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Background: As fat grafting is commonly used as a filler, Adipose-derived stem/stromal cells (ASC) have been reported to be key player in retention rate. Paracrine and differentiation potential of those cells confer them strong pro-angiogenic capacities. However, a full characterization of the influence of aging on ASC has not been reported yet. Here we've investigated the effect of age on paracrine function, stemness and angiogenic potential of ASC., Methods: ASC were extracted from young and old adult donors. We assessed stromal vascular fraction cell populations repartition, ASC stemness potential, capability to differentiate into mesenchymal lineages as well as their secretome. Angiogenic potential was assessed using a sprouting assay, an indirect co-culture of ASC and dermal microvascular endothelial cells (EC). Total vascular sprout length was measured, and co-culture soluble factors were quantified. Pro-angiogenic factors alone or in combination as well as ASC-conditioned medium (CM) were added to EC to assess sprouting induction., Results: Decrease of endothelial cells yield and percentage is observed in cells extracted from adipose tissue of older patients, whereas ASC percentage increased with age. Clonogenic potential of ASC is stable with age. ASC can differentiate into adipocytes, chondrocytes and osteoblasts, and aging does not alter this potential. Among the 25 analytes quantified, high levels of pro-angiogenic factors were found, but none is significantly modulated with age. ASC induce a significantly longer vascular sprouts compared to fibroblasts, and no difference was found between young and old ASC donors on that parameter. Higher concentrations of FGF-2, G-CSF, HGF and IL-8, and lower concentrations of VEGF-C were quantified in EC/ASC co-cultures compared to EC/fibroblasts co-cultures. EC/ASC from young donors secrete higher levels of VEGF-A compared to old ones. Neither soluble factor nor CM without cells are able to induce organized sprouts, highlighting the requirement of cell communication for sprouting. CM produced by ASC supporting development of long vascular sprouts promote sprouting in co-cultures that establish shorter sprouts., Conclusion: Our results show cells from young and old donors exhibit no difference in all assessed parameters, suggesting all patients could be included in clinical applications. We emphasized the leading role of ASC in angiogenesis, without impairment with age, where secretome is a key but not sufficient actor., Competing Interests: Declarations Competing interests The authors declare no competing interests. Ethics approval and consent to participate Written informed consent was obtained from the patient and surgical residue was collected according to French regulation and declared to research ministry., (© 2024. The Author(s).)

The field of organ transplantation is experiencing a transformative shift with the rise of Advanced Therapy Medicinal Products (ATMPs), which include gene therapies, somatic cell therapies, and tissue-engineered products. These therapies offer new, potentially curative treatments for longstanding medical challenges, impacting numerous patients. However, their adoption is hindered by complex regulatory frameworks, high production costs, and inconsistent access across Europe. The ESOT ATMP Task Force's position paper analyzes these challenges from research to clinical application, advocating for a coordinated strategy to position Europe as a leader in ATMP development. It proposes specific actions such as streamlining regulatory pathways to accelerate approvals, boosting funding for ATMP research, and creating specialized facilities for development and implementation. The paper also highlights the critical roles of patient engagement and real-world evidence in optimizing clinical and regulatory practices., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Berishvili, Piemonti, de Koning, Lindstedt, Scholz, Scott, Auxenfans, Johnson, Martin, Gunther, Mey, Potena and Thaunat.)

