods: C6 glioma cells were treated with QUE-NL and assayed for cell survival, apoptosis, and necrosis. Levels of reactive oxygen species production and loss of mitochondrial membrane potential (ΔΨm) were also determined by flow cytometry assay to assess the effects of QUE-NL. ATP levels and lactate dehydrogenase activity were measured, and Western blotting was used to assay cytochrome C release and caspase expression., Results: QUE-NL induced type III (necrotic) programmed cell death in C6 glioma cells in a dose-dependent and time-dependent manner. High concentrations of QUE-NL induced cell necrosis, which is distinct from apoptosis and autophagy, whereas liposomes administered alone induced neither significant apoptosis nor necrosis in C6 glioma cells. QUE-NL-induced ΔΨm loss and cytochrome C release had no effect on caspase activation, but decreased ATP levels and increased lactate dehydrogenase activity indicated that QUE-NL stimulated necrotic cell death., Conclusion: C6 glioma cells treated with QUE-NL showed a cellular pattern associated with necrosis without apoptosis and was independent of caspase activity. Nonapoptotic cell death induced by high concentrations of QUE-NL for controlling caspase-independent type III programmed cell death may provide the basis for novel therapeutic approaches to overcome avoidance of apoptosis by malignant cells.

Cancer polyarthritis is an uncommon, paraneoplastic manifestation of some solid and hematological malignancies. Herein, we report the case of a 55-year-old woman who recently experienced polyarthritis for 2 months. On admission, the patient developed cough, expectoration, and fever. According to the clinical manifestations and the findings in radiological examinations and sputum cultures, pneumonia was considered. No evidence of lung cancer was noted by repeated computed tomography scan of lung, single bronchoscopy, or computed tomography-guided lung biopsy. Ultimately, the second bronchoscopy with biopsy was carried out, and lung adenocarcinoma was confirmed by pathological examination. Symptoms of polyarthritis starting 2 months before the symptoms of her lung cancer in the present case leads us to believe that polyarthritis may be a manifestation of paraneoplastic syndrome. Arthritis resolved after anticancer therapy. Our report indicates that polyarthritis of unknown cause may be suspected as a manifestation of malignancies., (Copyright © 2011. Published by Elsevier B.V.)

The extracellular matrix (ECM) plays important roles in influencing cellular behavior such as attachment, differentiation, and proliferation. However, in conventional culture and tissue engineering strategies, single proteins are frequently utilized, which do not mimic the complex extracellular microenvironment seen in vivo. In this study we report a method to decellularize brain tissue using detergents. This decellularized brain matrix is rich in glycosaminoglycans and contains collagen I, collagen III, collagen IV, collagen V, collagen VI, perlecan, and laminin. By further processing the material into a liquid form, the brain matrix can be used as a cell culture coating. Neurons derived from human induced pluripotent stem cells plated on the brain matrix express neuronal markers and assume neuronal morphology. Additionally, the same material can potentially be used as a scaffold for tissue engineering as it reassembles upon injection in vivo to form a gel. Thus, our work demonstrates the ability to use decellularized brain ECM for cell culture and tissue engineering applications.

Cyclosporine (CsA) is a substrate of cytochrome P450 (CYP) 3A5 and has a narrow therapeutic range with large inter-individual variability. CYP3A5*3 polymorphism is reported to be functional and may contribute to the inter-individual variability. The objective of this meta-analysis was to accurately estimate the effect of CYP3A5*3 allele on CsA dose-adjusted blood concentration. A computerized literature search was conducted in PubMed. A total of 12 and 6 studies meeting the inclusion criteria were, respectively, included in meta-analysis about dose-adjusted trough concentration (C(0)/D) and dose-adjusted peak concentration (C(2)/D). The combined weighted mean difference (WMD) between CYP3A5 expressers (*1/*3 + *1/*1) and non-expressers (*3/*3) was significant in C(2)/D (WMD = -12.73 (ng ml(-1))/(mg kg(-1)), 95% confidence interval (CI) -25.23 to -0.22, P = 0.046), whereas it was marginally significant in C(0)/D (WMD = -3.75 (ng ml(-1))/(mg kg(-1)), 95% CI -7.58 to 0.07, P = 0.054). Exclusion of an outlier study greatly increased the association of CYP3A5 polymorphism with C(0)/D to be significant (WMD = -4.92 (ng ml(-1))/(mg kg(-1)), 95% CI: -8.27 to -1.58, P = 0.011). This meta-analysis showed that CYP3A5*3 polymorphism is associated with CsA dose-adjusted concentration in renal transplant recipients. Patients carrying the CYP3A5*3/*3 genotype will require a lower dose of CsA to reach target levels compared with the CYP3A5*1/*1 or *1/*3 carriers.

