Micronized cellular adipose matrix purified with a bladed connector contains abundant functional adipose stem cells.

INTRODUCTION: A specialized device equipped with a sharp blade filter has been developed to enable more efficient purification of a micronized cellular adipose matrix (MCAM) containing stem cells. The aim of this study is to compare the characteristics and functions of the population of stromal cells (mSVF) and cultured cells (mASCs) purified using this device with those of cSVF and cASCs obtained through conventional enzymatic purification. METHODS: Cell viability, proliferation capacity and yield were assessed. Characterization of stem cell potency was performed by analyzing cell surface markers including CD34, a marker of activated adipose-derived stem cells. The trilineage differentiation potential was evaluated using RT-PCR and histology. RESULTS: The yield rate of mSVF obtained from MCAM was significantly higher than that with the conventional method, although use of the device resulted in a slight decrease in cell viability. After culture, mASCs exhibited a remarkable clonogenic potential and significantly higher cell proliferation potential than cASCs. The mASCs also displayed a distinct pattern of ASC cell surface markers, increased expression of genes related to CD34, high pluripotency, and a high trilineage differentiation ability. CONCLUSION: The specialized device enhanced the yield of SVF and produced cells with high proliferation rates and characteristics that include expression of stem cell markers.

Optimizing mesenchymal stem cell extracellular vesicles for chronic wound healing: Bioengineering, standardization, and safety.

Chronic wounds represent a significant global burden, afflicting millions with debilitating complications. Despite standard care, impaired healing persists due to factors like persistent inflammation and impaired tissue regeneration. Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) offer an innovative regenerative medicine approach, delivering stem cell-derived therapeutic cargo in engineered nanoscale delivery systems. This review examines pioneering bioengineering strategies to engineer MSC-EVs into precision nanotherapeutics for chronic wounds. Emerging technologies like CRISPR gene editing, microfluidic manufacturing, and biomimetic delivery systems are highlighted for their potential to enhance MSC-EV targeting, optimize therapeutic cargo enrichment, and ensure consistent clinical-grade production. However, key hurdles remain, including batch variability, rigorous safety assessment for potential tumorigenicity, immunogenicity, and biodistribution profiling. Crucially, collaborative frameworks harmonizing regulatory science with bioengineering and patient advocacy hold the key to expediting global clinical translation. By overcoming these challenges, engineered MSC-EVs could catalyze a new era of off-the-shelf regenerative therapies, restoring hope and healing for millions afflicted by non-healing wounds.

Chronic Pain Following Breast Reconstruction: A Scoping Review.

BACKGROUND: Breast cancer survival rates have increased significantly, underscoring the importance of enhancing long-term health-related quality of life. Breast reconstruction following mastectomy has emerged as a common approach that contributes to improved health-related quality of life. Nonetheless, chronic pain following breast reconstruction is a prevalent issue that has a negative impact on overall well-being. METHODS: To examine recent findings on chronic pain after breast reconstruction and progress in pain management, we performed a review of the literature through independent searches using the MEDLINE database within NIH National Library of Medicine PubMed. RESULTS: The review suggested that autologous reconstruction causes chronic postsurgical pain, especially at specific donor sites, whereas implant-based reconstruction does not seem to increase the risk of chronic pain. Moreover, certain operational and patient factors are also associated with chronic pain. Appropriate pain management can reduce chronic pain and prevent the transition from acute to chronic pain. CONCLUSION: This scoping review evaluated the characteristics of long-term chronic pain after breast reconstruction. The findings provide patients with important treatment information and will assist with their decision on their preferred treatment.

Pain-relieving Effects of Autologous Fat Grafting in Breast Cancer Surgery: A Scoping Review.

