Proton irradiation suppresses angiogenic genes and impairs cell invasion and tumor growth

The energy deposition characteristics of proton radiation have attracted considerable attention in light of its implications for carcinogenesis risk in space travel, as well for application to cancer treatment. In space, it is the principle component of the galactic cosmic radiation to which astronauts will be exposed. For treatment, an increasing number of proton facilities are being established to exploit the physical advantages of this radiation type. However, the possibility that there may also be biologically based advantages to proton exposure has not been considered in either context. We demonstrate here that high-energy proton irradiation can inhibit expression of major pro-angiogenic factors and multiple angiogenesis-associated processes, including invasion and endothelial cell proliferation, which is prominent in cancer progression. Dose-dependent suppression of angiogenic signaling was demonstrated for both cancer and nontransformed cells. Pan-genomic microarray analysis and RT-PCR revealed that post-irradiation (0.5, 1.0 and 2.0 Gy), critical pro-angiogenic signaling factors including: vascular endothelial growth factor (VEGF), interleukin 6 and 8 (IL-6, IL-8) and hypoxia-inducible factor-1 alpha (HIF-1A), were significantly downregulated. Co-culture studies demonstrated that endothelial cell proliferation and invasion were inhibited by culturing with irradiated cancer or fibroblast cells, which suggests that proton irradiation may, in addition to direct action, contribute to angiogenesis suppression through modulation of paracrine signalings from targeted cells. Addition of recombinant IL-8 or VEGF partially restored these functions in vitro, while in vivo, an attenuated tumor growth rate was demonstrated for proton-irradiated human lung cancer cells. Taken together, these findings provide novel pre-clinical evidence that proton irradiation may, in addition to its physical targeting advantages, have important biological ramifications that should be a consideration in the optimization of proton therapy.     

Apicomplexa genes involved in the host cell invasion: the Cpa135 protein family

The availability of a bulk of genomic data of Apicomplexa parasites is a unique opportunity to identify groups of related proteins that are characteristic of this phylum. The Cpa135 protein of Cryptosporidium parvum is expressed and secreted through the apical complex at the invasive stage of sporozoite. This protein is characterised by an LCCL domain, a common trait of various secreted proteins within Apicomplexa. Using the Cpa135 as a "virtual template", we have identified Cpa135 orthologous genes in four apicomplexan species (Plasmodium falciparum, Theileria parva, Toxoplasma gondii and Eimeria tenella). In addition, the architecture of the deduced proteins shows that the Cpa135-related proteins are a distinct family among the apicomplexan LCCL proteins.     

肿瘤细胞侵袭研究进展

肿瘤细胞侵袭和转移是癌医学和癌生物学最重要的难题,癌症主要因其肿瘤细胞的侵袭和转移而成为致命的疾病,虽然侵袭和转移的机制仍不清楚,但肿瘤细胞侵袭一直是研究热点,本文就近年来对肿瘤细胞侵袭研究的新进展进行综述,以期为寻找治疗肿瘤的新方案提供参考.     

Local proteolytic activity in tumor cell invasion and metastasis

Proteolytic cleavage of extracellular matrix (ECM) is a critical regulator of many physiological and pathological events. It affects fundamental processes such as cell growth, differentiation, apoptosis and migration. Most proteases are produced as inactive proenzymes that undergo proteolytic cleavage for activation. Proteolytic activity is additionally modified by endogenous inhibitors. Mechanisms that localize and concentrate protease activity in the pericellular microenvironment of cells are prerequisites for processes like angiogenesis, bone development, inflammation and tumor cell invasion. Methods that enable real-time, high-resolution imaging and precise quantification of local proteolytic activity in vitro and in vivo remain major challenges. These methods will play an important role in the understanding of basic principles e.g. in cancer cell invasion, the identification of new therapeutical targets and hence drug design. This review highlights mechanisms and functions of local proteolytic activity with special emphasis on tumor cell invasion and metastasis, and focuses on techniques for the investigation of this process.     

Dominant versus recessive behavior of a cold- and a heat-sensitive mammalian cell cycle variant in heterokaryons

A heat-sensitive (hs, arrested at 39.5 degrees C, termed 21-Ta) and a cold-sensitive (cs, arrested at 33 degrees C, termed 21-Fb) clonal cell cycle variant were isolated from the same clone of the P-815 murine mastocytoma line. At the respective nonpermissive temperatures, both the hs and the cs variant were reversibly arrested in G1 phase, and numbers of cells forming colonies upon reincubation at the permissive temperature remained nearly constant for at least 6 days. Cells arrested in G1 by incubation at the respective nonpermissive temperatures were fused to cells of another P-815 clone (31-S) that had been arrested by serum deprivation. Upon reincubation in medium containing 10% serum for 48 h at 39.5 degrees C, 21-Ta x 31-S heterokaryons, similar to 31-S x 31-S homokaryons, entered the S phase, whereas at 33 degrees C, 21-Fb x 31-S heterokaryons, similar to 21-Fb x 21-Fb homokaryons, remained arrested in G1, indicating a recessive expression of the hs and a dominant expression of the cs phenotype.     

