Changes in DNA methylation and transgenerational mobilization of a transposable element (mPing) by the topoisomerase II inhibitor, etoposide, in rice

ABSTRACT: BACKGROUND: Etoposide (epipodophyllotoxin) is a chemical commonly used as an anti-cancer drug which inhibits DNA synthesis by blocking topoisomerase II activity. Previous studies in animal cells have demonstrated that etoposide constitutes a genotoxic stress which may induce genomic instability including mobilization of normally quiescent transposable elements (TEs). However, it remained unknown whether similar genetically mutagenic effects could be imposed by etoposide in plant cells. Also, no information is available with regard to whether the drug may cause a perturbation of epigenetic stability in any organism. RESULTS: To investigate whether etoposide could generate genetic and/or epigenetic instability in plant cells, we applied etoposide to germinating seeds of six cultivated rice (Oryza sativa L.) genotypes including both subspecies, japonica and indica. Based on the methylation-sensitive gel-blotting results, epigenetic changes in DNA methylation of three TEs (Tos17, Osr23 and Osr36) and two protein-encoding genes (Homeobox and CDPK-related genes) were detected in the etoposide-treated plants (S0 generation) in four of the six studied japonica cultivars, Nipponbare, RZ1, RZ2, and RZ35, but not in the rest japonica cultivar (Matsumae) and the indica cultivar (93-11). DNA methylation changes in the etoposide-treated S0 rice plants were validated by bisulfite sequencing at both of two analyzed loci (Tos17 and Osr36). Transpositional activity was tested for eight TEs endogenous to the rice genome in both the S0 plants and their selfed progenies (S1 and S2) of one of the cultivars, RZ1, which manifested heritable phenotypic variations. Results indicated that no transposition occurred in the etoposide-treated S0 plants for any of the TEs. Nonetheless, a MITE transposon, mPing, showed rampant mobilization in the S1 and S2 progenies descended from the drug-treated S0 plants. CONCLUSIONS: Our results demonstrate that etoposide imposes a similar genotoxic stress on plant cells as it does on animal and human cells, which may induce transgenerational genomic instability by instigating transpositional activation of otherwise dormant TEs. In addition, we show for the first time that etoposide may induce epigenetic instability in the form of altered DNA methylation patterns in eukaryotes. However, penetration of the genotoxic effects of etoposide on plant cells, as being reflected as genetic and epigenetic instability, appears to be in a strictly genotype- and/or generation-dependent manner.     

Liquid-Solid Phase-Inversion PLGA Implant for the Treatment of Residual Tumor Tissue after HIFU Ablation

BackgroundHIFU has been shown to be a more suitable alternative for the treatment of primary solid tumors and metastatic diseases than other focal heat ablation techniques due to its noninvasive and extracorporeal nature. However, similar to other focal heat ablation techniques, HIFU is still in need of refinements due to tumor recurrence.MethodsIn this work, we investigated the effectiveness of an adjunct treatment regimen using doxorubicin (DOX)-loaded, injectable, in situ-forming, and phase-inverting PLGA as the second line of defense after HIFU ablation to destroy detrimental residual tumors and to prevent tumor recurrence. All of the statistical analyses were performed using the Statistical Package for the Social Sciences 18.0(SPSS, Inc., Chicago, IL, USA), and p< 0.05 was considered statistically significant. All of the results are presented as the means ± STDEV (standard deviation). For multiple comparisons, ANOVA (differences in tumor volumes, growth rates, apoptosis, proliferation indexes, and Bcl-2 and Bax protein levels) was used when the data were normally distributed with homogenous variance, and rank sum tests were used otherwise. Once significant differences were detected, Student-t tests were used for comparisons between two groups.ResultsOur results revealed that DOX diffused beyond the ablated tissue regions and entered tumor cells that were not affected by the HIFU ablation. Our results also show that HIFU in concert with DOX-loaded PLGA led to a significantly higher rate of tumor cell apoptosis and a lower rate of tumor cell proliferation in the areas beyond the HIFU-ablated tissues and consequently caused significant tumor volume shrinkage (tumor volumes:0.26±0.1,1.09±0.76, and 1.42±0.9cm³ for treatment, sham, and no treatment control, respectively).ConclusionsFrom these results, we concluded that the intralesional injection of DOX-loaded PLGA after HIFU ablation is significantly more effective than HIFU alone for the treatment of solid tumors.     

