Oxygen tension affects terminal differentiation of corneal limbal epithelial cells

Oxygen concentration has been shown to be crucial in the proliferation and differentiation of various types of cells, while the impact of oxygen tension on the lineage commitment of epithelial cells remains elusive. In this study, we investigated the effect of hypoxia on the differentiation of corneal limbal epithelium using an ex vivo squamous metaplasia model. Under normoxic conditions when exposed to air, the hyperproliferation and abnormal epidermal-like differentiation of human corneal limbal epithelium was induced, whereas when exposed to air under hypoxic conditions, although we observed augmented proliferation, the abnormal differentiation was inhibited. The Notch signaling pathway was activated in hypoxic cultures, whereas the p38 MAPK signaling pathway was downregulated. The addition of Notch inhibitor under hypoxic conditions restored the activation of p38 MAPK and resulted in the recidivation of limbal epithelial cells to epidermal-like differentiation. Moreover, the epidermal-like differentiation of rabbit limbal epithelial cells was also blocked under hypoxic conditions in corneal epithelial cell sheets engineered ex vivo. We concluded that hypoxia can prevent abnormal differentiation while enhancing the proliferation of corneal limbal epithelial cells. Hypoxia coupled with air exposure can be used in the tissue engineering of corneal limbal epithelium.     

Retinoic acid as a vaccine adjuvant enhances CD8+ T cell response and mucosal protection from viral challenge

Vaccine-induced memory T cells localized at mucosal sites can provide rapid protection from viral infection. All-trans-retinoic acid (ATRA) has been shown to act physiologically to induce the expression of gut-homing receptors on lymphocytes. We tested whether the administration of exogenous ATRA during a systemic vaccination of mice could enhance the generation of mucosal CD8(+) T cell immunity, which might represent a strategy for establishing better protection from viral infection via mucosal routes. ATRA induced the expression of CCR9 and α4β7 on both mouse and human CD8(+) T cells activated in vitro. The administration of ATRA to mice during in vivo priming with a replication-defective recombinant adenovirus vector expressing the lymphocytic choriomeningitis virus glycoprotein (LCMVgp) (Ad5gp) increased numbers of both effector and memory T cells in intestinal mucosal tissues and showed higher frequencies of systemic central memory-like T cells that exhibited enhanced proliferation during boosting immunization with recombinant modified vaccinia virus Ankara expressing LCMVgp (MVAgp). Mice that received ATRA during Ad5gp vaccination were more resistant to intravaginal challenge by recombinant vaccinia virus expressing LCMVgp (VVgp), reflecting in part stronger T cell recall responses in situ. Thus, ATRA appears to be useful as an adjuvant during vaccination to increase memory T cell responses and protection from viral infection at mucosal sites and may facilitate the development of more effective vaccines against mucosally transmitted pathogens such as HIV.     

Luxury uptake of magnesium by peas,Pisum sativum

Increasing the magnesium (Mg) concentration of vegetables (biofortification) will often require ‘luxury’ uptake where the whole‐plant concentration of Mg (c_p) is greater than required for maximum yield. Our aim was to quantify some of the physiological factors influencing luxury uptake of Mg to aid subsequent development of agronomic techniques for biofortification. Peas, Pisum sativum, were used as a test species. A sand culture experiment related vegetative growth and c_p for plants grown with a range of Mg and potassium (K) supply rates. We developed a model of Mg uptake including feedback control exerted by c_p. The model was parameterised with results from a solution culture experiment and then used to explore ways to increase luxury uptake of Mg. Feedback control of Mg uptake by c_p was weak. Biomass did not increase if the Mg concentration exceeded 0.11% in the whole plant or 0.13% in the shoots. Values obtained in the field are often larger than this. Our results indicate that luxury uptake of Mg by peas is readily achieved, provided that there is ample supply of Mg²⁺ to the root surfaces. In field soils though, transport of Mg²⁺ to the roots may limit uptake and cation exchange processes restrict the ability of Mg fertilisers to substantially increase Mg uptake. Increasing root growth will usually increase Mg uptake, but c_p may not rise if biomass is also increased.     

Autocatalytic activation of a malarial egress protease is druggable and requires a protein cofactor

Malaria parasite egress from host erythrocytes (RBCs) is regulated by discharge of a parasite serine protease called SUB1 into the parasitophorous vacuole (PV). There, SUB1 activates a PV‐resident cysteine protease called SERA6, enabling host RBC rupture through SERA6‐mediated degradation of the RBC cytoskeleton protein β‐spectrin. Here, we show that the activation of Plasmodium falciparum SERA6 involves a second, autocatalytic step that is triggered by SUB1 cleavage. Unexpectedly, autoproteolytic maturation of SERA6 requires interaction in multimolecular complexes with a distinct PV‐located protein cofactor, MSA180, that is itself a SUB1 substrate. Genetic ablation of MSA180 mimics SERA6 disruption, producing a fatal block in β‐spectrin cleavage and RBC rupture. Drug‐like inhibitors of SERA6 autoprocessing similarly prevent β‐spectrin cleavage and egress in both P. falciparum and the emerging zoonotic pathogen P. knowlesi. Our results elucidate the egress pathway and identify SERA6 as a target for a new class of antimalarial drugs designed to prevent disease progression.     

