Paradise, Christopher R.

Stem cell-derived β (SC-β) cells are a promising therapy for type 1 diabetes (T1D), but their long-term efficacy is limited by immune-mediated graft rejection. Platelet-derived exosome product (PEP) has emerged as a novel immunomodulatory agent, although its effect on SC-β cell xenograft rejection remains unclear. Here, we evaluated PEP in stimulated human peripheral blood mononuclear cells (PBMCs) and in an immunocompetent mouse model of SC-β cell transplantation under the kidney capsule. Immune-related gene expression was assessed by quantitative PCR, immune cell infiltration by immunofluorescence, and graft function by circulating human insulin. In vitro, PEP reduced NK-cell-associated gene expression, including NK1.1, EOMES, and KLRK1, and increased IL-10 expression. In vivo, untreated SC-β cell xenografts showed progressive immune infiltration and loss of detectable human insulin by day 14, whereas PEP-treated grafts retained detectable insulin through day 14 across co-transplantation, pretreatment, and systemic administration strategies. PEP co-transplantation delayed, but did not prevent, xenograft rejection, with graft loss observed by day 21. This delay was associated with reduced NK1.1+ cell infiltration and lower expression of selected inflammatory and rejection-associated markers, including NK1.1, Nos2, and Nlrp3. These findings suggest that PEP modulates graft-associated immune responses and may serve as an adjunct immunomodulatory strategy for stem cell-based therapies for type 1 diabetes. Further studies are needed to evaluate sustained graft durability, safety, and translational efficacy.

ABSTRACT Femoroacetabular impingement (FAI) of the hip results from altered morphology of the proximal femur and/or acetabulum, creating abnormal contact between these structures during normal hip range of motion. The etiology of this structural phenotype is unknown. This study aimed to identify common single‐nucleotide variants (SNVs) among two families with seemingly heritable FAI. Additionally, this study examined biologic networks among proteins encoded by identified SNVs and surveyed the literature regarding the role of these proteins in musculoskeletal development. Methodologically, blood samples were collected from affected and unaffected members of two separate families with a pattern of heritable FAI. Exome sequencing identified SNVs, and a protein–protein interaction network assessment, in addition to a literature review, was performed to better understand the role of variant‐associated genes in musculoskeletal development. Results showed 33 and 43 variant‐associated genes affected in Family 1 and Family 2, respectively. Although no overlap in variant‐associated genes was found between families, network analysis showed that variant‐associated genes are functionally interconnected and are known to interact with each other. Furthermore, the variant‐associated genes are broadly associated with musculoskeletal development. Ultimately, genetic analysis of two families of multiple generations of FAI revealed variations in genes regulating musculoskeletal development, with several being critical local regulators of bone morphogenesis.

Summary: Exosomes, or extracellular vesicles, represent the latest cell-free addition to the regenerative medicine toolkit. In vitro preclinical studies have demonstrated the safety and efficacy of exosomes, which vary based on source and biomanufacturing, for a myriad of potential therapeutic applications relevant to skin and soft tissue reconstruction. Primary search was performed in September 2021 on the MEDLINE database via PubMed and Ovid, with focus on articles about therapeutic application of exosomes or extracellular vesicles. In total, 130 articles met criteria for applicability, including early-stage clinical trials, preclinical research studies with in vivo application, and articles applicable to plastic and reconstructive surgery and dermatology. Most studies used animal models of human disease processes, using either animal donor cells to isolate exosomes, or human donor cells in animal models. Exosome technology has catapulted as an acellular therapeutic vehicle with off-the-shelf accessibility. These features eliminate prior threshold for broad adoption of regenerative cell-based therapies into surgical and medical practice. To date, there are no exosome products approved by the US Food and Drug Administration. This review highlights exosomes as the new frontier in regenerative medicine and outlines its preclinical therapeutic applications for cutaneous repair and restoration.

