Minimally invasive scar release by autologous adipose tissue transfer for post-traumatic neuropathic pain.

Introduction: Addressing post traumatic lower limb neuropathic pain is challenging across medical specialties. To address this potentially devastating condition, several invasive and non-invasive approaches have been proposed with inconsistent results. Adipose fat transfer (AFT), also known as fat grafting, is a regenerative medicine technique in which a patient's own fat is harvested from one area of the body (usually through liposuction) and then injected into another area for various purposes, such as aesthetic contour enhancement or reconstruction and regeneration of scarred tissues. Methods: We analyze the effects of fat grafting for neuropathic pain combined with neuroma excision (hybrid technique, hAFT) or alone (AFT). A retrospective review was conducted on 22 patients with neuropathic lower limb pain, after trauma or orthopedic surgery treated with hAFT (n = 9) or AFT (n = 13). Results: Reduction in VAS scale more than 50 % was observed in 6 patients (66 %) treated with hybrid technique and in eleven patients (85 %) treated with AFT alone. Among these, complete pain reduction (>91 %) was achieved in 33.3 % of hAFT and 54 % of AFT technique. A 3.2 points reduction in VAS was found in the hAFT group versus 5.8 points in the AFT group (p = 0.035). Conclusion: This pioneering use of AFT emerges as a minimally invasive breakthrough, promising significant improvement in reconstructing scarred subcutaneous tissue and managing neuropathic pain.

Minimally invasive nerve and artery sparing surgical approach for temporal migraines.

Background: Temporal migraines (TM) present with throbbing, pulsating headaches in the temporal area. Different surgical techniques ranging from resecting the auriculotemporal nerve (ATN) and or ligating the superficial temporal artery (STA) have shown similar good results to decrease TM symptoms. No conclusive data supports a specific disease of the STA in TM patients. A minimally invasive technique is proposed to preserve both vascular and nerve structures. Methods: Patients with drug resistant TM were selected and treated with two techniques: nerve sparing and nerve and artery sparing. The study included 57 patients with TM, with an average age of 47.5 years. TM improvement was quantified after at least one year of follow up time. STA biopsies were sent for histological analysis. Results: Forty-two patients underwent nerve-sparing decompression, with a therapeutic success rate of 78.6%, corresponding to 22.1 days with migraine per month decreasing to 6.2. Histological analysis of the STA showed varying degrees of endofibrosis in 75% of the samples. Histological results do not correlate with the intensity of symptoms before or after surgery. Fifteen patients underwent nerve and artery sparing arteriolysis, with an overall therapeutic success rate of 86.6% of which 80% had >90% improvement. The average migraine days dropped from 24 to 2.5 days per month in this group. Conclusion: Minimally invasive nerve sparing approaches are an effective and safe treatment to improve drug resistant TM symptoms. Endofibrosis of the STA was present in 75% of the cases, but it was found to be unrelated to pre-operative symptoms and outcome. Results are promising, but the limited numbers of patients treated with artery and nerve sparing technique needs further investigations.

Minimally Invasive Nerve- and Muscle-Sparing Surgical Decompression for Occipital Neuralgia.

BACKGROUND: Occipital neuralgia is a well-defined type of headache, and its treatment algorithm is still debated across medical specialties. From the analysis of the literature, it appears that surgical decompression of the occipital nerves is the most effective invasive approach to improve the quality of life of patients with occipital neuralgia refractory to medications. The authors describe here a minimally invasive nerve- and muscle-sparing technique to decompress the occipital nerves. METHODS: The results in terms of reduction of migraine days per month, use of medications, pain evaluation, and decrease in Migraine Headache Index were analyzed by means of a retrospective chart review of 87 patients who underwent nerve- and muscle-sparing surgical decompression of the greater and lesser monolateral or bilateral occipital nerves in their institution and were followed up for at least 12 months. The surgical technique is described in detail. RESULTS: Surgical decompression significantly reduced occipital neuralgia burden (at least 50% improvement) in 91% of patients, with 45% reporting a complete remission of occipital pain. Days with pain per month decreased by 80%, chronic background pain intensity decreased by 81%, and pain intensity during crisis decreased by 76%. Accordingly, drug use dropped by approximately 70%. Only minor complications were reported in four patients. CONCLUSIONS: The described technique could contribute to and further support surgical decompression as the first option among the invasive approaches to treat occipital neuralgia. Results corroborate previous findings, adding a less-invasive, nerve- and muscle-sparing approach. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Selective denervation of the corrugator supercilii muscle for the treatment of idiopatic trigeminal neuralgia purely paroxysmal distributed in the supraorbital and suprathrochlear dermatomes.

