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Germination of pollen grains is a crucial step in plant reproduction. However, the molecular mechanisms involved remain unclear. We investigated the role of PECTIN METHYLESTERASE48 (PME48), an enzyme implicated in the remodeling of pectins in Arabidopsis (Arabidopsis thaliana) pollen. A combination of functional genomics, gene expression, in vivo and in vitro pollen germination, immunolabeling, and biochemical analyses was used on wild-type andAtpme48mutant plants. We showed thatAtPME48is specifically expressed in the male gametophyte and is the second most expressedPMEin dry and imbibed pollen grains. Pollen grains from homozygous mutant lines displayed a significant delay in imbibition and germination in vitro and in vivo. Moreover, numerous pollen grains showed two tips emerging instead of one in the wild type. Immunolabeling and Fourier transform infrared analyses showed that the degree of methylesterification of the homogalacturonan was higher inpme48−/− pollen grains. In contrast, the PME activity was lower inpme48−/−, partly due to a reduction of PME48 activity revealed by zymogram. Interestingly, the wild-type phenotype was restored inpme48−/− with the optimum germination medium supplemented with 2.5 mM calcium chloride, suggesting that in the wild-type pollen, the weakly methylesterified homogalacturonan is a source of Ca²⁺ necessary for pollen germination. Although pollen-specific PMEs are traditionally associated with pollen tube elongation, this study provides strong evidence that PME48 impacts the mechanical properties of the intine wall during maturation of the pollen grain, which, in turn, influences pollen grain germination.

Pectins are major components of primary cell wall that play a crucial role in plant development. After biosynthesis, pectins are secreted in the cell wall by Golgi-derived vesicles under a highly methylesterified form and are de-methylesterified by pectin methylesterases (PME). It is hypothesized that PME might be regulated by pectin methylesterase inhibitor (PMEI). In this paper, we show by isoelectric focalisation and subsequent zymogram that kiwi PMEI was able to inhibit Arabidopsis PME activity by forming a complex. The complexes were stable under a wide range of ionic strength and pH. Moreover, PMEI might be able to form a complex with basic PMEs including three PMEs strongly expressed in root and four PMEs expressed in pollen grains. Finally, exogenous treatment with kiwi PMEI was able to reduce the activity of cell wall resident PMEs with persistent effects such as an increase of the root growth and a dramatic effect on pollen tube stability.

Background/aimsTo describe the ocular manifestations associated with epidermolysis bullosa acquisita (EBA).MethodsThis retrospective study was conducted at a tertiary bullous disease clinic. Consecutive patients were enrolled with biopsy proven diagnosis of EBA, with ocular involvement and a follow-up of at least 36 months. A multidisciplinary team of dermatologists, ENT specialists and ophthalmologists evaluated all patients. Immunological workup included direct (including immune-electron microscopy) and indirect immunofluorescence. Ophthalmological examination included best-corrected visual acuity (BCVA) and slit-lamp examination with grading of conjunctival fibrosis using the Tauber classification.ResultsNine patients (five females, four males) were included. The mean age at diagnosis was 32 years (range 1–52 years). Follow-up ranged from 3 to 18 years (mean 10.7 years). Conjunctival fibrosis was present in all affected eyes and was stage III or greater in 60% of patients. Eight patients (14 eyes) had corneal involvement most frequently associated with trichiasis-associated mechanical irritation or extensive cicatrising conjunctivitis. Corneal lesions developed on three eyes of three patients without eyelid disease or severe fibrosis or any identifiable triggering factor. Eyelids were affected in six patients, with trichiasis being the most common feature (affecting three patients, four eyes). Corneal-related blindness occurred in at least one eye in 44% of the patients.ConclusionEBA may be associated with devastating ocular manifestations. Most patients develop severe cicatrising conjunctivitis. A subset of patients may present with isolated corneal lesions. Further studies are warranted to assess the effects of systemic treatments on the evolution of ocular manifestations.

