SLC27A2 regulates miR-411 to affect chemo-resistance in ovarian cancer.

Although platinum-based chemotherapies have long been used as standard treatment in ovarian cancer, cisplatin resistance is a major problem that restricts its use. Herein, we investigate the biological function of SLC27A2 and its underlying mechanisms in regulating chemo-resistance in ovarian cancer. The findings show that SLC27A2 down-regulation in primary ovarian cancer tissues correlates with chemo-resistance and poor patient survival in our patient cohort. Significantly, we demonstrate that up-regulation of SLC27A2 by lentivirus-mediated p-SLC27A2 sensitizes ovarian cancer cells to cisplatin in vitro and in vivo via apoptosis. Mechanistic investigation reveals that miR-411 is the most strikingly over-expressed gene in response to ectopic expression of SLC27A2, but under-expressed in recurrent ovarian cancer tissues. Lower miR-411 expression contributes to ovarian cancer chemo-resistance in vitro and in vivo. Furthermore, SLC27A2 directly binds specific sites in the miR-411 promoter region and promoter activity decreases after mutation of putative SLC27A2-binding sites. This indicates that SLC27A2 is required for the transcriptional induction of miR-411. The luciferase assays also confirm that miR-411 directly targets ABCG2 in ovarian cancer, and overall findings establish the SLC27A2-miR-411-ABCG2 pathway in the regulation of ovarian cancer chemo-resistance with potential therapeutic applications.

Isolation and characterization of the Chrysanthemum nitrate transporter CmNRT1.

In this study, the nitrate transporter gene CmNRT1 was isolated from the chrysanthemum variety 'Nannongxuefeng'. The full-length cDNA contains an open reading frame of 1761 bp encoding 587 residues. Using qRT-PCR, we found that CmNRT1 was induced by 10 mM NO3(-) in roots and shoots. Two Arabidopsis thaliana transgenic plants expressing CmNRT1 were selected for functional analyses. Root (15)N influx in wild-type and transgenic A. thaliana lines under 10 or 0.2 mM (15)NO3 was tested. Our results indicate that CmNRT1 encodes a constitutive component for a low-affinity transporter.

Salicylic acid-induced changes in physiological parameters and genes of the flavonoid biosynthesis pathway in Artemisia vulgaris and Dendranthema nankingense during aphid feeding.

Phloem-feeding aphids cause serious damage to plants. The mechanisms of plant-aphid interactions are only partially understood and involve multiple pathways, including phytohormones. In order to investigate whether salicylic acid (SA) is involved and how it plays a part in the defense response to the aphid Macrosiphoniella sanbourni, physiological changes and gene expression profiles in response to aphid inoculation with or without SA pretreatment were compared between the aphid-resistant Artemisia vulgaris 'Variegata' and the susceptible chrysanthemum, Dendranthema nankingense. Changes in levels of reactive oxygen species, malondialdehyde (MDA), and flavonoids, and in the expression of genes involved in flavonoid biosynthesis, including PAL (phenylalanine ammonia-lyase), CHS (chalcone synthase), CHI (chalcone isomerase), F3H (flavanone 3-hydroxylase), F3'H (flavanone 3'-hydroxylase), and DFR (dihydroflavonol reductase), were investigated. Levels of hydrogen peroxide, superoxide anions, MDA, and flavonoids, and their related gene expression, increased after aphid infestation and SA pretreatment followed by aphid infestation; the aphid-resistant A. vulgaris exhibited a more rapid response than the aphid-susceptible D. nankingense to SA treatment and aphid infestation. Taken together, our results suggest that SA could be used to increase aphid resistance in the chrysanthemum.

Synergistic effects of glycated chitosan with high-intensity focused ultrasound on suppression of metastases in a syngeneic breast tumor model.