Background: Neural induction of human pluripotent stem cells often yields heterogeneous cell populations that can hamper quantitative and comparative analyses. There is a need for improved differentiation and enrichment procedures that generate highly pure populations of neural stem cells (NSC), glia and neurons. One way to address this problem is to identify cell-surface signatures that enable the isolation of these cell types from heterogeneous cell populations by fluorescence activated cell sorting (FACS)., Methodology/principal Findings: We performed an unbiased FACS- and image-based immunophenotyping analysis using 190 antibodies to cell surface markers on naïve human embryonic stem cells (hESC) and cell derivatives from neural differentiation cultures. From this analysis we identified prospective cell surface signatures for the isolation of NSC, glia and neurons. We isolated a population of NSC that was CD184(+)/CD271(-)/CD44(-)/CD24(+) from neural induction cultures of hESC and human induced pluripotent stem cells (hiPSC). Sorted NSC could be propagated for many passages and could differentiate to mixed cultures of neurons and glia in vitro and in vivo. A population of neurons that was CD184(-)/CD44(-)/CD15(LOW)/CD24(+) and a population of glia that was CD184(+)/CD44(+) were subsequently purified from cultures of differentiating NSC. Purified neurons were viable, expressed mature and subtype-specific neuronal markers, and could fire action potentials. Purified glia were mitotic and could mature to GFAP-expressing astrocytes in vitro and in vivo., Conclusions/significance: These findings illustrate the utility of immunophenotyping screens for the identification of cell surface signatures of neural cells derived from human pluripotent stem cells. These signatures can be used for isolating highly pure populations of viable NSC, glia and neurons by FACS. The methods described here will enable downstream studies that require consistent and defined neural cell populations.

TOX3 is a nuclear protein containing a high mobility group (HMG)-box domain, which regulates Ca(2+)-dependent transcription in neurons through interaction with the cAMP-response-element-binding protein (CREB). TOX3 appears to be associated with breast cancer susceptibility and was previously shown to be expressed downstream of a cytoprotective cascade together with CITED1, a transcriptional regulator that does not bind directly to DNA. In the present study we show that TOX3 is predominantly expressed in the brain, forms homodimers and interacts with CITED1. TOX3 overexpression protects neuronal cells from cell death caused by endoplasmic reticulum stress or BAX overexpression through the induction of anti-apoptotic transcripts and repression of pro-apoptotic transcripts, which correlates with enhanced transcription involving isolated estrogen-responsive elements and estrogen-responsive promoters. However, both functions cannot be inhibited with the anti-estrogen fulvestrant and are only attenuated by mutation of estrogen-responsive elements. TOX3 also interacts with native CREB and induces the CREB-responsive BCL-2 promoter, which can be inhibited by coexpression of CITED1. Coexpression of CREB, by contrast, abolishes TOX3-mediated transcription from the estrogen-responsive complement C3 promoter. Our results suggest that TOX3 can enhance transcriptional activation from different cytoprotective promoters and that this is dependent on the predominance of either phosphorylated CREB or CITED1 within the transcriptionally active complex.

The influence of reaction atmosphere and TiO2 on photochemical decomposition of perfluorooctanoic acid (PFOA) in aqueous periodate was investigated using a type of low-pressure mercury lamps emitted at 254 nm. PFOA photolysis was slight with 254 nm light irradiation under nitrogen, whereas significant decomposition PFOA obtained with the addition of IO4-. In addition, oxygen restrained photochemical decomposition of PFOA. In UV/TiO2/IO4- system, PFOA degradation ratio was 54%, 15% lower than that for UV/IO4- system. *OH radicals generated from UV/TiO2 system exhausted a lot of IO4-, resulting in lower degree of IO3* production. IO3* was high reactive radical which great excitated PFOA decomposition. The accumulation of short-chain perfluorocarbonxylic acids (PFCAs) as products were identified with HPLC/MS. PFCAs bearing shorter perfluoroalkyl groups were formed in a stepwise way from PFCAs that bear longer perfluoroalkyl groups.