Vitamin D Attenuates Fibrotic Properties of Fibrous Dysplasia-Derived Cells for the Transit towards Osteocytic Phenotype.

Fibrous dysplasia (FD) poses a therapeutic challenge due to the dysregulated extracellular matrix (ECM) accumulation within affected bone tissues. In this study, we investigate the therapeutic potential of 1,25-dihydroxyvitamin D3 (1,25(OH)2D3) in managing FD by examining its effects on FD-derived cells in vitro. Our findings demonstrate that 1,25(OH)2D3 treatment attenuates the pro-fibrotic phenotype of FD-derived cells by suppressing the expression of key pro-fibrotic markers and inhibiting cell proliferation and migration. Moreover, 1,25(OH)2D3 enhances mineralization by attenuating pre-osteoblastic cellular hyperactivity and promoting maturation towards an osteocytic phenotype. These results offer valuable insights into potential treatments for FD, highlighting the role of 1,25(OH)2D3 in modulating the pathological properties of FD-derived cells.

Patient-Derived Organoids Recapitulate Pathological Intrinsic and Phenotypic Features of Fibrous Dysplasia.

Fibrous dysplasia (FD) is a rare bone disorder characterized by the replacement of normal bone with benign fibro-osseous tissue. Developments in our understanding of the pathophysiology and treatment options are impeded by the lack of suitable research models. In this study, we developed an in vitro organotypic model capable of recapitulating key intrinsic and phenotypic properties of FD. Initially, transcriptomic profiling of individual cells isolated from patient lesional tissues unveiled intralesional molecular and cellular heterogeneity. Leveraging these insights, we established patient-derived organoids (PDOs) using primary cells obtained from patient FD lesions. Evaluation of PDOs demonstrated preservation of fibrosis-associated constituent cell types and transcriptional signatures observed in FD lesions. Additionally, PDOs retained distinct constellations of genomic and metabolic alterations characteristic of FD. Histological evaluation further corroborated the fidelity of PDOs in recapitulating important phenotypic features of FD that underscore their pathophysiological relevance. Our findings represent meaningful progress in the field, as they open up the possibility for in vitro modeling of rare bone lesions in a three-dimensional context and may signify the first step towards creating a personalized platform for research and therapeutic studies.

Cyclic High Negative-Pressure External Volume Expansion Reduces Daily Device Application Time With Similar Effects on Recipient Site Preparation in a Murine Model.

Introduction: Recipient site preparation using external volume expansion (EVE) increases graft survival in large-volume fat grafting. To improve patient compliance with using the device, we tested a new cyclic high negative-pressure (CHNP) mode that involves 1 h/day at -55 mm Hg, cycled between 1-second negative-pressure activation, followed by a 2-second deactivation period in an animal model. Material and Method: A miniaturized EVE device was applied to 30 8-week-old male Sprague-Dawley rats. The rats were assigned to 3 groups (no pressure for the control group, conventional -25 mm Hg for 8 h/day for conventional EVE, and CHNP mode for the CHNP group). After 28 days, micro-computed tomography was performed and skin biopsy specimens were obtained. Results: The CHNP group showed a 6.6-fold increase and the conventional EVE group showed a 4.4-fold increase in volume compared to the control group. Hematoxylin and eosin staining showed a similar increase in subcutaneous tissue thickness in both EVE groups, compared to the control group. Masson's trichome and proliferating cell nuclear antigen staining showed significantly higher collagen deposition and subdermal adipocytes in EVE groups. Immunohistochemistry against platelet endothelial cell adhesion molecule 1 showed 2.5- and 2.7-times higher vessel density in the conventional and CHNP EVE groups, respectively. There was no statistically significant difference in subcutaneous tissue thickness, collagen deposition, subdermal adipocyte proliferation, and vessel density between the 2 EVE groups. Conclusion: CHNP produced comparable results in recipient site preparation (subcutaneous tissue thickening and angiogenesis) compared to the conventional protocol, while markedly reducing the daily wear-time from 8 hours to 1 hour. Although further clinical data must be acquired, our new pressure setting seems promising and provides a more patient-friendly pre-expansion environment.