Stimulation of the host immune system is crucial in cancer treatment. In particular, nonspecific immunotherapies, when combined with other traditional therapies such as radiation and chemotherapy, may induce immunity against primary and metastatic tumors. In this study, we demonstrate that a novel, non-toxic immunoadjuvant, glycated chitosan (GC), decreases the motility and invasion of mammalian breast cancer cells in vitro and in vivo. Lung metastatic ratios were reduced in 4T1 tumor-bearing mice when intratumoral GC injection was combined with local high-intensity focused ultrasound (HIFU) treatment. We postulate that this treatment modality stimulates the host immune system to combat cancer cells, as macrophage accumulation in tumor lesions was detected after GC-HIFU treatment. In addition, plasma collected from GC-HIFU-treated tumor-bearing mice exhibited tumor-specific cytotoxicity. We also investigated the effect of GC on epithelial-mesenchymal transition-related markers. Our results showed that GC decreased the expression of Twist-1 and Slug, proto-oncogenes commonly implicated in metastasis. Epithelial-cadherin, which is regulated by these genes, was also upregulated. Taken together, our current data suggest that GC alone can reduce cancer cell motility and invasion, whereas GC-HIFU treatment can induce immune responses to suppress tumor metastasis in vivo.

Biodistribution of phenylboric acid derivative entrapped lipiodol and 4-borono-2-18F-fluoro-L-phenylalanine-fructose in GP7TB liver tumor bearing rats for BNCT.

A new phenylboric acid derivative entrapped lipiodol (PBAD-lipiodol) was developed as a boron carrier for the boron neutron capture therapy (BNCT) of hepatoma in Taiwan. The biodistribution of both PBAD-lipiodol and BPA-fructose was assayed in GP7TB hepatoma-bearing rat model. The highest uptake of PBAD-lipiodol was found at 2h post injection. The application of BNCT for the hepatoma treatment in tumor-bearing rats is suggested to be 2-4h post PBAD-lipiodol injection.

A simple model for quantification of the radiobiological effectiveness of the 10B(n,alpha)7Li capture reaction in BNCT.

A simple model has been developed for predicting radiobiological effectiveness of the neutron capture reaction in boron neutron capture therapy. This model was derived from the relationship between the cell survival from the boron capture reaction, the intracellular boron concentration, and the thermal neutron fluence. We found that the cell-killing effect of the boron capture reaction was well described using a power function of the intracellular boron concentration. Hence the relationship between cell survival from the boron capture reaction, intracellular boron concentration, and the thermal neutron fluence could be determined using a simple mathematical equation. We consider that our current approach is more appropriate and realistic than the conventional theoretical mathematical model used to estimate the radiobiological effectiveness of the neutron capture reaction in boron neutron capture therapy.

Efficacy of Re-188-labelled sulphur colloid on prolongation of survival time in melanoma-bearing animals.

UNLABELLED: In this study, the effectiveness of a 188Re labeled sulfur colloid with two particle size ranges was used to evaluate the effectiveness of this agent on melanoma tumors in mice in terms of animal lifespan. METHODS: Two separate group of animals were used for investigating biodistribution and survival time. A total of 188 B16F10-melanoma-bearing BDF(1) mice were injected intraperitoneally with 3.7 MBq (0.1mCi)/2mL of radiolabeled sulfur colloid ten days after intraperitoneal inoculation of 5x10(5) B16F10 melanoma cells/2ml. For group 1, 30 mice were sacrificed at 1, 4, 24, 48 and 72 hours for biodistribution studies. In group 2, 158 mice were divided into 9 groups (n=16 approximately 18/groups)each receiving respectively tumor alone, tumor with normal saline, cold colloid or hot colloid with 16, 23, 31, 46, 62, or 124 MBq activity. Each of these colloid groups was further divided into two groups, one receiving smaller particle sizes (<3 microm:80.4 +/-7.2%, colloid 1) and the other receiving larger particle sizes (<3 microm:12.3+/-1.0%, colloid 2). The animals were checked daily until death and their survival recorded. RESULTS: Colloid 2 showed higher accumulation in almost all tissues, the highest accumulation organ was tumor ( approximately 40%), then spleen ( approximately 20%), stomach ( approximately 15%), diaphragm ( approximately 3%), and liver ( approximately 2%). There was a significant increase in survival time with increasing amount of the larger-particle-size colloid. Administered levels of 16-31 MBq/mouse were most efficacious and with higher amounts the survival times decreased significantly below that of the controls. There was a significant difference in the dose-response curves for the two preparations. Protection factors (1/Relative-risk) of nearly 5 were achieved using the larger colloid size, and nearly 30 using the smaller colloid size. An amount of 16-31 MBq of the colloid 2 was the optimal activity in these studies. On the one hand, the survival data agreed well with the biodistribution data, where higher accumulation was found in tumor with colloid 2. CONCLUSION: Rhenium-188 offers on-site availability, medium half-life, higher beta-particle energy of 2.12 MeV for therapy and emission of 155keV gamma photon suitable for imaging. The present study demonstrated that 188Re-sulfur colloid is an effective agent in controlling tumor cells in the abdominal cavity in animals.

