Evaluation of the Treatment Effects of Conditioned Medium from Human Orbital Adipose-Derived Stem Cells in a Corneal Alkali Burn Rabbit Model.

Purpose: This study aimed to evaluate the effects of a new treatment-conditioned medium from human orbital adipose-derived stem cells (OASC-CM)-on corneal recovery after alkali burns in a rabbit model. Methods: The corneal alkali burn rabbit model was established and treated with OASC-CM, conditioned medium from human abdominal subcutaneous adipose-derived stem cells (ABASC-CM), and fresh control culture medium (con-CM) three times a day for 7 days, respectively. Subsequently, the treatment effects were evaluated and compared through clinical, histological, immunohistochemical, and cytokine evaluations. Results: Clinically, OASC-CM alleviated corneal opacity and edema and promoted recovery of corneal epithelium defect. Histologically and immunohistochemically, OASC-CM inhibited neovascularization, conjunctivalization, and immuno-inflammatory reaction, while promoting corneal regeneration and rearrangement. Increased secretion of interleukin-10 and inhibited protein levels of cluster of differentiation 45, interferon-γ, and tumor necrosis factor-α were observed in the alkali-burned cornea after OASC-CM treatment, which might be the relevant molecular mechanism. Conclusions: OASC-CM showed significant effects on the recovery of rabbit corneal alkali burns and eliminated immunological and ethical limitations, representing a new option for corneal wound treatment.

The Efficacy of Magnesium Sulfate (MgSO4) Wet Dressing in Reducing Eyelid Swelling and Bruising after Blepharoplasty: A Randomized, Controlled, and Observer-Blinded Assessment Study.

The purpose of this study was to evaluate the effects of wet dressing with 50% magnesium sulfate (MgSO4) solution on decreasing eyelid swelling and bruising after blepharoplasty. Fifty-eight patients (23 male and 35 female) who underwent bilateral blepharoplasty were enrolled in our randomized clinical trial. One side of the periorbital area (upper and lower eyelids) per patient received a wet dressing with 50% MgSO4 solution randomly, and the other side was cooled with an ice pack from the first postoperative day for two consecutive days (30 minutes per time and twice a day). The eyelid edema and ecchymosis were evaluated and classified using respective graded scales. Degrees of eyelid edema were similar after surgery in both groups (p > 0.05) and were significantly decreased with time. Compared with the cooled ones, less swelling was observed in the eyelids treated by MgSO4 wet compress on postoperative day 5 (p < 0.01). Both the incidence and area of ecchymosis were lower in the MgSO4 group than those in the cooling group (p < 0.01 and p < 0.05, respectively). Moreover, the majority of patients (39/58, 67.2%) indicated a preference for MgSO4 wet dressing over ice cooling. MgSO4 wet dressing can be conveniently applied to alleviate eyelid swelling and reduce recovery time after blepharoplasty.

Trends in Adipose-Derived Stem Cell-Conditioned Medium: A Bibliometric and Visualized Review.

Adipose-derived stem cell-conditioned medium (ADSC-CM) has been widely studied and used as a stem cell-based cell-free therapy. Due to the explosion of scientific publications in this field, it is difficult to review all relevant publications systematically, not mention quantitively. In this study, we combined bibliometrics with the conventional review method to summarize, analyze, and visualize the characteristics of nearly all published articles related to ADSC-CM using CiteSpace-a bibliometrics software. We applied this software to quantitively and vividly show (a) annual publications and citations; (b) distributions and co-occurrence networks of countries/regions, authors, journals, and institutions; (c) keyword co-occurrence networks and clusters in different time periods; (d) cocitation networks of references; and (e) ongoing challenges and new topics in ADSC-CM. Altogether, we found that ADSC-CM is at a hot stage with an increasing number of publications and citations, extensive and close scientific collaborations, and dense cocited networks. Impact statement To our best knowledge, it is the first bibliometric and visualized review in the field of adipose-derived stem cell-conditioned medium (ADSC-CM). This review systematically and quantitatively revealed the developments, challenges, and emerging hot spots of ADSC-CM, providing a panoramic view to assist researchers to decide the direction of their future study in the fields of ADSCs and CM derived from stem cells.

