Search Results (16)

The exposure of endothelial cells to high glucose concentrations promotes angiogenesis. The present study investigated whether this pro-angiogenic effect of glucose is suitable to improve the capability of nanofat to vascularize implanted dermal substitutes. Nanofat was processed from white adipose tissue originating from green fluorescent protein (GFP)⁺ C57BL/6J donor mice and incubated for 1 h in Hank’s Balanced Salt Solution with or without (control) a high level of glucose (30 mM). The pretreated nanofat was seeded onto dermal substitutes, which were analyzed by intravital fluorescence microscopy, histology and immunohistochemistry in dorsal skinfold chambers of GFP⁻ C57BL/6J mice to assess their vivo performance over a period of 14 days. A high level of glucose-pretreated nanofat did not induce a stronger immune response when compared to the control. However, it improved the vascularization of the implants, as shown by a significantly higher density of blood-perfused microvessels in the border zones (~3.6-fold increase) and more CD31⁺/GFP⁺ microvessels (~3-fold increase) inside the implants. Accordingly, high glucose-pretreated nanofat levels also enhanced the tissue integration of the dermal substitutes, as indicated by the deposition of more type I collagen (~2.9-fold increase). These findings suggest that the short-term exposure of nanofat to a high level of glucose represents a promising and clinically feasible strategy to enhance its regenerative properties when seeded onto dermal substitutes. Full article

Autologous fat grafting of human lipoaspirate (LA) is increasingly used in reconstructive and cosmetic surgery for lipofilling and stem cell-rich “nanofat” reinjection for regenerative medicine. While commercial devices (e.g., REVOLVE and Puregraft) are available, many surgeons use non-standardized manual washing techniques, leading to inconsistent graft retention (20–80%). Moreover, no system can unite washing directly with mechanical processing to produce a nanofat-like product directly from raw LA. We developed a novel preparation device (PD) that is designed for peristaltic pump-driven washing of LA and can be seamlessly combined with our previously developed Emulsification and Micronization Device (EMD) into an automated closed-loop platform. Human LA samples were washed with the PD and compared to standard manual washing via visual colorimetric analysis. We then evaluated the mechanical processing of PD-washed LA using our EMD and assessed cell count, viability, and stromal vascular fraction-derived subpopulations (i.e., mesenchymal stem cells, endothelial progenitor cells (EPCs), pericytes, transit-amplifying (TA) progenitor cells, and supra-adventitial adipose stromal cells). Recirculating LA through the PD for at least one minute resulted in sufficient mixing, producing LA with equivalent color and quality to manual washing. Integrating the EMD within a platform enabled both washing and mechanical processing under peristaltic flow, enriching key subpopulations compared to manual methods. Thus, our fluidic platform effectively washes LA in a closed-loop system, minimizing LA tissue manipulation and opportunity for contamination while also simplifying the workflow for mechanical processing. Further refinement and automation of this platform would enhance the reproducibility and quality of small-volume fat grafts, cell-assisted lipotransfer, and stem/progenitor cell injections to promote wound healing and angiogenesis. Full article

Background: Histological and immunohistochemical analyses of adipose tissue are essential for evaluating the quality and functionality of lipoaspirates in regenerative medicine and fat grafting procedures. These methods provide insights into tissue viability, cellular subtypes, and extracellular matrix (ECM) composition—all factors influencing graft retention and clinical outcomes. Purpose: This scoping review aims to summarize the most commonly used staining methods and their applications in the histology and immunohistochemistry of adipose tissue. By exploring qualitative and quantitative markers, we seek to guide researchers in selecting the appropriate methodologies for addressing experimental and translational research. Methods: A systematic search was conducted using PubMed, Ovid, and the Cochrane Library databases from inception to 2024, employing Boolean operators (“lipoaspirate” OR “fat graft” OR “gauze rolling” OR “decantation” OR “coleman fat” OR “celt” OR “nanofat” OR “lipofilling” OR “human fat” AND “histol*”). Studies were included if they utilized histology or immunohistochemistry on undigested human adipose tissue or its derivatives. The inclusion criteria focused on peer-reviewed, English-language studies reporting quantitative and qualitative data on adipose tissue markers. Results: Out of 166 studies analyzed, hematoxylin–eosin (H&E) was the most frequently employed histological stain (152 studies), followed by Masson Trichrome and Sudan III. Immunohistochemical markers such as CD31, CD34, and perilipin were extensively used to distinguish stromal vascular fraction (SVF) cells, adipocytes, and inflammatory processes. Studies employing semiquantitative scoring demonstrated enhanced comparability, particularly for fibrosis, necrosis, and oil cyst evaluation. Quantitative analyses focused on SVF cell density, mature adipocyte integrity, and ECM composition. Methodological inconsistencies, particularly in preparation protocols, were observed in 25 studies. Conclusions: This review highlights the critical role of histological and immunohistochemical methods in adipose tissue research. H&E staining remains the cornerstone for general tissue evaluation in the clinical context, while specialized stains and immunohistochemical markers allow for detailed analyses of specific cellular and ECM components in experimental research. Standardizing preparation and evaluation protocols will enhance interstudy comparability and support advancements in adipose tissue-based therapies. Full article