Introduction: La préparation du site receveur utilisant l'expansion de volume externe (EVE) augmente la survie d'une greffe dans une greffe de tissu adipeux de grand volume. Pour améliorer l'observance de l'utilisation du dispositif par le patient, nous avons testé un nouveau mode cyclique à forte pression négative (CHNP) qui implique 1 heure par jour à −55 mm Hg, dans un cycle entre une activation de pression négative 1-s suivie d'une période de désactivation de 2-s dans un modèle animal. Matériel et Méthode: Un dispositif EVE miniaturisé a été appliqué à 30 rats mâles Sprague-Dawley âgés de 8 semaines. Les rats ont été répartis en trois groupes (pas de pression dans le groupe témoin, pression conventionnelle de −25 mm Hg pendant 8 h/jour pour l'EVE conventionnelle et forte pression cyclique négative pour le groupe CHNP). Après 28 jours, une micro-tomodensitométrie (TDM) a été réalisée et des échantillons de biopsie de peau ont été prélevés. Résultats: Le groupe CHNP avait une augmentation de 6,6 fois, et le groupe d'EVE conventionnelle présentait une augmentation de 4,4 fois le volume comparativement au groupe contrôle. La coloration à l'hématoxyline-éosine a mis en évidence une augmentation similaire de l'épaisseur du tissu sous-cutané dans les 2 groupes EVE, par rapport au groupe contrôle. Le trichrome de Masson et la coloration pour l'antigène nucléaire de prolifération cellulaire (PCNA) ont montré un dépôt de collagène significativement plus important et des adipocytes sous-dermiques plus nombreux dans les groupes EVE. L'immunohistochimie contre les molécules d'adhésion-1 des cellules endothéliales d'origine plaquettaire a montré une densité vasculaire plus élevée de 2,5 fois et 2,7 fois dans, respectivement, les groupes EVE conventionnelle et EVE CHNP. Il n'y a pas eu de différence statistiquement significative concernant l'épaisseur du tissu sous-cutané, le dépôt de collagène, la prolifération des adipocytes sous-dermiques et la densité des vaisseaux sanguins entre les deux groupes EVE. Conclusion: La forte pression négative cyclique a obtenu des résultats comparables pour la préparation d'un site receveur (épaississement du tissu sous-cutané et angiogenèse) comparativement au protocole conventionnel, tout en ayant une durée de port quotidien nettement réduite de 8 heures à 1 heure. Des données cliniques supplémentaires doivent être obtenues, mais notre nouveau cadre de pression semble prometteur et offre un environnement préexpansion plus agréable pour le patient.

A Rare Skeletal Disorder, Fibrous Dysplasia: A Review of Its Pathogenesis and Therapeutic Prospects.

Fibrous dysplasia (FD) is a rare, non-hereditary skeletal disorder characterized by its chronic course of non-neoplastic fibrous tissue buildup in place of healthy bone. A myriad of factors have been associated with its onset and progression. Perturbation of cell-cell signaling networks and response outputs leading to disrupted building blocks, incoherent multi-level organization, and loss of rigid structural motifs in mineralized tissues are factors that have been identified to participate in FD induction. In more recent years, novel insights into the unique biology of FD are transforming our understandings of its pathology, natural discourse of the disease, and treatment prospects. Herein, we built upon existing knowledge with recent findings to review clinical, etiologic, and histological features of FD and discussed known and potential mechanisms underlying FD manifestations. Subsequently, we ended on a note of optimism by highlighting emerging therapeutic approaches aimed at either halting or ameliorating disease progression.

Plasminogen-derived peptide promotes adipogenic differentiation of preadipocytes in vitro and in vivo.