Anoikis patterns via machine learning strategy and experimental verification exhibit distinct prognostic and immune landscapes in melanoma.

BACKGROUND: Anoikis is a cell death programmed to eliminate dysfunctional or damaged cells induced by detachment from the extracellular matrix. Utilizing an anoikis-based risk stratification is anticipated to understand melanoma's prognostic and immune landscapes comprehensively. METHODS: Differential expression genes (DEGs) were analyzed between melanoma and normal skin tissues in The Cancer Genome Atlas (TCGA) and Genotype-Tissue Expression data sets. Next, least absolute shrinkage and selection operator, support vector machine-recursive feature elimination algorithm, and univariate and multivariate Cox analyses on the 308 DEGs were performed to build the prognostic signature in the TCGA-melanoma data set. Finally, the signature was validated in GSE65904 and GSE22155 data sets. NOTCH3, PIK3R2, and SOD2 were validated in our clinical samples by immunohistochemistry. RESULTS: The prognostic model for melanoma patients was developed utilizing ten hub anoikis-related genes. The overall survival (OS) of patients in the high-risk subgroup, which was classified by the optimal cutoff value, was remarkably shorter in the TCGA-melanoma, GSE65904, and GSE22155 data sets. Low-risk patients exhibited low immune cell infiltration and high expression of immunophenoscores and immune checkpoints. They also demonstrated increased sensitivity to various drugs, including dasatinib and dabrafenib. NOTCH3, PIK3R2, and SOD2 were notably associated with OS by univariate Cox analysis in the GSE65904 data set. The clinical melanoma samples showed remarkably higher protein expressions of NOTCH3 (P = 0.003) and PIK3R2 (P = 0.009) than the para-melanoma samples, while the SOD2 protein expression remained unchanged. CONCLUSIONS: In this study, we successfully established a prognostic anoikis-connected signature using machine learning. This model may aid in evaluating patient prognosis, clinical characteristics, and immune treatment modalities for melanoma.

Conditioned medium from the bone marrow mesenchymal stem cells modulates immune response via signal transduction and activator of transcription 6 signaling pathway in an allergic rhinitis mouse model.

BACKGROUND: Allergic rhinitis (AR) is a common immune disease of the nasal mucosa characterized with immunoglobulin E (IgE)-mediated allergic inflammation after exposure to allergens in susceptible population. Previous reports have demonstrated that the bone marrow mesenchymal stem cells (BMSCs) could reduce allergic inflammation. However, there is little knowledge about whether the culture supernatant of BMSCs (conditioned medium, CM) has similar anti- inflammatory potential in treating AR. OBJECTIVE: The study aimed to evaluate the immunoregulatory effects of conditioned medium derived from BMSCs (BMSC-CM) on allergic inflammation in an AR mouse model. MATERIAL AND METHODS: The AR murine model was induced by repeated sensitization and challenges with ovalbumin (OVA). Subsequently the allergic symptoms of AR mice, cytokine levels, the histopathological features of the nasal mucosa and T helper 1 (Th1) : T helper 2 (Th2) cells ratio were evaluated. RESULTS: Treatment with BMSC-CM was found as effective as BMSCs in reducing allergic symptoms and inhibiting eosinophilic infiltration in the nasal mucosa. After BMSC-CM or BMSCs administration, the OVA-specific IgE and interleukin 4 levels in serum decreased and interferon gamma level increased compared with AR mice treated with uncultured fresh medium. Flow cytometry analysis revealed a decrease in Th1:Th2 cells ratio after OVA-sensitization and the ratio was reversed by BMSC-CM and BMSCs treatments. Furthermore, the data revealed that BMSC-CM suppressed the production of signal transduction and activator of transcription 6 (STAT6) at messenger RNA and protein levels in the nasal mucosa. CONCLUSION: BMSC-CM could ameliorate allergic inflammation and regulate the balance of Th cells, and the underlying mechanism was closely related to STAT6 signaling pathway. The immunoregulatory effects of BMSCs could be achieved through paracrine function, and nasal dripping of BMSC-CM might be a novel approach for the treatment of AR.

