CD24 flags anastasis in melanoma cells.

Anastasis is a phenomenon observed in cancer cells, where cells that have initiated apoptosis are able to recover and survive. This molecular event is increasingly recognized as a potential contributor to cancer metastasis, facilitating the survival and migration of tumor cells. Nevertheless, the identification of a specific surface marker for detecting cancer cells in anastasis remained elusive. Here we report our observation that the cell surface expression of CD24 is preferentially enriched in a non-adherent FSClowSSChigh melanoma subpopulation, which is generally considered a non-viable population in cultivated melanoma cell lines. More than 90% of non-adherent FSClowSSChighCD24+ve metastatic melanoma cells exhibited bonafide features of apoptosis on the cell surface and in the nucleus, marking apoptotic or seemingly apoptotic subpopulations of the in vitro cultivated metastatic melanoma cell lines. Unexpectedly, however, the CD24+ve subpopulation, despite being apoptotic, showed evidence of metabolic activity and exhibited proliferative capacities, including anchorage-independent growth, when inoculated in soft agarose growth medium. These findings indicate that apoptotic FSClowSSChighCD24+ve melanoma subpopulations are capable of reversing the progression of apoptosis. We report CD24 as the first novel cell surface marker for anastasis in melanoma cells.

Exosomes from SHED-MSC regulate polarization and stress oxidative indexes in THP-1 derived M1 macrophages.

OBJECTIVE: The inhibition of M1 macrophages may be interesting for targeted therapy with mesenchymal stem cell-derived Exosomes (MSC-EXOs). This study aimed to investigate the stem cells of human exfoliated deciduous teeth-derived EXOs (SHED-MSC-EXOs) effect on regulating the pro- and anti-oxidant indexes and inhibiting M1 macrophage polarization. Besides, an in-silico analysis of SHED-MSC-EXO miRNAs as the highest frequency of small RNAs in the exosomes was performed to discover the possible mechanism. METHODS: The flow cytometry analysis of CD80 and CD86 as M1-specific markers confirmed the polarization of macrophages derived from THP-1 cells. After exosome isolation, characterization, and internalization, THP-1-derived M1 macrophages were treated with SHED-MSC-EXOs. M1-specific markers and pro- and anti-oxidant indexes were evaluated. For in-silico analysis of SHED-MSC-EXOs miRNAs, initial miRNA array data of SHED-EXOs is collected from GEO, and the interaction of the miRNAs in M1 macrophage polarization (M1P), mitochondrial oxidative stress (MOS) and LPS-induced oxidative stress (LOS) were analyzed by miRWalk 3.0 server. Outcomes were filtered by 75th percentile signal intensity, score cut-off ≥0.95, minimum free energy (MEF)≤ -20 kcal/mol, and seed = 1. RESULTS: It shows a decrease in the expression of CD80 and CD81, a reduction in pro-oxidant indicators, and an increase in the anti-oxidant indexes (P < 0.05). Computational analysis showed that eight microRNAs of SHED-MSC-EXO miRNAs can bind to and interfere with the expression of candidate genes in the M1P, MOS, and LOS pathways simultaneously. CONCLUSION: SHED-MSCs-EXOs can be utilized to treat conditions related to M1 macrophage-induced diseases (M1IDs) due to their unique physical properties and ability to penetrate target cells easily.

Young small extracellular vesicles rejuvenate replicative senescence by remodeling Drp1 translocation-mediated mitochondrial dynamics.