Background: Chronic pain is relatively common after breast cancer surgery, including breast reconstruction. Autologous fat grafting (AFG) has gained attention as a novel method for breast reconstruction, and recent clinical studies have also shown effects of AFG on alleviation of chronic pain after breast cancer surgery. Our objective was to conduct a scoping review of studies that have examined these effects with clearly defined clinical outcomes. Methods: A literature search was conducted using three databases: PubMed, MEDLINE, and Google Scholar, following PRISMA guidelines and the Arkesy and O'Malley framework. The search focused on clinical studies of the effects of AFG on chronic pain after breast cancer surgery. All studies reporting functional outcomes, return to work, and secondary surgery in a repeat operation were identified. Results: Of the 148 studies identified in the search, 11 studies with a total of 684 patients were included in the review. The average volume of fat grafted was approximately 128 mL over an average of 1.6 sessions. The most common time point for assessment was 1 year post-AFG. In all studies with an evidence level of 3 or lower, AFG showed positive results in alleviating pain after breast cancer surgery. However, one of the three randomized controlled trials did not show clinically significant effects. Conclusions: Most of the studies examined in this review suggested pain-relieving effects of AFG. However, there was one randomized controlled trial in which these effects were not confirmed, indicating a need for further accumulation of cases and performance of new, well-designed randomized controlled trials.

A novel surgical correction using a combination of wave-like incision and Z-axis Z-plasty for congenital constriction band syndrome.

BACKGROUND: A congenital constriction band (CCB) is a relatively common anomaly among limb malformations. However, the number of cases treated at a given center is limited and differences in techniques used by surgeons at each facility have a significant impact on the treatment outcomes. Several surgical methods have been reported, but a standard technique that yields consistent satisfactory results is still needed. Here, we introduce a novel technique for the treatment of CCB syndrome that uses a combination of wave-like skin incision and Z-axis Z-plasty. PATIENTS AND METHODS: A wave-like incision was used for skin incision with the goals of achieving an accordion effect and a less noticeable post-operative scar. After the fibrous constriction band was completely excised, a Z-plasty of sufficient size in the Z-axis direction was performed on the dermis-inclusive adipose tissue. For over 10 years, this technique was applied to 11 sites in 6 patients. A retrospective study of the characteristics of these cases was performed, including age at surgery, gender, type of deformity, degree of constriction, types of examination, number of surgeries, and post-operative outcomes. RESULTS: In all patients, a normal contour of the limbs was achieved and the hourglass-like deformity caused by the constriction band was satisfactorily improved. No additional corrective surgeries were needed, there was no abnormal growth of the treated limbs, and scarring was aesthetically acceptable in all cases. CONCLUSION: The novel technique described here uses deep subcutaneous fat to correct the hourglass-like deformity and restores a normal limb contour, while the wave-like skin incision minimizes post-operative scarring.

Multivariate Analysis of Risk Factors for Complications in Pediatric Tissue Expansion.

BACKGROUND: Tissue expanders represent one of the main surgical options for skin reconstruction in cases of tumors, traumalike burn injury, scar contracture, and alopecia. However, the tissue expander device is also associated with complications such as infection and extrusion. The aim of this study was to analyze risk factors for major complications of use of tissue expanders in pediatric patients using multivariate analysis. METHODS: A retrospective, single-center observational study was performed over 10 years in pediatric patients who were treated with tissue expanders for tumors, nevus, scars, burn reconstruction, and alopecia from April 2012 to March 2022. The primary outcome was overall complications per operation and expander, including infection and extrusion. Ten predictor variables were included as risk factors based on previous studies and as new factors considered important from clinical experience. Univariate and multivariate logistic regression analyses were performed to identify risk factors for major complications such as expander infection or extrusion. RESULTS: The study included 44 patients who underwent 92 operations using 238 tissue expanders. The overall complication rate per expander was 14.3%. Univariate logistic regression analysis identified associations of younger age, number of expanders used per operation, history of infection, and tissue expander locations with a higher complication rate. In multivariate logistic regression analysis, younger age (odds ratio, 1.14; P = 0.043) was associated with a high likelihood of expander complications. CONCLUSIONS: Younger age is an independent risk factor for tissue expander complications in pediatric patients. This factor should be considered in preoperative planning and discussions with the patient's family.

Improving stability of human three dimensional skin equivalents using plasma surface treatment.