Anti-Parkinson's Disease Activity of Sanghuangprous vaninii Extracts in the MPTP-Induced Zebrafish Model.

Parkinson's disease (PD) is a devastating disease of the central nervous system that occurs mainly in the elderly age group, affecting their quality of life. The PD pathogenesis is not yet fully understood and lacks the disease-modifying treatment strategies. Sanghuangprous vaninii (S. vaninii) is a perennial fungus with a plethora of pharmacological activities including anti-cancer and antioxidant activity and so on. However, no study till date has reported its neuroprotective effect against symptoms that are similar to PD in pre-clinical investigation. In the current study, we investigated anti-PD-like effects of S. vaninii mycelium extracts (SvMEs) on MPTP-induced PD in zebrafish. We observed that the loss of dopaminergic neurons and neurovascular reduction were reversed by using SvMEs in the zebrafish brain in a concentration-independent manner. Moreover, it also relieved locomotor impairments in MPTP-induced PD zebrafish. In addition, SvMEs exerted significant antioxidant activity in vitro, which was also demonstrated in vivo on ktr4:NTR-hKikGR zebrafish. Upon investigating the underlying mechanism, we found that SvMEs may alleviate oxidant stress and accelerate α-synuclein degradation and then alleviate PD-like symptoms. Antioxidant-related genes (sod1, gss, gpx4a, gclm, and cat) implied that the SvMEs exhibited anti-PD activity due to the antioxidation mechanism. Finally, upon analysis of chemical composition of SvMEs by liquid chromatography-mass spectrometry, we identified 10 compounds that are plausibly responsible for the anti-PD-like effect of SvMEs. On the limiting part, the finding of the study would have been more robust had we investigated the protein expression of genes related to PD and oxidative stress and compared the effects of SvMEs with any standard anti-PD therapy. Despite this, our results indicated that SvMEs possess anti-PD effects, indicating SvMEs as a potential candidate that is worth exploring further in this avenue.

Glabellar wrinkle correction by autologous fat transplantation and evidence of human adipose tissue regeneration in a nude mouse model.

BACKGROUND: Fat grafts are increasingly applied in augmenting soft tissue defects and correcting static wrinkles, but the outcome is always unpredictable because of high absorption rate. Perilipin1 (Plin1) and perilipin2 (Plin2), two perilipin family proteins on lipid droplets in adipocytes, are reported to be used as biomarkers to evaluate adipocyte regeneration in vivo. AIMS: The aim of this study was to assess the efficacy of glabellar wrinkle correction with autologous fat grafting and to observe human adipose regeneration in a nude mouse model. PATIENTS/METHODS: Ten patients with 16 moderate or severe glabellar wrinkles underwent wrinkle correction by subcutaneously injecting autologous lipoaspirates obtained from the abdomen. The injection dose was 0.05 ml per 5 mm. The aspirated adipose tissue (0.05 ml) was also injected under the scalps of nude mice. Fat grafts were explanted at 3, 7, 15, 30, and 120 days after transplantation and the dynamic cellular changes were evaluated by HE staining and immunostaining of Plin1 and Plin2. RESULTS: Among the sixteen wrinkle lines, thirteen were obviously improved 4 months after procedure. Eight of the ten patients were satisfied with their wrinkle correction effects. The fat grafts demonstrated a continuous changing process from degeneration to regeneration in the mouse model without significant absorption and necrosis. It was Plin1, not Plin2 that was expressed in mature adipocytes. After transplantation, Plin1 expression was lost in dead fat cells while Plin2 expression was activated in newly regenerated adipocytes. After 120 days, all the surviving adipocytes were negative for Plin2 but positive for Plin1 again. CONCLUSIONS: Micro-volume fat transplantation was an easy and safe method to improve glabellum wrinkle lines, and the regenerative process of human adipose tissue could be verified in the mouse model.

Perilipin2 is an Earlier Marker Than Perilipin1 for Identifying Adipocyte Regeneration in Fat Grafts.

