Transgenic rice endosperm as a bioreactor for molecular pharming

Plants provide a promising expression platform for producing recombinant proteins with several advantages in terms of high expression level, lower production cost, scalability, and safety and environment-friendly. Molecular pharming has been recognized as an emerging industry with strategic importance that could play an important role in economic development and healthcare in China. Here, this review represents the significant advances using transgenic rice endosperm as bioreactor to produce various therapeutic recombinant proteins in transgenic rice endosperm and large-scale production of OsrHSA, and discusses the challenges to develop molecular pharming as an emerging industry with strategic importance in China.     

Asymmetric Addition of Triethylaluminium to Aromatic Aldehydes Catalyzed by Titanium‐(5,5’‐Biquinoline‐6,6’‐Diol) Complexes

A novel convenient procedure for the resolution of 5,5’‐biquinoline‐6,6’‐diol (BIQOL) was achieved by separating the corresponding diastereomeric mixture of (S)‐(+)‐camphorsulfonates on a semiprepared XDB‐C8 column followed by hydrolysis. The efficient asymmetric addition of triethylaluminium to aromatic aldehydes catalyzed by Ti‐(+)/(–)BIQOL complexes under mild conditions is described. The reactions led to the formation of 1‐arylpropan‐1‐ol in up to 87.5% ee. Chirality 26:268‐271, 2014. © 2014 Wiley Periodicals, Inc.     

Intermittent Preventive Treatment of Malaria in Pregnancy with Mefloquine in HIV-Negative Women: A Multicentre Randomized Controlled Trial

Background

Intermittent preventive treatment in pregnancy (IPTp) with sulfadoxine-pyrimethamine (SP) is recommended by WHO to prevent malaria in African pregnant women. The spread of SP parasite resistance has raised concerns regarding long-term use for IPT. Mefloquine (MQ) is the most promising of available alternatives to SP based on safety profile, long half-life, and high efficacy in Africa. We evaluated the safety and efficacy of MQ for IPTp compared to those of SP in HIV-negative women.

Methods and Findings

A total of 4,749 pregnant women were enrolled in an open-label randomized clinical trial conducted in Benin, Gabon, Mozambique, and Tanzania comparing two-dose MQ or SP for IPTp and MQ tolerability of two different regimens. The study arms were: (1) SP, (2) single dose MQ (15 mg/kg), and (3) split-dose MQ in the context of long lasting insecticide treated nets. There was no difference on low birth weight prevalence (primary study outcome) between groups (360/2,778 [13.0%]) for MQ group and 177/1,398 (12.7%) for SP group; risk ratio [RR], 1.02 (95% CI 0.86–1.22; p = 0.80 in the ITT analysis). Women receiving MQ had reduced risks of parasitemia (63/1,372 [4.6%] in the SP group and 88/2,737 [3.2%] in the MQ group; RR, 0.70 [95% CI 0.51–0.96]; p = 0.03) and anemia at delivery (609/1,380 [44.1%] in the SP group and 1,110/2743 [40.5%] in the MQ group; RR, 0.92 [95% CI 0.85–0.99]; p = 0.03), and reduced incidence of clinical malaria (96/551.8 malaria episodes person/year [PYAR] in the SP group and 130/1,103.2 episodes PYAR in the MQ group; RR, 0.67 [95% CI 0.52–0.88]; p = 0.004) and all-cause outpatient attendances during pregnancy (850/557.8 outpatients visits PYAR in the SP group and 1,480/1,110.1 visits PYAR in the MQ group; RR, 0.86 [0.78–0.95]; p = 0.003). There were no differences in the prevalence of placental infection and adverse pregnancy outcomes between groups. Tolerability was poorer in the two MQ groups compared to SP. The most frequently reported related adverse events were dizziness (ranging from 33.9% to 35.5% after dose 1; and 16.0% to 20.8% after dose 2) and vomiting (30.2% to 31.7%, after dose 1 and 15.3% to 17.4% after dose 2) with similar proportions in the full and split MQ arms. The open-label design is a limitation of the study that affects mainly the safety assessment.