Preclinical evaluation of locoregional delivery of radiolabeled iododeoxyuridine and thymidylate synthase inhibitor in a hepatoma model.

UNLABELLED: We report improved incorporation of the radiolabeled-thymidine analog [125I/131I]5-iodo-2'-deoxyuridine ([125I/131I]IdUrd) into DNA by the addition of Thymitaq, a thymidylate synthase inhibitor, as a strategy of molecular radiotherapy for hepatoma treatment. METHODS: The synergistic effect of combination [125I]IdUrd and Thymitaq in clonogenic survival and DNA incorporation was shown on the human hepatoma cell line Hep3B. Radiobiodistribution of intrahepatic arterially injected [125I]IdUrd and Thymitaq was studied in a rat N1S1 hepatoma model. In vivo therapeutic effects of locoregional delivery of both drugs were evaluated in mouse subcutaneous hepatoma and ascitic hepatoma models. RESULTS: In a clonogenic assay, Thymitaq showed a synergistic effect with [125I]IdUrd but not cold IdUrd. Thymitaq had a dose-dependent modulation effect on DNA-[125I]IdUrd incorporation. The biodistribution study indicated a slower clearance rate of [125I]IdUdR in the hepatoma as well as an initially higher uptake of [125I]IdUrd into DNA when the [125I]IdUrd was combined with Thymitaq. In vivo studies showed a superior therapeutic effect of combination Thymitaq and [125I]IdUrd in both subcutaneous and ascites tumor models, but the combination of [131I]IdUrd and [125I]IdUrd may be more effective than Auger electron emitters alone for the treatment of subcutaneous tumor. CONCLUSION: The strategy of locoregional delivery of [125I/131I]IdUrd to a tumor site through an intrahepatic arterial, intratumoral, or intraperitoneal route in combination with Thymitaq is promising and may also have a favorable therapeutic index in vivo.

Sensitization of a tumor, but not normal tissue, to the cytotoxic effect of ionizing radiation using Panax notoginseng extract.

The aim of the present study was to investigate any sensitization effect of the Panax notoginseng extract (PNE) and the purified Saponin (Rb1) on the radiation response of an experimental tumor (KHT sarcoma) in mice, in comparison with any effects on a normal tissue (bone marrow). PNE at a concentration of 0.1-100 mg/kg produced an increase in tumor radiosensitivity. The sensitization effect was maximal at 10 mg/kg and at 30 minutes after injection. Higher doses were toxic to the bone marrow stem cells. Similarly Rb1 at a concentration 0.001 to 1 mg/kg also produced an increase in tumor radiosensitivity, with maximum effect at 1 mg/kg. Higher doses were not toxic to the bone marrow stem cells in this case. Radiosensitization factors were calculated as ratios of D0 (the radiosensitivity parameter), and these were highly significant for the tumor and very similar for both compounds at the doses used, namely 1.18-1.19. There was no significant effect for bone marrow stem cells (sensitization factors of 0.99 +/- 0.01 for both compounds). The differential effect on tumor, and the magnitude of the radiosensitization, suggest that further purified or synthetic versions of this extract may be useful not only in vascular-related diseases but also in cancer therapy.

Differential gene expression in a DNA double-strand-break repair mutant XRS-5 defective in Ku80: analysis by cDNA microarray.

The ability of cells to rejoin DNA double-strand breaks (DSBs) usually correlates with their radiosensitivity. This correlation has been demonstrated in radiosensitive cells, including the Chinese hamster ovary mutant XRS-5. XRS-5 is defective in a DNA end-binding protein, Ku80, which is a component of a DNA-dependent protein kinase complex used for joining strand breaks. However, Ku80-deficient cells are known to be retarded in cell proliferation and growth as well as other yet to be identified defects. Using custom-made 600-gene cDNA microarray filters, we found differential gene expressions between the wild-type and XRS-5 cells. Defective Ku80 apparently affects the expression of several repair genes, including topoisomerase-I and -IIA, ERCC5, MLH1, and ATM. In contrast, other DNA repair-associated genes, such as GADD45A, EGR1 MDM2 and p53, were not affected. In addition, for large numbers of growth-associated genes, such as cyclins and clks, the growth factors and cytokines were also affected. Down-regulated expression was also found in several categories of seemingly unrelated genes, including apoptosis, angiogenesis, kinase and signaling, phosphatase, stress protein, proto-oncogenes and tumor suppressors, transcription and translation factors. A RT-PCR analysis confirmed that the XRS-5 cells used were defective in Ku80 expression. The diversified groups of genes being affected could mean that Ku80, a multi-functional DNA-binding protein, not only affects DNA repair, but is also involved in transcription regulation. Our data, taken together, indicate that there are specific genes being modulated in Ku80- deficient cells, and that some of the DNA repair pathways and other biological functions are apparently linked, suggesting that a defect in one gene could have global effects on many other processes.