BACKGROUND: Both perilipin1 (Plin1) and perilipin2 (Plin2) play a crucial role in regulating lipid droplet (LD) formation in fat cells. Plin2 is expressed early in the adipocyte differentiation process but is replaced by Plin1 after cell maturation. In free fat grafts, only a small number of adipocytes remain alive or are replaced by newly regenerated fat cells. It is known that Plin1-positive adipocytes participate in regeneration, but the characteristics of Plin2 expression during this process are still poorly understood. OBJECTIVES: The aim of this study was to investigate whether Plin2 is a more precise early marker for detecting adipocyte regeneration in fat grafts than Plin1. METHODS: Autologous fat tissue (120 mg) harvested from inguinal fat pads was injected under the scalps of C57 mice. Samples were explanted at days 3, 7, 15, and 30 after transplantation. Changes in sample size and weight were evaluated. Hematoxylin-eosin staining, real-time polymerase chain reaction, and immunostaining of Plin1 and Plin2 expression were performed. RESULTS: Plin1, but not Plin2, expression was detected in the freshly harvested fat, but the latter was activated after grafting. Newly regenerated Plin2-positive adipocytes increased from day 3 to day 7 and then declined, whereas the number of Plin1-positive fat cells decreased first and began to increase after day 15. The expression levels of Plin1 and Plin2 mRNA demonstrated similar changes over time. At day 30, adipocytes lost Plin2 expression and were positive for Plin1 again. CONCLUSIONS: Our experiments showed convincing evidence that Plin2 expression could be used to detect early adipocyte regeneration in grafted fat tissue.

Influence of Recipient Site on the Function and Survival of Fat Grafts.

BACKGROUND: Autologous fat grafting has become an increasingly common procedure for soft tissue augmentation throughout the body. However, the long-term outcome is always unpredictable because of inconsistent graft survival. Based on the "law of use and disuse," we speculate that the volume loss of fat grafts will occur when transferred into a site where there is less fat. The purpose of this study is to investigate the cause of high resorption rate from the perspective of fat function after transplantation. METHODS: Adipose aspirates obtained from routine liposuction were injected into the dorsal site of athymic mice, which have no subcutaneous fat layer. The fat grafts were explanted at days 7, 15, and 30 after transplantation. Changes in fat function were evaluated by measuring the adipocyte size and the expression level of adipose differentiation-related protein. RESULTS: After grafting, adipose tissue was replaced by fibrosis, inflammation, and vacuolar tissues gradually over time. The size of fat cells decreased sharply from day 0 to day 7, increased at day 15, and further declined at day 30. Adipose differentiation-related protein expression experienced a dramatic increase at day 7 and then continuously decreased until day 30. CONCLUSIONS: Assuming that the extrinsic factors influencing fat function and distribution remain stable, capabilities of the redistributed fat to handle free fatty acid and store lipid substance are reduced, leading to substantial tissue atrophy and volume decline after grafting.

Secretome from bone marrow mesenchymal stem cells: A promising, cell-free therapy for allergic rhinitis.

Allergic rhinitis (AR), characterized by the symptoms of sneezing, rhinorrhea, itchiness and nasal blockage, is a type I allergic disease of nasal mucosa, which is mainly mediated by IgE after exposure to allergens. At present, general drug therapy is limited to alleviating allergic symptoms but fails to regulate the allergic reaction; the recurrence of symptoms and the side effects of the drugs make many patients with AR resist treatments and bring serious impacts on the quality of life. Bone marrow mesenchymal stem cells (BMSCs) are a population of adult stem cells with multipotential differentiation capability, low immunogenicity, and immunoregulatory effects. The unique immunoregulatory properties of BMSCs make them hold great promise in the treatment of chronic inflammation and immune disorders through a paracrine mechanism of anti-inflammatory and anti-allergic effects. The stem cell secretome is defined as the set of molecules secreted to the extracellular space. The secretome such as conditioned media (CM) obtained from BMSCs contains various bioactive molecules and vesicular elements, which may act as therapeutic mediators to support their immunoregulatory effects. Therefore, we hypothesize that the BMSCs secretome may represent a promising treatment for AR by anti-allergic effects via the paracrine mechanism.

Identification and Characterization of Skeletal Muscle Stem Cells from Human Orbicularis Oculi Muscle.

