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.

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.

Efficacy for Whitlockite for Augmenting Spinal Fusion.

Whitlockite (WH) is the second most abundant inorganic component of human bone, accounting for approximately 25% of bone tissue. This study investigated the role of WH in bone remodeling and formation in a mouse spinal fusion model. Specifically, morphology and composition analysis, tests of porosity and surface area, thermogravimetric analysis, an ion-release test, and a cell viability test were conducted to analyze the properties of bone substitutes. The MagOss group received WH, Group A received 100% beta-tricalcium phosphate (ß-TCP), Group B received 100% hydroxyapatite (HAp), Group C received 30% HAp/70% ß-TCP, and Group D received 60% HAp/40% ß-TCP (n = 10 each). All mice were sacrificed 6 weeks after implantation, and micro-CT, hematoxylin and eosin (HE) staining, and Masson trichome (MT) staining and immunohistochemistry were performed. The MagOss group showed more homogeneous and smaller grains, and nanopores (<500 nm) were found in only the MagOss group. On micro-CT, the MagOss group showed larger fusion mass and better graft incorporation into the decorticate mouse spine than other groups. In the in vivo experiment with HE staining, the MagOss group showed the highest new bone area (mean: decortication group, 9.50%; A, 15.08%; B, 15.70%; C, 14.76%; D, 14.70%; MagOss, 22.69%; p < 0.0001). In MT staining, the MagOss group demonstrated the highest new bone area (mean: decortication group, 15.62%; A, 21.41%; B, 22.86%; C, 23.07%; D, 22.47%; MagOss, 26.29%; p < 0.0001). In an immunohistochemical analysis for osteocalcin, osteopontin, and CD31, the MagOss group showed a higher positive area than other groups. WH showed comparable bone conductivity to HAp and ß-TCP and increased new bone formation. WH is likely to be used as an improved bone substitute with better bone conductivity than HAp and ß-TCP.

The Effects of Local Treatment of PTH(1-34) and Whitlockite and Hydroxyapatite Graft to the Calvarial Defect in a Rat Osteoporosis Model.

With the aging population, there is a rising incidence of senile diseases, notably osteoporosis, marked by fractures, prolonged recovery, and elevated mortality rates, underscoring the urgency for effective treatments. In this study, we applied the method of absorbing parathyroid hormone (PTH), a treatment for osteoporosis, into graft materials. Two types of graft materials with different properties, whitlockite (WH) and hydroxyapatite (HAP), were used. After forming calvarial defects in osteoporotic rats, WH and HAP grafts were implanted, with PTH applied directly to the graft sites. Micro-CT analysis was employed to assess bone regeneration, while tissue sections were stained to elucidate the regeneration process and bone cell dynamics. The results showed that bone regeneration was higher in the grafts that were actively degraded by osteoclasts in the early stage of regeneration. When PTH was applied, osteoclast activity increased, leading to enhanced bone regeneration. Furthermore, the activation of osteoclasts resulted in the penetration and formation of new bone within the degraded graft, which exhibited higher osseointegration. Therefore, for osteoporotic bone defects, bone grafts that can be easily degraded by osteoclasts are more suitable. Additionally, treatment with PTH can activate osteoclasts around the bone graft in the early stages of regeneration, inducing higher bone regeneration and improving osseointegration.

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 comparative study of regulatory perspectives on innovative medical devices in Korea and the United States.

INTRODUCTION: As the 4th Industrial Revolution era emerges, medical devices that apply technologies such as big data, artificial intelligence, and 3D printers are on the rise. In April 2019, Korea introduced the Act on Nurturing Medical Devices Industry and Supporting Innovative Medical Devices to shorten the market entry time by conducting step-by-step screening through the designation of innovative medical devices, priority screening systems, and special permission screening systems. AREAS COVERED: In this study, the Breakthrough Device Program of the United States, which has been implemented since 2016, and Korea's innovative medical device designation system were compared. EXPERT OPINION: Compared to the United States, Korea seems complicated because it has one more step in reviewing the innovative medical device group, but in terms of content, the two countries designate innovative medical devices on a similar basis. Neither country has established properly innovative medical device health insurance. Thus, a new insurance benefit scoring system based on actual evidence will have to be established. The role of experts in analyzing these data will be important and the voices of both innovative medical device manufacturers and medical field experts must be accepted.

Synergistic Effect of Whitlockite Scaffolds Combined with Alendronate to Promote Bone Regeneration.