The pollen tube is a fast tip-growing cell carrying the two sperm cells to the ovule allowing the double fertilization process and seed setting. To succeed in this process, the spatial and temporal controls of pollen tube growth within the female organ are critical. It requires a massive cell wall deposition to promote fast pollen tube elongation and a tight control of the cell wall remodeling to modify the mechanical properties. In addition, during its journey, the pollen tube interacts with the pistil, which plays key roles in pollen tube nutrition, guidance and in the rejection of the self-incompatible pollen. This review focuses on our current knowledge in the biochemistry and localization of the main cell wall polymers including pectin, hemicellulose, cellulose and callose from several pollen tube species. Moreover, based on transcriptomic data and functional genomic studies, the possible enzymes involved in the cell wall remodeling during pollen tube growth and their impact on the cell wall mechanics are also described. Finally, mutant analyses have permitted to gain insight in the function of several genes involved in the pollen tube cell wall biosynthesis and their roles in pollen tube growth are further discussed.

Increasing evidences demonstrate the role of sensory innervation in bone metabolism, remodeling and repair, however neurovascular coupling in bone is rarely studied. Using microfluidic devices as an indirect co-culture model to mimic in vitro the physiological scenario of innervation, our group demonstrated that sensory neurons (SNs) were able to regulate the extracellular matrix remodeling by endothelial cells (ECs), in particular through sensory neuropeptides, i.e. calcitonin gene-related peptide (CGRP) and substance P (SP). Nonetheless, still little is known about the cell signaling pathways and mechanism of action in neurovascular coupling. Here, in order to characterize the communication between SNs and ECs at molecular level, we evaluated the effect of SNs and the neuropeptides CGRP and SP on ECs. We focused on different pathways known to play a role on endothelial functions: calcium signaling, p38 and Erk1/2; the control of signal propagation through Cx43; and endothelial functions through the production of nitric oxide (NO). The effect of SNs was evaluated on ECs Ca 2+ influx, the expression of Cx43, endothelial nitric oxide synthase (eNOS) and nitric oxide (NO) production, p38, ERK1/2 as well as their phosphorylated forms. In addition, the role of CGRP and SP were either analyzed using respective antagonists in the co-culture model, or by adding directly on the ECs monocultures. We show that capsaicin-stimulated SNs induce increased Ca 2+ influx in ECs. SNs stimulate the increase of NO production in ECs, probably involving a decrease in the inhibitory eNOS T495 phosphorylation site. The neuropeptide CGRP, produced by SNs, seems to be one of the mediators of this effect in ECs since NO production is decreased in the presence of CGRP antagonist in the co-culture of ECs and SNs, and increased when ECs are stimulated with synthetic CGRP. Taken together, our results suggest that SNs play an important role in the control of the endothelial cell functions through CGRP production and NO signaling pathway.

Protein-protein interactions (PPIs) may represent one of the next major classes of therapeutic targets. So far, only a minute fraction of the estimated 650,000 PPIs that comprise the human interactome are known with a tiny number of complexes being drugged. Such intricate biological systems cannot be cost-efficiently tackled using conventional high-throughput screening methods. Rather, time has come for designing new strategies that will maximize the chance for hit identification through a rationalization of the PPI inhibitor chemical space and the design of PPI-focused compound libraries (global or target-specific). Here, we train machine-learning-based models, mainly decision trees, using a dataset of known PPI inhibitors and of regular drugs in order to determine a global physico-chemical profile for putative PPI inhibitors. This statistical analysis unravels two important molecular descriptors for PPI inhibitors characterizing specific molecular shapes and the presence of a privileged number of aromatic bonds. The best model has been transposed into a computer program, PPI-HitProfiler, that can output from any drug-like compound collection a focused chemical library enriched in putative PPI inhibitors. Our PPI inhibitor profiler is challenged on the experimental screening results of 11 different PPIs among which the p53/MDM2 interaction screened within our own CDithem platform, that in addition to the validation of our concept led to the identification of 4 novel p53/MDM2 inhibitors. Collectively, our tool shows a robust behavior on the 11 experimental datasets by correctly profiling 70% of the experimentally identified hits while removing 52% of the inactive compounds from the initial compound collections. We strongly believe that this new tool can be used as a global PPI inhibitor profiler prior to screening assays to reduce the size of the compound collections to be experimentally screened while keeping most of the true PPI inhibitors. PPI-HitProfiler is freely available on request from our CDithem platform website, www.CDithem.com.

