A novel strategy for amplification full-length cDNA and promoter sequences has been developed using bioinformatics technology and multiplexed PCR methods in this study. The amplification of 3' ends ...of cDNA is performed according to the modified classic 3' RACE techniques, therein the more efficient and effective oligo(dT)-anchor primer with hairpin structure is specially designed. For the amplification of 5' ends of cDNA, two or three-round TAIL-PCR or touch-down PCR using arbitrary degenerate (AD) and sequence-specific reverse (SPR) primers is performed until the 5' sequence of multi-assembled fragment reaches the exon1 region identified by aligning this fragment to reference genome database. Then another TAIL-PCR or touch-down PCR using genomic DNA as template is conducted to obtain the remaining 5' and promoter sequences. The 5' end sites of cDNA are predicted by aligning finally assembled fragment to homologous reference genes of other species, and screening the relative locations of common characteristic cis-elements in silico on promoter. The putative 5' ends are further validated by primers corresponding to these predicted sites in cDNAs. This method is suitable for researchers to isolate limited full-length cDNA sequences due to its operability, inexpensiveness, efficiency and speediness.
The objective of the work described here was to assess the value of the combination of pre-operative multimodal data-including clinical data, contrast-enhanced ultrasound (CEUS) information and liver ...stiffness measurement (LSM) based on 2-D shear wave elastography (SWE)-in predicting early (within 1 y) and late (after 1 y) recurrence of hepatocellular carcinoma (HCC) after curative treatment.
We retrospectively included 101 patients with HCC who met the Milan criteria and received curative treatment. The multimodel data from clinical parameters, LSM by 2-D SWE and CEUS enhancement patterns were collected. The association between different variables in HCC recurrence was accessed using a Cox proportional hazard model. On the basis of the independent factors of early recurrence, models with different source variables were established (Clinical Model, CEUS-Clinical Model, SWE-Clinical Model, CEUS-SWE-Clinical Model). The goodness-of-fit of models was evaluated and the performance trends of different models were calculated by time-dependent area under the curve (AUC).
Two-dimensional SWE, CEUS enhancement patterns and clinical parameters (spleen length, multiple tumors, α-fetoprotein, albumin and prothrombin time) were independently associated with early recurrence (all p values <0.05). Multiple tumors and a decrease in albumin independently contributed to the late recurrence. The model fit of CEUS-SWE-Clinical Model was superior to other models in predicting early recurrence (all p values <0.05). The AUCs of the CEUS-Clinical Model were higher from 2 mo to 7 mo, while the SWE-Clinical Model had higher AUCs from 9 mo to 12 mo.
CEUS enhancement patterns and 2-D SWE were independent predictors of HCC early recurrence as the two factors contributed to the predictive performance at different times. The multimodal model, which included diverse data in predicting early HCC recurrence, had the best goodness-of-fit.
To study
adaption to water hypoxia, the changes in physiological levels, innate immune responses, redox balance of
.amblycephala during hypoxia were investigated in the present study. When
were ...exposed to different dissolved oxygen (DO) including control (DO: 5.5 mg/L) and acute hypoxia (DO: 3.5 and 1.0 mg/L, respectively), hemoglobin (Hb), methemoglobin (MetHb), glucose, Na
, succinatedehydrogenase (SDH), lactate, interferon alpha (IFNα), and lysozyme (LYZ), except hepatic glycogen and albumin gradually increased with the decrease of DO level. When
were exposed to different hypoxia time including 0.5 and 6 h (DO: 3.5 mg/L), and then reoxygenation for 24 h after 6 h hypoxia, Hb, MetHb, glucose, lactate, and IFNα, except Na
, SDH, hepatic glycogen, albumin, and LYZ increased with the extension of hypoxia time, while the above investigated indexes (except albumin, IFNα, and LYZ) decreased after reoxygenation. On the other hand, the liver SOD, CAT, hydrogen peroxide (H
O
), and total ROS were all remained at lower levels under hypoxia stress. Finally, Hif-1α protein in the liver, spleen, and gill were increased with the decrease of oxygen concentration and prolongation of hypoxia time. Interestingly, one Hsp70 isoforms mediated by internal ribozyme entry site (IRES) named junior Hsp70 was only detected in liver, spleen and gill. Taken together, these results suggest that hypoxia affects
physiology and reduces liver oxidative stress. Hypoxia-reoxygenation stimulates
immune parameter expressions, while Hsp70 response to hypoxia is tissue-specific.