BACKGROUND: Due to the increasing aging of society, the number of patients suffering from senile diseases is increasing. Patients suffering from osteoporosis, which is a representative senile disease, take a long time to recover from fractures, and the resulting mortality rate is very high. Alendronate (Ald), which is widely used as a treatment for osteoporosis, alleviates osteoporosis by inhibiting osteoclasts. In addition, whitlockite (WH) promotes the osteogenic differentiation of bone cells and improves bone regeneration. Therefore, we intended to bring about a synergistic effect by using these substances together. METHODS: In this study, a scaffold composed of gelatin/heparin was fabricated and applied to effectively use WH and Ald together. A scaffold was constructed using gelatin and heparin was used to effectively utilize the cations released from WH. In addition, it formed a porous structure for effective bone regeneration. In vitro and in vivo osteoclast inhibition, osteogenic differentiation, and bone regeneration were studied using the prepared scaffolds. RESULTS: The inhibition of osteoclast was much higher when WH and Ald were applied in combination rather than individually. The highest level of osteogenic differentiation was observed when both substances were applied simultaneously. In addition, when applied to bone regeneration through the mouse calvarial defect model, combined treatment showed excellent bone regeneration. CONCLUSION: Therefore, this study showed the synergistic effect of WH and Ald, and it is suggested that better bone regeneration is possible by applying this treatment to bones with fractures that are difficult to regenerate.

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.

The Effect of Whitlockite as an Osteoconductive Synthetic Bone Substitute Material in Animal Bony Defect Model.

This study aimed to evaluate the biomechanical properties in vitro and the bone regeneration of whitlockite (WH) compared with hydroxyapatite (HA) or ß-tricalcium phosphate (ß-TCP)-based material. We investigated the morphology and phase composition of the bone grafts using a scanning electron microscope and X-ray diffractometer patterns and tested the compressive strength. Four circular defects of 8 mm in diameter were created on the calvaria of twelve rabbits. One defect was left empty, and each of the other defects was filled with WH, HA, and ß-TCP. At 4 and 8 weeks, the specimens were harvested to evaluate for the new bone formation and the remaining bone grafts. Regarding the biomechanical properties, the three grafts had a similar micropore size, and WH showed nanopores. The compressive strength of WH was higher than HA and ß-TCP without statistical significance. The radiological and histomorphometric analyses demonstrated that the new bone formation was similar among the groups. The remaining bone graft of the WH group was greater than that of the HA and ß-TCP groups at 4 weeks (p < 0.05), and the total bone area of the WH, HA, and ß-TCP groups was greater than that of the other (p < 0.01). WH has excellent volumetric stability and osteoconductivity compared with HA and ß-TCP.

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.

Bioactive calcium phosphate materials and applications in bone regeneration.

BACKGROUND: Bone regeneration involves various complex biological processes. Many experiments have been performed using biomaterials in vivo and in vitro to promote and understand bone regeneration. Among the many biomaterials, calcium phosphates which exist in the natural bone have been conducted a number of studies because of its bone regenerative property. It can be directly contributed to bone regeneration process or assist in the use of other biomaterials. Therefore, it is widely used in many applications and has been continuously studied. MAINBODY: Calcium phosphate has been widely used in bone regeneration applications because it shows osteoconductive and in some cases osteoinductive features. The release of calcium and phosphorus ions regulates the activation of osteoblasts and osteoclasts to facilitate bone regeneration. The control of surface properties and porosity of calcium phosphate affects cell/protein adhesion and growth and regulates bone mineral formation. Properties affecting bioactivity vary depending on the types of calcium phosphates such as HAP, TCP and can be utilized in various applications because of differences in ion release, solubility, stability, and mechanical strength. In order to make use of these properties, different calcium phosphates have been used together or mixed with other materials to complement their disadvantages and to highlight their advantages. Calcium phosphate has been utilized to improve bone regeneration in ways such as increasing osteoconductivity for bone ingrowth, enhancing osteoinductivity for bone mineralization with ion release control, and encapsulating drugs or growth factors. CONCLUSION: Calcium phosphate has been used for bone regeneration in various forms such as coating, cement and scaffold based on its unique bioactive properties and bone regeneration effectiveness. Additionally, several studies have been actively carried out to improve the efficacy of calcium phosphate in combination with various healing agents. By summarizing the properties of calcium phosphate and its research direction, we hope that calcium phosphate can contribute to the clinical treatment approach for bone defect and disease.

Gene discovery analysis from mouse embryonic stem cells based on time course microarray data.