Butachlor, a suspected carcinogen, alters growth and transformation characteristics of mouse liver cells.

Butachlor is a widely used herbicide in Asia and South America. Previous investigations have indicated that it is a suspected carcinogen. To understand more about the biological effects of butachlor on cultured cells and the mechanism(s) of its carcinogenicity, we studied the alteration of the growth characteristics that was induced by butachlor in normal mouse liver cells (BNL CL2). This study demonstrates that butachlor decreases the population-doubling time of BNL CL2 cells, suggesting that it stimulates cell proliferation. To support this finding, a thymidine incorporation assay was conducted and a similar result that butachlor stimulates cell proliferation was elucidated. In addition, we show that butachlor increases the saturation density of the BNL CL2 cells. When combined with the tumor initiator N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), butachlor transforms cells efficiently, as demonstrated by loss of contact inhibition. These findings indicate that butachlor alters the growth characteristics of BNL CL2 cells and suggest that butachlor may induce malignant transformation through stimulation of cell proliferation, alteration of cell cycle regulation, and suppression of cell density-dependent inhibition of proliferation.

Chromosomal damage in long-term residents of houses contaminated with cobalt-60.

Chromosomal translocations in people who have lived in houses contaminated with radiation were substantially raised compared with controls. Retrospective biological dosimetry indicated cumulative exposures less than 1.0 Gy, which were lower than values derived from physical measurements.

Preliminary study of dose equivalent evaluation for residents in radioactivity contaminated rebar buildings.

It has recently been found that several resident and office buildings in Taiwan were constructed with 60Co-contaminated reinforcing steel bar (rebar). Both governmental officials and the residents of such buildings have been concerned about this finding. In order to respond to the situation, the government has adopted a number of remedial measures, including full-scale radiation survey, dose evaluation and physical examinations of residents. This article presents three methods for evaluating the dose equivalents of the residents living in the contaminated rebar buildings by means of gamma-ray survey, necklace-type thermoluminescence dosimeters (TLDs) and the human lymphocyte chromosome aberration analyses. The results reveal that the dose evaluation by gamma-ray survey is rather conservative. Generally for the residents whose annual dose equivalents are greater than 5 mSv (0.5 rem) by gamma-ray survey, the dose equivalents from necklace-type TLDs are only within the range of 20 to 50% of the evaluated values mentioned above. For chromosome analyses, at least 500 lymphocyte cells were scored and analyzed for each resident. Most of the chromosome analysis data show that the dose equivalents received by residents are lower than the detection limit of the method (100 mSv) and quite different from the estimated dose obtained from either gamma-ray survey or necklace-type TLD measurements.

Assessment of dose and risk to the body following conventional and spiral computed tomography.