EFFECTS OF LIGHT ON PHOTOSYNTHESIS,GRAZING, AND POPULATION DYNAMICS OF THE HETEROTROPHIC DINOFLAGELLATE PFIESTERIA PISCICIDA (DINOPHYCEAE)1

In studying how environmental factors control the population dynamics of Pfiesteria piscicida Steidinger et Burkholder, we examined the influence of light regime on kleptoplastidic photosynthesis, growth, and grazing. Prey (Rhodomonas sp.)‐saturated growth rate of P. piscicida increased (0.67 ± 0.03 d^?1 to 0.91 ± 0.11 d^?1) with light intensity varying from 0 to 200 μmol photons·m^?2·s^?1. No significant effect was observed on grazing, excluding the possibility that light enhanced P. piscicida growth through stimulating grazing. Light‐grown P. piscicida exhibited a higher gross growth efficiency (0.78 ± 0.10) than P. piscicida incubated in the dark (0.32 ± 0.16), and photosynthetic inhibitors significantly decreased growth of recently fed populations. These results demonstrate a role of kleptoplastidic photosynthesis in enhancing growth in P. piscicida. However, when the prey alga R. sp. was depleted, light's stimulating effect on P. piscicida growth diminished quickly, coinciding with rapid disappearance of Rhodomonas‐derived pigments and RUBISCO from P. piscicida cells. Furthermore, the effect of light on growth was reversed after extended starvation, and starved light‐grown P. piscicida declined at a rate significantly greater than dark‐incubated cultures. The observed difference in rates of decline appeared to be attributable to light‐dependent cannibalism. Using a 5‐chloromethylfluorescein diacetate staining technique, cannibalistic grazing was observed after 7 days of starvation, at a rate four times greater under illumination than in the dark. The results from this study suggest that kleptoplastidy enhances growth of P. piscicida only in the presence of algal prey. When prey is absent, P. piscicida populations may become vulnerable to light‐stimulated cannibalism.     

Intratumoral expression of IL-17 and its prognostic role in gastric adenocarcinoma patients

In this study, we characterized the intratumoral expression of IL-17 and CD8(+) TILs in gastric adenocarcinoma patients after resection and determined the correlation between the survival probability of gastric adenocarcinoma patients and the expression of IL-17 in tumor. Expression of IL-17 and CD8 was assessed by immunohistochemistry, and the prognostic effects of intratumoral IL-17 expression and CD8(+) TILs were evaluated by Cox regression and Kaplan-Meier analysis. Immunohistochemical detection revealed the presence of IL-17 and CD8(+) cells in gastric adenocarcinoma tissue samples (90.6%, 174 out of 192 patients and 96.9%, 186 out of 192 patients, respectively). We have also found that intratumoral IL-17 expression was significantly correlated with age (p=0.004) and that the number of CD8(+)TILs was significantly correlated with UICC staging (p=0.012) and the depth of tumor invasion (p=0.022). The five-year overall survival probability among patients intratumorally expressing higher levels of IL-17 was significantly better than those expressing lower levels of IL-17 (p=0.036). Multivariate Cox proportional hazard analyses revealed that intratumoral IL-17 expression (HR: 0.521; 95% CI: 0.329-0.823; p=0.005) was an independent factor affecting the five-year overall survival probability. We conclude that low levels of intratumoral IL-17 expression may indicate poor prognosis in gastric adenocarcinoma patients.     