BACKGROUND: Human mesenchymal stem cells have attracted interest in regenerative medicine and are being tested in many clinical trials. In vitro expansion is necessary to provide clinical-grade quantities of mesenchymal stem cells; however, it has been reported to cause replicative senescence and undefined dysfunction in mesenchymal stem cells. Quality control assessments of in vitro expansion have rarely been addressed in ongoing trials. Young small extracellular vesicles from the remnant pulp of human exfoliated deciduous teeth stem cells have demonstrated therapeutic potential for diverse diseases. However, it is still unclear whether young small extracellular vesicles can reverse senescence-related declines. RESULTS: We demonstrated that mitochondrial structural disruption precedes cellular dysfunction during bone marrow-derived mesenchymal stem cell replication, indicating mitochondrial parameters as quality assessment indicators of mesenchymal stem cells. Dynamin-related protein 1-mediated mitochondrial dynamism is an upstream regulator of replicative senescence-induced dysfunction in bone marrow-derived mesenchymal stem cells. We observed that the application of young small extracellular vesicles could rescue the pluripotency dissolution, immunoregulatory capacities, and therapeutic effects of replicative senescent bone marrow-derived mesenchymal stem cells. Mechanistically, young small extracellular vesicles could promote Dynamin-related protein 1 translocation from the cytoplasm to the mitochondria and remodel mitochondrial disruption during replication history. CONCLUSIONS: Our findings show that Dynamin-related protein 1-mediated mitochondrial disruption is associated with the replication history of bone marrow-derived mesenchymal stem cells. Young small extracellular vesicles from human exfoliated deciduous teeth stem cells alleviate replicative senescence by promoting Dynamin-related protein 1 translocation onto the mitochondria, providing evidence for a potential rejuvenation strategy.

The Exosomes of Stem Cells from Human Exfoliated Deciduous Teeth Suppress Inflammation in Osteoarthritis.

Hyaluronic acid injection is commonly used clinically to slow down the development of osteoarthritis (OA). A newly developed therapeutic method is to implant chondrocytes/stem cells to regenerate cartilage in the body. The curative effect of stem cell therapy has been proven to come from the paracrine of stem cells. In this study, exosomes secreted by stem cells from human exfoliated deciduous teeth (SHED) and hyaluronic acid were used individually to evaluate the therapeutic effect in slowing down OA. SHED was cultured in a serum-free medium for three days, and the supernatant was collected and then centrifuged with a speed difference to obtain exosomes containing CD9 and CD63 markers, with an average particle size of 154.1 nm. SW1353 cells were stimulated with IL-1ß to produce the inflammatory characteristics of OA and then treated with 40 µg/mL exosomes and hyaluronic acid individually. The results showed that the exosomes successfully inhibited the pro-inflammatory factors, including TNF-α, IL-6, iNOS, NO, COX-2 and PGE2, induced by IL-1ß and the degrading enzyme of the extrachondral matrix (MMP-13). Collagen II and ACAN, the main components of the extrachondral matrix, were also increased by 1.76-fold and 2.98-fold, respectively, after treatment, which were similar to that of the normal joints. The effect can be attributed to the partial mediation of SHED exosomes to the NF-κB pathway, and the ability of exosomes to inhibit OA is found not inferior to that of hyaluronic acid.

Distribution characteristics and influencing factors of Haemaphysalis longicornis around goat sheds in Jinan city, East China.

As one of the most important disease vectors worldwide, ticks can transmit a number of pathogenic organisms to humans and domestic animals and cause a variety of important natural focal diseases and zoonoses. Domestic livestock play a vital role in the dispersal of ticks from the field environment to the human settlement, contributing to the prevalence of tick-borne diseases. Identification of the tick control region could contribute a vital role in strategic planning and cost-effective tick control measures. However, little is known about the spatial distribution characteristics of ticks around livestock sheds, which will lead to abusage and overuse of insecticides. Therefore, this study aimed to explore spatial distribution characteristics and correlation factors of ticks around goat sheds. A total of 3898 ticks were collected from eight goat sheds from April to June in Jinan city. All the sampled ticks belonged to the same species, namely Haemaphysalis longicornis, and 88.8% of them were nymphs. A significant positive correlation was noted between free-living ticks and parasitic ticks (r = 0.411, P < 0.001). However, there was a significant negative correlation between number of free-living ticks and distance from the goat sheds (r = -0.622, P < 0.001). Within 20 m from the goat sheds, 2211 ticks were collected respectively, representing 56.7% of the total free-living ticks. At a distance of 30 m, 57.6% decline in the tick density was found with a significant difference (q = 5.534, P < 0.001). In conclusion, focusing control efforts near the goat sheds should be recommend for tick prevention and control.

A Collaborative Approach for Metal Pollution Assessment in Production System of Plastic-Shed Vegetables Near Industrial Areas.