Soft tissue defects caused by adipose tissue loss can result in various conditions such as lipodystrophy in congenital diseases, trauma secondary to ageing, and mastectomy in breast cancer; fat grafting is commonly performed to restore these defects. Although various enrichment strategies have been studied, novel therapeutics that are cost-effective, safe, technologically easy to manufacture, and minimally invasive are required. In this study, we identified a novel peptide derived from plasminogen, named plasminogen-derived peptide-1 (PLP-1), which showed adipogenic differentiation potential and led to an increase in the expression levels of adiponectin, C1Q and collagen domain containing protein, fatty acid-binding protein 4, and CCAAT/enhancer-binding protein-alpha. In vivo experiments confirmed an increase in the rate of adipocyte differentiation and the expression levels of CD31 in the PLP-1-treated mice groups. These results suggested that PLP-1 plays an important role in promoting the differentiation of preadipocytes and may be useful for developing therapeutic approaches to treat adipose tissue defects.

Melanoma Detection by AFM Indentation of Histological Specimens.

Melanoma is visible unlike other types of cancer, but it is still challenging to diagnose correctly because of the difficulty in distinguishing between benign nevus and melanoma. We conducted a robust investigation of melanoma, identifying considerable differences in local elastic properties between nevus and melanoma tissues by using atomic force microscopy (AFM) indentation of histological specimens. Specifically, the histograms of the elastic modulus of melanoma displayed multimodal Gaussian distributions, exhibiting heterogeneous mechanical properties, in contrast with the unimodal distributions of elastic modulus in the benign nevus. We identified this notable signature was consistent regardless of blotch incidence by sex, age, anatomical site (e.g., thigh, calf, arm, eyelid, and cheek), or cancer stage (I, IV, and V). In addition, we found that the non-linearity of the force-distance curves for melanoma is increased compared to benign nevus. We believe that AFM indentation of histological specimens may technically complement conventional histopathological analysis for earlier and more precise melanoma detection.

Efficacy study of the new polycaprolactone thread compared with other commercialized threads in a murine model.

BACKGROUND: Polydioxanone (PDO) threads, poly-L-lactic acid (PLLA) threads, and polycaprolactone (PCL) threads have been used for lifting and antiaging purposes. The new PCL threads that have less residual monomer compared to the previous PCL are developed. AIMS: The efficacy of threads regarding collagen synthesis and wrinkle improvement was evaluated in vivo model. METHODS: In this study, threads were inserted into 30 six-week-old male SKH-1 hairless mice. One of four threads was implanted at either side of the spine of each mouse. Biopsy specimens obtained at 1, 4, and 8 weeks were examined using hematoxylin and eosin (H&E) and Herovici's stain. Additionally, immunoblot analysis was performed using primary antibody for collagen type III and transforming growth factor-ß (TGF-ß) and visualized by chemiluminescence and densitometric quantification. Finally, skin replicas were used to calculate total wrinkle area (mm2 ). RESULTS: Neocollagenesis was significantly increased by 50% in the new PCL and pre-existing PCL groups at 8 weeks (p value < 0.001). Additionally, new-PCL-implanted mice showed a significant increase in collagen type III and TGF-ß expressions at 8 weeks (p value < 0.001). The number of inflammatory cells was also increased in the skin of PCL-implanted mice at 8 weeks. Finally, wrinkles were reduced about 20% in the new PCL group at 8 weeks. CONCLUSIONS: The new PCL thread exhibited a superior skin rejuvenation effect. This suggests that the material processing technology can be applied not only to the thread but also to various products such as dermal filler and cosmetics.

Development of Poly(HEMA-Am) Polymer Hydrogel Filler for Soft Tissue Reconstruction by Facile Polymerization.

