Matrix metalloproteinases (MMP) are important regulators of tumor progression and angiogenesis. MMPs generate both proangiogenic and antiangiogenic fragments, such as vascular endothelial growth ...factor and endostatin. The in vivo activation of MMPs and endostatin generation occur mainly in the extracellular environment by interactions of different cell types. Therefore, these processes are necessary to study in the extracellular space in vivo. Sex steroids play a dominant role in breast carcinogenesis, by largely unknown mechanisms. In the present study, we used in vivo microdialysis to directly quantify MMP-2 and MMP-9 activity and sample endostatin from both stroma (murine) and tumor (human) cells in vivo in solid MCF-7 tumors in nude mice. We found that tamoxifen in combination with estradiol increased tumor MMP-2/MMP-9 in vivo activity, endostatin levels, and decreased tumor vascularization compared with estradiol treatment only. The stroma-derived endostatin was three to five times higher than cancer cell-generated endostatin. After inhibition of MMP-2/MMP-9, endostatin levels decreased, providing evidence that these proteases are highly involved in the generation of endostatin. Our results support the previously reported concept that MMPs may serve as negative regulators of angiogenesis. The regulation of endostatin generation by modulation of MMP-2/MMP-9 activities suggests a previously unrecognized mechanism of estradiol and tamoxifen, which may have implications for the pathogenesis of breast cancer.
The genetic diversity and population structure of breeding germplasm is central knowledge for crop improvement. To gain insight into the genetic potential of the germplasm used for potato breeding in ...a Nordic breeding program as well as all available accessions from the Nordic genebank (NordGen), 133 potato genotypes were genotyped using the Infinium Illumina 20 K SNP array. After SNP filtering, 11 610 polymorphic SNPs were included in the analysis. In addition, data from three important breeding traits - percent dry matter and uniformity of tuber shape and eye - were scored to measure the variation potato cultivars and breeding clones. The genetic diversity among the genotypes was estimated using principal coordinate analysis based on the genetic distance between individuals, as well as by using the software STRUCTURE. Both methods suggest that the collected breeding material and the germplasm from the gene-bank are closely related, with a low degree of population structure between the groups. The phenotypic distribution among the genotypes revealed significant differences, especially between farmer's cultivars and released cultivars and breeding clones. The percent heterozygosity was similar between the groups, with a mean average of 58-60%. Overall, the breeding germplasm and the accessions from the Nordic genebank seems to be closely related with similar genetic background. The genetic potential of available Nordic potato breeding germplasm is low, and for genetic hybridization purposes, genotypes from outside the Nordic region should be employed.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Skin (contact) allergy, the most prevalent form of immunotoxicity in humans, is caused by low molecular weight chemicals (haptens) that penetrate stratum corneum and modify endogenous proteins. The ...fate of haptens after cutaneous absorption, especially what protein(s) they react with, is largely unknown. In this study the fluorescent hapten tetramethylrhodamine isothiocyanate (TRITC) was used to identify hapten-protein conjugates in the local lymph nodes after topical application, as they play a key role in activation of the adaptive immune system. TRITC interacted with dendritic cells but also with T and B cells in the lymph nodes as shown by flow cytometry. Identification of the most abundant TRITC-modified protein in lymph nodes by tandem mass spectrometry revealed TRITC-modification of the N-terminal proline of macrophage migration inhibitory factor (MIF) - an evolutionary well-conserved protein involved in cell-mediated immunity and inflammation. This is the first time a hapten-modified protein has been identified in lymph nodes after topical administration of the hapten. Most haptens are electrophiles and can therefore modify the N-terminal proline of MIF, which has an unusually reactive amino group under physiological conditions; thus, modification of MIF by haptens may have an immunomodulating role in contact allergy as well as in other immunotoxicity reactions.
Summary
Background
Citronellol is a frequently used fragrance compound in consumer products. It is present in fragrance mix II, which is used for screening of contact allergy to fragrances. Because ...of its chemical structure, citronellol could be susceptible to autoxidation.