A collaborative assessment approach, including impact index of comprehensive quality (IICQ), food pollution index (FPI), and single factor pollution index (PI), was used to simultaneously select priority metal pollutants and assess metal contamination status in the plastic-shed soil (PSS)-vegetable system of the industrial towns situated in the Yangtze River Delta, China. Overall, significant Cr increment as well as Cd and Cu pollution in PSS existed, which was related to anthropogenic activities, especially industrial wastewater irrigation. The evaluation using PI and FPI demonstrated that priority metal pollutants were Cu and Cd in PSS while Cr and Cd in vegetables. Additionally, the estimation using IICQ method revealed that 23.3% and 13.3% of the sampling sites were sub-moderately and heavily contaminated by metals, respectively. These sites especially with heavy pollution need priority pollution management. These data will be beneficial to metal pollution control in PSS-vegetable system around industrial areas.

Comparative proteomic analysis of three major extracellular vesicle classes secreted from human primary and metastatic colorectal cancer cells: Exosomes, microparticles, and shed midbody remnants.

Cell-derived extracellular vesicles (EVs) are evolutionary-conserved secretory organelles that, based on their molecular composition, are important intercellular signaling regulators. At least three classes of circulating EVs are known based on mechanism of biogenesis: exosomes (sEVs/Exos), microparticles (lEVs/MPs), and shed midbody remnants (lEVs/sMB-Rs). sEVs/Exos are of endosomal pathway origin, microparticles (lEVs/MPs) from plasma membrane blebbing and shed midbody remnants (lEVs/sMB-Rs) arise from symmetric cytokinetic abscission. Here, we isolate sEVs/Exos, lEVs/MPs, and lEVs/sMB-Rs secreted from human isogenic primary (SW480) and metastatic (SW620) colorectal cancer (CRC) cell lines in milligram quantities for label-free MS/MS-based proteomic profiling. Purified EVs revealed selective composition packaging of exosomal protein markers in SW480/SW620-sEVs/Exos, metabolic enzymes in SW480/SW620-lEVs/MPs, while centralspindlin complex proteins, nucleoproteins, splicing factors, RNA granule proteins, translation-initiation factors, and mitochondrial proteins selectively traffic to SW480/SW620- lEVs/sMB-Rs. Collectively, we identify 39 human cancer-associated genes in EVs; 17 associated with SW480-EVs, 22 with SW620-EVs. We highlight oncogenic receptors/transporters selectively enriched in sEVs/Exos (EGFR/FAS in SW480-sEVs/Exos and MET, TGFBR2, ABCB1 in SW620-sEVs/Exos). Interestingly, MDK, STAT1, and TGM2 are selectively enriched in SW480-lEVs/sMB-Rs, and ADAM15 to SW620-lEVs/sMB-Rs. Our study reveals sEVs/Exos, lEVs/MPs, and lEVs/sMB-Rs have distinct protein signatures that open potential diagnostic avenues of distinct types of EVs for clinical utility.

SheddomeDB 2023: A Revision of an Ectodomain Shedding Database Based on a Comprehensive Literature Review and Online Resources.

Ectodomain shedding of membrane proteins is a proteolytic event involved in several biological phenomena, including inflammation, development, diseases, and cancer progression. Though ectodomain shedding is a post-translational modification that plays an important role in cellular regulation, this biological phenomenon is seriously underannotated in public protein databases. Given the importance of the shedding events, we conducted a comprehensive literature review for membrane protein shedding and constructed the database, SheddomeDB in 2017. In response to user feedback, novel shedding findings, more associated biomedical events, and the advance in web technology, we revised SheddomeDB to a new version, SheddomeDB 2023. The revised SheddomeDB 2023 includes 481 protein entries across seven species; all the content was manually verified and curated. The content of SheddomeDB 2023 mainly came from a comprehensive literature survey by our newly developed semiautomated screening tool. We also integrated verified and updated cleavage and secretome information from other databases into the revision. In addition, SheddomeDB 2023 features a graphical presentation of cleavage information and a user-friendly interface for searching and browsing entries in the database. This revised comprehensive database of ectodomain shedding is expected to benefit biomedical researchers across different disciplines.

Current concepts of microRNA-mediated regulatory mechanisms in human pulp tissue-derived stem cells: a snapshot in the regenerative dentistry.