Manual processing of lipoaspirate can enhance stem cell concentration, thereby improving the take rate, which still represents a major challenge in autologous fat transfer. However, since the preparation consists of many manual steps that are difficult to standardize, we investigated the influence of residual tumescent solution on the macroscopic and microscopic outcome of the mechanically processed lipoaspirate. Additionally, we investigated whether sedimentation followed by vacuum filtration of the aqueous phase could accelerate processing by replacing the initial centrifugation step. Samples with more than 5% remaining aqueous phase show no clearly defined oil phase, preventing any volume reduction. In contrast, all centrifuged samples produced a clear oil phase. The remaining tissue, as confirmed by both histology and viability assays, was superior to nanofat. Although sedimentation and filtration in the LipoCollector did not sufficiently separate enough aqueous phase from the lipoaspirate, tissue viability was significantly higher compared to our control container. Our findings indicate that centrifugation remains essential for effective aqueous phase separation and further mechanical processing, while the automatic filtration may enhance processing efficiency. These results indicate that further work is needed to simplify mechanical processing, as the outcome can be significantly influenced by parameters such as tumescent impurities. Full article

Heat preconditioning has been shown to promote nutritive perfusion and tissue survival in autologous fat grafting as well as in flap and breast surgery. However, its impact on the vascularization properties of nanofat has not been investigated so far. Therefore, we exposed nanofat from donor mice to a temperature of 43 °C for 1 h and assessed the effects of this heat stress on cell viability and the expression of heat shock proteins (HSPs) and angiogenesis-related factors. Moreover, dermal substitutes seeded with heat-preconditioned and non-preconditioned control nanofat were implanted into dorsal skinfold chambers of recipient mice to study their vascularization and tissue integration in vivo by means of repeated intravital fluorescence microscopy, histology and immunohistochemistry. Heat preconditioning upregulated the expression of HSPs in nanofat without affecting cell viability. Moreover, it resulted in the downregulation of many pro-angiogenic factors and the increased expression of anti-angiogenic factors, indicating a shift towards an anti-angiogenic phenotype. Accordingly, implanted dermal substitutes seeded with heat-preconditioned nanofat exhibited a reduced vascularization and were not better integrated into the host tissue when compared to controls. These findings indicate that heat preconditioning cannot be recommended for enhancing the vascularization capacity of nanofat. Full article

Dermal substitutes require sufficient tissue integration and vascularization to be successfully covered with split-thickness skin grafts. To rapidly achieve this, we provide the proof of principle for a novel vascularization strategy with high translational potential. Nanofat was generated from subcutaneous adipose tissue of green fluorescence protein (GFP)⁺ C57BL/6J donor mice and seeded onto small samples (4 mm in diameter) of the clinically approved dermal substitute Integra^®. These samples and non-seeded controls were then implanted into full-thickness skin defects in the dorsal skinfold chamber of C57BL/6J wild-type mice and analyzed by intravital fluorescence microscopy, histology and immunohistochemistry over a 14-day period. Nanofat-seeded dermal substitutes exhibited an accelerated vascularization, as indicated by a significantly higher functional microvessel density on days 10 and 14 when compared to controls. This was primarily caused by the reassembly of GFP⁺ microvascular fragments inside the nanofat into microvascular networks. The improved vascularization promoted integration of the implants into the surrounding host tissue, which finally exhibited an increased formation of a collagen-rich granulation tissue. There were no marked differences in the inflammatory host tissue reaction to nanofat-seeded and control implants. These findings demonstrate that nanofat significantly improves the in vivo performance of dermal substitutes without affecting their biocompatibility. Full article

