Adipose-Derived Regenerative Cell Therapy for Burn Wound Healing: A Comparison of Two Delivery Methods.
ABSTRACT: Objective: The use of noncultured autologous stromal vascular fraction or clinical grade adipose-derived regenerative cells (ADRCs) is a promising strategy to promote wound healing and tissue repair. Nevertheless, issues regarding the optimal mode of administration remain unclear. The purpose of this study was to compare the effects of local injection and topical spray delivery of ADRCs in a porcine model of thermal burns. Approach: Full-thickness thermal burns were created on the dorsum of 10 Gottingen minipigs. Two days following injury, wounds underwent fascial excision and were randomized to receive control vehicle or freshly isolated autologous ADRCs delivered by either multiple injections into or surrounding the wound bed, or by spray onto the wound surface (0.25 × 106 viable cells/cm2). Healing was evaluated by planimetry, histopathology, and immunohistochemistry at day 7, 12, 16, 21, and 28 posttreatment. Results:In vitro analysis demonstrated that there was no substantial loss of cell number or viability attributable to the spray procedure. Planimetric assessment revealed that delivery of ADRCs by either local injection or topical spray increased wound reepithelialization relative to control at day 14. No significant difference in wound reepithelialization was observed between both delivery approaches. In addition, on day 7 posttreatment, blood vessel density was greater in wounds receiving local or topical spray ADRCs than in the wounds treated with vehicle control. Histopathologic analysis suggests that ADRC treatment may modulate the inflammatory response by reducing neutrophil infiltration at day 7 and 12 posttreatment, irrespective of the route of administration. Conclusions: These data demonstrate that local injection and spray delivery of ADRCs modulate inflammation and improve wound angiogenesis and epithelialization. Importantly, both delivery routes exhibited similar effects on wound healing. Given the greater ease-of-use associated with topical spray delivery, these data support the use of a spray system for autologous ADRC delivery.
Project description:Introduction:The purpose of this study was to evaluate whether cryopreserved (frozen) adipose-derived regenerative cells (ADRCs) have a therapeutic effect on burn wound healing as well as freshly isolated (fresh) ADRCs. Methods:Full thickness burns were created on dorsum of nude mice and burn wound was excised. The wound was covered by artificial dermis with; (i) fresh ADRCs, (ii) frozen ADRCs, and (iii) PBS (control). The assessment for wound healing was performed by morphological, histopathological and immunohistochemical analyses. Results:In vivo analyses exhibited the significant therapeutic effect of frozen ADRCs on burn wound healing up to the similar or higher level of fresh ADRCs. There were significant differences of wound closure, epithelized tissue thickness, and neovascularization between the treatment groups and control group. Although there was no significant difference of therapeutic efficacy between fresh ADRC group and frozen ADRC group, frozen ADRCs improved burn wound healing process in dermal regeneration with increased great type I collagen synthesis compared with fresh ADRCs. Conclusions:These findings indicate that frozen ADRCs allow us to apply not only quickly but also for multiple times, and the cryopreserved ADRCs could therefore be useful for the treatment of burn wounds in clinical settings.
Project description:Paralysis of one vocal fold leads to glottal gap and vocal fold insufficiency that has significant impact upon a patient's quality of life. Fillers have been tested to perform intracordal injections, but they do not provide perdurable results. Early data suggest that enriching fat grafts with adipose-derived regenerative cells (ADRCs) promote angiogenesis and modulate the immune response, improving graft survival. The aim of this study is to propose ADRC-enriched adipose tissue grafts as effective filler for the paralyzed vocal fold to use it for functional reconstruction of the glottal gap.This is the first phase I-IIA clinical trial (phase I/IIA clinical trial, unicentric, randomized, controlled, and two parallel groups), to evaluate the safety of a new therapy with ADRC-enriched fat grafting (ADRC: group I) for laryngoplasty after unilateral vocal fold paralysis. Control group patients received centrifuged autologous fat (CAF: group II) grafts. Overall mean age is 52.49 ± 16.60 years. Group I (ADRC): 7 patients (3 males and 4 females), 52.28 ± 20.95 year. Group II (CAF): 7 patients (3 males and 4 females), 52.71 ± 12.59 year.VHI-10 test showed that preoperative mean score was 24.21 ± 8.28. Postoperative mean score was 6.71 ± 6.75. Preoperative result in group I was 21.14 ± 3.58 and postoperative result was 3.14 ± 3.53. Preoperative result for group II was 27.29 ± 10.66. Postoperative score in group II was 10.29 ± 7.52. Wilcoxon and the Student t-tests showed that the patient's self-perception of posttreatment improvement is larger when ADRCs are used. Comparing pre- and posttreatment voice quality analysis, group I showed a p = 0.053. Group II showed a p = 0.007. There would be no significant differentiation between pre- and posttreatment results. This is true for group II and limited for group I.This prospective trial demonstrates the safety and efficacy of the treatment of glottal gap defects utilizing ADRC-enriched fat grafts. This trial is registered with NCT02904824.
