BACKGROUND: Decellularized extracellular matrix (dECM) is a non-cellular scaffold with various functions in tissue engineering and regenerative medicine. Elastin is related to tissue elasticity and scarless wound healing, abundantly found in lung and blood vessel tissues. We studied the characteristics of blood vessel-derived dECM (VdECM) and its effect in wound healing. METHODS: VdECM was prepared from porcine blood vessel tissue. Weight percentages of elastin of VdECM and atelocollagen were analyzed. Migratory potential of VdECM was tested by scratch assay. VdECM in hydrogel form was microscopically examined, tested for fibroblast proliferation, and examined for L/D staining. Cytokine array of various growth factors in adipocyte-derived mesenchymal stem cell (ASC) media with VdECM was done. Animal wound model showed the wound healing effect of VdECM hydrogel in comparison to other topical agents. RESULTS: VdECM contained 6.7 times more elastin than atelocollagen per unit weight. Microscopic view of 0.35% VdECM hydrogel showed consistent distribution. Compared to 3% atelocollagen, 0.35% VdECM showed superior results in fibroblast migration. Fluorescent microscopic findings of L/D assay had highest percentage of cell survival in 1% VdECM compared to atelocollagen. Growth factor expression was drastically amplified when VdECM was added to ASC media. In the animal study model, epithelialization rate in the VdECM group was higher than that of control, oxytetracycline, and epidermal growth factor ointments. CONCLUSION VdECM contains a high ratio of elastin to collagen and amplifies expressions of many growth factors. It promotes fibroblast migration, proliferation, and survival, and epithelialization comparable to other topical agents.

BACKGROUND: The repair of large bone defects remains a significant challenge in clinical practice and requires bone grafts or substitute materials. In this study, we developed a unique hybrid bone scaffold comprising a three dimensional (3D)-printed metal plate for weight bearing and a biodegradable polymer tube serving as bone conduit. We assessed the long-term effect of the hybrid bone scaffold in repairing radial bone defects in a beagle model. METHODS: Bone defects were created surgically on the radial bone of three beagle dogs and individually-tailored scaffolds were used for reconstruction with or without injection of autologous bone and decellularized extracellular matrix (dECM). The repaired tissue was evaluated by X-ray, micro-computed tomography, and histological observation 6 months after surgery. The functional integrity of hybrid bone scaffold-mediated reconstructions was assessed by gait analysis. RESULTS: In vivo analysis showed that the hybrid bone scaffolds maintained the physical space and bone conductivity around the defect. New bone was formed adjacent to the scaffolds. Addition of autologous bone and dECM in the polymer tube improved healing by enhancing bone induction and osteoconduction. Furthermore, the beagles’ gait appeared normal by 4 months. CONCLUSION The future of bone healing and regeneration is closely related to advances in tissue engineering. Bone production using autologous bone and dECM loaded on 3D-printed hybrid bone scaffolds can successfully induce osteogenesis and provide mechanical force for functional bone regeneration, even in large bone defects.

BACKGROUND: Verapamil is used in the treatment of hypertension, angina pectoris, cardiac arrhythmia, hypertrophic scars, and keloids to block transmembrane calcium ion flux. Verapamil has antioxidant activity, which enhances the production of nitric oxide (NO). NO promotes the proliferation of fibroblasts, keratinocytes, endothelial cells, and epithelial cells during wound healing. In this study, we investigated the effect of verapamil and its antioxidant properties on the enhancement of acute wound healing via NO. METHODS: A full-thickness wound healing model was created on the rat dorsal with a silicone ring. The wound closure rate was estimated every 2 days for 14 days. A histological study was performed to evaluate wound healing.Immunofluorescence staining was analyzed for angiogenesis. The expressions of collagen type I (COL I), collagen type III (COL III), and vascular endothelial growth factor (VEGF) were assessed by Western blot. Real-time polymerase chain reaction (qRT-PCR) was performed to examine the expression of endothelial NO synthase and inducible NO synthase, which are related to antioxidant activity in the process of wound healing. RESULTS: The wound closure rate was faster in the verapamil group compared to the control and silicone groups.Histologic analysis revealed capillaries and stratum basale in the verapamil group. Immunofluorescence staining was shown vessel formation in the verapamil group. Western blot and qRT-PCR analysis revealed high expression levels of COL I, VEGF, eNOS, and FGF in the verapamil. CONCLUSION Verapamil’s antioxidant activity enhances NO production in acute wound healing. We suggest that verapamil can be used to promote acute wound healing.