Transcription and replication of the influenza A virus (IAV) genome take place in the nucleus of infected cells, which rely on host factors to aid viral ribonucleoprotein (vRNP) to cross the nuclear ...pore complex (NPC) and complete the bidirectional nucleocytoplasmic trafficking. Here, we showed that nucleoporin 85 (NUP85), a component of NPC, interacted with RNP subunits polymerase basic 1 (PB1) and polymerase basic 2 (PB2) in an RNA-dependent manner during IAV infection. Knockdown of NUP85 delayed the nuclear import of vRNP, PB1 and PB2, inhibiting polymerase activity and ultimately suppressing viral replication. Further analysis revealed that NUP85 assisted the binding of PB1 to nuclear transport factor Ran-binding protein 5 (RanBP5) and the binding of PB2 to nuclear transport factor importin α1 and importin α7. We also found that NUP85 expression was downregulated upon IAV infection. Together, our study demonstrated that NUP85 positively regulated IAV infection by interacting with viral PB1 and PB2, which may provide new insight into the process of vRNP nuclear import and a novel target for effective antivirals.
Nitrite stress is a major environmental factor that limits aquatic animal growth, reproduction and survival. Even so, some shrimps still can withstand somewhat high concentrations of nitrite ...environment. However, few studies have been conducted about the tolerance molecular mechanism of Litopenaeus vannamei in the high concentration nitrite. To identify the genes and pathways involved in the regulation of nitrite tolerance, we performed comparative transcriptomic analysis in the L. vannamei nitrite-tolerant (NT) and nitrite-sensitive (NS) families, and untreated shrimps were used as the control group. After 24 h of nitrite exposure (NaNO2, 112.5 mg/L), a total of 1521 and 868 differentially expressed genes (DEGs) were obtained from NT compared with NS and control group, respectively. Functional enrichment analysis revealed that most of these DEGs were involved in immune defense, energy metabolism processes and endoplasmic reticulum (ER) stress. During nitrite stress, energy metabolism in NT was significantly enhanced by activating the related genes expression of oxidative phosphorylation (OXPHOS) pathway and tricarboxylic acid (TCA) cycle. Meanwhile, some DEGs involved in innate immunity- related genes and pathways, and ER stress responses also were highly expressed in NT. Therefore, we speculate that accelerated energy metabolism, higher expression of immunity and ER related genes might be the important adaptive strategies for NT in relative to NS under nitrite stress. These results will provide new insights on the potential tolerant molecular mechanisms and the breeding of new varieties of nitrite tolerant L. vannamei.
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•Transcriptomic responses to nitrite stress were explored in L. vannamei with different tolerance.•Most of DEGs were involved in immune defense, energy metabolism and endoplasmic reticulum stress.•The activation of OXPHOS pathway and TCA cycle might be related to nitrite tolerance.•Enhanced expressions of innate immunity and ER stress related-genes might be an important defensive mechanism.
To correlate the microbiological and clinical features of infections caused by Nocardia species.
We determined the species and drug susceptibility of 138 Nocardia strains isolated from 132 patients ...at the University of Texas M. D. Anderson Cancer Center (Houston, TX) from 2002 through 2012 and analyzed the clinical features.
The 132 patients included 82 men and 50 women with a mean age of 59.1 years. All except two had underlying cancer, and 47 (35.6%) also received a stem cell transplant. These patients experienced 136 episodes of Nocardia infection, including pulmonary infection, abscess of deep skin and soft tissue, bacteremia and dissemination, and brain abscess. The 138 Nocardia strains involved 27 species, of which 20 species have been described since 2000. Common species included Nocardia nova, Nocardia cyriacigeorgica, Nocardia farcinica, and Nocardia abscessus, together accounting for 59.4%. N nova caused most bacteremia cases, whereas N farcinica caused most of the skin and brain infections. Infections with a few recent species likely represented first confirmation or report of human infections. Antimicrobial susceptibility tests of 117 strains showed that they were all susceptible to trimethoprim-sulfamethoxazole and linezolid but variably susceptible to other drugs depending on species. Most patients who were treated for the infection showed improvement or resolution.
Diverse Nocardia species can cause secondary infections in patients with cancer. Timely species identification and antimicrobial susceptibility tests may guide treatment.
Introduction
Poliovirus receptor (PVR) is a tumor promoter and a regulatory checkpoint that enhances immunosuppression. We investigated PVR expression by applying immunohistochemistry (IHC) staining. ...A positive association existed between PVR expression and cytotoxic T lymphocyte-associated antigen 4 (CTLA4) expression in patients with surgically resected non-small-cell lung cancer (NSCLC). PVR expression is a prognosis predictor of lung adenocarcinoma.