INTRODUCTION: Idiopatic trigeminal neuralgia purely paroxysmal (ITNp) distributed in the supraorbital and suprathrochlear dermatomes (SSd), refractory to conventional treatments have been linked to the hyperactivity of the corrugator supercilii muscle (CSM). In these patients, the inactivation of the CSM via botulinum toxin type A (BTA) injections has been proven to be safe and effective in reducing migraine burden. The main limitation of BTA is the need of repetitive injections and relative high costs. Based on the study of the motor innervation of the CSM, we describe here an alternative approach to improve these type of migraines, based on a minimally invasive denervation of the CSM. MATERIALS AND METHODS: Motor innervation and feasibility of selective CSM denervation was first studied on fresh frozen cadavers. Once the technique was safely established, 15 patients were enrolled. To be considered eligible, patients had to meet the following criteria: positive response to BTA treatment, migraine disability assessment score > 24, > 15 migraine days/month, no occipital/temporal trigger points and plausible reasons to discontinue BTA treatment. Pre- and post- operative migraine headache index (MHI) were compared, and complications were classified following the Clavien-Dindo classification (CDC). RESULTS: Fifteen patients (9 females and 6 males) underwent the described surgical procedure. The mean age was 41 ± 10 years. Migraine headache episodes decreased from 24 ± 4 day/month to 2 ± 2 (p < 0.001) The MHI decreased from 208 ± 35 to 10 ± 11 (p < 0.001). One patient (7%) had a grade I complication according to the CDC. No patient needed a second operative procedure. CONCLUSIONS: Our findings suggest that the selective CSM denervation represents a safe and minimally invasive approach to improve ITNp distributed in the SSd associated with CSM hyperactivation. TRIAL REGISTRATION: The data collection was conducted as a retrospective quality assessment study and all procedures were performed in accordance with the ethical standards of the national research committee and the 1964 Helsinki Declaration and its later amendments.

Chronic Knee and Ankle Pain Treatment through Selective Microsurgical Approaches: A Minimally Invasive Option in the Treatment Algorithm for Refractory Lower Limb Pain.

BACKGROUND: Injury or compression of a sensory nerve is an under-reported source of disabling pain in the lower limb. It is known that peripheral nerve microsurgeons can reconstruct and rewire injured nerves to relieve chronic pain but this option remains not completely understood and ignored by most orthopaedic surgeons, neurologists, and pain therapists. In this paper, we describe our experience with knee and ankle peripheral nerve surgery to improve the condition of patients suffering from chronic, posttraumatic lower limb pain. PATIENTS AND METHODS: Between 2015 and 2018, a retrospective investigation was performed including patients who underwent ankle and knee nerve surgery for posttraumatic chronic pain with at least 1 year of follow-up. Previous surgeries or type of trauma, pain location and characteristics, type of operation (reconstruction, decompression, or denervation), and number of nerves operated were listed. Selective neuropathy was confirmed by ultrasound-guided nerve blocks. Outcome was assessed combining the visual analogue scale (VAS) score at rest and during movement, functional indexes, drug use, and ability to work. RESULTS: A total of 34 patients (14 knee and 20 ankle) were included in this study. A statistically significant difference (p < 0.05%) in postoperative pain at rest and during activity was seen in both groups of patients. Good to excellent outcomes were recorded in 92 and 80% of patients treated, respectively, at the knee and the ankle levels. No major complications occurred, while a secondary procedure due to neuroma recurrence was necessary in seven patients (three knees and four ankles). CONCLUSION: Peripheral nerve microsurgery is a useful and minimally invasive tool to be added in the algorithm of treatment of chronic knee and ankle pain. Physicians should be trained to suspect a neuropathic origin of pain in absence of musculoskeletal signs of pathology, especially following trauma or surgeries.

Embryonic stem cell extracts improve wound healing in diabetic mice.