In developing three-dimensional (3D) human skin equivalents (HSEs), preventing dermis and epidermis layer distortion due to the contraction of hydrogels by fibroblasts is a challenging issue. Previously, a fabrication method of HSEs was tested using a modified solid scaffold or a hydrogel matrix in combination with the natural polymer coated onto the tissue culture surface, but the obtained HSEs exhibited skin layer contraction and loss of the skin integrity and barrier functions. In this study, we investigated the method of HSE fabrication that enhances the stability of the skin model by using surface plasma treatment. The results showed that plasma treatment of the tissue culture surface prevented dermal layer shrinkage of HSEs, in contrast to the HSE fabrication using fibronectin coating. The HSEs from plasma-treated surface showed significantly higher transepithelial electrical resistance compared to the fibronectin-coated model. They also expressed markers of epidermal differentiation (keratin 10, keratin 14 and loricrin), epidermal tight junctions (claudin 1 and zonula occludens-1), and extracellular matrix proteins (collagen IV), and exhibited morphological characteristics of the primary human skins. Taken together, the use of plasma surface treatment significantly improves the stability of 3D HSEs with well-defined dermis and epidermis layers and enhanced skin integrity and the barrier functions.

Design, synthesis, and biological evaluation of phenylcyclopropylamine-entinostat conjugates that selectively target cancer cells.

Small molecule-based selective cancer cell-targeting can be a desirable anticancer therapeutic strategy. Aiming to discover such small molecules, we previously developed phenylcyclopropylamine (PCPA)-drug conjugates (PDCs) that selectively release anticancer agents in cancer cells where lysine-specific demethylase 1 (LSD1) is overexpressed. In this work, we designed PCPA-entinostat conjugates for selective cancer cell targeting. PCPA-entinostat conjugate 12 with a 4-oxybenzyl group linker released entinostat in the presence of LSD1 in in vitro assays and selectively inhibited the growth of cancer cells in preference to normal cells, suggesting the potential of PCPA-entinostat conjugates as novel anticancer drug delivery small molecules.

Evaluation of adipogenesis over time using a novel bioabsorbable implant without the addition of exogenous cells or growth factors.

Background: Breast reconstruction is crucial for patients who have undergone mastectomy for breast cancer. Our bioabsorbable implants comprising an outer poly-l-lactic acid mesh and an inner component filled with collagen sponge promote and retain adipogenesis in vivo without the addition of exogenous cells or growth factors. In this study, we evaluated adipogenesis over time histologically and at the gene expression level using this implant in a rodent model. Methods: The implants were inserted in the inguinal and dorsal regions of the animals. At 1, 3, 6, and 12 months post-operation, the weight, volume, and histological assessment of all newly formed tissue were performed. We analyzed the formation of new adipose tissue using multiphoton microscopy and RNA sequencing. Results: Both in the inguinal and dorsal regions, adipose tissue began to form 1 month post-operation in the peripheral area. Angiogenesis into implants was observed until 3 months. At 6 months, microvessels matured and the amount of newly generated adipose tissue peaked and was uniformly distributed inside implants. The amount of newly generated adipose tissue decreased from 6 to 12 months but at 12 months, adipose tissue was equivalent to the native tissue histologically and in terms of gene expression. Conclusions: Our bioabsorbable implants could induce normal adipogenesis into the implants after subcutaneous implantation. Our implants can serve as a novel and safe material for breast reconstruction without requiring exogenous cells or growth factors.

An Effective Approach to Collecting Lymph Vessels Using a Small Diameter Wire for Lymphaticovenous Anastomosis in Upper Limb Lymphedema.

Lymphaticovenous anastomosis (LVA), which involves connecting lymphatic vessels to veins under a microscope, is widely performed around the world as a surgical treatment for lymphedema due to its relatively low patient burden and consistent efficacy. One of the crucial points of LVA is identifying suitable lymphatic vessels from among many lymphatic vessels and connecting them at the most effective site, but in practice, this is not easy to do. To overcome this issue, we have been able to effectively drain lymph by connecting lymphatic vessels to veins just before the occluded site using a lymphatic wire that was just recently developed in Japan. The device guides the subcutaneous deep collecting lymphatic vessels from the peripheral relatively superficial lymphatic vessels. While this special wire is typically adapted in the lower limbs, we have confirmed its effectiveness in upper limb lymphedema as well. Overall, this approach shows promise for improving the accuracy and success rates of LVA procedures, which can have significant benefits for patients with upper limb lymphedema. In this article, we share our experience using lymphatic wire for upper limb lymphedema cases.

Induction of MYCN-amplified neuroblastoma differentiation through NMYC suppression using PPAR-γ antagonist.