Introduction: Peripheral nerve injury (PNI) is increasingly prevalent and challenging to treat despite advances in microsurgical techniques. In this context, adipose tissue derivatives, such as adipose-derived stem cells, nanofat, and stromal vascular fraction have been gaining attention as potential allies in peripheral nerve regeneration. Objectives: This study aims to explore the use of adipose tissue derivatives in nerve regeneration following peripheral nerve transection in murine models. Thus, we assess and synthesize the key techniques and methods used for evaluating the obtained nerve regeneration to guide future experimental research and clinical interventions. Methodology: A systematic review was conducted in February 2024, adhering to the Cochrane and PRISMA 2020 guidelines, using the PubMed, SciELO, and LILACS databases. The focus was on experimental studies involving adipose tissue derivatives in nerve regeneration in animal models post-transection. Only experimental trials reporting nerve regeneration outcomes were included; studies lacking a comparator group or evaluation methods were excluded. Results: Out of 273 studies initially identified from MEDLINE, 19 were selected for detailed analysis. The average study included 32.5 subjects, with about 10.2 subjects per intervention subgroup. The predominant model was the sciatic nerve injury with a 10 mm gap. The most common intervention involved unprocessed adipose-derived stem cells, utilized in 14 articles. Conclusions: This review underscores the significant potential of current methodologies in peripheral nerve regeneration, particularly highlighting the use of murine models and thorough evaluation techniques. Full article

Background and Objectives: This three-year clinical trial aimed to demonstrate that only the signaling vesicles produced by ADSCa, containing mRNA, microRNA, growth factors (GFs), and bioactive peptides, provide an advantage over classical therapy with adipose disaggregate to make the tissue regeneration technique safer due to the absence of interfering materials and cells, while being extremely minimally invasive. The infiltration of disaggregated adipose nanofat, defined by the Tonnard method, for the regeneration of the dermis and epidermis during physiological or pathological aging continues to be successfully used for the presence of numerous adult stem cells in suspension (ADSCa). An improvement in this method is the exclusion of fibrous shots and cellular debris from the nanofat to avoid inflammatory phenomena by microfiltration. Materials and Methods: A small amount of adipose tissue was extracted after surface anesthesia and disaggregated according to the Tonnard method. An initial microfiltration at 20/40 microns was performed to remove fibrous shots and cellular debris. The microfiltration was stabilized with a sterile solution containing hyaluronic acid and immediately ultrafiltered to a final size of 0.20 microns to exclude the cellular component and hyaluronic acid chains of different molecular weights. The suspension was then injected into the dermis using a mesotherapy technique with microinjections. Results: This study found that it is possible to extract signaling microvesicles using a simple ultrafiltration system. The Berardesca Scale, Numeric Rating Scale (NRS), and Modified Vancouver Scale (MVS) showed that it is possible to obtain excellent results with this technique. The ultrafiltrate can validly be used in a therapy involving injection into target tissues affected by chronic and photoaging with excellent results. Conclusions: This retrospective clinical evaluation study allowed us to consider the results obtained with this method for the treatment of dermal wrinkles and facial tissue furrows as excellent. The method is safe and an innovative regenerative therapy as a powerful and viable alternative to skin regeneration therapies, antiaging therapies, and chronic inflammatory diseases because it lacks the inflammatory component produced by cellular debris and fibrous sprouts and because it can exclude the mesenchymal cellular component by reducing multiple inflammatory cytokine levels. Full article

The treatment of wounds using the body’s own resources is a promising approach to support the physiological regenerative process. To advance this concept, we evaluated the effect of nanofat (NF) on wound healing. For this purpose, full-thickness skin defects were created in dorsal skinfold chambers of wild-type mice. These defects were filled with NF generated from the inguinal subcutaneous adipose tissue of green fluorescent protein (GFP)⁺ donor mice, which was stabilized using platelet-rich plasma (PRP). Empty wounds and wounds solely filled with PRP served as controls. Wound closure, vascularization and formation of granulation tissue were repeatedly analyzed using stereomicroscopy, intravital fluorescence microscopy, histology and immunohistochemistry over an observation period of 14 days. PRP + NF-treated wounds exhibited accelerated vascularization and wound closure when compared to controls. This was primarily due to the fact that the grafted NF contained a substantial fraction of viable GFP⁺ vascular and lymph vessel fragments, which interconnected with the GFP⁻ vessels of the host tissue. Moreover, the switch from inflammatory M1- to regenerative M2-polarized macrophages was promoted in PRP + NF-treated wounds. These findings indicate that NF markedly accelerates and improves the wound healing process and, thus, represents a promising autologous product for future wound management. Full article