BACKGROUND: Computed tomography (CT) is one of the most frequently used examination procedures in diagnostic radiology and the dose given to the patients is higher than in general radiographic procedures. In this study LiF chip thermoluminescent dosimeters (TLD-100) were placed in each relative organ or tissue position, including head, chest and abdomen, in a Rando phantom. CT was performed using both conventional and spiral modes, and effective dose and effective dose equivalent were assessed for each organ or tissue scanned. METHODS: The TLD reader used in this experiment was controlled at a nitrogen flow rate of 450 ml/min, preheat time of 14 seconds, reading time of 16 seconds and annealing time of 16 seconds. This CT scanner can be used to perform both conventional and spiral tomography. Operating conditions for spiral tomography were 120 kV, 80 mA for scout film, and 120 kV, 200 mA, 1 sec/slice for each scanning. However, for conventional tomography, the operating conditions were 120 kV, 80 mA for scout film and 120 kV, 160 mA, 1.5 sec/slice for each scanning. These operating conditions are satisfactory to most clinical applications, and therefore were adopted for the present studies. RESULTS: Results showed that, in both effective dose and effective dose and effective dose equivalent, conventional tomography was higher than spiral tomography. The average effective doses for each part were measured to be 1.89 and 4.95 mSv for the head, 30.01 and 40.65 mSv for the chest, and 12.85 and 19.62 mSv for the abdomen of spiral and conventional CT, respectively. Higher carcinogenic risk was assessed in organs such as liver, lung, stomach and bone marrow, other organs had a relatively lower incidence of risk. CONCLUSIONS: The main purpose of this study was to obtain distribution values of effective dose and effective dose equivalent, and to know the probability of carcinogenic effect upon each organ or tissue after CT scanning. Results showed the average effective dose for spiral CT to be less than conventional CT, and the dose in the body surface was generally lower than the dose in the central region.

Effects of field size on the survival of pig epidermal colony-forming in situ after electron irradiation.

The survival of colony-forming cells in pig epidermis after irradiation was measured using different electron field sizes. The sensitivity of the colony-forming cells was characterized by D(o) = 2.5-3.0 Gy. There was an effect of field size, described approximately by: Dose (to give five colonies/cm2) (Gy) = (31 +/- 2) x Area-(0.048 +/- 0.015). The effect of field size was less than found previously using tolerance to skin reactions (area exponent = 0.16). This work indicates for the first time that the effects of different large field sizes in skin can be detected at the level of colony-forming cell survival.

Development of a novel Bacillus subtilis cloning system employing its neutral protease as screen marker.

Part of the pUC19 polylinker sequence (33 bp) was inserted into the pro-peptide-coding region of the Bacillus subtilis neutral protease-encoding gene to replace a 93-bp FspI-HindIII fragment. This in-frame sequence replacement had little effect on the expression and secretion of the neutral protease. This plasmid can therefore be used as a cloning vector, and recombinant clones can be directly identified on skim milk indicator plates by the loss of a clear ring (or halo) around the colonies. This novel cloning system offers several advantages over existing B. subtilis cloning vectors: (i) convenient direct screening of recombinants; (ii) the use of inexpensive indicator; (iii) no restriction on the use of host strains; and (iv) the availability of seven frequently used unique cloning sites: BamHI, XbaI, SalI, PstI, SphI, HindIII, and EcoRI. This system also has the potential to be used as an expression/secretion vector.

Residual skin injury after repeated irradiation: differences observed using healing, macrocolony, and microcolony endpoints.

Following three repeated tolerance doses to mouse tail skin, residual injury was characterized by a 35% reduction in the iso-effective dose compared to age-matched controls, using healing or macrocolony endpoints. In contrast, the reduction was only 9%, measured using microcolony formation. The colony data showed that the reduction was a constant dose, not a dose-modifying effect. The residual injury is interpreted as due to a reduced density of microcolony-forming cells in the epidermis, and these are less capable of macrocolony formation and hence of re-epithelialization in the repeatedly-irradiated epidermis.

Re-irradiation of mouse skin: similarity of dose reductions for healing and macrocolony endpoints.

The amount of residual injury in mouse tail skin, assessed by the decrease in re-irradiation dose for equal effect, was similar whether assessed using healing or colony endpoints (17-21% after single priming doses). There were tendencies towards an increased sensitivity of the colony-forming cells by a factor of about 2, and less residual injury after multifractionated priming doses. These observations are compatible with a lower alpha/beta ratio characterising the response to dose fractionation for residual injury than for the acute healing response.

The radiosensitivity of microcolony- and macrocolony-forming cells in mouse tail epidermis.

A new microcolony technique is described for measuring the survival of colony-forming cells in mouse tail epidermis. The survival curve is characterised by D0 = 2.70 +/- 0.12 Gy. The number of microcolonies per cm2 is similar to the number of macrocolonies after high doses, which shows for the first time that all microcolonies (greater than or equal to 32 cells) in epidermis develop into macrocolonies. At low doses the number of macrocolonies underestimates the number of colony-forming cells because of coalescence of microcolonies to form macrocolonies. This results in a lower apparent sensitivity of macrocolony-forming cells by a factor of about 1.5. About 3% of basal cells in tail epidermis appear to be capable of colony formation.