BACKGROUND: Angiogenesis and vasculogenesis are essential processes for successful tissue regeneration in tissue engineering and regenerative medicine. The adipose-derived stromal vascular fraction (SVF) is not only a source of adipose stem cells (ASC) but also a suitable source of microvascular endothelial cells because it is a rich capillary network. So, we propose a new hypothesis for isolating adipose-derived human microvascular endothelial cells (HMVEC-A) from the SVF and developed a dual isolation system that isolates two cell types from one tissue.METHOD: To isolate HMVEC-A, we analyzed the supernatant discarded when ASC is isolated from the adipose-derived SVF. Based on this analysis, we assumed that the SVF adherent to the bottom of the culture plate was divided into two fractions: the stromal fraction as the ASC-rich fraction, and the vascular fraction (VF) as the endothelial cells-rich fraction floating in the culture supernatant. VF isolation was optimized and the efficiency was compared, and the endothelial cells characteristics of HMVEC-A were confirmed by flow cytometric analysis, immunocytochemistry (ICC), a DiI-acetylated low-density lipoprotein (DiI-Ac-LDL) uptake, and in vitro tube formation assay. RESULTS: Consistent with the hypothesis, we found a large population of HMVEC-A in the VF and isolated these HMVEC-A by our isolation method. Additionally, this method had higher yields and shorter doubling times than other endothelial cells isolation methods and showed typical morphological and phenotypic characteristics of endothelial cells. CONCLUSION Cells obtained by the method according to our hypothesis can be applied as a useful source for studies such as tissue-to-tissue networks, angiogenesis and tissue regeneration, patient-specific cell therapy, and organoid chips.

BACKGROUND: Verapamil is used in the treatment of hypertension, angina pectoris, cardiac arrhythmia, hypertrophic scars, and keloids to block transmembrane calcium ion flux. Verapamil has antioxidant activity, which enhances the production of nitric oxide (NO). NO promotes the proliferation of fibroblasts, keratinocytes, endothelial cells, and epithelial cells during wound healing. In this study, we investigated the effect of verapamil and its antioxidant properties on the enhancement of acute wound healing via NO. METHODS: A full-thickness wound healing model was created on the rat dorsal with a silicone ring. The wound closure rate was estimated every 2 days for 14 days. A histological study was performed to evaluate wound healing.Immunofluorescence staining was analyzed for angiogenesis. The expressions of collagen type I (COL I), collagen type III (COL III), and vascular endothelial growth factor (VEGF) were assessed by Western blot. Real-time polymerase chain reaction (qRT-PCR) was performed to examine the expression of endothelial NO synthase and inducible NO synthase, which are related to antioxidant activity in the process of wound healing. RESULTS: The wound closure rate was faster in the verapamil group compared to the control and silicone groups.Histologic analysis revealed capillaries and stratum basale in the verapamil group. Immunofluorescence staining was shown vessel formation in the verapamil group. Western blot and qRT-PCR analysis revealed high expression levels of COL I, VEGF, eNOS, and FGF in the verapamil. CONCLUSION Verapamil’s antioxidant activity enhances NO production in acute wound healing. We suggest that verapamil can be used to promote acute wound healing.

BACKGROUND: Angiogenesis and vasculogenesis are essential processes for successful tissue regeneration in tissue engineering and regenerative medicine. The adipose-derived stromal vascular fraction (SVF) is not only a source of adipose stem cells (ASC) but also a suitable source of microvascular endothelial cells because it is a rich capillary network. So, we propose a new hypothesis for isolating adipose-derived human microvascular endothelial cells (HMVEC-A) from the SVF and developed a dual isolation system that isolates two cell types from one tissue.METHOD: To isolate HMVEC-A, we analyzed the supernatant discarded when ASC is isolated from the adipose-derived SVF. Based on this analysis, we assumed that the SVF adherent to the bottom of the culture plate was divided into two fractions: the stromal fraction as the ASC-rich fraction, and the vascular fraction (VF) as the endothelial cells-rich fraction floating in the culture supernatant. VF isolation was optimized and the efficiency was compared, and the endothelial cells characteristics of HMVEC-A were confirmed by flow cytometric analysis, immunocytochemistry (ICC), a DiI-acetylated low-density lipoprotein (DiI-Ac-LDL) uptake, and in vitro tube formation assay. RESULTS: Consistent with the hypothesis, we found a large population of HMVEC-A in the VF and isolated these HMVEC-A by our isolation method. Additionally, this method had higher yields and shorter doubling times than other endothelial cells isolation methods and showed typical morphological and phenotypic characteristics of endothelial cells. CONCLUSION Cells obtained by the method according to our hypothesis can be applied as a useful source for studies such as tissue-to-tissue networks, angiogenesis and tissue regeneration, patient-specific cell therapy, and organoid chips.