One of the most studied class of non-coding RNAs is microRNAs (miRNAs) which regulate more than 60% of human genes. A network of miRNA gene interactions participates in stem cell self-renewal, proliferation, migration, apoptosis, immunomodulation, and differentiation. Human pulp tissue-derived stem cells (PSCs) are an attractive source of dental mesenchymal stem cells (MSCs) which comprise human dental pulp stem cells (hDPSCs) obtained from the dental pulp of permanent teeth and stem cells isolated from exfoliated deciduous teeth (SHEDs) that would be a therapeutic opportunity in stomatognathic system reconstruction and repair of other damaged tissues. The regenerative capacity of hDPSCs and SHEDs is mediated by osteogenic, odontogenic, myogenic, neurogenic, angiogenic differentiation, and immunomodulatory function. Multi-lineage differentiation of PSCs can be induced or inhibited by the interaction of miRNAs with their target genes. Manipulating the expression of functional miRNAs in PSCs by mimicking miRNAs or inhibiting miRNAs emerged as a therapeutic tool in the clinical translation. However, the effectiveness and safety of miRNA-based therapeutics, besides higher stability, biocompatibility, less off-target effects, and immunologic reactions, have received particular attention. This review aimed to comprehensively overview the molecular mechanisms underlying miRNA-modified PSCs as a futuristic therapeutic option in regenerative dentistry.

Stem cells from human exfoliated deciduous teeth rejuvenate the liver in naturally aged mice by improving ribosomal and mitochondrial proteins.

BACKGROUND AIMS: Aging is accompanied by a decline in cellular proteome homeostasis, mitochondrial, and metabolic function. Mesenchymal stromal cell (MSC) therapies have been reported to extend lifespan and delay some age-related pathologies, yet the anti-aging rate and mechanisms remain unclear. Here, we investigated the effects and mechanism by transplantation of stem cells from human exfoliated deciduous teeth (SHED) into the naturally aged mice model. METHODS: SHED were cultured in vitro and injected into mice by caudal vein. The in vivo imaging uncovered that SHED labeled by DiR dye mainly migrated to the liver, spleen, and lung organs of wild-type mice. As the main metabolic organ and SHED homing place, the liver was selected for proteomics and aging clock algorithm (LiverClock) analysis, which was constructed to estimate the proteomic pattern related to liver age state. RESULTS: After 6 months of continuous SHED injections, the liver proteomic pattern was reversed from senescent (∼30 months) to a youthful state (∼3 months), accompanied with upregulation of hepatocytes marker genes, anti-aging protein Klotho, a global improvement of liver functional pathways proteins, and a dramatic regulation of ribosomal and mitochondrial proteins, including upregulation of translation elongation and ribosome-sparing proteins Rpsa and Rplp0; elongation factors Eif4a1, Eef1b2, Eif5a; protein-folding chaperones Hsp90aa and Hspe1; ATP synthesis proteins Atp5b, Atp5o, Atp5j; and downregulation of most ribosomal proteins, suggesting that the proteome homeostasis destruction and mitochondria dysfunction in the aged mice liver might be relieved after SHED treatment. CONCLUSIONS: SHED treatment could dramatically relieve the senescent state of the aged liver, affect ribosome component proteins and upregulate the ribosomal biogenesis proteins in the aged mice liver. These results may help understand the improvements and mechanisms of SHED treatment in anti-aging.

SHED-derived exosomes improve the repair capacity and osteogenesis potential of hPDLCs.

OBJECTIVE: Exosomes secreted by stem cells are recognized as a critical component in tissue regeneration during stem cell-based therapy. Considering the limited sources and bone regeneration efficiency of human periodontal ligament cells (hPDLCs), we explored whether exosomes secreted by stem cells from human exfoliated deciduous teeth (SHED-exo) could improve the pluripotency and regenerative potential of hPDLCs. METHODS AND MATERIALS: In hPDLCs, cell proliferation, migration, cell cycle, apoptosis, and osteogenic differentiation were detected after cells were exposed to SHED-exo (SHED-exo group), blank (control group), or control supernatant without exo (Csup group), via CCK-8, scratch analysis, flow cytometric, real-time PCR, and so on. Exosomes sequencing was performed to compare and analyze miRNAs contented in SHED-exo and hPDLC-exo. RESULTS: As compared to control or Csup, SHED-exo significantly increased migration, apoptosis, and proliferation, promoted cell cycle transition from G1 to S phase in hPDLCs, and enhanced Runx2 expression and mineralization. In addition, it may be explained by the significant differences in miRNA contented in SHED-exo and hPDLC-exo. CONCLUSION: Exosomes from SHED can improve cell proliferation, migration, cell cycle, apoptosis, and osteogenic differentiation of hPDLCs, which highlights the therapeutic value of this bioactive component in the regeneration of periodontal tissues using hPDLCs in clinical practice.