The number of breast reconstruction surgeries has been increasing due to the increase in mastectomies. Surgical implants (the standard polydimethylsiloxane (PDMS) implants) are widely used to reconstruct breast tissues, however, it can cause problems such as adverse immune reactions, fibrosis, rupture, and additional surgery. Hence, polymeric fillers have recently garnered increasing attention as strong alternatives for breast reconstruction materials. Polymeric fillers offer noninvasive methods of reconstruction, thereby reducing the possible adverse effects and simplifying the treatment. In this study, we synthesized a 2-hydroxylethylmethacrylate (HEMA) and acrylamide (Am) copolymer (Poly(HEMA-Am)) by redox polymerization to be used as a biocompatible filler material for breast reconstruction. The synthesized hydrogel swelled in phosphate buffered saline (PBS) shows an average modulus of 50 Pa, which is a characteristic similar to that of the standard dermal acrylamide filler. To investigate the biocompatibility and cytotoxicity of the Poly(HEMA-Am) hydrogel, we evaluated an in vitro cytotoxicity assay on human fibroblasts (hFBs) and human adipose-derived stem cells (hADSCs) with the hydrogel eluate, and confirmed a cell viability of over 80% of the cell viability with the Poly(HEMA-Am) hydrogel. These results suggest our polymeric hydrogel is a promising filler material in soft tissue augmentation including breast reconstruction.

KCl Mediates K(+) Channel-Activated Mitogen-Activated Protein Kinases Signaling in Wound Healing.

BACKGROUND: Wound healing is an interaction of a complex signaling cascade of cellular events, including inflammation, proliferation, and maturation. K(+) channels modulate the mitogen-activated protein kinase (MAPK) signaling pathway. Here, we investigated whether K(+) channel-activated MAPK signaling directs collagen synthesis and angiogenesis in wound healing. METHODS: The human skin fibroblast HS27 cell line was used to examine cell viability and collagen synthesis after potassium chloride (KCl) treatment by Cell Counting Kit-8 (CCK-8) and western blotting. To investigate whether K(+) ion channels function upstream of MAPK signaling, thus affecting collagen synthesis and angiogenesis, we examined alteration of MAPK expression after treatment with KCl (channel inhibitor), NS1619 (channel activator), or kinase inhibitors. To research the effect of KCl on angiogenesis, angiogenesis-related proteins such as thrombospondin 1 (TSP1), anti-angiogenic factor, basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF), pro-angiogenic factor were assayed by western blot. RESULTS: The viability of HS27 cells was not affected by 25 mM KCl. Collagen synthesis increased dependent on time and concentration of KCl exposure. The phosphorylations of MAPK proteins such as extracellular-signal-regulated kinase (ERK) and p38 increased about 2.5-3 fold in the KCl treatment cells and were inhibited by treatment of NS1619. TSP1 expression increased by 100%, bFGF expression decreased by 40%, and there is no significant differences in the VEGF level by KCl treatment, TSP1 was inhibited by NS1619 or kinase inhibitors. CONCLUSIONS: Our results suggest that KCl may function as a therapeutic agent for wound healing in the skin through MAPK signaling mediated by the K(+) ion channel.

Mitochondrial dysfunction of immortalized human adipose tissue-derived mesenchymal stromal cells from patients with Parkinson's disease.

Mitochondrial dysfunction in dopaminergic neurons of patients with idiopathic and familial Parkinson's disease (PD) is well known although the underlying mechanism is not clear. We established a homogeneous population of human adipose tissue-derived mesenchymal stromal cells (hAD-MSCs) from human adult patients with early-onset hereditary familial Parkin-defect PD as well as late-onset idiopathic PD by immortalizing cells with the hTERT gene to better understand the underlying mechanism of PD. The hAD-MSCs from patients with idiopathic PD were designated as "PD", from patients with Parkin-defect PD as "Parkin" and from patients with pituitary adenomas as "non-PD" in short. The pGRN145 plasmid containing hTERT was introduced to establish telomerase immortalized cells. The established hTERT-immortalized cell lines showed chromosomal aneuploidy sustained stably over two-years. The morphological study of mitochondria in the primary and immortalized hAD-MSCs showed that the mitochondria of the non-PD were normal; however, those of the PD and Parkin were gradually damaged. A striking decrease in mitochondrial complex I, II, and IV activities was observed in the hTERT-immortalized cells from the patients with idiopathic and Parkin-defect PD. Comparative Western blot analyses were performed to investigate the expressions of PD specific marker proteins in the hTERT-immortalized cell lines. This study suggests that the hTERT-immortalized hAD-MSC cell lines established from patients with idiopathic and familial Parkin-defect PD could be good cellular models to evaluate mitochondrial dysfunction to better understand the pathogenesis of PD and to develop early diagnostic markers and effective therapy targets for the treatment of PD.