Conclusions

Women taking MQ IPTp (15 mg/kg) in the context of long lasting insecticide treated nets had similar prevalence rates of low birth weight as those taking SP IPTp. MQ recipients had less clinical malaria than SP recipients, and the pregnancy outcomes and safety profile were similar. MQ had poorer tolerability even when splitting the dose over two days. These results do not support a change in the current IPTp policy.

Trial registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT 00811421","term_id":"NCT00811421"}}NCT 00811421; Pan African Clinical Trials Registry PACTR 2010020001429343 Please see later in the article for the Editors'' Summary     

TGF-β1 诱导人牙周膜细胞细胞骨架重排的机制*

摘要目的：牙周病是由多种因素引起的,特别是人牙周膜细胞的缺失。转化生长因子-β1（TGF-β1）是一种多功能细胞因子,在治 疗牙周病中发挥重要的作用,但很少有人清楚地研究TGF-β1 对人牙周膜细胞的影响。因此,本研究的目的是探讨TGF-β1诱导 人牙周膜细胞细胞骨架重排的信号通路。方法：人牙周膜细胞取自健康的前磨牙,并向同步化处理的细胞中加入10 ng / ml 的 TGF-β1,并通过相差显微镜观察它们的形态学变化。通过免疫组化和共聚焦显微镜观察F-肌动蛋白重排。用Western blot 分析蛋 白表达情况。结果：我们发现TGF-β1 诱导人牙周膜细胞细胞骨架重排,激活ROCK 蛋白的表达,并增加p-lIMK 和p-cofilin 的 蛋白表达。ROCK 抑制剂Y-27632 使ROCK,p-lIMK 和p-cofilin 的蛋白表达下降。结论：TGF-β1可以诱导人牙周膜细胞细胞骨架 重排,并且是通过上凋ROCK,p-lIMK 和p-cofilin 的活性完成的。本研究可以增强对TGF-β1 在治疗牙周疾病方面的作用机制的 了解。     

Knockdown of VEGFR2 Inhibits Proliferation and Induces Apoptosis in Hemangioma-Derived Endothelial Cells

Angiogenesis is a process of development and growth of new capillary blood vessels from pre-existing vessels. Angiogenic growth factors play important roles in the development and maintenance of some malignancies, of which vascular endothelial growth factor (VEGF)/VEGFR2 interactions are involved in proliferation, migration, and survival of many cancer cells. The aim of this study was to investigate the function of VEGFR2 in human hemangiomas (HAs). Using immunohistochemistry assay, we examined the expression levels of VEGF, VEGFR2, Ki-67, glucose transporter-1 (Glut-1), phosphorylated protein kinase B (p-AKT) and p-ERK in different phases of human HAs. Positive expression of VEGF, VEGFR2, Ki-67, Glut-1, p-AKT and p-ERK was significantly increased in proliferating phase HAs, while decreased in involuting phase HAs (P=0.001; P=0.003). In contrast, cell apoptotic indexes were decreased in proliferating phase HAs, but increased in involuting phase HAs (P<0.01). Furthermore, we used small hairpin RNA (shRNA)-mediated VEGFR2 knockdown in primary HA-derived endothelial cells (HemECs) to understand the role of VEGF/VEGFR2 signaling. Knockdown of VEGFR2 by Lv-shVEGFR2 inhibited cell viability and induced apoptosis in primary HemECs companied with decreased expression of p-AKT, p-ERK, p-p38MAPK and Ki-67 and increased expression of caspase-3 (CAS-3); Overexpression of VEGFR2 promoted cell viability and blocked apoptosis in Lv-VEGFR2-transfected HemECs. Taken together, our findings demonstrate that, increased expression of VEGFR2 is involved in the development of primary HemECs possibly through regulation of the AKT and ERK pathways, suggesting that VEGFR2 may be a potential therapeutic target for HAs.Key words: vascular endothelial growth factor receptor 2, hemangioma, proliferation, apoptosis     

The dual role of phytoene synthase genes in carotenogenesis in carrot roots and leaves