Neuroblastomas are the most common extracranial solid tumors in children and have a unique feature of neuronal differentiation. Peroxisome proliferator-activated receptor (PPAR)-γ is reported to have neuroprotective effects in addition to having antitumor effects in various cancers. Thus, we aimed to clarify the role of PPAR-γ agonist and antagonist in malignant neuroblastomas, which also possess neuronal features. In MYCN-amplified neuroblastoma CHP212 cells, treatment with the PPAR-γ antagonist GW9662 induced growth inhibition in a dose-dependent manner. In addition, the PPAR-γ antagonist treatment changed cell morphology with increasing expression of the neuronal differentiation marker tubulin beta 3 (TUBB3) and induced G1 phase arrest and apoptosis in MYCN-amplified neuroblastoma. Notably, the PPAR-γ antagonist treatment significantly decreased expression of NMYC, B-cell lymphoma 2 (BCL2) and bromodomain-containing protein 4 (BRD4). It is implied that BRD4, NMYC, BCL2 suppression by the PPAR-γ antagonist resulted in cell growth inhibition, differentiation, and apoptosis induction. In our in vivo study, the PPAR-γ antagonist treatment induced CHP212 cells differentiation and resultant tumor growth inhibition. Our results provide a deeper understanding of the mechanisms of tumor cell differentiation and suggest that PPAR-γ antagonist is a new therapeutic and prevention option for neuroblastomas.

ECM proteins and cationic polymers coating promote dedifferentiation of patient-derived mature adipocytes to stem cells.

Reprogramming of mature adipocytes is an attractive research area due to the plasticity of these cells. Mature adipocytes can be reprogrammed in vitro, transforming them into dedifferentiated fat cells (DFATs), which are considered a new type of stem cell, and thereby have a high potential for use in tissue engineering and regenerative medicine. However, there are still no reports or findings on in vitro controlling the dedifferentiation. Although ceiling culture performed in related studies is a relatively simple method, its yield is low and does not allow manipulation of mature adipocytes to increase or decrease the dedifferentiation. In this study, to understand the role of physicochemical surface effects on the dedifferentiation of patient-derived mature adipocytes, the surfaces of cell culture flasks were coated with extracellular matrix, basement membrane proteins, and cationic/anionic polymers. Extracellular matrix such as fibronectin and collagen type I, and basement membrane proteins such as collagen type IV and laminin strongly promoted dedifferentiation of mature adipocytes, with laminin showing the highest effect with a DFAT ratio of 2.98 (±0.84). Interestingly, cationic polymers also showed a high dedifferentiation effect, but anionic polymers did not, and poly(diallyl dimethylammonium chloride) showed the highest DFAT ratio of 2.27 (±2.8) among the cationic polymers. Protein assay results revealed that serum proteins were strongly adsorbed on the surfaces of the cationic polymer coating, including inducing high mature adipocyte adhesion. This study demonstrates for the first time the possibility of regulating the transformation of mature adipocytes to DFAT stem cells by controlling the physicochemical properties of the surface of conventional cell culture flasks.

Discovery of Selective Histone Deacetylase 1 and 2 Inhibitors: Screening of a Focused Library Constructed by Click Chemistry, Kinetic Binding Analysis, and Biological Evaluation.

Histone deacetylase 1 and 2 (HDAC1/2) inhibitors are potentially useful as tools for probing the biological functions of the isoforms and as therapeutic agents for cancer and neurodegenerative disorders. To discover potent and selective inhibitors, we screened a focused library synthesized by using click chemistry and obtained KPZ560 as an HDAC1/2-selective inhibitor. Kinetic binding analysis revealed that KPZ560 inhibits HDAC2 through a two-step slow-binding mechanism. In cellular assays, KPZ560 induced a dose- and time-dependent increase of histone acetylation and showed potent breast cancer cell growth-inhibitory activity. In addition, gene expression analyses suggested that the two-step slow-binding inhibition by KPZ560 regulated the expression of genes associated with cell proliferation and DNA damage. KPZ560 also induced neurite outgrowth of Neuro-2a cells and an increase in the spine density of granule neuron dendrites of mice. The unique two-step slow-binding character of o-aminoanilides such as KPZ560 makes them interesting candidates as therapeutic agents.

Long term observation of de novo adipogenesis using novel bioabsorbable implants with larger size in a porcine model.