Project description:Adipose-derived regenerative cell (ADRC) is a promising alternative source of autologous somatic stem cells for the repair of damaged tissue. This study aimed to assess the safety and feasibility of autologous ADRC implantation for therapeutic angiogenesis in patients with critical limb ischaemia (CLI). A clinical pilot study—Therapeutic Angiogenesis by Cell Transplantation using ADRCs (TACT-ADRC) study—was initiated in Japan. Adipose tissue was obtained by ordinary liposuction method. Isolated ADRCs were injected into the ischaemic limb. We performed TACT-ADRC procedure in five patients with CLI. At 6 months, no adverse events related to the TACT-ADRC were observed. No patients required major limb amputation, and ischaemic ulcers were partly or completely healed during the 6-month follow-up. In all cases, significant clinical improvements were seen in terms of rest pain and 6-min walking distance. Numbers of circulating CD34+ and CD133+ cells markers of progenitor cell persistently increased after ADRC implantation. The ratio of VEGF-A165b (an anti-angiogenic isoform of VEGF) to total VEGF-A in plasma significantly decreased after ADRC implantation. In vitro experiments, cultured with ADRC-conditioned media (CM) resulted in increased total VEGF-A and decreased VEGF-A165b in C2C12 cells, but not in macrophages. ADRC-CM also increased CD206+ cells expression and decreased TNF-? in macrophages. Autologous ADRC implantation was safe and effective in patients with CLI and could repair damaged tissue via its ability to promote angiogenesis and suppress tissue inflammation.
Project description:Introduction:Adipose tissue stromal cells contain a substantial number of mesenchymal stem cells. As such, their application to regeneration of miscellaneous impaired organs has attracted much attention. Methods:We designed a clinical study to investigate freshly isolated autologous adipose tissue-derived stromal/stem (regenerative) cell (ADRC) therapy for liver cirrhosis and conducted treatment in four cirrhotic patients. ADRCs were isolated from autologous subcutaneous adipose tissue obtained by the liposuction method, followed with use of the Celution system adipose tissue dissociation device. The primary endpoint is assessment of safety one month after treatment. We also characterized the obtained ADRCs. Results:Two patients had type C cirrhosis, one had nonalcoholic steatohepatitis-cirrhosis, and one had type B cirrhosis. No serious adverse events were observed during the 1-month study period after freshly isolated ADRC infusion. Serum albumin concentrations were maintained or improved during this period as well as during the succeeding follow-up of approximately 1 year in two patients and 6 months in another patient. Liver regeneration-related factors, namely hepatocyte growth factor and interleukin-6, were elevated 1 day after ADRC treatment in all patients. The obtained freshly isolated ADRCs were expanded in culture and found to express mesenchymal stem cell markers. Gene expression profile analysis of ADRCs was shown to involve inflammatory features, suggesting that characteristics of the obtained ADRCs were related to immunomodulatory biological effects. Conclusion:This clinical study treatment for liver cirrhosis using ADRCs was proven to be safely conductible, and can be further investigated in future for regeneration/repair of liver cirrhosis.
Project description:INTRODUCTION:Skin fragility and recurrent wounds are hallmarks of hereditary epidermolysis bullosa (EB). Treatment options to accelerate wound healing are urgently needed. Oleogel-S10 contains a betulin-rich triterpene extract from birch bark. In this study, we tested the wound healing properties of topical Oleogel-S10 in patients with dystrophic EB. METHODS:We conducted an open, blindly evaluated, controlled, prospective phase II pilot trial in patients with dystrophic EB (EudraCT number 2010-019945-24). Healing of wounds treated with and without topical Oleogel-S10 was compared. Primary efficacy variable was faster reepithelialization as determined by 2 blinded experts. The main secondary outcome variable of the study was percentage of wound epithelialization. RESULTS:Twelve wound pairs of 10 patients with dystrophic EB were evaluated. In 5 of 12 cases, both blinded reviewers considered epithelialization of the intervention wounds as superior. In 3 cases, only one reviewer considered Oleogel-S10 as superior and the other one as equal to control. Measurements of wound size showed a trend towards accelerated wound healing with the intervention but without reaching statistical significance. CONCLUSION:Our results indicate a potential for faster reepithelialization of wounds in patients with dystrophic EB when treated with Oleogel-S10 but larger studies are needed to confirm significance.
Project description:Acute kidney injury (AKI) represents a major clinical problem with high mortality and limited causal treatments. The use of cell therapy has been suggested as a potential modality to improve the course and outcome of AKI.We investigated the possible renoprotection of freshly isolated, uncultured adipose tissue-derived stem and regenerative cells (ADRCs) before and after cryopreservation in a rat ischemia-reperfusion (I-R) model of AKI.We demonstrated that ADRC therapy drastically reduced mortality (survival 100% vs. 57%, ADRC vs. controls, respectively) and significantly reduced serum creatinine (sCr on Day 3: 3.03 ± 1.58 vs. 7.37 ± 2.32 mg/dL, ADRC vs. controls, respectively). Histological analysis further validated a significantly reduced intratubular cast formation, ameliorated acute tubular epithelial cell necrosis and mitigated macrophage infiltration. Furthermore, a reduced RNA expression of CXCL2 and IL-6 was found in the ADRC group which could explain the reduced macrophage recruitment. Use of cryopreserved ADRCs resulted in an equally high survival (90% vs. 33% in the control group) and similarly improved renal function (sCr on Day 3: 4.64 ± 2.43 vs. 7.24 ± 1.40 mg/dL in controls).Collectively, these results suggest a potential clinical role for ADRC therapy in patients with AKI. Importantly, cryopreservation of ADRCs could offer an autologous treatment strategy for patients who are at high risk for AKI during planned interventions.
