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Papillary and reticular dermis show distinct extracellular matrix (ECM) and vascularization corresponding to their specific functions. These characteristics are associated with gene expression patterns of fibroblasts freshly isolated from their native microenvironment. In order to assess the relevance of these fibroblast subpopulations in a tissue engineering context, we investigated their contribution to matrix production and vascularization using cell sheet culture conditions. We first performed RNA-seq differential expression analysis to determine whether several rounds of cell amplification and high-density culture affected their gene expression profile. Bioinformatics analysis revealed that expression of angiogenesis-related and matrisome gene signatures were maintained, resulting in papillary and reticular ECMs that differ in composition and structure. The impact of secreted or ECM-associated factors was then assessed using two independent 3D angiogenesis assays: -1/ a fibrin hydrogel-based assay allowing investigation of diffusible secreted factors, -2/ a scaffold-free cell-sheet based assay for investigation of fibroblast-produced microenvironment. These analyses revealed that papillary fibroblasts secrete highly angiogenic factors and produce a microenvironment characterised by ECM remodelling capacity and dense and branched microvascular network, whereas reticular fibroblasts produced more structural core components of the ECM associated with less branched and larger vessels. These features mimick the characteristics of both the ECM and the vasculature of dermis subcompartments. In addition to showing that skin fibroblast populations differentially regulate angiogenesis via both secreted and ECM factors, our work emphasizes the importance of papillary and reticular fibroblasts for engineering and modelling dermis microenvironment and vascularization. STATEMENT OF SIGNIFICANCE: Recent advances have brought to the forefront the central role of microenvironment and vascularization in tissue engineering for regenerative medicine and microtissue modelling. We have investigated the role of papillary and reticular fibroblast subpopulations using scaffold-free cell sheet culture. This approach provides differentiated cells conditions allowing the production of their own microenvironment. Analysis of gene expression profiles and characterisation of the matrix produced revealed strong and specific angiogenic properties that we functionally characterized using 3D angiogenesis models targeting the respective role of either secreted or matrix-bound factors. This study demonstrates the importance of cell-generated extracellular matrix and questions the importance of cell source and the relevance of hydrogels for developing physio-pathologically relevant tissue engineered substitutes., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Adèle Mauroux, Laëtitia Marchand, Sylvie Bordes, and Brigitte Closs are employees of SILAB. Florence Ruggiero and Laurent Muller declare the receipt of a grant from SILAB. PJ, NBJ, HB, BG, SH, CAR, AL, PM, CM and SG state no conflict of interest, (Copyright © 2023 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

The ultrasonographic assessment of the caudal vena cava-to-aorta ratio (CVC:Ao) appears to be a promising method for early recognition of alterations of intravascular volume status in veterinary medicine. The primary objective of this study was to establish the reference intervals of the CVC:Ao ratio with ultrasound in nonsedated healthy dogs. Secondary objectives were to determine the influence of the respiratory cycle and to evaluate correlations between ultrasonographic measurements, signalment and physical exam findings. Ultrasonographic measurements of Ao and CVC diameters were successfully obtained for all sixty dogs included. No evidence of a difference was observed between the measurements of Ao and CVC diameters, and CVC:Ao ratio between inspiration and expiration (P = .373, P = .318, and P = .537, respectively). The reference interval for CVC:Ao ratio (95% CI), generated from US measurements performed at any moment of the respiratory cycle was defined as 0.93 (0.91-0.95) -1.32 (1.30-1.34). The CVC:Ao ratio was significantly negatively correlated with age (r = -0.341, P = .008) and positively correlated with respiratory rate (r = 0.423, P < .001), but not with heart rate (P = .573) or arterial systolic blood pressure (P = .166). A low inter- and intraoperator variability in repeated measurements was observed for each operator and between operators. The ultrasonographic measurement of the CVC:Ao ratio appears as a simple method with low inter- and intraoperator variability using the ultrasonographic protocol described in the current study. With the reference interval established in the present study in healthy nonsedated dogs, further studies should evaluate the utility of this simple method in assessing and monitoring volume status in hypo- and hypervolemic dogs., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