Purpose
To investigate the prognostic significance of PVR expression and CTLA4 expression for surgically resected NSCLC.
Patients and methods
The medical records of 108 Chinese patients with primary NSCLC who underwent surgery were retrospectively reviewed. The expression of PVR and CTLA4 were measured through IHC. Clinical characteristics, the association between PVR and CTLA4, and the prognostic significance of PVR were analyzed.
Results
A significant positive association was observed between PVR and CTLA4 expression in NSCLC (
P
= 0.016). PVR had a high positive rate among females, nonsmokers, and patients with adenocarcinoma and advanced lung cancer. The overall survival (OS) of patients with negative PVR expression was significantly longer than that of patients with positive PVR expression (
P
= 0.049), especially among females (
P
= 0.03) and nonsmokers (
P
= 0.025). Multivariate analysis results showed that advanced tumor stage and PVR expression were independent prognosis predictors of poor OS.
Conclusion
PVR can potentially serve as a prognostic predictor and biomarker for NSCLC and cancer anti-CTLA4 immunotherapy response.
Hepatolenticular degeneration, also known as Wilson's disease (WD), is an autosomal recessive inheritance nervous disorder of copper metabolism. The treatment of hepatolenticular degeneration ...emphasizes the combination of medical therapy and dietary therapy, such as a high zinc, low copper and sulfhydryl rich diet. Food therapy of WD based on trace elements is presented in this paper.
The negative relationship between maternal high-density lipoprotein-cholesterol (HDL-c) level during pregnancy and infant birth weight has been found. Syncytialization (differentiation and fusion) of ...trophoblast cells is important to fetal development. HDL has an antioxidant effect, and has been proved to protect trophoblast functions including hormone secretion and invasion. However, HDL is susceptible to oxidation, and high concentrations of HDL impair cell growth and oxidized HDL (oxHDL) inhibits cell proliferation and migration. Moreover, the effects of HDL and oxHDL on trophoblast syncytialization have not been characterized. The aim of this study was to investigate the effects of HDL and oxHDL on trophoblast syncytialization.
Human choriocarcinoma trophoblasts (BeWo cells) were treated with human HDL or oxHDL and then induced to differentiate by forskolin in syncytialization assays. Expression levels of mRNAs and proteins regulating syncytialization were detected by real-time PCR and western blotting, respectively.
Treatments of HDL at high concentrations reduced human chorionic gonadotropin (hCG) secretion, placental alkaline phosphatase activity and fusion rates, and decreased the expressions of GCM1 and ERVW-1 mRNA as well as phospho-MAPK1/3 (p-MAPK1/3) and total MAPK1/3 protein in the forskolin-induced syncytialization of BeWo cells. Furthermore, treatment of oxHDL (20 μg/ml) decreased hCG secretion, but increased the expression of p-MAPK1/3 protein.
These data suggested that both HDL at high concentrations and oxHDL inhibited BeWo cells syncytialization, and might be harmful to placental and fetal development.
•High-density lipoprotein at high concentrations inhibited BeWo cells syncytialization.•Oxidized high-density lipoprotein inhibited BeWo cells syncytialization.•Inhibited syncytialization might have inverse effects on placental and fetal development.
Safety evaluation of toppling rock slopes developing in reservoir areas is crucial. To reduce the uncertainty of safety evaluation, this study developed a composite cloud model, which improved the ...combination weights of the decision-making trial and evaluation laboratory (DEMATEL) and criteria importance through intercriteria correlation (CRITIC) methods. A safety evaluation system was developed according to in situ monitoring data. The backward cloud generator was used to calculate the numerical characteristics of a cloud model of quantitative indices, and different virtual clouds were used to synthesize some clouds into a generalized one. The synthesized numerical characteristics were calculated to comprehensively evaluate the safety of toppling rock slopes. A case study of a toppling rock slope near the Huangdeng Hydropower Station in China was conducted using monitoring data collected since operation of the hydropower project began. The results indicated that the toppling rock slope was moderately safe with a low safety margin. The composite cloud model considers the fuzziness and randomness of safety evaluation and enables interchange between qualitative and quantitative knowledge. This study provides a new theoretical method for evaluating the safety of toppling rock slopes. It can aid in the predication, control, and even prevention of disasters.
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