Objectives
To compare the behaviour of citronellol with that of the structurally similar compounds linalool and geraniol, in terms of ability to autoxidize, the products formed, and the sensitization potencies of these.
Methods
Citronellol was exposed to air, and autoxidation was followed by gas chromatography–mass spectrometry (GC–MS) analysis after derivatization of thermolabile compounds. The sensitizing potencies of the oxidation mixture and its major oxidation compounds were examined with the local lymph node assay.
Results
The concentration of citronellol decreased while the sensitization potency increased in air‐exposed samples over time, with hydroperoxides being identified as the major oxidation products and main skin sensitizers.
Conclusions
The present study shows the impact of the absence of the 2,3‐double bond in the citronellol structure on the oxidation pathways for formation of oxidation products. The study also shows the usefulness of our new GC–MS method for quantification of the citronellol oxidation products, especially the hydroperoxides. The investigated citronellol hydroperoxides could be important allergens, owing to the high concentrations detected and frequent exposure to citronellol in the population.
A simple and sensitive method was developed using hydrophilic interaction liquid chromatography coupled to tandem mass spectrometry for determination of monosaccharides liberated from marine ...polysaccharides by acidic hydrolysis. Optimal separation of diastereomeric monosaccharides including hexoses, pentoses, and deoxyhexoses was achieved using an aminopropyl bonded column with mobile phase containing ternary solvents (acetonitrile/methanol/water) in conjunction with MS/MS in SRM mode. Mechanisms for fragmentation of deprotonated monosaccharides with regard to cross-ring cleavage were proposed. Matrix effects from coeluting interferences were observed and isotopic-labeled internal standard was used to compensate for the signal suppression. The method demonstrated excellent instrumental limits of detection (LOD), ranging from 0.7 to 4.2 pg. Method LODs range from 0.9 to 5.1 nM. The proposed method was applied to the analysis of polysaccharides in seawater collected from the open leads of the central Arctic Ocean in the summer of 2008.
The breeding of new cultivars is a powerful approach to increase both the quantity and quality of potato harvest per land unit. The aim of this research was to determine using multi-site testing the ...progress made by the genetic enhancement of potato in Sweden in the last 1.5 decades by comparing advanced breeding clones (T
4
upwards) bred in Sweden (Svensk potatisförädling hereafter) versus available released cultivars in Europe and grown in its Nordic Region. The multi-site testing results show that potato breeding based in Scandinavia offers to the growers of the Nordic Region of Europe cultivars for prevailing farming environments and end-user needs rather than relying, as happens today in the market, on foreign cultivars. These cultivars bred elsewhere are not always very suitable for the challenging Nordic agroecosystems, as shown by the results of the multi-site testing herein. Such an approach on relying on foreign cultivars may be advocated for not funding potato breeding in, and for Fennoscandia by those ignoring the results shown by this research.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
To determine the safety, tolerability, and efficacy of combination gefitinib and methotrexate to treat ectopic pregnancy.
We performed a phase I, single-arm (nonrandomized), open-label study. Twelve ...women with ectopic pregnancies were administered methotrexate (50 mg/m, intramuscular) and 250 mg oral gefitinib in a dose-escalation protocol: one dose (day 1) n=3; three doses (days 1-3) n=3; seven doses (days 1-7) n=6. Efficacy was examined by comparing human chorionic gonadotrophin (hCG) decline and time to resolution with historic controls administered methotrexate only.
Common side effects were transient acneiform rash in 67% (8/12) and diarrhea in 42% (5/12) of participants. There was no clinical or biochemical evidence of serious pulmonary, renal, hepatic, or hematologic toxicity. Of six participants with a pretreatment serum hCG level between 1,000 and 3,000 international units/L, hCG levels declined significantly faster than in the control group. Median serum hCG levels by day 7 after treatment were less than one fifth of levels observed among 71 historic controls treated with methotrexate alone (median interquartile range hCG in participants 261 55-1,445 international units/L compared with controls 1,426 940-2,573; P=.008). Median time for the ectopic pregnancies to resolve with combination therapy was 34% shorter compared with methotrexate alone (21 days compared with 32 days; P=.018).