S-Allylcysteine reduces breast tumor cell adhesion and invasion

Previous studies show that aqueous garlic extract and its derivatives (e.g. S-allylcysteine [SAC]) prevent carcinogen-induced breast tumorigenesis. However, investigations testing the effect of SAC on later stages of breast tumorigenesis and/or metastasis have produced mixed results. Here we show that SAC significantly reduced anchorage-dependent and -independent growth of MDA-MB-231 breast tumor cells in a dose- and time-dependent fashion, and sub-lethal SAC-treatment altered mammary tumor cell adhesion and invasion through components of the extracellular matrix. We provide evidence to suggest increased expression of E-cadherin and reduced MMP-2 expression and activity are partially responsible for inhibition of mammary tumor cell invasion by SAC. Because E-cadherin and MMP-2 are important in cancer metastasis, these results suggest a link between SAC induction of E-cadherin and reduction of MMP2 activity with the inhibition of cell motility and invasion; thus providing evidence that events leading to breast cancer metastasis are repressed by sub-lethal SAC-treatment.     

Screening of genes involved in cell migration in Dictyostelium

A single cell of wild-type Dictyostelium discoideum forms a visible colony on a plastic dish in several days, but due to enhanced cell migration, amiB-null mutant cells scatter over a large area and do not form noticeable colonies. Here, with an aim to identify genes involved in cell migration, we isolated suppresser mutants of amiB-null mutants that restore the ability to form colonies. From REMI (restriction enzyme-mediated integration)-mutagenized pool of double-mutants, we identified 18 responsible genes from them. These genes can be categorized into several biological processes. One cell line, Sab16 (Suppressor of amiB) was chosen for further analysis, which had a disrupted phospholipase D pldB gene. To confirm the role of pldB gene in cell migration, we knocked out the pldB gene and over-expressed gfp-pldB in wild-type cells. GFP-PLDB localized to plasma membrane and on vesicles, and in migrating cells, at the protruding regions of pseudopodia. Migration speed of vegetative pldB-null cells was reduced to 73% of that of the wild-type. These results suggest that PLDB plays an important role in migration in Dictyostelium cells, and that our screening system is useful for the identification of genes involved in cell migration.     

Dominant and recessive informational suppressors of a missense mutation in Coprinus

Summary Twenty-one suppressor gene mutations which suppress the met-5.1 missense mutation of Coprinus were separated into six groups (A-F) on the basis of dominance or recessiveness, linkage to the met-5 locus, comlementation in heterozygous cells and growth behaviour. The actual number of suppressor loci could not be determined because crosses between suppressed mutants were inviable. The allele specificity of group A, C, D and F suppressors was confirmed by appropriate crosses. Group B and E suppressors were not tested because of close linkage to the met-5 locus. No evidence for functional suppression of met-5 mutations was obtained thus it is likely that all the suppressors cause translational corelation of met-5.1. Suppressors in four groups (C-F) have properties expected of tRNA structural gene mutations: the group C mutation is dominant, the other mutations are recessive but do not complement in heterozygous cells. The relative efficiencies of the tRNA species involved was assessed by comparing the degree to which the different sup ⁺ mutations depressed the growth rate on methionine supplemented medium. The dominant mutation depressed growth to the greatest extent and is, therefore, the most efficient suppressor. The least efficient suppressors did not depress growth at all. When growth was compared on minimal medium it was found that the more efficient the suppressor the less well it restored growth. The mutations in groups A and B depressed growth more than the tRNA mutations but affect some other component in translation because they are recessive and complement normally. It is suggested that they may act to alter tRNA modifying enzymes.     

ArfGAP family proteins in cell adhesion, migration and tumor invasion

The identification of several ArfGAP proteins as binding partners of paxillin, an integrin signaling and scaffolding protein, has suggested the existence of molecular links between integrin functions and intracellular traffic, as proposed by MS Bretscher long ago. Among the paxillin-binding ArfGAPs, AMAP1 has recently been strongly implicated in tumor invasion as well as malignancy, owing to its highly augmented expression in tumors and its direct involvement in invasive activities. Another ArfGAP, Git2, was found to be a component of the Gbetagamma-mediated directional sensing machinery, while simultaneously playing an essential role in the suppressive control of superoxide production, which is mediated by vesicle transport in GPCR-stimulated neutrophils. These emerging molecular mechanisms may further delineate key processes regulating intracellular traffic as principal controls of cell motility and invasive activities.