Introduction: The regeneration of adipose tissue in patients after breast cancer surgery would be desirable without the use of growth factors or cells to avoid potential recurrence and metastasis. We reported that prolate spheroidal-shaped poly-L-lactic acid (PLLA) mesh implants of approximately 18-mm polar diameter and 7.5-mm greatest equatorial diameter containing collagen sponge (CS) would be replaced by regenerated adipose tissue after implantation, thereby suggesting an innovative method for breast reconstruction. Our study aimed to evaluate the adipose tissue regeneration ability of implant aggregates in a porcine model. Methods: We prepared implant aggregates consisting of thirty PLLA mesh implants containing CS packed in a woven poly (glycolic acid) bag. The implant aggregates were inserted under the mammary glands in the porcine abdomen for a year. Single and double groups were classified by inserting either one or two implant aggregates on each side of the abdomen, respectively. Results: In both groups, the volume of the implant aggregates decreased over time, and the formation of adipose tissue peaked between 6 and 9 months. Histologically, the formation of adipose tissue was confirmed in the area that was in contact with native adipose tissue. Conclusions: Our implant aggregates could induce the autologous adipose tissue after long term implantation in vivo, without the use of any growth factor or cell treatment, presenting a potential novel method of breast reconstruction.

An easy-to-use semi-automatic volumetric evaluation for secondary bone grafting in patients with unilateral alveolar cleft.

Surgical intervention for alveolar bone formation is important in patients with alveolar cleft; however, the treatment methods and materials are still controversial. A precise evaluation method for postoperative bone formation is important for comparing outcomes and establishing the best treatment protocol. The purpose of this study is to establish a new method of evaluating surgical outcomes for patients with alveolar cleft. Computed tomography datasets from 20 patients who underwent secondary alveolar bone grafting were obtained before and 1 year after surgery. Six anatomical landmarks were used to superimpose the preoperative and mirrored preoperative volume and postoperative volume data. The cleft region was segmented by subtracting the preoperative from mirrored preoperative volume data, and the failed osteogenesis region was segmented by subtracting the postoperative volume data from the cleft region; subsequently, the bone formation ratio was calculated. Two observers performed this method using a free software 3D slicer and the average evaluation times were 12.7 and 13.2 min for observers 1 and 2, respectively. Method reliability was determined by evaluating intraclass correlation coefficients. The intra-observer intraclass correlation coefficients were 0.97 and 0.96 for observers 1 and 2, respectively. The inter-observer intraclass correlation coefficient was 0.97. Our method is practical for assessing bone formation after treatment, which does not require specific knowledge or software and can be used by ordinary physicians.

Interposition grafting of collagen-gelatin sponge impregnated with basic fibroblast growth factor in primary palatoplasty.

Introduction: An oronasal fistula is a challenging post-operative complication of palatoplasty due to impaired velopharyngeal function or its high recurrence rate. Muscle repositioning, a key procedure in palatoplasty, causes dead space at the junction between the hard and soft palates. Consequently, thin oral and nasal mucosae are prone to break down and form fistulas. In this study, we used basic fibroblast growth factor-impregnated collagen gelatin sponge (bFGF-CGS) in primary palatoplasty to reduce fistula formation. Methods: This retrospective study assessed the complications and efficacy of bFGF-CGS to reduce fistula formation. Patients who underwent primary palatoplasty with bFGF-CGS were included. The same number of patients who underwent primary palatoplasty without bFGF-CGS was included as a control group. The outcomes included post-operative oronasal fistula formation, delayed healing, bleeding, and infection. Results: Both groups included 44 patients. Except for age at palatoplasty, there were no statistically significant demographic differences between the two groups; however, the rates of fistula formation in the study and control group were 2.3% and 13.6%, respectively. There were no infections among the patients. Conclusions: The grafting of bFGF-CGS in primary palatoplasty was safe and probably effective in reducing post-operative oronasal fistula formation.

Preoperative visualization of midline-crossing subcutaneous arteries in transverse abdominal flaps using photoacoustic tomography.