How pathogenic bacteria infect and kill their host is currently widely investigated. In comparison, the fate of pathogens after the death of their host receives less attention. We studied Bacillus thuringiensis (Bt) infection of an insect host, and show that NprR, a quorum sensor, is active after death of the insect and allows Bt to survive in the cadavers as vegetative cells. Transcriptomic analysis revealed that NprR regulates at least 41 genes, including many encoding degradative enzymes or proteins involved in the synthesis of a nonribosomal peptide named kurstakin. These degradative enzymes are essential in vitro to degrade several substrates and are specifically expressed after host death suggesting that Bt has an active necrotrophic lifestyle in the cadaver. We show that kurstakin is essential for Bt survival during necrotrophic development. It is required for swarming mobility and biofilm formation, presumably through a pore forming activity. A nprR deficient mutant does not develop necrotrophically and does not sporulate efficiently in the cadaver. We report that necrotrophism is a highly regulated mechanism essential for the Bt infectious cycle, contributing to spore spreading.

Background Non-metastatic breast cancer treatment is mainly based on surgery, with or without chemotherapy, radiotherapy and/or hormone therapy. To reduce the risk of hormone receptor positive (HR+) disease recurrence, hormone therapy is prescribed for at least 5 years. It may induce adverse drug reactions (ADRs) as joint pain, sexual dysfunction, weight increase, fatigue, mood disorders and vasomotor symptoms. Around 30–40% of patients withhold hormone therapy within 5 years after initiation. Based on encouraging results of mobile health in patient follow-up, we developed a web-application addressed for breast cancer patients initiating adjuvant hormonal therapy and aimed to assess its impact on hormone therapy adherence, ADRs management, and health-related quality of life. Methods The WEBAPPAC trial is a randomized, open-label, prospective, single-center phase 3 study aiming to assess the interest of a web-application support as compared to standard management among breast cancer patients initiating hormone therapy. The main endpoint is the proportion of patients with hormone therapy adherence failure within 18 months after treatment start, in each arm. Eligible patients will be 1:1 randomized between the WEBAPPAC web-application support (experimental arm,) or standard support (control arm), with stratification on type of hormone therapy (Aromatase inhibitor or Tamoxifen). We plan to enroll 438 patients overall. Failure to hormone therapy will be assessed using the Morisky 8-item self-questionnaire (MMSA8), patient adherence logbook, and medical consultations. Secondary outcomes include hormone therapy adherence at 6 months, pain (Visual Analogue Scale and Brief Pain Inventory), quality of life (EORTC QLQ-C30 and BR23 self-questionnaires), anxiety and depression (Hospital and Depression Scale), and return to work and/or daily activities. The user experience with the WEBAPPAC web-application will be assessed using the System Usability Scale (SUS) questionnaire. Discussion Hormone therapy discontinuation or adherence failure in breast cancer patients may be indirectly related to an increased risk of recurrence. A better control of medication adherence, through the detection of side effects and some proposed actions trying to reduce them, appears therefore essential to limit the risk of disease recurrence. The WEBAPPAC web-application thus aims better monitoring and allowing higher level of responsiveness in case of ADRs, thus improving treatment adherence. Trial registration NCT04554927, registered September 18, 2020. Protocol version Version 2.1 dated from December 21, 2021.