Skeletal muscle stem cell (SMSC) transplantation has shown great therapeutical potential in repairing muscle loss and dysfunction, but the muscle acquisition is usually a traumatic procedure causing pain and morbidity to the donor. In this study, we investigated the feasibility of isolating SMSCs from human orbicularis oculi muscle (OOM), which is routinely removed and discarded during ophthalmic cosmetic surgeries. OOM fragments were harvested from 18 female healthy donors undergoing upper eyelid plasties. Plastic-adherent cells were isolated from the muscles using a two-step plating method combined with collagenase digestion. A total of 15 cell cultures were successfully established from the muscle samples. These adherent cells were positive for the specific markers of SMSCs and could be directed toward the osteogenic, adipogenic, chondrogenic, and myogenic phenotypes in the presence of lineage-specific inductive media. Moreover, after cultured in the myogenic inductive medium for 3 weeks, the muscle cells were injected into the tibialis anterior muscles of nude mice and the cell fate was detected using a DiI-labeling technique. In vivo myogenesis was evidenced by the expression of DiI fluorescence after cell transplantation. The donor cells could be found in the satellite cell position and incorporated into the host myofibers. Our results demonstrated that human OOM represents a novel source of myogenic precursors with stem cell-like properties, which may provide a foundation for the SMSC-based therapeutics of skeletal muscle diseases.

In Vitro and In Vivo Osteogenesis of Human Orbicularis Oculi Muscle-Derived Stem Cells.

BACKGROUND: Cell-based therapies for treating bone defects require a source of stem cells with osteogenic potential. There is evidence from pathologic ossification within muscles that human skeletal muscles contain osteogenic progenitor cells. However, muscle samples are usually acquired through a traumatic biopsy procedure which causes pain and morbidity to the donor. Herein, we identified a new alternative source of skeletal muscle stem cells (SMSCs) without conferring morbidity to donors. METHODS: Adherent cells isolated from human orbicularis oculi muscle (OOM) fragments, which are currently discarded during ophthalmic cosmetic surgeries, were obtained using a two-step plating method. The cell growth kinetics, immunophenotype and capabilities of in vitro multilineage differentiation were evaluated respectively. Moreover, the osteogenically-induced cells were transduced with GFP gene, loaded onto the porous ß-tricalcium phosphate (ß-TCP) bioceramics, and transplanted into the subcutaneous site of athymic mice. Ectopic bone formation was assessed and the cell fate in vivo was detected. RESULTS: OOM-derived cells were fibroblastic in shape, clonogenic in growth, and displayed phenotypic and behavioral characteristics similar to SMSCs. In particular, these cells could be induced into osteoblasts in vitro evidenced by the extracellular matrix calcification and enhanced alkaline phosphatase (ALP) activity and osteocalcin (OCN) production. New bone formation was found in the cell-loaded bioceramics 6 weeks after implantation. By using the GFP-labeling technique, these muscle cells were detected to participate in the process of ectopic osteogenesis in vivo. CONCLUSION: Our data suggest that human OOM tissue is a valuable and noninvasive resource for osteoprogenitor cells to be used in bone repair and regeneration.

Development of orbital adipose-derived stem cells as a model for studying the formation of baggy lower eyelids.

In old population, there is often a protrusion of the orbital fat pad underlying the skin in the lower eyelids, giving an aspect of palpable pouches. It is generally thought that orbital fat hyperplasia is the main contributing factor to the formation of baggy lower eyelids, and resection of excessive orbital fat pad is routinely performed during the eyelid cosmetic surgery. In our clinical study, however, it was revealed that the adipocytes in orbital fat tissue from older people became smaller compared to those from the young individuals. Based on this finding, we hypothesize that the orbital fat size may not increase, but decrease with age, and the declined fat depot volume is related to the reduced fat cell size and impaired differentiation of preadipocytes into fat cells. Adipose-derived stem cells (ASCs) are a population of postnatal stem cells residing in the fat tissue, capable of differentiating into preadipocytes and subsequently into mature fat cells throughout the lifespan. As preadipocytes are a substantial component of fat tissue and can greatly influence the fat composition and function, we speculate that orbital adipose-derived stem cells can be used as an excellent model to determine effects of aging on orbital fat. By evaluating the age-related changes in preadipocyte number, replication, and differentiation, we can reveal alterations in orbital fat cellularity and function with age, and investigate the relationship between orbital fat volume and the of baggy lower eyelid formation.