Introduction: Ductal carcinoma in situ (DCIS), characterized by a proliferation of neoplastic cells confined within the mammary ducts, is distinctly isolated from the surrounding stroma by an almost uninterrupted layer of myoepithelial cells (MECs) and by the basement membrane. Heightened interactions within the adipose microenvironment, particularly in obese patients, may play a key role in the transition from DCIS to invasive ductal carcinoma (IDC), which is attracting growing interest in scientific research. Adipose tissue undergoes metabolic changes in obesity, impacting adipokine secretion and promoting chronic inflammation. This study aimed to assess the interactions between DCIS, including in situ cancer cells and MECs, and the various components of its inflammatory adipose microenvironment (adipocytes and macrophages)., Methods: To this end, a 3D co-culture model was developed using bicellular bi-fluorescent DCIS-like tumoroids, adipose cells, and macrophages to investigate the influence of the inflammatory adipose microenvironment on DCIS progression., Results: The 3D co-culture model demonstrated an inhibition of the expression of genes involved in apoptosis ( BAX, BAG1, BCL2, CASP3, CASP8 , and CASP9 ), and an increase in genes related to cell survival ( TP53 , JUN , and TGFB1 ), inflammation ( TNF-α, PTGS2, IL-6R ), invasion and metastasis (TIMP1 and MMP-9) in cancer cells of the tumoroids under inflammatory conditions versus a non-inflammatory microenvironment. On the contrary, it confirmed the compromised functionality of MECs, resulting in the loss of their protective effects against cancer cells. Adipocytes from obese women showed a significant increase in the expression of all studied myofibroblast-associated genes (myoCAFs), such as FAP and α-SMA . In contrast, adipocytes from normal-weight women expressed markers of inflammatory fibroblast phenotypes (iCAF) characterized by a significant increase in the expression of LIF and inflammatory cytokines such as TNF-α, IL-1β, IL-8 , and CXCL-10 . These changes also influenced macrophage polarization, leading to a pro-inflammatory M1 phenotype. In contrast, myoCAF-associated adipocytes, and the cancer-promoting microenvironment polarized macrophages towards an M2 phenotype, characterized by high CD163 receptor expression and IL-10 and TGF-β secretion., Discussion: Reciprocal interactions between the tumoroid and its microenvironment, particularly in obesity, led to transcriptomic changes in adipocytes and macrophages, may participate in breast cancer progression while disrupting the integrity of the MEC layer. These results underlined the importance of adipose tissue in cancer progression., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Habanjar, Nehme, Goncalves-Mendes, Cueff, Blavignac, Aoun, Decombat, Auxenfans, Diab-Assaf, Caldefie-Chézet and Delort.)

Excess weight and obesity are the fifth leading cause of death globally, and sustained efforts from health professionals and researchers are required to mitigate this pandemic-scale problem. Polyphenols and flavonoids found in Aspalathus linearis -a plant widely consumed as Rooibos tea-are increasingly being investigated for their positive effects on various health issues including inflammation. The aim of our study was to examine the effect of Rooibos extract on obesity and the associated low-grade chronic inflammatory state by testing antioxidant activity, cytokine secretions, macrophage polarization and the differentiation of human adipocytes through the development of adipospheroids. Rooibos extract significantly decreased ROS production and the secretion of pro-inflammatory cytokines (IFN-γ, IL-12, IL-2 and IL-17a) in human leukocytes. Additionally, Rooibos extract down-regulated LPS-induced macrophage M1 polarization, shown by a significant decrease in the expression of pro-inflammatory cytokines: TNFα , IL-8 , IL-6 , IL-1β and CXCL10 . In addition, Rooibos inhibited intracellular lipid accumulation and reduced adipogenesis by decreasing the expression of PPARγ , Ap2 and HSL in adipospheroids. A significant decrease in leptin expression was noted and this, more interestingly, was accompanied by a significant increase in adiponectin expression. Using a co-culture system between macrophages and adipocytes, Rooibos extract significantly decreased the expression of all studied pro-inflammatory cytokines and particularly leptin , and increased adiponectin expression. Thus, adding Rooibos tea to the daily diet is likely to prevent the development of obesity associated with chronic low-level inflammation.

Summary: Over the past 30 years, there has been a dramatic increase in the use of autologous fat grafting for soft-tissue augmentation and to improve facial skin quality. Several studies have highlighted the impact of aging on adipose tissue, leading to a decrease of adipose tissue volume and preadipocyte proliferation and increase of fibrosis. Recently, there has been a rising interest in adipose tissue components, including adipose-derived stem/stromal cells (ASCs) because of their regenerative potential, including inflammation, fibrosis, and vascularization modulation. Because of their differentiation potential and paracrine function, ASCs have been largely used for fat grafting procedures, as they are described to be a key component in fat graft survival. However, many parameters as surgical procedures or adipose tissue biology could change clinical outcomes. Variation on fat grafting methods have led to numerous inconsistent clinical outcomes. Donor-to-donor variation could also be imputed to ASCs, tissue inflammatory state, or tissue origin. In this review, the authors aim to analyze (1) the parameters involved in graft survival, and (2) the effect of aging on adipose tissue components, especially ASCs, that could lead to a decrease of skin regeneration and fat graft retention., Clinical Relevance Statement: This review aims to enlighten surgeons about known parameters that could play a role in fat graft survival. ASCs and their potential mechanism of action in regenerative medicine are more specifically described., (Copyright © 2022 by the American Society of Plastic Surgeons.)