A recombinant mimetics of the HIV-1 gp41 prehairpin fusion intermediate fused with human IgG Fc fragment elicits neutralizing antibody response in the vaccinated mice

HIV-1 gp41 prehairpin fusion intermediate (PFI) composed of three N-terminal heptad repeats (NHR) plays a crucial role in viral fusion and entry and represents an attractive target for anti-HIV therapeutics (e.g., enfuvirtide) and vaccines. In present study, we constructed and expressed two recombinant gp41 PFI mimetics, designated N46Fd and N46FdFc. N46Fd consists of N46 (residues 536-581) in gp41 NHR and foldon (Fd), a trimerization motif. N46FdFc is composed of N46Fd fused with human IgG Fc fragment as an immunoenhancer. We immunized mice with N46 peptide, N46Fd and N46FdFc, respectively, and found that only N46FdFc elicited neutralizing antibody response in mice against infection by HIV-1 strains IIIB (clade B, X4), 92US657 (clade B, R5), and 94UG103 (clade A, X4R5). Anti-N46FdFc antibodies inhibited PIE7 binding to PFI, blocked gp41 six-helix bundle formation, and suppressed HIV-1 mediated cell-cell fusion. These findings provide an important clue for developing recombinant gp41 PFI mimetics-based HIV vaccines.     

Deficiency in the anti‐aging gene Klotho promotes aortic valve fibrosis through AMPKα‐mediated activation of RUNX2

Fibrotic aortic valve disease (FAVD) is an important cause of aortic stenosis, yet currently there is no effective treatment for FAVD due to its unknown etiology. The purpose of this study was to investigate whether deficiency in the anti‐aging Klotho gene (KL) promotes high‐fat‐diet‐induced FAVD and to explore the underlying molecular mechanism. Heterozygous Klotho‐deficient (KL^+/?) mice and WT littermates were fed with a high‐fat diet (HFD) or normal diet for 13 weeks, followed by treatment with the AMPKα activator (AICAR) for an additional 2 weeks. A HFD caused a greater increase in collagen levels in the aortic valves of KL^+/? mice than of WT mice, indicating that Klotho deficiency promotes HFD‐induced aortic valve fibrosis (AVF). AMPKα activity (pAMPKα) was decreased, while protein expression of collagen I and RUNX2 was increased in the aortic valves of KL^+/? mice fed with a HFD. Treatment with AICAR markedly attenuated HFD‐induced AVF in KL^+/? mice. AICAR not only abolished the downregulation of pAMPKα but also eliminated the upregulation of collagen I and RUNX2 in the aortic valves of KL^+/? mice fed with HFD. In cultured porcine aortic valve interstitial cells, Klotho‐deficient serum plus cholesterol increased RUNX2 and collagen I protein expression, which were attenuated by activation of AMPKα by AICAR. Interestingly, silencing of RUNX2 abolished the stimulatory effect of Klotho deficiency on cholesterol‐induced upregulation of matrix proteins, including collagen I and osteocalcin. In conclusion, Klotho gene deficiency promotes HFD‐induced fibrosis in aortic valves, likely through the AMPKα–RUNX2 pathway.     

LED照明对植物体内功能性化学物质积累的影响

LED照明对植物生长发育作用的研究已进行了多年, 但之前学者们多从植物形态学角度进行研究。近年来, 该项研究已逐步转向LED照明对植物体内功能性化学物质积累的影响这一领域。该文就这方面研究的最新进展进行了评述, 并指出了目前存在的问题及今后研究的思路和可能方向, 以供生产和研究部门科研人员参考。     

食管穿孔外科治疗22例临床分析

目的:探讨食管穿孔的诊断、临床特征与外科治疗方法.方法:回顾分析1999年-2009年我院收治的食管穿孔22例.结果:本组病例4例保守治疗,2例行颈部脓肿切开引流,8例行单纯食管穿孔修补术,3例行纵隔及胸腔脓肿清除术并引流,1例行食管下段切除、胃代食管弓上吻合术,2例行食管修补并肺叶切除,2例行空肠双管造瘘加纵隔胸腔引流,2例因经济原因放弃治疗出院.治愈19例,治愈率86.3%,死亡1例,死亡率4.5%.结论:早期诊断和及时采取正确的处理措施是提高本病治愈率,降低死亡率的关键.