The antioxidant Trolox helps recovery from the familial Parkinson's disease-specific mitochondrial deficits caused by PINK1- and DJ-1-deficiency in dopaminergic neuronal cells.

The nature of mitochondrial dysfunction in dopaminergic neurons in familial Parkinson's disease (PD) is unknown. We characterized the pathophenotypes of dopaminergic neuronal cells that were deficient in PINK1 or DJ-1, genes with mutations linked to familial PD. Both PINK1- and DJ-1-deficient dopaminergic neurons had the increased production of ROS, severe mitochondrial structural damages and complex I deficits. A striking decrease in complex IV activity was also prominent by the PINK1-deficiency. The complex I deficits were relatively PD-specific and were significantly improved by an antioxidant Trolox. These data suggest that mitochondrial deficits are severe in dopaminergic neurons in familial PD and antioxidant-mediated functional recovery is feasible.

DJ-1 mediates paraquat-induced dopaminergic neuronal cell death.

There are two causes of Parkinson's disease (PD): environmental insults and genetic mutations of PD-associated genes. Environmental insults and genetic mutations lead to mitochondrial dysfunction, and a combination of mitochondrial dysfunction and increased oxidative stress in dopaminergic neurons is thought to contribute to the pathogenesis of PD. Among the PD-associated genes, DJ-1 acts as a redox sensor for oxidative stress and has been also proposed to maintain mitochondrial complex I activity. To understand molecular functions of DJ-1 in the cell, we have generated DJ-1 null cells from the DJ-1(-/-) mouse embryos. Using these null cells, we investigated the susceptibility to an environmental toxin, paraquat, which is known to inhibit mitochondrial complex I. Interestingly, we found that DJ-1 null cells showed a resistance to paraquat-induced apoptosis, including reduced poly (ADP-ribose) polymerase and procaspase-3. Also DJ-1 null cells generated less superoxide than SN4741 cells by paraquat treatment. Consistent with the reduced paraquat sensitivity, DJ-1 null cells showed reduced complex I activity, which was partially rescued by ectopic DJ-I expression. In summary, our results suggest that DJ-1 is critical to maintain mitochondrial complex I and complex I could be a key target in interaction of paraquat toxicity and DJ-1 for giving rise to PD.

Up-regulation of Idh3alpha causes reduction of neuronal differentiation in PC12 cells.

The PC12 is the widely used cell line to study neuronal differentiation. We had extensively investigated the details of protein expression in differentiated PC12 cells by proteomic analysis. The cells were incubated at the presence of nerve growth factor. We had analyzed the expression changes in the differentiating PC12 cells by 2-dimensional electrophoresis and the identification of the proteins using MALDI-TOF MS. By comparing expression pattern in the time course, we identified the candidate genes which are associated with neuronal differentiation. Among these genes, we performed real-time PCR analysis to validate Idh3alpha expression by the time course. To identify the function of Idh3alpha in neuronal differentiation stage, the transfection of Idh3alpha to PC12 cells was performed. As a result, we proved that up-regulation of Idh3alpha causes reduction in neural differentiation of PC12 cells. Based on these data, we suggest that Idh3alpha plays a role to the neuronal differentiation.