甲肝病毒在人血白细胞中体外连续传代和在分类白细胞中增殖的研究

1985年我们采用间接免疫荧光法(IF法)检测出甲肝患者外周血白细胞中有甲肝抗原(HAAg)存在,继而又将HAAg阳性白细胞直接种入PLC/PRF/5细胞,分离到两株甲肝病毒(HAV)NJ—3株和H—1株。为了弄清白细胞所携带的病毒究竟仅为吸附吞饮,抑或能在其中复制增殖,我们将分离到的HAV用正常人血白细胞进行体外增殖试验,现将结果报告如下。     

DNA Topological Context Affects Access to Eukaryotic DNA Topoisomerase I

Abstract

We have analyzed the reactivity of a 217 base pair segment of the intrinsically curved Crithidia fasciculata kinetoplast DNA towards eukaryotic DNA topoisomerase I. The substrates were open [linear fragment and nicked circle] and closed minidomains [closed relaxed circle and circles with linking differences of ?1 and ?2], We interpreted the results with the aid of a model that was used to predict the structures of the topoisomers. The modelling shows that the ΔLk(?l) form is unusually compact because of the curvature in the DNA. To determine the role of sequence-directed curvature in both the experimental and modeling studies, controls were examined in which the curved Crithidia sequence was replaced by an uncurved sequence obtained from the plasmid pBR322.

Reactivity of the Crithidia DNA [as analyzed both by the cleavage and the topoisomerization reactions] markedly varied among the DNA forms: (i) the hierarchy of overall reactivity observed is: linear fragment > nicked circular, closed circular [ΔLk(O)], interwound [ΔLk(?2)] > bent interwound [ΔLk(?l)]; (ii) the intensity of several cleavage positions differs among DNA forms.

The results show that eukaryotic DNA topoisomerase I is very sensitive to the conformation of the substrates and that its reactivity is modulated by the variation of the compactness of the DNA molecule. The C. fasciculata sequence contains a highly curved segment that determines the conformation of the closed circle in a complex way.     

The 1973 WHO Classification Is More Suitable than the 2004 WHO Classification for Predicting Prognosis in Non-Muscle-Invasive Bladder Cancer

Background

Predicting the recurrence and progression of Non-muscle-invasive bladder cancer(NMIBC) is critical for urologist. Histological grade provides significant prognostic information, especially for prediction of progression. Currently, the 1973 and the 2004 WHO classification co-exist. Which system is better for predicting rumor recurrence and progression still a matter for debate.

Methodology/Principal Findings

348 patients diagnosed with Non-muscle invasive bladder cancer were enrolled in our retrospective study. Paraffin sections were assessed by an experienced urological pathologist according to both the 1973 and 2004 WHO classifications. Tumor recurrence and progression was followed-up in all patients. During follow-up, corresponding 5-year recurrence-free survival rates of G1, G2 and G3 were 82.1%, 55.9%, 32.1% and the 5-year progression-free survival rates were 95.9%, 84.4% and 43.3%, respectively. The 5-year recurrence-free survival rates of papillary urothelial neoplasm of low malignant potential (PUNLMP), low-grade papillary urothelial carcinoma(LGPUC) and high-grade papillary urothelial carcinoma (HGPUC) were 69.8%, 67.1% and 42.0% respectively and the 5-year progression-free survival rates were 100%, 90.9% and 54.8% respectively. In multivariate analysis, the 1973 WHO classification significantly associated with both tumor recurrence and progression(p = 0.010 and p = 0.022, respectively); the 2004 WHO classification correlated with tumor progression(p = 0.019), while was not proved to be a variable that can predict the risk of recurrence(p = 0.547). Kaplan-Meier plots showed that both the 1973 WHO and the 2004 WHO classifications were significantly associated with progression-free survival (p<0.0001, log-rank test). For prediction of recurrence, significant differences were observed between the tumor grades classified using the 1973 WHO grading system (p<0.0001, log-rank test), while a significant overlap was observed between PUNLMP and LG plots using the 2004 WHO grading system(p = 0.616, log-rank test).

Conclusion/Significance

Both the 1973 WHO and the 2004 WHO Classifications are effective in predicting tumor progression in Non-muscle invasive bladder cancer, while the 1973 WHO Classification is more suitable for predicting tumor recurrence.     

HepaCAM inhibits clear cell renal carcinoma 786-0 cell proliferation via blocking PKCε translocation from cytoplasm to plasma membrane

p62/sequestosome-1 is a multifunctional adapter protein implicated in selective autophagy, cell signaling pathways, and tumorigenesis, and plays an important role at the crossroad between autophagy and cancer. But, the connection between autophagy and cancer is complex and in some cases contradictory. Human colorectal cancer tissues from patients were analyzed for expression of p62 and Microtubule-associated protein light chain 3 (LC3, an autophagosome marker) using immunostaining, western blotting, real-time PCR, and confocal microscopy. To study the effects of p62 on autophagy and cell growth, shRNA for p62 was applied and cell growth curve was monitored in human colorectal cancer cell. In vivo experiments were done using the mouse xenograft model. We showed that up-regulated expression of p62 and LC3 in colorectal cancer tissues. We also demonstrated that specifically knockdown the expression of p62 showed significantly inhibitory effects not only on autophagy activation, but also on tumor growth both in vitro and xenograft tumors model. The ectopic overexpression of p62 and autophagy activation contributes to colorectal tumorigenesis. p62 and autophagy will be therapy targets for the treatment of colorectal cancer.