With the aim of designing a preclinical study evaluating an intracerebral cell-based therapy for stroke, an observational study was performed in the rat suture model of ischemic stroke. Objectives were threefold: (i) to characterize neurofunctional and imaging readouts in the first weeks following transient ischemic stroke, according to lesion subtype (hypothalamic, striatal, corticostriatal); (ii) to confirm that intracerebral administration does not negatively impact these readouts; and (iii) to calculate sample sizes for a future therapeutic trial using these readouts as endpoints. Our results suggested that the most relevant endpoints were side bias (staircase test) and axial diffusivity (AD) (diffusion tensor imaging). Hypothalamic-only lesions did not affect those parameters, which were close to normal. Side bias in striatal lesions reached near-normal levels within 2 weeks, while rats with corticostriatal lesions remained impaired until week 14. AD values were decreased at 4 days and increased at 5 weeks post-surgery, with a subtype gradient: hypothalamic < striatal < corticostriatal. Intracerebral administration did not impact these readouts. After sample size calculation (18-147 rats per group according to the endpoint considered), we conclude that a therapeutic trial based on both readouts would be feasible only in the framework of a multicenter trial., (© 2022. The Author(s).)

Intraoperative three-dimensional fabrication of living tissues could be the next biomedical revolution in patient treatment., Approach: We developed a surgery-ready robotic three-dimensional bioprinter and demonstrated that a bioprinting procedure using medical grade hydrogel could be performed using a 6-axis robotic arm in vivo for treating burn injuries., Results: We conducted a pilot swine animal study on a deep third-degree severe burn model. We observed that the use of cell-laden bioink as treatment substantially affects skin regeneration, producing in situ fibroblast growth factor and vascular endothelial growth factor, necessary for tissue regeneration and re-epidermalization of the wound., Conclusions: We described an animal study of intraoperative three-dimensional bioprinting living tissue. This emerging technology brings the first proof of in vivo skin printing feasibility using a surgery-ready robotic arm-based bioprinter. Our positive outcome in skin regeneration, joined with this procedure's feasibility, allow us to envision the possibility of using this innovative approach in a human clinical trial in the near future., (Copyright © 2022 The Authors. Published by Wolters Kluwer Health, Inc. on behalf of The American Society of Plastic Surgeons.)

Background: In human subcutaneous adipose tissue, the superficial fascia distinguishes superficial and deep microenvironments showing extensions called retinacula cutis. The superficial subcutaneous adipose tissue has been described as hyperplastic and the deep subcutaneous adipose tissue as inflammatory. However, few studies have described stromal-vascular fraction (SVF) content and adipose-derived stromal/stem cells (ASCs) behavior derived from superficial and deep subcutaneous adipose tissue. In this study, we analyzed a third conjunctive microenvironment: the retinacula cutis superficialis derived from superficial subcutaneous adipose tissue., Methods: The samples of abdominal human subcutaneous adipose tissue were obtained during plastic aesthetic surgery in France (Declaration DC-2008-162) and Brazil (Protocol 145/09)., Results: The SVF content was characterized in situ by immunofluorescence and ex vivo by flow cytometry revealing a high content of pre-adipocytes rather in superficial subcutaneous adipose tissue microenvironment. Adipogenic assays revealed higher percentage of lipid accumulation area in ASCs from superficial subcutaneous adipose tissue compared with retinacula cutis superficialis (p < 0.0001) and deep subcutaneous adipose tissue (p < 0.0001). The high adipogenic potential of superficial subcutaneous adipose tissue was corroborated by an up-regulation of adipocyte fatty acid-binding protein (FABP4) compared with retinacula cutis superficialis (p < 0.0001) and deep subcutaneous adipose tissue (p < 0.0001) and of C/EBPα (CCAAT/enhancer-binding protein alpha) compared with retinacula cutis superficialis (p < 0.0001) and deep subcutaneous adipose tissue (p < 0.0001) microenvironments. Curiously, ASCs from retinacula cutis superficialis showed a higher level of adiponectin receptor gene compared with superficial subcutaneous adipose tissue (p = 0.0409), widely known as an anti-inflammatory hormone. Non-induced ASCs from retinacula cutis superficialis showed higher secretion of human vascular endothelial growth factor (VEGF), compared with superficial (p = 0.0485) and deep (p = 0.0112) subcutaneous adipose tissue and with adipogenic-induced ASCs from superficial (p = 0.0175) and deep (p = 0.0328) subcutaneous adipose tissue. Furthermore, ASCs from retinacula cutis superficialis showed higher secretion of Chemokine (C-C motif) ligand 5 (CCL5) compared with non-induced (p = 0.0029) and induced (p = 0.0089) superficial subcutaneous adipose tissue., Conclusions: This study highlights the contribution to ASCs from retinacula cutis superficialis in their angiogenic property previously described for the whole superficial subcutaneous adipose tissue besides supporting its adipogenic potential for superficial subcutaneous adipose tissue., (© 2021. The Author(s).)