Project description:Effective reepithelialization after injury is essential for correct wound healing. The upregulation of keratinocyte alpha3beta1 integrin during reepithelialization suggests that this adhesion molecule is involved in wound healing; however, its precise role in this process is unknown. We have shown here that retarded reepithelialization in Itga3(-/-) mouse skin wounds is due predominantly to repressed TGF-beta1-mediated responses. Specifically, expression of the inhibitor of TGF-beta1-signaling Smad7 was elevated in Itga3(-/-) keratinocytes. Indeed, in vivo blockade of Smad7 increased the rate of reepithelialization in Itga3(-/-) and WT wounds to similar levels. Our data therefore indicate that the function of alpha3beta1 integrin as a mediator of keratinocyte migration is not essential for reepithelialization but suggest instead that alpha3beta1 integrin has a major new in vivo role as an inhibitor of Smad7 during wound healing. Moreover, our study may identify a previously undocumented function for Smad7 as a regulator of reepithelialization in vivo and implicates Smad7 as a potential novel target for the treatment of cutaneous wounds.
Project description:Impaired wound healing is a major source of morbidity in diabetic patients. Poor outcome has, in part, been related to increased inflammation, poor angiogenesis, and deficiencies in extracellular matrix components. Despite the enormous impact of these chronic wounds, effective therapies are lacking. Here, we showed that the topical application of recombinant matricellular protein angiopoietin-like 4 (ANGPTL4) accelerated wound reepithelialization in diabetic mice, in part, by improving angiogenesis. ANGPTL4 expression is markedly elevated upon normal wound injury. In contrast, ANGPTL4 expression remains low throughout the healing period in diabetic wounds. Exogenous ANGPTL4 modulated several regulatory networks involved in cell migration, angiogenesis, and inflammation, as evidenced by an altered gene expression signature. ANGPTL4 influenced the expression profile of endothelial-specific CD31 in diabetic wounds, returning its profile to that observed in wild-type wounds. We showed ANGPTL4-induced nitric oxide production through an integrin/JAK/STAT3-mediated upregulation of inducible nitric oxide synthase (iNOS) expression in wound epithelia, thus revealing a hitherto unknown mechanism by which ANGPTL4 regulated angiogenesis via keratinocyte-to-endothelial-cell communication. These data show that the replacement of ANGPTL4 may be an effective adjunctive or new therapeutic avenue for treating poor healing wounds. The present finding also confirms that therapeutic angiogenesis remains an attractive treatment modality for diabetic wound healing.
Project description:Burn injuries are a leading cause of morbidity including prolonged hospitalization, disfigurement, and disability. Currently there is no Food and Drug Administration-approved burn therapeutics. A clinical distinction of burn injuries from other acute wounds is the event of the so-called secondary burn wound progression within the first week of the injury, in which a burn expands horizontally and vertically from its initial boundary to a larger area. Therefore, an effective therapeutics for burns should show dual abilities to prevent the burn wound progression and thereafter promote burn wound healing. Herein we report that topically applied F-5 fragment of heat shock protein-90? is a dual functional agent to promote burn wound healing in pigs. First, F-5 prevents burn wound progression by protecting the surrounding cells from undergoing heat-induced caspase 3 activation and apoptosis with increased Akt activation. Accordingly, F-5-treated burn and excision wounds show a marked decline in inflammation. Thereafter, F-5 accelerates burn wound healing by stimulating the keratinocyte migration-led reepithelialization, leading to wound closure. This study addresses a topical agent that is capable of preventing burn wound progression and accelerating burn wound healing.
Project description:Wound reepithelialization is an evolutionarily conserved process in which skin cells migrate as sheets to heal the breach and is critical to prevent infection but impaired in chronic wounds. Integrin heterodimers mediate attachment between epithelia and underlying extracellular matrix and also act in large signaling complexes. The complexity of the mammalian wound environment and evident redundancy among integrins has impeded determination of their specific contributions to reepithelialization. Taking advantage of the genetic tools and smaller number of integrins in Drosophila, we undertook a systematic in vivo analysis of integrin requirements in the reepithelialization of skin wounds in the larva. We identify ?PS2-?PS and ?PS3-?PS as the crucial integrin dimers and talin as the only integrin adhesion component required for reepithelialization. The integrins rapidly accumulate in a JNK-dependent manner in a few rows of cells surrounding a wound. Intriguingly, the integrins localize to the distal margin in these cells, instead of the frontal or lamellipodial distribution expected for proteins providing traction and recruit nonmuscle myosin II to the same location. These findings indicate that signaling roles of integrins may be important for epithelial polarization around wounds and lay the groundwork for using Drosophila to better understand integrin contributions to reepithelialization.