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The ultimate aim of fluorescence microscopy is to achieve high-resolution imaging of increasingly larger biological samples. Extended depth of field presents a potential solution to accelerate imaging of large samples when compression of information along the optical axis is not detrimental to the interpretation of images. We have implemented an extended depth of field (EDF) approach in a random illumination microscope (RIM). RIM uses multiple speckled illuminations and variance data processing to double the resolution. It is particularly adapted to the imaging of thick samples as it does not require the knowledge of illumination patterns. We demonstrate highly-resolved projective images of biological tissues and cells. Compared to a sequential scan of the imaged volume with conventional 2D-RIM, EDF-RIM allows an order of magnitude improvement in speed and light dose reduction, with comparable resolution. As the axial information is lost in an EDF modality, we propose a method to retrieve the sample topography for samples that are organized in cell sheets., (© 2024. The Author(s).)

Secondary lymphedema (LD) corresponds to a severe lymphatic dysfunction leading to the accumulation of fluid and fibrotic adipose tissue in a limb. Here, we identified apelin (APLN) as a powerful molecule for regenerating lymphatic function in LD. We identified the loss of APLN expression in the lymphedematous arm compared to the normal arm in patients. The role of APLN in LD was confirmed in APLN knockout mice, in which LD is increased and associated with fibrosis and dermal backflow. This was reversed by intradermal injection of APLN-lentivectors. Mechanistically, APLN stimulates lymphatic endothelial cell gene expression and induces the binding of E2F8 transcription factor to the promoter of CCBE1 that controls VEGF-C processing. In addition, APLN induces Akt and eNOS pathways to stimulate lymphatic collector pumping. Our results show that APLN represents a novel partner for VEGF-C to restore lymphatic function in both initial and collecting vessels. As LD appears after cancer treatment, we validated the APLN-VEGF-C combination using a novel class of nonintegrative RNA delivery LentiFlash® vector that will be evaluated for phase I/IIa clinical trial., (© 2024. The Author(s).)

p57 Kip2 is a cyclin/CDK inhibitor and a negative regulator of cell proliferation. Here, we report that p57 regulates intestinal stem cell (ISC) fate and proliferation in a CDK-independent manner during intestinal development. In the absence of p57, intestinal crypts exhibit an increased proliferation and an amplification of transit-amplifying cells and of Hopx + ISCs, which are no longer quiescent, while Lgr5 + ISCs are unaffected. RNA sequencing (RNA-seq) analyses of Hopx + ISCs show major gene expression changes in the absence of p57. We found that p57 binds to and inhibits the activity of Ascl2, a transcription factor critical for ISC specification and maintenance, by participating in the recruitment of a corepressor complex to Ascl2 target gene promoters. Thus, our data suggest that, during intestinal development, p57 plays a key role in maintaining Hopx + ISC quiescence and repressing the ISC phenotype outside of the crypt bottom by inhibiting the transcription factor Ascl2 in a CDK-independent manner., Competing Interests: Declaration of interests The authors declare no competing interests., (Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved.)