NRF2 is a master regulator of the antioxidative response that was recently proposed as a potential regulator of extracellular matrix (ECM) gene expression. Fibroblasts are major ECM producers in all connective tissues, including the dermis. A better understanding of NRF2-mediated ECM regulation in skin fibroblasts is thus of great interest for skin homeostasis maintenance and aging protection. In this study, we investigate the impact of NRF2 downregulation on matrisome gene expression and ECM deposits in human primary dermal fibroblasts. RNA-sequencing‒based transcriptome analysis of NRF2 silenced dermal fibroblasts shows that ECM genes are the most regulated gene sets, highlighting the relevance of the NRF2-mediated matrisome program in these cells. Using complementary light and electron microscopy methods, we show that NRF2 deprivation in dermal fibroblasts results in reduced collagen I biosynthesis and impacts collagen fibril deposition. Moreover, we identify ZNF469, a putative transcriptional regulator of collagen biosynthesis, as a target of NRF2. Both ZNF469 silenced fibroblasts and fibroblasts derived from Brittle Corneal Syndrome patients carrying variants in ZNF469 gene show reduced collagen I gene expression. Our study shows that NRF2 orchestrates matrisome expression in human skin fibroblasts through direct or indirect transcriptional mechanisms that could be prioritized to target dermal ECM homeostasis in health and disease., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Most former industrial sites are contaminated by mixtures of trace elements and organic pollutants. Levels of pollutants do not provide information regarding their biological impact, bioavailability and possible interactions between substances. There is genuine interest in combining chemical analyses with biological investigations. We studied a brownfield where several industrial activities were carried out starting in the 1970s, (incineration of pyralene transformers, recovery of copper by burning cables in the open air). Four representative plots showing different levels of polychlorobiphenyls were selected. Organic and trace metal levels were measured together with soil pedological characteristics. The bacterial community structure and functional diversity were assessed by 16S metagenomics with deep sequencing and community-level physiological profiling. Additionally, a vegetation survey was performed. Polychlorobiphenyls (8 mg.kg -1 to 1500 mg.kg -1 ) were from 2.4 × 10 3 -fold to 6 × 10 5 -fold higher than the European background level of 2.5 μg.kg -1 . Polychlorinated dibenzo-p-dioxins and dibenzofurans ranged from 0.5 to 8.0 μg.kg -1 . The soil was also contaminated with trace metals, i.e., Cu > 187, Zn > 217 and Pb > 372 mg.kg -1 . Location within the study area, trace metal content and soil humidity were stronger determinants than organic pollutants of bacterial community structures and activities. Thus, the highest biological activity and the greatest bacteriological richness were observed in the plot that was less contaminated with trace metals, despite the high level of organic pollutants in the plot. Moreover, trace element pollution was associated with a relatively low presence of Actinobacteria and Rhizobia. The plot with the highest metal contamination was rich in metal-resistant bacteria such as Sphingomonadales, Geodermatophilaceae and KD4-96 (Chloroflexi phylum). Acidobacteria and Sphingomonadales, capable of resisting trace metals and degrading persistent organic pollutants, were dominant in the plots that had accumulated metal and organic contamination, but bacterial activity was lower in these plots than in the other plots., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

How some animals regenerate missing body parts is not well understood. Taking advantage of the zebrafish caudal fin model, we performed a global unbiased time-course transcriptomic analysis of fin regeneration. Biostatistics analyses identified extracellular matrix (ECM) as the most enriched gene sets. Basement membranes (BMs) are specialized ECM structures that provide tissues with structural cohesion and serve as a major extracellular signaling platform. While the embryonic formation of BM has been extensively investigated, its regeneration in adults remains poorly studied. We therefore focused on BM gene expression kinetics and showed that it recapitulates many aspects of development. As such, the re-expression of the embryonic col14a1a gene indicated that col14a1a is part of the regeneration-specific program. We showed that laminins and col14a1a genes display similar kinetics and that the corresponding proteins are spatially and temporally controlled during regeneration. Analysis of our CRISPR/Cas9-mediated col14a1a knockout fish showed that collagen XIV-A contributes to timely deposition of laminins. As changes in ECM organization can affect tissue mechanical properties, we analyzed the biomechanics of col14a1a -/- regenerative BM using atomic force microscopy (AFM). Our data revealed a thinner BM accompanied by a substantial increase of the stiffness when compared to controls. Further AFM 3D-reconstructions showed that BM is organized as a checkerboard made of alternation of soft and rigid regions that is compromised in mutants leading to a more compact structure. We conclude that collagen XIV-A transiently acts as a molecular spacer responsible for BM structure and biomechanics possibly by helping laminins integration within regenerative BM., (Copyright © 2018 International Society of Matrix Biology. Published by Elsevier B.V. All rights reserved.)