Sodium citrate effectively used in shed mediastinal blood autotransfusion after cardiac surgery.

BACKGROUND: We used sodium citrate as an alternative anticoagulation agent to heparin in the procedure of autologous blood transfusion with patients with postoperative haemorrhage after CPB. The aim of study was to evaluate the efficacy and safety of sodium citrate used in shed mediastinal blood autotransfusion after cardiac surgery. METHODS: Ninety-three patients were divided into two groups in this study. In the control group, 52 patients' shed mediastinal blood was discarded. The reinfusion group consisted of 41 patients receiving a reinfusion of washed autologous red cells from shed mediastinal blood. Each 400 mL shed blood sample was anticoagulated by 140 mL of 1.6% diluted sodium citrate in the washing procedure using a blood recovery machine. Hemoglobin (Hb), hematocrit (Hct), and electrolyte concentrations in both the patients and shed mediastinal blood were measured before and after this procedure. RESULTS: The mean volume of autotransfused shed blood was 239.5 ± 54.6 mL.The Hct of the washed red cells was 56.8 ± 6.1%. Significantly, fewer units of allogeneic blood were required per patient in the reinfusion group at 24 h postoperatively (2.91 ± 1.34 vs 4.03 ± 0.19 U, p = 0.002). At 24 h postoperatively, Hb and Hct levels were higher in the reinfusion group than in the control group. The calcium ion concentration was very low in the shed mediastinal blood, 0.25 ± 0.08 mmol/L, and was lower after washing, 0.15 ± 0.04 mmol/L. CONCLUSIONS: Sodium citrate, as an alternative anticoagulant agent, can be used in autologous shed mediastinal blood transfusion after CPB cardiac surgery. This procedure can effectively reduce the amount of allogeneic blood for patients with haemorrhage.

From Teeth to Therapy: A Review of Therapeutic Potential within the Secretome of Stem Cells from Human Exfoliated Deciduous Teeth.

Stem cells derived from human exfoliated deciduous teeth (SHED) have emerged as an alternative stem cell source for cell therapy and regenerative medicine because they are readily available, pose fewer ethical concerns, and have low immunogenicity and tumourigenicity. SHED offer a number of advantages over other dental stem cells, including a high proliferation rate with the potential to differentiate into multiple developmental lineages. The therapeutic effects of SHED are mediated by multiple mechanisms, including immunomodulation, angiogenesis, neurogenesis, osteogenesis, and adipogenesis. In recent years, there is ample evidence that the mechanism of action of SHED is mainly due to its paracrine action, releasing a wide range of soluble factors such as cytokines, chemokines, and trophic factors (also known as 'secretome') into the local tissue microenvironment to promote tissue survival and recovery. This review provides an overview of the secretome derived from SHED and highlights the bioactive molecules involved in tissue regeneration and their potential applications in regenerative medicine.

Stem Cells Derived from Human Exfoliated Deciduous Teeth Functional Assessment: Exploring the Changes of Free Fatty Acids Composition during Cultivation.

The metabolic regulation of stemness is widely recognized as a crucial factor in determining the fate of stem cells. When transferred to a stimulating and nutrient-rich environment, mesenchymal stem cells (MSCs) undergo rapid proliferation, accompanied by a change in protein expression and a significant reconfiguration of central energy metabolism. This metabolic shift, from quiescence to metabolically active cells, can lead to an increase in the proportion of senescent cells and limit their regenerative potential. In this study, MSCs from human exfoliated deciduous teeth (SHEDs) were isolated and expanded in vitro for up to 10 passages. Immunophenotypic analysis, growth kinetics, in vitro plasticity, fatty acid content, and autophagic capacity were assessed throughout cultivation to evaluate the functional characteristics of SHEDs. Our findings revealed that SHEDs exhibit distinctive patterns of cell surface marker expression, possess high self-renewal capacity, and have a unique potential for neurogenic differentiation. Aged SHEDs exhibited lower proliferation rates, reduced potential for chondrogenic and osteogenic differentiation, an increasing capacity for adipogenic differentiation, and decreased autophagic potential. Prolonged cultivation of SHEDs resulted in changes in fatty acid composition, signaling a transition from anti-inflammatory to proinflammatory pathways. This underscores the intricate connection between metabolic regulation, stemness, and aging, crucial for optimizing therapeutic applications.