Breast cancer is the most common cancer among women worldwide. Overweight and obesity are now recognized as established risk factors for this pathology in postmenopausal women. These conditions are also believed to be responsible for higher recurrence and mortality rates. Reciprocal interactions have been described between adipose and cancer cells. An adipose microenvironment favors a greater proliferation of cancer cells, their invasion and even resistance to anti-cancer treatments. In addition, the chronic low-grade inflammation observed in obese individuals is believed to amplify these processes. Among the cell types present in the breast, myoepithelial cells (MECs), located at the interface of the epithelial cells and the stroma, are considered "tumor suppressor" cells. During the transition from ductal carcinoma in situ to invasive cancer, disorganization or even the disappearance of MECs is observed, thereby enhancing the ability of the cancer cells to migrate. As the adipose microenvironment is now considered as a central actor in the progression of breast cancer, our objective was to evaluate if it could be involved in MEC functional modifications, leading to the transition of in situ to invasive carcinoma, particularly in obese patients. Through a co-culture model, we investigated the impact of human adipose stem cells from women of normal weight and obese women, differentiated or not into mature adipocytes, on the functionality of the MECs by measuring changes in viability, apoptosis, gene, and miRNA expressions. We found that adipose cells (precursors and differentiated adipocytes) could decrease the viability of the MECs, regardless of the original BMI. The adipose cells could also disrupt the expression of the genes involved in the maintenance of the extracellular matrix and to amplify the expression of leptin and inflammatory markers. miR-122-5p and miR-132-3p could also be considered as targets for adipose cells. The metabolite analyses revealed specific profiles that may be involved in the growth of neoplastic cells. All of these perturbations could thus be responsible for the loss of tumor suppressor status of MECs and promote the transition from in situ to invasive carcinoma., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Delort, Cholet, Decombat, Vermerie, Dumontet, Castelli, Fenaille, Auxenfans, Rossary and Caldefie-Chezet.)

Clinical grade cultured epithelial autograft (CEA) are routinely used to treat burns covering more than 60% of the total body surface area. However, although the epidermis may be efficiently repaired by CEA, the dermal layer, which is not spared in deep burns, requires additional treatment strategies. Our aim is to develop an innovative method of skin regeneration based on in situ 3D bioprinting of freshly isolated autologous skin cells. We describe herein bioink formulation and cell preparation steps together with experimental data validating a straightforward enzyme-free protocol of skin cell extraction. This procedure complies with both the specific needs of 3D bioprinting process and the stringent rules of good manufacturing practices. This mechanical extraction protocol, starting from human skin biopsies, allows harvesting a sufficient amount of both viable and growing keratinocytes and fibroblasts. We demonstrated that a dermis may be reconstituted in vitro starting from a medical grade bioink and mechanically extracted skin cells. In these experiments, proliferation of the extracted cells can be observed over the first 21 days period after 3D bioprinting and the analysis of type I collagen exhibited a de novo production of extracellular matrix proteins. Finally, in vivo experiments in a murine model of severe burn provided evidences that a topical application of our medical grade bioink was feasible and well-tolerated. Overall, these results represent a valuable groundwork for the design of future 3D bioprinting tissue engineering strategies aimed at treating, in a single intraoperative step, patients suffering from extended severe burns., (© 2020 John Wiley & Sons Ltd.)