Combination gefitinib and methotrexate has potential as a treatment for ectopic pregnancy but is commonly associated with minor side effects such as transient rash and diarrhea. The treatment requires validation of safety and efficacy in a larger trial.
Australian New Zealand Clinical Trials Registry, www.anzctr.org, AC'TRN12610000684022.
: II.
A novel set of protein descriptors has been developed to increase the understanding of protein behavior on chromatographic media. The protein descriptors are pH-dependent and based on electrostatic ...and hydrophobic properties of mainly the surface of the proteins as revealed by their three-dimensional structure. Interpretable and predictive quantitative structure property relationship (QSPR) models were then obtained for protein retention in ion exchange chromatography at different pH values. In most cases the calculated average surface potential could be directly related to retention times. Moreover, the high retention of human lactoferrin observed in cation exchange even at high pH values could be modeled by adding descriptors of the charge asymmetry.
Short-interfering RNAs (siRNAs) have engendered much enthusiasm for their ability to silence the expression of specific genes. However, it is now well established that siRNAs, depending on their ...sequence, can be variably sensed by the innate immune system through recruitment of toll-like receptors 7 and 8 (TLR7/8). Here, we aimed to identify sequence-based modifications allowing for the design of bifunctional siRNAs with both proinflammatory and specific silencing activities, and with potentially increased therapeutic benefits. We found that the introduction of a micro-RNA (miRNA)-like nonpairing uridine-bulge in the passenger strand robustly increased immunostimulatory activity on human immune cells. This sequence modification had no effect on the silencing efficiency of the siRNA. Increased immunostimulation with the uridine-bulge design was specific to human cells, and conserved silencing efficiency required a Dicer-substrate scaffold. The increased cytokine production with the uridine-bulge design resulted in enhanced protection against Semliki Forest virus (SFV) infection, in viral assays. Thus, we characterize a design scaffold applicable to any given siRNA sequence, that results in increased innate immune activation without affecting gene silencing. Our data suggest that this sequence modification coupled with structural modification differentially recruits human TLR8 over TLR7, and could have potential application in antiviral therapies.
Placenta has the highest expression of epidermal growth factor (EGF) receptor of all tissues, a cell signaling pathway promoting survival and growth. Therefore, EGF receptor inhibition could ...potentially treat ectopic pregnancy. We undertook preclinical studies to examine whether gefitinib (orally available EGF receptor inhibitor) with or without methotrexate inhibits placental cell growth.
Gefitinib and methotrexate were added to placental cells and their ability inhibit cell growth, block EGF receptor signaling, and induce apoptosis (programmed cell death) was examined. They were also administered to two animal mouse models to examine their effects on placental tissue in vivo.
Epidermal growth factor receptor was highly expressed in placental tissue from ectopic pregnancies. Combining gefitinib with methotrexate potently inhibited growth of placental cells, including placental cell lines (JEG3, BeWo cells) and cells isolated from first-trimester placenta. These drugs were additive in blocking EGF receptor signaling and inducing apoptosis. Gefitinib and methotrexate administered together were more potent in decreasing the volume of human placental cells xenografted subcutaneously onto mice compared with either alone. By day 19 after xenografting, mean (± standard error of the mean), xenograft volumes were: 821 (± 68) mm after gefitinib treatment, 901 (± 204) mm after methotrexate treatment, and 345 (±137) mm after both drugs were given (P<.01 for both comparisons of single therapy compared with combination therapy). Combining these agents doubled rates of fetal resorption in pregnant mice compared with each drug alone.
Combining gefitinib with methotrexate potently inhibits placental cell growth in vitro and in mouse models. The combination may have potential in treating ectopic pregnancies.
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