AIMS/HYPOTHESIS: Impaired wound healing significantly impacts morbidity and mortality in diabetic patients, necessitating the development of novel treatments to improve the wound healing process. We here investigated the topical use of acellular embryonic stem cell extracts (EXTs) in wound healing in diabetic db/db mice. METHODS: Wounds were induced in diabetic db/db mice, which were subsequently treated with EXTs, with 3T3 fibroblast cell line protein extracts (3T3XTs) or with saline as a control. Pathology and mechanistic assays were then performed. RESULTS: The in vivo topical administration of EXTs facilitates wound closure, contraction and re-epithelialization. Moreover, EXTs reduced the number of wound-infiltrating CD45+ inflammatory cells and increased the rate of repair and of angiogenesis as compared to controls. Interestingly, the EXT effect was partly enhanced by the use of a collagen-based biocompatible scaffold. In vivo, topical administration of EXTs increased the percentage of regulatory T cells in the wounded tissue, while in vitro EXT treatment reduced T cell-mediated IFN-γ production. Proteomic screening revealed 82 proteins differentially segregating in EXTs as compared to 3T3 extracts, with APEX1 identified as a key player for the observed immunomodulatory effect of EXTs. CONCLUSIONS: EXTs are endowed with immunoregulatory and anti-inflammatory properties; their use improves wound healing in diabetic preclinical models.

The Greater Occipital Nerve and Obliquus Capitis Inferior Muscle: Anatomical Interactions and Implications for Occipital Pain Syndromes.

BACKGROUND: The compression/injury of the greater occipital nerve has been identified as a trigger of occipital headaches. Several compression points have been described, but the morphology of the myofascial unit between the greater occipital nerve and the obliquus capitis inferior muscle has not been studied yet. METHODS: Twenty fresh cadaveric heads were dissected, and the greater occipital nerve was tracked from its emergence to its passage around the obliquus capitis inferior. The intersection point between the greater occipital nerve and the obliquus capitis inferior, and the length and thickness of the obliquus capitis inferior, were measured. In addition, the nature of the interaction and whether the nerve passed through the muscle were also noted. RESULTS: All nerves passed either around the muscle loosely (type I), incorporated in the dense superficial muscle fascia (type II), or directly through a myofascial sleeve within the muscle (type III). The obliquus capitis inferior length was 5.60 ± 0.46 cm. The intersection point between the obliquus capitis inferior and the greater occipital nerve was 6.80 ± 0.68 cm caudal to the occiput and 3.56 ± 0.36 cm lateral to the midline. The thickness of the muscle at its intersection with the greater occipital nerve was 1.20 ± 0.25 cm. Loose, tight, and intramuscular connections were found in seven, 31, and two specimens, respectively. CONCLUSIONS: The obliquus capitis inferior remains relatively immobile during traumatic events, like whiplash injuries, placing strain as a tethering point on the greater occipital nerve. Better understanding of the anatomical relationship between the greater occipital nerve and the obliquus capitis inferior can be clinically useful in cases of posttraumatic occipital headaches for diagnostic and operative planning purposes.

Double Triangular Cartilage Excision Otoplasty.

BACKGROUND: Prominent ears have a negative impact on patients' psychosocial well-being. There are numerous surgical correction techniques described, but the majority have high complication and recurrence rates. In this article, the authors present a stitchless technique that takes advantage of different approaches to minimize complications and recurrences. METHODS: The authors prospectively studied patients who underwent a bilateral double triangular cartilage excision otoplasty in an outpatient setting. This prominent ear setback technique is based on a specific cartilage excision to mechanically collapse the ear without using any sutures through the cartilage. Postoperatively, surgical complications were classified according to the Clavien-Dindo classification. The patient-reported outcome was analyzed after completion of a questionnaire 6 months after surgery, and another clinical follow-up appointment was arranged 12 months after surgery. RESULTS: Sixty patients (120 ears) with an average age of 24 years were studied, and the overall complication rate was 6 percent. Three ears (2.5 percent) developed superficial cutaneous necrosis at the anterior concha, and one ear (1 percent) required an additional correction for unsatisfactory pinna rotation. Also, 2.5 percent of the patients experienced a grade I complication, and 1 percent of the patients experienced a grade IIIa complication according to the Clavien-Dindo classification. No wound infections, hematomas, or hypertrophic scars were observed, and the patient-reported outcome showed satisfaction with the results. CONCLUSIONS: The authors' findings show that the double triangular cartilage excision otoplasty is a safe procedure with low complication/recurrence rates. This stitchless technique should be included in each surgeon's repertoire as an alternative to previously published surgical techniques. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Minimal Undermining Suspension Technique (MUST): Combined Eyebrow and Mid-face Lift via Temporal Access.