Itraconazole-loaded microemulsions: formulation, characterization, and dermal delivery using shed snakeskin as the model membrane.

Microemulsions (MEs) were developed for dermal delivery of 1% w/w itraconazole (ITZ). Solubility of ITZ in various oils was investigated and clove oil was selected as oil phase. Pseudoternary phase diagrams were constructed by titration method. The system containing clove oil as oil phase, Tween®80 as surfactant, and 1:1 mixture of water and polyethylene glycol 400 as aqueous phase provided the largest ME region. It was selected for the formulation development of ITZ-loaded MEs. Physicochemical stability was evaluated at 4 °C, room temperature (25 °C), and 45 °C for three months. In vitro permeation and retention studies were assessed using shed snakeskin as a model membrane. Antifungal activity was investigated by agar diffusion method. Results indicated that incorporation of ITZ in the selected MEs did not affect physical properties. Physicochemical data after storage periods revealed that the most suitable storage temperature was 4 °C. Skin permeation and retention data indicated that water-in-oil (w/o) ITZ-loaded MEs had superior dermal delivery of ITZ than oil-in-water (o/w) ITZ-loaded ME and ITZ-oily solution. Moreover, w/o ITZ-loaded MEs showed larger inhibition zones against C. albicans and T. rubrum than a commercial gel. Therefore, w/o ITZ-loaded MEs possibly provided effective dermal delivery and antifungal activity to treat superficial fungal infections.

Stemness maintenance of stem cells derived from human exfoliated deciduous teeth (SHED) in 3D spheroid formation through the TGF-ß/Smad signaling pathway.

Mesenchymal stem cells (MSCs) have shown great potential as important therapeutic tools for dental pulp tissue engineering, with the maintenance and enhancement of their stemness being crucial for successful therapeutic application in vivo and three-dimensional (3D) spheroid formation considered a reliable technique for enhancing their pluripotency. Human exfoliated deciduous tooth stem cells (SHED) were cultured in a low attachment plate to form aggregates for five days. Then, the resulting spheroids were analyzed for pluripotent marker expression, paracrine secretory function, proliferation, signaling pathways involved, and distribution of key proteins within the spheroids. The results indicated that 3D spheroid formation significantly increased the activation of the transforming growth factor beta (TGF-ß)/Smad signaling pathway and upregulated the secretion and mRNA expression levels of TGF-ß, which in turn enhanced the expression of pluripotency markers in SHED spheroids. The activation of the TGF-ß/Smad signaling pathway through 3D spheroid formation was found to preserve the stemness properties of SHED. Thus, understanding the mechanisms behind pluripotency maintenance of SHED culture through 3D spheroid formation could have implications for the therapeutic application of MSCs in regenerative medicine and tissue engineering.

Tegument Ultrastructure and Morphology of the Capsule Surrounding the Tetrathyridia of the Genus Mesocestoides Vaillant, 1863 in the Liver of the Root Vole.

The ultrastructure of the tegument of encapsulated tetrathyridia of the genus Mesocestoides Vaillant, 1863 (Cestoda, Cyclophyllidea, Mesocestoididae) from the liver of root voles Microtus oeconomus (Pallas, 1776) and the structure of the three-layered capsule surrounding them were studied for the first time. Several types of extracellular structures were noted on the surface of the tetrathyridia tegument: vesicles, fine granular material, and vacuoles. In addition, the phenomenon of shedding microtriches, which have expanded parts, was found. Host cells in contact with extracellular material show signs of destruction. A characteristic feature of the capsules surrounding the tetrathyridia is the reticular structure of the fibrous layer containing both native and degenerating inflammatory cells.

Metformin pre-treatment of stem cells from human exfoliated deciduous teeth promotes migration and angiogenesis of human umbilical vein endothelial cells for tissue engineering.