Carrot (Daucus carota L.) is an important food crop and is useful for studying carotenogenesis due to the quantity and diversity of carotenoids in its roots. Phytoene synthase catalyzes the first committed step in the carotenoid biosynthesis pathway, and its overexpression is the main driving force in the orange phenotype. At present, we lack fundamental knowledge of the role of these genes and their effects on carotenoid accumulation in leaves. In the present study, three backcross inbred lines (BC2S4) with different colored roots derived from a cross between the orange inbred line (Af) and related wild species were used to investigate the role of the duplicated DcPSY genes in root carotenogenesis. Promoter analysis showed that DcPSY genes have diverged substantially in their regulatory sequences after gene duplication. Expression levels of DcPSY1 and DcPSY2 were generally positively correlated with carotenoid content during root development. In mature leaves, total carotenoid content was higher than that in the roots, DcPSY1 expression increased extremely higher than DcPSY2 expression compared with roots, and DcPSY1 was more sensitive than DcPSY2 during leaf de-etiolation under sunlight. These results suggest that DcPSY1 seems to make an important contribution to carotenoid accumulation in the leaves and is important for photosynthesis and photoprotection, but they are not the determining factors of root color. This expands our understanding of the regulation of carotenoid biosynthesis in carrot.     

Integrated analyses of DNA methylation and hydroxymethylation reveal tumor suppressive roles of ECM1, ATF5, and EOMES in human hepatocellular carcinoma

Background

Differences in 5-hydroxymethylcytosine, 5hmC, distributions may complicate previous observations of abnormal cytosine methylation statuses that are used for the identification of new tumor suppressor gene candidates that are relevant to human hepatocarcinogenesis. The simultaneous detection of 5-methylcytosine and 5-hydroxymethylcytosine is likely to stimulate the discovery of aberrantly methylated genes with increased accuracy in human hepatocellular carcinoma.

Results

Here, we performed ultra-performance liquid chromatography/tandem mass spectrometry and single-base high-throughput sequencing, Hydroxymethylation and Methylation Sensitive Tag sequencing, HMST-seq, to synchronously measure these two modifications in human hepatocellular carcinoma samples. After identification of differentially methylated and hydroxymethylated genes in human hepatocellular carcinoma, we integrate DNA copy-number alterations, as determined using array-based comparative genomic hybridization data, with gene expression to identify genes that are potentially silenced by promoter hypermethylation.

Conclusions

We report a high enrichment of genes with epigenetic aberrations in cancer signaling pathways. Six genes were selected as tumor suppressor gene candidates, among which, ECM1, ATF5 and EOMES are confirmed via siRNA experiments to have potential anti-cancer functions.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-014-0533-9) contains supplementary material, which is available to authorized users.     

Differences in TCR-Vβ Repertoire and Effector Phenotype between Tumor Infiltrating Lymphocytes and Peripheral Blood Lymphocytes Increase with Age

Tumor infiltrating lymphocytes (TIL) reflect the host''s anti-tumor immune response, and can be a valuable predictor of prognosis. However, many properties of TIL are not fully understood. In the present study, TCR-Vβ repertoires of cancer patients were primarily analyzed by flow cytometry. Abnormally expressed TCR-Vβ subfamilies were generally found in both TIL and peripheral blood lymphocytes (PBL) of each patient. Of note, increased patient age was associated with increasingly biased TCR-Vβ repertoire in TIL but not in PBL, and the dispersion degree of the differences of TCR-Vβ subfamilies between TIL and PBL correlated positively with age (P = 0.007). Utilizing immunoscope analysis, we identified the age-related reduction in TCR-Vβ diversity, but polyclonal pattern was predominant in significantly expanded TCR-Vβ subfamilies. In addition, we found that older patients possessed a decreased ratio of CD8⁺CD62L⁺ non-effector cells in TIL compared to PBL, implying age-related increase of CD8⁺CD62L⁻ effector cells in TIL. The colocalization analysis of CD8 and CD3, however, suggested the suppressed activity of these effector cells in tumor microenvironment. These findings further elucidate the properties of TIL, showing an increasing difference between TIL and PBL with age, which may provide insight for the development of effective immunotherapies for cancer patients of different ages.