BACKGROUND: Less downtime following esthetic interventions leads to the popularity of injectable solutions for facial rejuvenation treatments. Surgical interventions for esthetic purposes are usually associated with higher complication rates and longer recovery times when compared to less invasive treatments. Here we present for the first time a minimally invasive surgical technique for a simultaneous mid-face and eyebrow lift using one small temporal incision. MATERIALS AND METHODS: We prospectively studied patients who underwent facial rejuvenation using a minimal undermining suspension technique (MUST) in an outpatient setting. Postoperatively, surgical complications were classified according to the Clavien-Dindo classification. Preoperatively and at 12-month post-intervention, patient-reported outcomes were described using the FACE-Q questionnaire. Pre- and postoperative pictures were compared using MERZ Aesthetic Scales. RESULTS: Fifty-five patients (50 females and 5 males) with a mean age of 47 years were studied. The overall complication rate was 19%, whereas 18 patients (15%) developed an ecchymosis of the orbicular temporal region and two patients (4%) developed a dimple caused by the anchor of the suture. No displacements of the palpebral rim or injuries to the facial nerve were observed. Recovery time was in average 7 days and no long-term complications were seen. Both, the FACE-Q and the MERZ Aesthetic Scales showed significant improvements at 12-month post-surgery (p < 0.01). CONCLUSIONS: The MUST can be used as a minimal invasive procedure for facial rejuvenation with a short downtime and low complication rate. The discreet temporal access and the MUST dissector allowed a safe dissection of anatomical tissue planes to perform safely a mid-face and eyebrow lift. Further studies are warranted to prove long-term outcomes. LEVEL OF EVIDENCE IV: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the A3 online Instructions to Authors. www.springer.com/00266 .

Anatomic and Compression Topography of the Lesser Occipital Nerve.

BACKGROUND: The surgical treatment of occipital headaches focuses on the greater, lesser, and third occipital nerves. The lesser occipital nerve (LON) is usually transected with relatively limited available information regarding the compression topography thereof and how such knowledge may impact surgical treatment. METHODS: Eight fresh frozen cadavers were dissected focusing on the LON in relation to 3 clinically relevant compression zones. The x axis was a line drawn through the occipital protuberance (OP) and the y axis, the posterior midline (PM). In addition, a prospectively collected cohort of 36 patients who underwent decompression of the LON is presented with their clinical results, including migraine headache index scores. RESULTS: The LON was found in compression zone 1, with a mean of 7.8 cm caudal to the OP and 6.3 cm lateral to the PM. The LON was found at the midpoint of compression zone 2, with an average of 5.5 cm caudal to the OP and 6.2 cm lateral to the PM. At compression zone 3, the medial-most LON branch was located approximately 1 cm caudal to the OP and 5.35 cm lateral to the PM, whereas the lateral-most branch was identified 1 cm caudal to the OP and 6.5 cm lateral to the PM. Of the 36 decompression patients analyzed, only 5 (14%) required neurectomy as the remainder achieved statistically significant improvements in migraine headache index scores postoperatively. CONCLUSION: The knowledge of LON anatomy can aid in nerve dissection and preservation, thereby leading to successful outcomes without requiring neurectomy.

Novel micropatterns mechanically control fibrotic reactions at the surface of silicone implants.

Over the past decade, various implantable devices have been developed to treat diseases that were previously difficult to manage such diabetes, chronic pain, and neurodegenerative disorders. However, translation of these novel technologies into clinical practice is often difficult because fibrotic encapsulation and/or rejection impairs device function after body implantation. Ideally, cells of the host tissue should perceive the surface of the implant being similar to the normal extracellular matrix. Here, we developed an innovative approach to provide implant surfaces with adhesive protein micropatterns. The patterns were designed to promote adhesion of fibroblasts and macrophages by simultaneously suppressing fibrogenic activation of both cell types. In a rat model, subcutaneously implanted silicone pads provided with the novel micropatterns caused 6-fold lower formation of inflammatory giant cells compared with clinical grade, uncoated, or collagen-coated silicone implants. We further show that micropatterning of implants resulted in 2-3-fold reduced numbers of pro-fibrotic myofibroblast by inhibiting their mechanical activation. Our novel approach allows controlled cell attachment to implant surfaces, representing a critical advance for enhanced biointegration of implantable medical devices.