An embryonic stem cell is a powerful tool for investigation of early development in vitro. The study of embryonic stem cell mediated neuronal differentiation allows for improved understanding of the mechanisms involved in embryonic neuronal development. We investigated expression profile changes using time course cDNA microarray to identify clues for the signaling network of neuronal differentiation. For the short time course microarray data, pattern analysis based on the quadratic regression method is an effective approach for identification and classification of a variety of expressed genes that have biological relevance. We studied the expression patterns, at each of 5 stages, after neuronal induction at the mRNA level of embryonic stem cells using the quadratic regression method for pattern analysis. As a result, a total of 316 genes (3.1%) including 166 (1.7%) informative genes in 8 possible expression patterns were identified by pattern analysis. Among the selected genes associated with neurological system, all three genes showing linearly increasing pattern over time, and one gene showing decreasing pattern over time, were verified by RT-PCR. Therefore, an increase in gene expression over time, in a linear pattern, may be associated with embryonic development. The genes: Tcfap2c, Ttr, Wnt3a, Btg2 and Foxk1 detected by pattern analysis, and verified by RT-PCR simultaneously, may be candidate markers associated with the development of the nervous system. Our study shows that pattern analysis, using the quadratic regression method, is very useful for investigation of time course cDNA microarray data. The pattern analysis used in this study has biological significance for the study of embryonic stem cells.

Proteomic analysis of methamphetamine-induced reinforcement processes within the mesolimbic dopamine system.

ABSTRACT Methamphetamine (MAP) is a commonly used, addictive drug, and a powerful stimulant that dramatically affects the central nervous system. In this study, we used the conditioned place preference (CPP) paradigm in order to study the reinforcing properties of MAP and the herewith associated changes in proteins within the mesolimbic dopamine system. A CPP was induced by MAP after three intermittent intraperitoneal injections (1 mg/kg) in rats and protein profiles in the nucleus accumbens, striatum, prefrontal cortex, cingulate cortex and hippocampus were compared with a saline-treated control group. In addition, a group of animals was run through extinction and protein profiles were compared with a non-extinguished group. Protein screening was conducted using two-dimensional electrophoresis analysis which identified 27 proteins in the group that showed MAP-induced CPP. Some of the proteins were confirmed by Western lot analysis. Identified proteins had functions related to the cytoskeleton, transport/endocytosis or exocytosis (e.g. profilin-2 and syntaxin-binding protein), and signal transduction, among others.

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.

A new method of fabricating a blend scaffold using an indirect three-dimensional printing technique.

Due to its simplicity and effectiveness, the physical blending of polymers is considered to be a practical strategy for developing a versatile scaffold having desirable mechanical and biochemical properties. In the present work, an indirect three-dimensional (i3D) printing technique was proposed to fabricate a 3D free-form scaffold using a blend of immiscible materials, such as polycaprolactone (PCL) and gelatin. The i3D printing technique includes 3D printing of a mold and a sacrificial molding process. PCL/chloroform and gelatin/water were physically mixed to prepare the blend solution, which was subsequently injected into the cavity of a 3D printed mold. After solvent removal and gelatin cross-linking, the mold was dissolved to obtain a PCL-gelatin (PG) scaffold, with a specific 3D structure. Scanning electron microscopy and Fourier transform infrared spectroscopy analysis indicated that PCL masses and gelatin fibers in the PG scaffold homogenously coexisted without chemical bonding. Compression tests confirmed that gelatin incorporation into the PCL enhanced its mechanical flexibility and softness, to the point of being suitable for soft-tissue engineering, as opposed to pure PCL. Human adipose-derived stem cells, cultured on a PG scaffold, exhibited enhanced in vitro chondrogenic differentiation and tissue formation, compared with those on a PCL scaffold. The i3D printing technique can be used to blend a variety of materials, facilitating 3D scaffold fabrication for specific tissue regeneration. Furthermore, this convenient and versatile technique may lead to wider application of 3D printing in tissue engineering.

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.

Neuronal autophagy and neurodegenerative diseases.

Autophagy is a dynamic cellular pathway involved in the turnover of proteins, protein complexes, and organelles through lysosomal degradation. The integrity of postmitotic neurons is heavily dependent on high basal autophagy compared to non-neuronal cells as misfolded proteins and damaged organelles cannot be diluted through cell division. Moreover, neurons contain the specialized structures for intercellular communication, such as axons, dendrites and synapses, which require the reciprocal transport of proteins, organelles and autophagosomes over significant distances from the soma. Defects in autophagy affect the intercellular communication and subsequently, contributing to neurodegeneration. The presence of abnormal autophagic activity is frequently observed in selective neuronal populations afflicted in common neurodegenerative diseases, such as Alzheimer's disease, Parkinson's disease, Huntington's disease and amyotrophic lateral sclerosis. These observations have provoked controversy regarding whether the increase in autophagosomes observed in the degenerating neurons play a protective role or instead contribute to pathogenic neuronal cell death. It is still unknown what factors may determine whether active autophagy is beneficial or pathogenic during neurodegeneration. In this review, we consider both the normal and pathophysiological roles of neuronal autophagy and its potential therapeutic implications for common neurodegenerative diseases.