Severe burn injuries remain a major health problem due to high rates of mortality, residual morbidity, and/or aesthetic damages. To find new therapies aimed at promoting a harmonious healing of skin burns, it is important to develop models which take into account the unique properties of the human skin. Based on previously described models of burn injury performed on human skin explants, we hypothesized that maintaining explants under constant tension forces would allow to more closely reproduce the pathophysiological processes of skin remodeling. We thus. Here, we set up and characterized an improved model of deep second-degree burn injury on ex vivo cultured human skin explants at air-liquid interface and maintained under conditions of constant tension forces. A spontaneous re-epithelialization of the lesion was observed 8 to 9 days post burn and was found to rely on the proliferation of basal keratinocytes at the wound edges. Collagen VII at the dermo-epidermal junction reformed along with the progression of re-epithelializatio and a synthesis of procollagen III was observed in the dermis at the wound site. These findings indicate that our model is suitable for the assessment of clinically-relevant therapies aimed at modulating the kinetics of re-epithelialization and/or the activation of fibroblasts following skin burn injuries. In this regard, we evaluated the use of a thermoreversible poloxamer hydrogel as a vehicle for topically-testable therapeutic molecules. Our data showed that, although useful for drug formulation, the p407/p188 poloxamer hydrogel induces a delay of skin re-epithelialization in humans skin explants submitted to experimental burn injury.

Bone morphogenetic protein 1 (BMP-1) is an important metalloproteinase that synchronizes growth factor activation with extracellular matrix assembly during morphogenesis and tissue repair. The mechanisms by which BMP-1 exerts these effects are highly context dependent. Because BMP-1 overexpression induces marked phenotypic changes in two human cell lines (HT1080 and 293-EBNA cells), we investigated how BMP-1 simultaneously affects cell-matrix interactions and growth factor activity in these cells. Increasing BMP-1 led to a loss of cell adhesion that depended on the matricellular glycoprotein thrombospondin-1 (TSP-1). BMP-1 cleaved TSP-1 between the VWFC/procollagen-like domain and the type 1 repeats that mediate several key TSP-1 functions. This cleavage induced the release of TSP-1 C-terminal domains from the extracellular matrix and abolished its previously described multisite cooperative interactions with heparan sulfate proteoglycans and CD36 on HT1080 cells. In addition, BMP-1-dependent proteolysis potentiated the TSP-1-mediated activation of latent transforming growth factor-β (TGF-β), leading to increased signaling through the canonical SMAD pathway. In primary human corneal stromal cells (keratocytes), endogenous BMP-1 cleaved TSP-1, and the addition of exogenous BMP-1 enhanced cleavage, but this had no substantial effect on cell adhesion. Instead, processed TSP-1 promoted the differentiation of keratocytes into myofibroblasts and stimulated production of the myofibroblast marker α-SMA, consistent with the presence of processed TSP-1 in human corneal scars. Our results indicate that BMP-1 can both trigger the disruption of cell adhesion and stimulate TGF-β signaling in TSP-1-rich microenvironments, which has important potential consequences for wound healing and tumor progression., (Copyright © 2020 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Although sulfur-rich thermal waters have ancestrally been used in the context of dermatological conditions, a global mapping of the molecular effects exerted by H 2 S on human keratinocytes is still lacking. To fill this knowledge gap, we subjected cultured human keratinocytes to distinct amounts of the non-gaseous hydrogen sulfur donor NaHS. We first checked that H 2 S accumulated in the cytoplasm of keratinocytes under our experimental conditions andused a combination of proteomics, genomics and biochemical approaches to unravel functionally relevant H 2 S targets in human keratinocytes. We found that the identified targets fall into two main categories: (i) the oxidative stress response molecules superoxide dismutase 2 (SOD2), NAD(P)H quinone dehydrogenase 1 (NQO1) and culin 3 (CUL3) and (ii) the chemokines interleukin-8 (IL-8) and CXCL2. Interestingly, NaHS also stimulated the caspase-1 inflammasome pathway, leading to increased secretion of the pro-inflammatory molecule interleukin-18 (IL-18). Interestingly, the secretion of interleukin-1 beta (IL-1β) was only modestly impacted by NaHS exposure despite a significant accumulation of IL-1β pro-form. Finally, we observed that NaHS significantly hampered the growth of human keratinocyte progenitors and stem cells cultured under clonogenic conditions or as epidermal cell sheets. We conclude that H 2 S exerts specific molecular effects on normal human keratinocytes., Competing Interests: Laurent Misery is a consultant for Avène, Uriage and La Roche-Posay.