Poly-N-acetyl glucosamine fibers induce angiogenesis in ADP inhibitor-treated diabetic mice.

BACKGROUND: It has been previously demonstrated that short-fiber poly-N-acetyl-glucosamine (sNAG) nanofibers specifically interact with platelets, are hemostatic, and stimulate diabetic wound healing by activating angiogenesis, cell proliferation, and reepithelialization. Platelets play a significant physiologic role in wound healing. The influence of altered platelet function by treatment with the ADP inhibitor Clopidogrel (CL) on wound healing and the ability of sNAG to repair wounds in diabetic mice treated with CL were studied. METHODS: Dorsal 1 cm2 skin wounds were excised on genetically diabetic 8-week to 12-week-old, Lep/r-db/db male mice, and wound healing kinetics were determined. Microscopic analysis was performed for angiogenesis (PECAM-1) and cell proliferation (Ki67). Mice were either treated with CL (P2Y12 ADP receptor antagonist, CL) or saline solution (NT). CL wounds were also treated with either a single application of topical sNAG (CL-sNAG) or were left untreated (CL-NT). RESULTS: CL treatment did not alter wound healing kinetics, while sNAG induced faster wound closure in CL-treated mice compared with controls. CL treatment of diabetic mice caused an augmentation of cell proliferation and reduced angiogenesis compared with nontreated wounds. However, sNAG reversed the effects of CL on angiogenesis and partially reversed the effect on cell proliferation in the wound beds. The sNAG-treated wounds in CL-treated mice showed higher levels of cell proliferation and not did inhibit angiogenesis. CONCLUSIONS: CL treatment of diabetic mice decreased angiogenesis and increased cell proliferation in wounds but did not influence macroscopic wound healing kinetics. sNAG treatment did not inhibit angiogenesis in CL-treated mice and induced faster wound closure; sNAG technology is a promising strategy to facilitate the healing of complex bleeding wounds in CL-treated diabetic patients.

Waveform modulation of negative-pressure wound therapy in the murine model.

BACKGROUND: Negative-pressure wound therapy applied with a porous foam interface has been shown to accelerate granulation-tissue formation when a cyclic application mode of suction is applied, but the optimal waveform has not been determined. The authors hypothesized that changes in the suction waveform applied to wounds would modulate the biological response of granulation tissue formation. METHODS: A vacuum-assisted closure device (Kinetic Concepts, Inc., San Antonio, Texas) was applied to full-thickness wounds in 48 male diabetic mice (C57BL/KsJ-Lepr db), which were treated with six different waveforms: square waveforms of 125 mmHg of suction for 2 minutes, alternating with 50 mmHg of suction for 2 minutes, 5 minutes, or 10 minutes; triangular waveform with a 7-minute period oscillating between 50 and 125 mmHg; and static suction at 125 mmHg or static suction at 0 mmHg (occlusive dressing). Wounds were quantitatively evaluated for granulation tissue thickness as well as the number of proliferating cells and the number of blood vessels of the newly formed granulation tissue. RESULTS: At 7 days, the continuous and triangular waveforms induced the thickest granulation tissue, with high rates of cellular proliferation and blood vessel counts compared with square wave and occlusive dressing control wounds. Decreasing square waveform frequency significantly increased granulation tissue thickness, cellular proliferation, and blood vessel counts. CONCLUSIONS: Waveform modulation has a significant effect on granulation tissue formation, angiogenesis, and cellular proliferation in excisional wounds in diabetic mice. The rapid change in pressure seen in our square wave model may be detrimental to granulation tissue formation.

Combination of stromal cell-derived factor-1 and collagen-glycosaminoglycan scaffold delays contraction and accelerates reepithelialization of dermal wounds in wild-type mice.