We aimed to compare AS03-adjuvanted inactivated trivalent influenza vaccine (TIV) with non-adjuvanted TIV for seasonal influenza prevention in elderly people. We did a randomised trial in 15 countries worldwide during the 2008–09 (year 1) and 2009–10 (year 2) influenza seasons. Eligible participants aged at least 65 years who were not in hospital or bedridden and were without acute illness were randomly assigned (1:1) to receive either AS03-adjuvanted TIV or non-adjuvanted TIV. Randomisation was done in an internet-based system, with a blocking scheme and stratification by age (65–74 years and 75 years or older). Participants were scheduled to receive one vaccine in each year, and remained in the same group in years 1 and 2. Unmasked personnel prepared and gave the vaccines, but participants and individuals assessing any study endpoint were masked. The coprimary objectives were to assess the relative efficacy of the vaccines and lot-to-lot consistency of the AS03-adjuvanted TIV (to be reported elsewhere). For the first objective, the primary endpoint was relative efficacy of the vaccines for prevention of influenza A (excluding A H1N1 pdm09) or B, or both, that was confirmed by PCR analysis in year 1 (lower limit of two-sided 95% CI had to be greater than zero to establish superiority). From Nov 15, to April 30, in both years, participants were monitored by telephone or site contact and home visits every week or 2 weeks to identify cases of influenza-like illness. After onset of suspected cases, we obtained nasal and throat swabs to identify influenza RNA with real-time PCR. Efficacy analyses were done per protocol. This trial is registered with ClinicalTrials.gov, number NCT00753272. We enrolled 43 802 participants, of whom 21 893 were assigned to and received the AS03-adjuvanted TIV and 21 802 the non-adjuvanted TIV in year 1. In the year 1 efficacy cohort, fewer participants given AS03-adjuvanted than non-adjuvanted TIV were infected with influenza A or B, or both (274 [1·27%, 95% CI 1·12–1·43] of 21 573 vs 310 [1·44%, 1·29–1·61] of 21 482; relative efficacy 12·11%, 95% CI −3·40 to 25·29; superiority not established). Fewer participants in the year 1 efficacy cohort given AS03-adjuvanted TIV than non-adjuvanted TIV were infected with influenza A (224 [1·04%, 95% CI 0·91–1·18] vs 270 [1·26, 1·11–1·41]; relative efficacy 17·53%, 95% CI 1·55–30·92) and influenza A H3N2 (170 [0·79, 0·67–0·92] vs 205 [0·95, 0·83–1·09]; post-hoc analysis relative efficacy 22·0%, 95% CI 5·68–35·49). AS03-adjuvanted TIV has a higher efficacy for prevention of some subtypes of influenza than does a non-adjuvanted TIV. Future influenza vaccine studies in elderly people should be based on subtype or lineage-specific endpoints. GlaxoSmithKline Biologicals SA.

Background Martinique shares with the other Caribbean countries specific public health issues, particularly in the diagnostic and therapeutic management of cancer patients. Mutualization of human and material resources by promoting cooperation is the most appropriate response to the challenges of the health systems of the Caribbean territories. Through the French PRPH-3 program, we propose to set up a collaborative digital platform adapted to the specificities of the Caribbean to strengthen professional links and skills in oncofertility and oncosexology and reduce inequalities in access to reproductive and sexual health care for cancer patients. Methods Within the context of this program, we have developed of an open-source platform based on a Learning Content Management System (LCMS), with an operating system developed by UNFM for low speed internet. LO libraries have been created and interaction between trainers and learners were done in asynchronous mode. This training management platform is based on: a TCC learning system (Training, Coaching, Communities); a web-hosting with pedagogical engineering appropriate to low bandwidth; a reporting system and a responsibility for processing. Results We have carried out a flexible, multilingual and accessible digital learning strategy functionality called e-MCPPO according to low-speed internet ecosystem. In close connection with the e-learning strategy we conceived (i) a multidisciplinary team; (ii) an appropriate training program for expert health professionals and (iii) a responsive design. Discussion and Conclusion This low-speed web-based infrastructure allows communities of experts to cooperate in creating, validating, publishing and managing academic learning content. The self-learning modules provide the digital layer for each learner to extend their skills. Learners, as well as trainers, would gradually take ownership of this platform and encourage its promotion. Innovation in this context is both technological (low-speed Internet broadcasting, free interactive software) and organizational (moderating educational resources). This collaborative digital platform is unique in its form and content. This challenge could contribute to the digital transformation of the Caribbean ecosystem for capacity building in this specifics topics.