Major limitations in the effective treatment of neurological cancer include systemic cytotoxicity of chemotherapy, inaccessibility, and inoperability. The capability to successfully target a drug to the tumor site(s) without incurring serious side effects—especially in the case of aggressive tumors, such as glioblastoma and neuroblastoma—would represent a significant breakthrough in therapy. Orthotopic systems, capable of storing and releasing proteins over a prolonged period at the site of a tumor, that utilize nanoparticles, liposomes, and hydrogels have been proposed. One candidate for drug delivery is Micro-Fragmented Adipose Tissue (MFAT). Easily obtained from the patient by abdominal subcutaneous liposuction (autologous), and with a high content of Mesenchymal Stem Cells (MSCs), mechanically derived nanofat is a natural tissue graft with a structural scaffold organization. It has a well-preserved stromal vascular fraction and a prolonged capacity to secrete anti-tumorigenic concentrations of pre-absorbed chemotherapeutics within extracellular vesicles. This review discusses current evidence supporting the potential of drug-modified MFAT for the treatment of neurological cancer with respect to recent preclinical and in vitro studies. Possible limitations and future perspectives are considered. Full article

Nanofat is a relatively novel technique in fat grafting that has gained significant interest in the fields of regenerative medicine, aesthetic and translational research. It involves the extraction of autologous fat from a patient, which is then transformed into “nanofat”, consisting of small fat particles with a diameter of less than 0.1 mm and containing high concentrations of stem cells and growth factors. This article focuses on the use of nanofat in facial rejuvenation and its potential for lipomodelling. Fat tissue is a “stem cell depot” and nanofat contains many stem cells that can differentiate into various cell types. The Lipogem technology, developed in 2013, enables the isolation of nanofat with an intact perivascular structure, utilizing the high concentration of mesenchymal stromal cells near the pericytes of the adipose vascular system. Nowadays nanofat is used primarily for cosmetic purposes particularly in rejuvenating and improving the appearance of the skin, especially the face. Indeed, it has wide applicability; it can be used to treat fine lines, wrinkles, acne scars, sun-damaged skin, scar repair, and as an alopecia treatment. However, further studies are needed to assess the long-term efficacy and safety of this technique. In conclusion, nanofat is a safe and minimally invasive option for tissue regeneration with considerable therapeutic potential. This study reviews the application and effects of nanofat in regenerative medicine and facial cosmetic surgery. Full article

Osteoarthritis (OA) is one of the main causes of disability worldwide and is caused by the progressive degeneration of joint tissues, ultimately leading to chronic pain and loss of function. Intraarticular delivery of mesenchymal stromal cells, such as adipose-derived stromal cells (ASCs), is being actively investigated due to their trophic properties observed in both preclinical and clinical studies. However, cell expansion and handling involve costly and time-consuming processes that limit their application. Recently, several devices and kits have been developed to isolate and process the stromal vascular fraction (SVF), a high biologically active compound of the adipose tissue, right at the patient’s bedside. In this study, we introduce a novel technique to obtain an SVF graft with a high content of ASCs for intraarticular injection directly from liposuction and with minimal equipment. In this technical note, we describe in detail the steps of the surgical technique as well as strategies to avoid common pitfalls and complications. Full article