BACKGROUND: Photoacoustic tomography is a noninvasive vascular imaging modality that uses near-infrared pulsed laser light and ultrasound to visualize vessels. We previously demonstrated the utility of photoacoustic tomography for anterolateral thigh flap surgery involving body-attachable vascular mapping sheets. However, it was not possible to obtain clear separate images of arteries and veins. In this study, we tried to visualize subcutaneous arteries that cross the midline of the abdomen, since these arteries are known to be important for obtaining large perfusion areas in transverse abdominal flaps. METHODS: Four patients scheduled to undergo breast reconstruction with abdominal flaps were examined. Photoacoustic tomography was performed preoperatively. The tentative arteries and veins were traced according to the S-factor, an approximate hemoglobin oxygen saturation parameter calculated using 2 laser excitation wavelengths (756 and 797 nm). Intraoperatively, arterial-phase indocyanine green (ICG) angiography was performed after abdominal flap elevation. Images of vessels speculated to be arteries by preoperative photoacoustic tomography were merged with those of intraoperative ICG angiography and analyzed in an 8 × 4-cm2 area below the umbilical region. RESULTS: The S-factor was used to visualize the midline-crossing subcutaneous arteries in all 4 patients. A matching analysis compared preoperative tentative arteries according to photoacoustic tomography with ICG angiography results in the 8 × 4-cm2 area below the umbilical region and indicated a 71.3-82.1% match (average: 76.9% match). CONCLUSIONS: This study demonstrates that the S-factor, a noninvasive, label-free imaging modality, can be used to successfully visualize subcutaneous arteries. This information can aid in selecting perforators for abdominal flap surgery.

Lower Lip Reconstruction Using a Sensory Anterolateral Thigh Flap as the First Choice.

Local flaps from the upper lip and cheeks have been the first choice for two-thirds to total resection of the lower lip. However, these local flap techniques involve many clinical problems, including small a mouth, drooling, scarring, and hypesthesia. The improvement of free anterolateral thigh (ALT) flap transfer can solve these problems with expansion of the application of free flaps for lower lip reconstruction. The patient in this case was a 56-year-old man with squamous cell carcinoma of the lower lip (cT3N1M0). Subtotal lower lip resection preserving both corners of the mouth with bilateral neck dissection was performed. Simultaneously, a sensory ALT flap was elevated with an 8 × 6 cm skin island and a lateral femoral cutaneous nerve. The lateral and medial sides of the fascia lata were processed into 1-cm-wide strings, which were tunneled through the orbicularis oris muscle of the upper lip and sutured to the orbicularis oris muscle at the mucosal side of the philtrum. The lateral femoral cutaneous nerve and right mental nerve were sutured. At 3 months, a second surgery was performed to replace the ALT flap on the white labial side with a clavicle full-thickness skin graft. This surgery achieved four important factors: opening and closing of the mouth, sensory function of the lower lip, cosmetic appearance, and minimization of donor-site damage. We believe the worldwide improvement of microsurgery techniques enables lower lip reconstruction using the sensory ALT flap to be selected as the first choice for two-thirds to total lower lip defects.

Deep Infantile Hemangioma in the Involuting Phase That Was Difficult to Diagnose before Surgery.

Infantile hemangioma (IH) is a common pediatric vascular tumor and is easily diagnosed in most cases based on the clinical course and appearance, but deep IHs are difficult to diagnose based on external appearance alone. Clinical and imaging findings are therefore important clues to the diagnosis of soft tissue tumors; however, a definitive diagnosis is decided based on the pathological examination of biopsy or resection specimens. A 1-year-old girl with a subcutaneous mass on her glabella was referred to our hospital. At 3 months of age, her mother noticed a tumor that swelled when she cried. It gradually enlarged, and ultrasonography and magnetic resonance imaging were performed at 12 months of age. Doppler ultrasonography showed a hypo-vascular mass. Magnetic resonance imaging revealed a subcutaneous mass with low-intensity on T1-weighted image and slightly high-intensity on T2-weighted image, with tiny flow voids. Computed tomography showed no frontal bone defect. The soft tissue tumor could not be diagnosed based on these imaging findings; thus, we decided to perform total resection under general anesthesia. A histopathological examination showed a highly cellular tumor with capillaries with opened small vascular channels and glucose transporter 1 positivity. Thus, it was diagnosed as deep IH transitioning from the proliferative phase to the involuting phase. Deep IHs are difficult to diagnose because characteristic imaging findings disappear during the involuting phase. We emphasize the importance of performing Doppler ultrasonography in the early phase (eg, at 6 months of age) for soft tissue tumors of infancy.