GEC-ESTRO APBI classification as a decision-making tool for the management of 2nd ipsilateral breast tumor event.

AIM: Second ipsilateral breast tumor event (2ndIBTE) occurring after primary radio-surgical treatment can be treated by either salvage mastectomy or 2nd conservative treatment (2ndCT) including an accelerated partial breast re-irradiation (APBrI). We analyzed the impact of the GEC-ESTRO APBI classification (GAC) on the oncological outcome after APBrI. MATERIALS AND METHODS: Between 2000 and 2016, 159 patients (pts) underwent a 2ndCT. After lumpectomy, APBrI was performed using either low-dose (30-55 Gy reference isodose) or high-dose rate brachytherapy (28-34 Gy). Oncological outcome including 3rdIBTE, regional (RFS) or metastasis-free survival (MFS), specific (SS) and overall survival (OS) was analyzed according to GAC. Univariate (UVA) and multivariate analyses (MVA) were conducted to identify significant prognostic factors for 3rdIBTE. RESULTS: With a median follow-up of 71 months (range 62-85 months), 60 pts (42%), 61 pts (42.7%) and 22 pts (15.4%) were classified as low-risk (LR), intermediate-risk (IR) and high-risk (HR), respectively. For the whole cohort, 6-year 3rdIBTE-free survival, RFS, MFS, SS and OS rates were 97.4, 96.4, 90.3, 92.9 and 91.2%, respectively. Six-year 3rdIBTE-free survival rates for LR, IR and HR were 100, 95.8 and 92.9%, respectively (p = 0.003), while no significant differences were found between the three GAC groups for RFS, MFS, SS. In UVA, lympho-vascular invasion (p = 0.009), positive margins (p = 0.0001) and GAC high-risk group (p = 0.001) were considered as significant prognostic factors for 3rdIBTE, while, in MVA, high-risk group (p = 0.009) was the only prognostic factor. CONCLUSION: In case of 2ndIBTE, GAC could be used as a decision helping tool to discuss conservative or radical treatment options. Patient information remains crucial in order to accurately define the salvage therapy modalities.

Nonredundant functions of Arabidopsis LecRK-V.2 and LecRK-VII.1 in controlling stomatal immunity and jasmonate-mediated stomatal closure.

Stomatal immunity restricts bacterial entry to leaves through the recognition of microbe-associated molecular patterns (MAMPs) by pattern-recognition receptors (PRRs) and downstream abscisic acid and salicylic acid signaling. Through a reverse genetics approach, we characterized the function of the L-type lectin receptor kinase-V.2 (LecRK-V.2) and -VII.1 (LecRK-VII.1). Analyses of interactions with the PRR FLAGELLIN SENSING2 (FLS2) were performed by co-immunoprecipitation and bimolecular fluorescence complementation and whole-cell patch-clamp analyses were used to evaluate guard cell Ca2+ -permeable cation channels. The Arabidopsis thaliana LecRK-V.2 and LecRK-VII.1 and notably their kinase activities were required for full activation of stomatal immunity. Knockout lecrk-V.2 and lecrk-VII.1 mutants were hyper-susceptible to Pseudomonas syringae infection and showed defective stomatal closure in response to bacteria or to the MAMPs flagellin and EF-Tu. By contrast, Arabidopsis over-expressing LecRK-V.2 or LecRK-VII.1 demonstrated a potentiated stomatal immunity. LecRK-V.2 and LecRK-VII.1 are shown to be part of the FLS2 PRR complex. In addition, LecRK-V.2 and LecRK-VII.1 were critical for methyl jasmonate (MeJA)-mediated stomatal closure, notably for MeJA-induced activation of guard cell Ca2+ -permeable cation channels. This study highlights the role of LecRK-V.2 and LecRK-VII.1 in stomatal immunity at the FLS2 PRR complex and in MeJA-mediated stomatal closure.

A high-throughput seed germination assay for root parasitic plants.

BACKGROUND: Some root-parasitic plants belonging to the Orobanche, Phelipanche or Striga genus represent one of the most destructive and intractable weed problems to agricultural production in both developed and developing countries. Compared with most of the other weeds, parasitic weeds are difficult to control by conventional methods because of their life style. The main difficulties that currently limit the development of successful control methods are the ability of the parasite to produce a tremendous number of tiny seeds that may remain viable in the soil for more than 15 years. Seed germination requires induction by stimulants present in root exudates of host plants. Researches performed on these minute seeds are until now tedious and time-consuming because germination rate is usually evaluated in Petri-dish by counting germinated seeds under a binocular microscope. RESULTS: We developed an easy and fast method for germination rate determination based on a standardized 96-well plate test coupled with spectrophotometric reading of tetrazolium salt (MTT) reduction. We adapted the Mosmann's protocol for cell cultures to germinating seeds and determined the conditions of seed stimulation and germination, MTT staining and formazan salt solubilization required to obtain a linear relationship between absorbance and germination rate. Dose-response analyses were presented as applications of interest for assessing half maximal effective or inhibitory concentrations of germination stimulants (strigolactones) or inhibitors (ABA), respectively, using four parameter logistic curves. CONCLUSION: The developed MTT system is simple and accurate. It yields reproducible results for germination bioassays of parasitic plant seeds. This method is adapted to high-throughput screenings of allelochemicals (stimulants, inhibitors) or biological extracts on parasitic plant seed germination, and strengthens the investigations of distinctive features of parasitic plant germination.

PrCYP707A1, an ABA catabolic gene, is a key component of Phelipanche ramosa seed germination in response to the strigolactone analogue GR24.

After a conditioning period, seed dormancy in obligate root parasitic plants is released by a chemical stimulus secreted by the roots of host plants. Using Phelipanche ramosa as the model, experiments conducted in this study showed that seeds require a conditioning period of at least 4 d to be receptive to the synthetic germination stimulant GR24. A cDNA-AFLP procedure on seeds revealed 58 transcript-derived fragments (TDFs) whose expression pattern changed upon GR24 treatment. Among the isolated TDFs, two up-regulated sequences corresponded to an abscisic acid (ABA) catabolic gene, PrCYP707A1, encoding an ABA 8'-hydroxylase. Using the rapid amplification of cDNA ends method, two full-length cDNAs, PrCYP707A1 and PrCYP707A2, were isolated from seeds. Both genes were always expressed at low levels during conditioning during which an initial decline in ABA levels was recorded. GR24 application after conditioning triggered a strong up-regulation of PrCYP707A1 during the first 18 h, followed by an 8-fold decrease in ABA levels detectable 3 d after treatment. In situ hybridization experiments on GR24-treated seeds revealed a specific PrCYP707A1 mRNA accumulation in the cells located between the embryo and the micropyle. Abz-E2B, a specific inhibitor of CYP707A enzymes, significantly impeded seed germination, proving to be a non-competitive antagonist of GR24 with reversible inhibitory activity. These results demonstrate that P. ramosa seed dormancy release relies on ABA catabolism mediated by the GR24-dependent activation of PrCYP707A1. In addition, in situ hybridization corroborates the putative location of cells receptive to the germination stimulants in seeds.