The combination of adipose-derived stem cells (ASCs) and dermal scaffolds has been shown to be an approach with high potential in soft tissue reconstruction. The addition of dermal templates to skin grafts can increase graft survival through angiogenesis, improve regeneration and healing time, and enhance the overall appearance. However, it remains unknown whether the addition of nanofat-containing ASCs to this construct could effectively facilitate the creation of a multi-layer biological regenerative graft, which could possibly be used for soft tissue reconstruction in the future in a single operation. Initially, microfat was harvested using Coleman’s technique, then isolated through the strict protocol using Tonnard’s technique. Finally, centrifugation, emulsification, and filtration were conducted to seed the filtered nanofat-containing ASCs onto Matriderm for sterile ex vivo cellular enrichment. After seeding, a resazurin-based reagent was added, and the construct was visualized using two-photon microscopy. Within 1 h of incubation, viable ASCs were detected and attached to the top layer of the scaffold. This experimental ex vivo note opens more dimensions and horizons towards the combination of ASCs and collagen–elastin matrices (i.e., dermal scaffolds) as an effective approach in soft tissue regeneration. The proposed multi-layered structure containing nanofat and dermal template (Lipoderm) may be used, in the future, as a biological regenerative graft for wound defect reconstruction and regeneration in a single operation and can also be combined with skin grafts. Such protocols may optimize the skin graft results by creating a multi-layer soft tissue reconstruction template, leading to more optimal regeneration and aesthetic outcomes. Full article

Adipose tissue contains adult mesenchymal stem cells that may modulate the metabolism when applied to other tissues. Stromal vascular fraction (SVF) can be isolated from adipose tissue mechanically and/or enzymatically. SVF was recently used to decrease the pain and improve the function of knee osteoarthritis (OA) patients. Primary and/or secondary OA causes inflammation and degeneration in joints, and regenerative approaches that may modify the natural course of the disease are limited. SVF may modulate inflammation and initiate regeneration in joint tissues by initiating a paracrine effect. Chemokines released from SVF may slow down degeneration and stimulate regeneration in joints. In this review, we overviewed articular joint cartilage structures and functions, OA, and macro-, micro-, and nano-fat isolation techniques. Mechanic and enzymatic SVF processing techniques were summarized. Clinical outcomes of adipose tissue derived tissue SVF (AD-tSVF) were evaluated. Medical devices that can mechanically isolate AD-tSVF were listed, and publications referring to such devices were summarized. Recent review manuscripts were also systematically evaluated and included. Transferring adipose tissues and cells has its roots in plastic, reconstructive, and aesthetic surgery. Micro- and nano-fat is also transferred to other organs and tissues to stimulate regeneration as it contains regenerative cells. Minimal manipulation of the adipose tissue is recently preferred to isolate the regenerative cells without disrupting them from their natural environment. The number of patients in the follow-up studies are recently increasing. The duration of follow up is also increasing with favorable outcomes from the short- to mid-term. There are however variations for mean age and the severity of knee OA patients between studies. Positive outcomes are related to the higher number of cells in the AD-tSVF. Repetition of injections and concomitant treatments such as combining the AD-tSVF with platelet rich plasma or hyaluronan are not solidified. Good results were obtained when combined with arthroscopic debridement and micro- or nano-fracture techniques for small-sized cartilage defects. The optimum pressure applied to the tissues and cells during filtration and purification of the AD-tSVF is not specified yet. Quantitative monitoring of articular joint cartilage regeneration by ultrasound, MR, and synovial fluid analysis as well as with second-look arthroscopy could improve our current knowledge on AD-tSVF treatment in knee OA. AD-tSVF isolation techniques and technologies have the potential to improve knee OA treatment. The duration of centrifugation, filtration, washing, and purification should however be standardized. Using gravity-only for isolation and filtration could be a reasonable approach to avoid possible complications of other methodologies. Full article

In recent years, lipofilling became a popular scar treatment method. Its beneficial outcomes have been partly attributed to the regenerative capacity of adipose-derived stem cells (ADSCs), suspended in an extracellular matrix—the stromal vascular fraction (SVF). The aim of this review was to verify if existing data support the clinical use of ADSC-related interventions in scar treatment. A systematic search of the literature was performed in July 2020 in five databases (Medline, Cochrane, Web of Science, Scopus and Embase). Articles written in English, except for reviews, letters and editorials, were identified and screened for eligibility. We looked for reports of any outcomes in scars treated with ADSCs or SVF. Data from selected articles were extracted and the quality of each study was assessed. Five hundred and fourteen studies were identified in the primary search, of which nineteen were eventually included in the systematic review. Extracted data pointed to beneficial microscopic, functional and aesthetic outcomes in a total of 665 patients. Six studies included comparative interventions—platelet-rich plasma or CO₂ fractional laser. Collected data give low-to-average quality evidence for beneficial effects of ADSC-related interventions in scar treatment. Some studies suggest that these interventions are noninferior to PRP or fractional CO₂ laser. Full article