BACKGROUND AIMS: Stem cells from human exfoliated deciduous teeth (SHED) play a significant role in tissue engineering and regenerative medicine. Angiogenesis is crucial in tissue regeneration and a primary target of regenerative medicine. As a first-line anti-diabetic drug, metformin demonstrates numerous valuable impacts on stem cells. This study aimed to explore metformin's impact and mechanism of action on SHED-mediated angiogenesis. METHODS: First, cell proliferation; flow cytometry; osteogenic, adipogenic and chondrogenic induction; and proteomics analyses were conducted to explore the role of metformin in SHED. Subsequently, migration and tube formation assays were used to evaluate chemotaxis and angiogenesis enhancement by SHED pre-treated with metformin under co-culture conditions in vitro, and relative messenger RNA expression levels were determined by quantitative reverse transcription polymerase chain reaction. Finally, nude mice were used for in vivo tube formation assay, and sections were analyzed through immunohistochemistry staining with anti-human CD31 antibody. RESULTS: Metformin significantly promoted SHED proliferation as well as osteogenic, adipogenic and chondrogenic differentiation. Proteomics showed that metformin significantly upregulated 124 differentially abundant proteins involved in intracellular processes, including various proteins involved in cell migration and angiogenesis, such as MAPK1. The co-culture system demonstrated that SHED pre-treated with metformin significantly improved the migration and angiogenesis of human umbilical vein endothelial cells. In addition, SHED pre-treated with metformin possessed greater ability to promote angiogenesis in vivo. CONCLUSIONS: In summary, the authors' findings illustrate metformin's mechanism of action on SHED and confirm that SHED pre-treated with metformin exhibits a strong capacity for promoting angiogenesis. This helps in promoting the application of dental pulp-derived stem cells pre-treated with metformin in regeneration engineering.

Stem cells from human exfoliated deciduous teeth affect mitochondria and reverse cognitive decline in a senescence-accelerated mouse prone 8 model.

BACKGROUND AIMS: Stem cell therapy is a novel therapy being explored for AD. The molecular mechanism of its effect is still unclear. The authors investigated the effects and mechanism by injection of SHEDs into an AD mouse model. METHODS: SHEDs were cultured in vitro and injected into AD SAMP8 mice by caudal vein, and SHEDs labeled via synthetic dye showed in vivo migration to the head. The cognitive ability of SAMP8 mice was evaluated via Barnes maze and new object recognition. The pathological indicators of AD, including Tau, amyloid plaques and inflammatory factors, were examined at the protein or RNA level. Next, macro-proteomics analysis and weighted gene co-expression network analysis (WGCNA) based on protein groups and behavioral data were applied to discover the important gene cluster involved in the improvement of AD by SHEDs, which was further confirmed in an AD model in both mouse and cell lines. RESULTS: SHED treatment improved the cognitive ability and pathological symptoms of SAMP8 mice. Proteomics analysis indicated that these improvements were tightly related to the mitochondria, which was proved through examination of the shape and function of mitochondria both in vivo (SAMP8 brain) and in vitro (SH-SY5Y cells). Finally, the core targets of SHEDs in the mitochondrial pathway, Hook3, Mic13 and MIF, were screened out and confirmed in vivo. CONCLUSIONS: SHED treatment significantly relieved AD symptoms, improved cognitive ability and reversed memory loss in an AD mouse model, possibly through the recovery of dysfunctional mitochondria. These results raise the possibility that SHED may ease the symptoms of AD by targeting the mitochondria.

Hydrogel supplemented with human platelet lysate enhances multi-lineage differentiation of mesenchymal stem cells.

Stem cells from human exfoliated deciduous teeth (SHED) can be used as a potential clinical material. But the use of xenogeneic ingredients will increase the risk of zoonotic disease transmission. Human platelet lysate (HPL) is a potential surrogate and used in human cell expansion with reliability in clinical applications. In this study, we synthesized chitosan/gelatin/gellan gum hydrogel supplemented with HPL and investigated the effect of 3D culture for SHED. TMT-tagged proteomics was used to decipher the secretome protein profiles of SHEDs and a total of 3209 proteins were identified, of which 23 were up-regulated and 192 were down-regulated. The results showed that hydrogel supplemented with HPL promoted SHED proliferation. After induction, the hydrogel coating contributed to osteogenic differentiation, adipogenic differentiation and differentiation into neural-like cells of SHED. SHED encapsulated in a hydrogel promotes migration and angiogenesis of HUVEC. In conclusion, our research found that hydrogel supplemented with HPL can be used as a method for SHED in standardized production and can contribute to the clinical application of SHED in cell therapy.