Background: Multiple effective treatments exist for correction of skin photoaging. Topical L-ascorbic acid (vitamin C), a well-known antioxidant and topical human platelet extract (HPE), is a novel off-the-shelf cosmetic ingredient that has shown positive results in recent clinical studies. HPE is a leukocyte-depleted allogeneic product derived from United States–sourced, pooled, apheresed platelets produced with consistent batch quality, purity, and effect. The authors sought to characterize the effect of topical HPE (plated) intense serum (Rion Aesthetics) compared with vitamin C (C E Ferulic with 15% L-ascorbic acid; SkinCeuticals) in skin rejuvenation of dorsal hands after 12 to 26 weeks of twice-daily use. Methods: This prospective, longitudinal study sought to compare the effectiveness of 2 known treatments for skin rejuvenation. Evaluations at baseline and 6, 12, and 26 weeks included photographic documentation to assess common skin concerns related to aging. Results: For age-related skin appearance on the dorsal hands, topical HPE was noninferior to topical vitamin C for improvement in brown spot fractional area, wrinkle fractional area, and improvement in luminosity at 12 weeks after twice-daily topical use. Conclusions: HPE performed as well as vitamin C to rejuvenate the skin on the dorsal hands after 12 to 26 weeks of twice-daily topical use. Both topical serums may yield similar or superior results versus invasive procedures, such as intense pulsed light, in reducing brown spots on the dorsal hands. These topical products work equally well in both sexes. Skin improvements lasted through 6 months. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, II.

AbstractBackgroundFractional carbon dioxide (CO2) laser resurfacing is used successfully for facial rejuvenation. Post procedure skincare is a variable that influences downtime caused by pain/tenderness, erythema, crusting, and bruising.AimsThe primary objective of this pilot study was to demonstrate the benefits of human platelet extract (HPE) (plated)™ CALM Serum, a new topical cosmetic product, following fractionated CO2 ablative laser resurfacing treatment to the entire face versus standard of care.MethodsIn a single‐center, randomized, evaluator‐blinded pilot study, a total of 18 subjects were randomized into two groups, CO2 facial resurfacing followed by post‐procedural standard of care (Stratacel silicone gel) or CO2 facial resurfacing with the addition of HPE renewosomes in the CALM Serum.ResultsCALM Serum demonstrated statistically significant less crusting at Day 10 compared to the control group (p = 0.0193) with less downtime in the first 14 days (p = 0.03). Subjects treated with CALM Serum had statistically significant brighter appearing skin at 14 days (p = 0.007) and more youthful looking skin on Days 14 and 30 (p = 0.003 and 0.04, respectively).ConclusionsThis study demonstrates that Renewosome™ technology provides statistically significant post‐laser clinical recovery over silicone gel for reducing crusting, and downtime. Subjects reported less diary days of symptoms of pain/tenderness, redness, crusting/flaking, bruising, and itching in the first 14 days compared to the control group. CALM also demonstrated statistically significant improvements in brighter and more youthful appearing skin. CALM is safe and well tolerated.

Abstract Background Exosomes are regenerative mediators for skin rejuvenation. Human platelet extract (HPE) is an allogeneic exosome product derived from US-sourced, leukocyte-reduced apheresed platelets with consistent purity and potency. Objectives The authors sought to better characterize the safety and tolerability of novel HPE (plated) Intensive Repair Serum (Rion Aesthetics, Rochester, MN) and its maximal effects on skin rejuvenation at 6 weeks. Methods This prospective, single-arm, non-randomized, longitudinal study investigated the safety and efficacy of HPE. Structured sub-analysis evaluated multifactorial improvement in skin health following standardized skin care regimen to determine the maximal effect. Evaluation at baseline and 6 weeks included participant questionnaires and photo documentation with VISIA-CR Generation 5 3D PRIMOS (Canfield Scientific Inc, Fairfield, NJ). Results VISIA-CR imaging yielded quantifiable and statistically significant improvements in overall skin health (skin health score). A greater score correlated to greater overall skin health, and there was a statistically significant mean delta improvement of 224.2 ± 112.8 (mean ± standard deviation, P ≤ 0.0001) in skin health score at 6 weeks compared with baseline. This correlated to reduction in redness, wrinkles, and melanin production across all cosmetic units (P = 0.005, P = 0.0023, P ≤ 0.0001, respectively) and significant improvements in luminosity and color evenness (P ≤ 0.001). Conclusions A topically applied platelet-derived exosome product, HPE, induced normalization to skin health at 4 to 6 weeks with improved various clinical measures of facial photodamage and cutaneous aging. It is safe, well-tolerated, and well-liked by participants. Level of Evidence: 4