Background: The stromal vascular fraction (SVF) isolated from adipose tissue, which contains stem cells as well as other cell types, has been applied in various research fields. Although different enzymatic concentrations and treatment durations have been applied to isolate the SVF, optimal conditions have not been established. Thus, we aimed to establish the optimal conditions for isolation of the SVF from adipose tissue by automated systems. Methods: The SVF was collected from removed adipose tissues of five donors during surgery. The SVF was treated with 0.1% or 0.2% collagenase type I for 20, 40, or 60 min. Then, colony forming unit (CFU) assays and flow cytometry were performed to characterize the adipose stem cells (ASCs). A cytokine array was used to investigate the correlation between colony-formation ability and the secretion of isolated ASCs. Results: Treatment with 0.1% collagenase type I for 60 min resulted in a higher SVF yield, whereas treatment with 0.1% collagenase for 40 min resulted in higher CFU values. In addition, expression of interleukin (IL)-6, IL-8, and monocyte chemoattractant protein-1 in the SVF was higher in the high-CFU group than in the low-CFU group. Conclusion The optimal conditions for isolation of the SVF from adipose tissue were treatment with 0.1% collagenase type I for 40 min. We identified the conditions required for efficient SVF isolation based on high CFU values, and our results will facilitate the development of automated systems.

Background: Nipple reconstruction is usually performed as a delayed procedure in patients with breast cancer who undergo skin-sparing mastectomy and breast reconstruction surgery using a deep inferior epigastric perforator (DIEP) flap. The authors designed this study to evaluate the utility of breast reconstruction based on a DIEP flap and immediate nipple reconstruction. Methods: A retrospective review was conducted of all patients who underwent breast reconstruction performed by a single plastic surgeon from October 2016 to June 2018. Through a questionnaire and chart review, we compared surgical results and complications in cases of single-stage nipple reconstruction after skin-sparing mastectomy (n=17) with patients who underwent delayed nipple reconstruction after skin-sparing mastectomy, modified radical mastectomy, or simple mastectomy (n=7). Results: In a subjective analysis using clinical photos, the immediate nipple reconstruction group had higher scores than their counterparts in an evaluation of the nipple-areolar complex (NAC) (NAC placement, 3.34 vs. 3.04; nipple projection, 3.05 vs. 3.03; nipple size, 3.30 vs. 3.29). No significant differences between the groups were found in terms of complications. Conclusions Simultaneous nipple reconstruction is a reliable surgical method with economic advantages. No differences were found in terms of outcomes and complications in comparison to delayed reconstruction. Therefore, surgeons can consider simultaneous nipple reconstruction without particular concerns about asymmetry or necrosis.

Salivary gland tumors usually appear in solitary mass in single salivary gland. The coexistence of tumors with different histological types occurring within a unilateral parotid gland is an extremely rare event. We experienced a case which two different types of malignant tumors developed at different time points in same gland; metachronous tumors. The second tumor was excised widely and reconstruction was performed by free tissue transfer. Sensory and motor nerve to the left cheek appeared to be intact, and circulation was adequate. This rare case was presented in this article.
Adenocarcinoma ; Cheek ; Neoplasms, Second Primary ; Parotid Gland ; Salivary Glands

BACKGROUND: Silica particles (SPs) induce cell proliferation and osteogenic differentiation. We reported that SPs in the scaffold induced early stage osteogenic differentiation. METHODS: A polycaprolactone (PCL) scaffold was fabricated with a 10 wt% SPs. The surface of PCL scaffold was coated with a 10 µg/mL collagen solution. Next, the scaffold was conjugated with 2 µM SPs, 2 µg/mL bone morphogenetic protein 2 (BMP2), or 2 µM BMP2-conjugated SPs (BCSPs). Green fluorescent protein-coupled BMP2 was applied to fabricate the scaffold. The fluorescence intensity was analyzed by confocal microscopy. The mRNA levels of the early osteogenic differentiation marker, alkaline phosphatase (ALP), were analyzed by real-time quantitative polymerase chain reaction. Levels of BMP2, RUNX2, ERK1/2, and AKT were assessed by western blotting. RESULTS: ALP mRNA levels were significantly higher in the BCSP-conjugated scaffold than in the other scaffolds. In the early stage of osteogenic differentiation, the protein levels of BMP2, RUNX2, ERK1/2, and AKT in cells were significantly higher in the BCSP-conjugated scaffold than in other scaffolds. Thus, the BCSP composite scaffold induced rapid osteogenic differentiation. CONCLUSION: These results suggest that BCSP composite can be used to promote early stage osteogenic differentiation and show promise as a material for use in scaffolds for bone regeneration.
Alkaline Phosphatase ; Blotting, Western ; Bone Morphogenetic Protein 2 ; Bone Morphogenetic Proteins ; Bone Regeneration ; Cell Proliferation ; Collagen ; Fluorescence ; Microscopy, Confocal ; Polymerase Chain Reaction ; RNA, Messenger ; Silicon Dioxide ; Stem Cells