miR-203 Acts as a Tumor Suppressor Gene in Osteosarcoma by Regulating RAB22A.

microRNAs (miRNAs), small noncoding RNAs of 19-25 nt, play an important roles in the pathological processes of tumorigenesis. The object of this study was to study the expression and function of miR-203 and to found its target gene in osteosarcoma. In our study, we found the expression level of miR-203 was significantly downregulated in osteosarcoma cell lines and tissues. In addition, overexpression of miR-203 inhibited the osteosarcoma cell proliferation and migration and inhibited Mesenchymal-to-Epithelial reversion Transition (MErT). Moreover, we identified RAB22A as a direct target of miR-203 and RAB22A overexpression blocks the roles of miR-203 in osteosarcoma cell. Furthermore, we demonstrated that RAB22A expression was upregulated in human osteosarcoma cell lines and tissues. Take together, our results demonstrated that miR-203 act as a tumor suppressor miRNA through regulating RAB22A expression and suggested its involvement in osteosarcoma progression and carcinogenesis.

Autologously generated tissue-engineered bone flaps for reconstruction of large mandibular defects in an ovine model.

The reconstruction of large craniofacial defects remains a significant clinical challenge. The complex geometry of facial bone and the lack of suitable donor tissue often hinders successful repair. One strategy to address both of these difficulties is the development of an in vivo bioreactor, where a tissue flap of suitable geometry can be orthotopically grown within the same patient requiring reconstruction. Our group has previously designed such an approach using tissue chambers filled with morcellized bone autograft as a scaffold to autologously generate tissue with a predefined geometry. However, this approach still required donor tissue for filling the tissue chamber. With the recent advances in biodegradable synthetic bone graft materials, it may be possible to minimize this donor tissue by replacing it with synthetic ceramic particles. In addition, these flaps have not previously been transferred to a mandibular defect. In this study, we demonstrate the feasibility of transferring an autologously generated tissue-engineered vascularized bone flap to a mandibular defect in an ovine model, using either morcellized autograft or synthetic bone graft as scaffold material.

Adipose-derived stem cells alleviate osteoporosis by enhancing osteogenesis and inhibiting adipogenesis in a rabbit model.

BACKGROUND AIMS: Osteoporosis (OP) is characterized by a reduction in bone quality, which is associated with inadequacies in bone marrow mesenchymal stromal cells (BMSCs). As an alternative cell source to BMSCs, adipose-derived stem cells (ASCs) have been investigated for bone repair because of their osteogenic potential and self-renewal capability. Nevertheless, whether autologous ASCs can be used to promote bone regeneration under osteoporotic conditions has not been elucidated. METHODS: The OP rabbit model was established by means of bilateral ovariectomy (OVX). Both BMSCs and ASCs were harvested from OVX rabbits and expanded in vitro. The effects of osteogenic-induced ASCs on the in vitro adipogenic and osteogenic capabilities of BMSCs were evaluated. Autologous ASCs were then encapsulated by calcium alginate gel and transplanted into the distal femurs of OVX rabbits (n = 12). Hydrogel without loading cells was injected into the contralateral femurs as a control. Animals were killed for investigation at 12 weeks after transplantation. RESULTS: Osteogenic-induced ASCs were able to promote osteogenesis and inhibit adipogenesis of osteoporotic BMSCs through activation of the bone morphogenetic protein 2/bone morphogenetic protein receptor type IB signal pathway. Local bone mineral density began to increase at 8 weeks after ASC transplantation (P < 0.05). At 12 weeks, micro-computed tomography and histological evaluation revealed more new bone formation in the cell-treated femurs than in the control group (P < 0.05). CONCLUSIONS: This study demonstrated that ASCs could stimulate proliferation and osteogenic differentiation of BMSCs in vitro and enhance bone regeneration in vivo, which suggests that autologous osteogenic-induced ASCs might be useful to alleviate OP temporally.