In the past years, the fruit fly Drosophila melanogaster has been extensively used to study the relationship between animals and their associated microbes. Compared to the one of wild populations, the microbiota of laboratory-reared flies is less diverse, and comprises fewer bacterial taxa; nevertheless, the main commensal bacteria found in fly microbiota always belong to the Acetobacteraceae and Lactobacillaceae families. The bacterial communities associated with the fly are environmentally acquired, and the partners engage in a perpetual re-association process. Adult flies constantly ingest and excrete microbes from and onto their feeding substrate, which are then transmitted to the next generation developing within this shared habitat. We wanted to analyze the potential changes in the bacterial community during its reciprocal transfer between the two compartments of the niche (i.e. the fly and the diet). To address this question, we used a diverse, wild-derived microbial community and analyzed its relationship with the fly population and the nutritive substrate in a given habitat. Here we show that the community was overall well maintained upon transmission to a new niche, to a new fly population and to their progeny, illustrating the stable association of a Drosophila-derived microbiota with its fly partner and the nutritional environment. These results highlight the preponderant role of the nutritional substrate in the dynamics of Drosophila/microbiota interactions, and the need to fully integrate this variable when performing such studies., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

Brominated flame retardants are suspected to act as disruptors of thyroid hormone signaling. This raises the concern that they might affect children's cognitive functions by influencing thyroid hormone signaling in the developing brain. We present here an in vitro analysis of the ability of the most common compounds, tetrabromobisphenol A (TBBPA) and BDE-209, to alter thyroid hormone response based on a model neural cell line and genome-wide analysis of gene expression., (Copyright © 2014. Published by Elsevier Ireland Ltd.)

(13)C spectroscopy combined with the injection of (13)C-labeled substrates is a powerful method for the study of brain metabolism in vivo. Since highly localized measurements are required in a heterogeneous organ such as the brain, it is of interest to augment the sensitivity of (13)C spectroscopy by proton acquisition. Furthermore, as focal cerebral lesions are often encountered in animal models of disorders in which the two brain hemispheres are compared, we wished to develop a bi-voxel localized sequence for the simultaneous bilateral investigation of rat brain metabolism, with no need for external additional references. Two sequences were developed at 9.4T: a bi-voxel (1)H-((13)C) STEAM-POCE (Proton Observed Carbon Edited) sequence and a bi-voxel (1)H-((13)C) PRESS-POCE adiabatically decoupled sequence with Hadamard encoding. Hadamard encoding allows both voxels to be recorded simultaneously, with the same acquisition time as that required for a single voxel. The method was validated in a biological investigation into the neuronal damage and the effect on the Tri Carboxylic Acid cycle in localized excitotoxic lesions. Following an excitotoxic quinolinate-induced localized lesion in the rat cortex and the infusion of U-(13)C glucose, two (1)H-((13)C) spectra of distinct (4x4x4mm(3)) voxels, one centred on the injured hemisphere and the other on the contralateral hemisphere, were recorded simultaneously. Two (1)H bi-voxel spectra were also recorded and showed a significant decrease in N-acetyl aspartate, and an accumulation of lactate in the ipsilateral hemisphere. The (1)H-((13)C) spectra could be recorded dynamically as a function of time, and showed a fall in the glutamate/glutamine ratio and the presence of a stable glutamine pool, with a permanent increase of lactate in the ipsilateral hemisphere. This bi-voxel (1)H-((13)C) method can be used to investigate simultaneously both brain hemispheres, and to perform dynamic studies. We report here the neuronal damage and the effect on the Tri Carboxylic Acid cycle in localized excitotoxic lesions.

The combination of localized 2D 1H MR correlation spectroscopy and Hadamard encoding allows the simultaneous acquisition of multiple volumes of interest without an increase in the experimental duration, compared to single-voxel acquisition. In the present study, 2D correlation spectra were acquired simultaneously within 20 to 40 min in two voxels located in each hemisphere of the rat brain. An intervoxel distance of 20% of the voxel size was sufficient to limit spatial contamination. The following cerebral metabolites gave detectable crosspeaks: N-acetylaspartate, the glutamate/glutamine pool, aspartate, phosphoethanolamine, glucose, glutathione, taurine, myo-inositols, lactate, threonine, gamma-aminobutyric acid, and alanine. Most of the metabolites were measured without contamination of other resonances., (Copyright 2001 Academic Press.)

31P High resolution nuclear magnetic resonance studies have been carried out on the P388D1 tumoral cell line and the BCG elicited alveolar rabbit macrophages both in sedimented cells and in perfused agarose-embedded cells. When the cells were sufficiently oxygenated, the phosphorylated sugars and ATP concentrations attained high levels. The intensity of the peak representing phosphorylated sugars varied inversely with ATP level when macrophagic cells were treated by NaF. The identities of the phosphorylated sugars were revealed by 1H and 31P NMR studies of the P 388D1 cells perchloric extracts.

We report the full structure of two elliptinium diribonucleosides monophosphate adducts: the oxidized form of this antitumor agent alkylates very selectively the pX ribose of ApX (X = G or U) leading to a spiro derivative, where the C10 atom of ellipticine skeleton is linked to both sugar oxygen atoms 2' and 3'. No other adducts could be detected, specially the expected ones corresponding to the usual alkylation sites of bases.