Abstract Background Adipose-derived mesenchymal stromal cells (AMSCs) represent attractive cellular therapeutics for treatment of various diseases including osteoarthritis and bone degeneration. Preparation and characterization of AMSCs show donor-to-donor variability that indicating that AMSCs are heterogeneous populations of cells. Cell surface markers to identify MSCs, such as CD44, CD90, and CD105, are relatively invariable among AMSCs and do not reflect the heterogeneity of AMSC populations. CD36, a multifunctional surface receptor associated with transport of oxidized low-density lipoproteins and fatty acids, is also expressed by AMSCs but this marker is considerable more variable. Therefore, we assessed whether CD36 can be used to identify subpopulations in AMSCs. Methods Clinical-grade AMSCs were isolated from patient fat biopsies, cultured under xenobiotic-free conditions using human platelet lysate and sorted by flow cytometry or magnetic cell sorting to characterize CD36 expression. CD36 enriched, CD36 depleted, and unsorted AMSCs were characterized using cellular and molecular techniques including proliferation, multilineage potential, RNA-sequencing and bioinformatics, metabolomics, and sensitivity to the CD36 inhibitor sulfosuccinimidyl oleate (SSO). Results CD36 exhibited biphasic expression on AMSCs grown in human platelet lysate. Enrichment of CD36+ AMSCs showed that CD36+ expression identifies a stable subpopulation. Transcriptomic analysis revealed enhancement of calcium transporter genes. Osteogenic differentiation of CD36+ enriched, CD36+ depleted, and unsorted AMSCs showed strikingly enhanced osteogenic calcium deposition and enhanced expression of alkaline phosphatase (ALPL) and osteoprotegrin (TNFRSF11B) in the CD36+ population. While lipid droplets were not altered, adipogenic differentiation showed upregulated gene expression of key adipogenic markers, including fatty acid-binding protein 4 (FABP4) and Adiponectin (ADIPOQ), with CD36+ enrichment. Treatment of CD36+ enriched AMSCs with the SSO showed reduced calcium deposition whereas CD36+ depleted AMSCs were unaffected. Conclusions CD36 exhibits variable expression amongst AMSCs. CD36+ enriched AMSCs are a stable subpopulation with enhanced osteogenic potential that linked to CD36 receptor function. These results may further refine production and clinical application of AMSC cellular therapeutics. Furthermore, the enhanced osteogenic potential of CD36+ AMSCs may be considered for therapeutic regeneration of bone.

ABSTRACTBromodomain (BRD) proteins are histone code interpreters that recognize acetylated lysines and link the dynamic state of chromatin with the transcriptional machinery. Here, we demonstrate that ablation of the Brd4 gene in primary mouse bone marrow–derived mesenchymal stem cells via a conditional Brd4fl/fl allele suppresses osteogenic lineage commitment. Remarkably, loss of Brd4 function also enhances expression of genes in engineered adenoviral vectors, including Cre recombinase and green fluorescent protein (GFP). Similarly, vector‐based expression of BMP2 mRNA and protein levels are enhanced upon Brd4 depletion in cells transduced with an adenoviral vector that expresses BMP2 (Ad‐BMP2). Importantly, Brd4 depletion in MC3T3‐E1 and human adipose‐derived mesenchymal stem cells (AMSCs) transduced with Ad‐BMP2 enhances osteogenic differentiation of naïve MC3T3‐E1 cells via paracrine mechanisms based on transwell and conditioned medium studies. Our studies indicate that Brd4 depletion enhances adenoviral transgene expression in mammalian cells, which can be leveraged as a therapeutic strategy to improve viral vector‐based gene therapies. © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Objective Cartilage formation is stimulated in mixtures of chondrocytes and human adipose–derived mesenchymal stromal cells (MSCs) both in vitro and in vivo. During coculture, human MSCs perish. The goal of this study is to elucidate the mechanism by which adipose tissue–derived MSC cell death occurs in the presence of chondrocytes. Methods Human primary chondrocytes were cocultured with human MSCs derived from 3 donors. The cells were cultured in monoculture or coculture (20% chondrocytes and 80% MSCs) in pellets (200,000 cells/pellet) for 7 days in chondrocyte proliferation media in hypoxia (2% O 2 ). RNA sequencing was performed to assess for differences in gene expression between monocultures or coculture. Immune fluorescence assays were performed to determine the presence of caspase-3, LC3B, and P62. Results RNA sequencing revealed significant upregulation of >90 genes in the 3 cocultures when compared with monocultures. STRING analysis showed interconnections between >50 of these genes. Remarkably, 75% of these genes play a role in cell death pathways such as apoptosis and autophagy. Immunofluorescence shows a clear upregulation of the autophagic machinery with no substantial activation of the apoptotic pathway. Conclusion In cocultures of human MSCs with primary chondrocytes, autophagy is involved in the disappearance of MSCs. We propose that this sacrificial cell death may contribute to the trophic effects of MSCs on cartilage formation.