While dermal substitutes can mitigate scarring and wound contraction, a significant drawback of current dermal replacement technologies is the apparent delay in vascular ingrowth compared with conventional skin grafts. Herein, we examined the effect of the chemokine stromal cell-derived factor-1 (SDF-1) on the performance of a porous collagen-glycosaminoglycan dermal analog in excisional wounds in mice. C57BL/6 mice with 1 cm × 1 cm dorsal full-thickness wounds were covered with a collagen-glycosaminoglycan scaffold, followed by four daily topical applications of 1 µg SDF-1 or phosphate-buffered saline vehicle. Some animals were also pretreated with five daily doses of 300 mg/kg granulocyte colony-stimulating factor. Animals treated with SDF-1 and no granulocyte colony-stimulating factor reepithelialized 36% faster than vehicle controls (16 vs. 25 days), and exhibited less wound contraction on postwounding day 18 (∼ 35% greater wound area) plus three-fold longer neoepidermis formed than controls. Conversely, granulocyte colony-stimulating factor promoted contraction and no epidermal regeneration. Early (postwounding Day 3) inflammatory cell infiltration in the SDF-1-treated group was 86% less, while the fraction of proliferating cells (positive Ki67 staining) was 32% more, when compared with controls. These results suggest that SDF-1 simultaneously delays contraction and promotes reepithelialization and may improve the wound-healing performance of skin substitutes.

The mobilization and effect of endogenous bone marrow progenitor cells in diabetic wound healing.

Diabetic patients suffer from impaired wound healing, characterized by only modest angiogenesis and cell proliferation. Stem cells may stimulate healing, but little is known about the kinetics of mobilization and function of bone marrow progenitor cells (BM-PCs) during diabetic wound repair. The objective of this study was to investigate the kinetics of BM-PC mobilization and their role during early diabetic wound repair in diabetic db/db mice. After wounding, circulating hematopoietic stem cells (Lin(-)c-Kit(+)Sca-1(+)) stably increased in the periphery and lymphoid tissue of db/db mice compared to unwounded controls. Peripheral endothelial progenitor cells (CD34(+)VEGFR(+)) were 2.5- and 3.5-fold increased on days 6 and 10 after wounding, respectively. Targeting the CXCR4-CXCL12 axis induced an increased release and engraftment of endogenous BM-PCs that was paralleled by an increased expression of CXCL12/SDF-1α in the wounds. Increased levels of peripheral and engrafted BM-PCs corresponded to stimulated angiogenesis and cell proliferation, while the addition of an agonist (GM-CSF) or an antagonist (ACK2) did not further modulate wound healing. Macroscopic histological correlations showed that increased levels of stem cells corresponded to higher levels of wound reepithelialization. After wounding, a natural release of endogenous BM-PCs was shown in diabetic mice, but only low levels of these cells homed in the healing tissue. Higher levels of CXCL12/SDF-1α and circulating stem cells were required to enhance their engraftment and biological effects. Despite controversial data about the functional impairment of diabetic BM-PCs, in this model our data showed a residual capacity of these cells to trigger angiogenesis and cell proliferation.

Use of the parabiotic model in studies of cutaneous wound healing to define the participation of circulating cells.

Previous experimental studies to assess the contribution of blood-borne circulating (BBC) cells to cutaneous wound healing have relied on discontinuous pulsing of labeled BBC elements or bone marrow transplant protocols. Such approaches do not allow the examination of stable BBC cells that have matured in a physiologically normal host. We have used a parabiotic murine model for cutaneous wound healing to evaluate the relative contribution of stable populations of peripheral blood cells expressing the green fluorescent protein (GFP) transgene in otherwise normal animals. Circulating cells (mature and immature) expressing the GFP transgene were easily detected and quantified in wounds of GFP- parabiotic twins during all evaluated stages of the healing response. Using multiple antibody probes, the relative contribution of various subsets of BBC cells could be comparatively assessed. In early wounds, some cells expressing mesenchymal epitopes were documented to be of hematopoietic origin, indicating the utility of this model in assessing cell plasticity in the context of tissue regeneration and repair. Application of this approach enables further investigation into the contribution of peripheral blood in normal and abnormal healing responses.

In vivo acceleration of skin growth using a servo-controlled stretching device.

Tension is a principal force experienced by skin and serves a critical role in growth and development. Optimal tension application regimens may be an important component for skin tissue engineering and dermatogenesis. In this study, we designed and tested a novel servo-controlled skin-stretching device to apply predetermined tension and waveforms in mice. The effects of static and cyclical stretching forces were compared in 48 mice by measuring epidermal proliferation, angiogenesis, cutaneous perfusion, and principal growth factors using immunohistochemistry, real-time reverse transcriptase-polymerase chain reaction, and hyperspectral imaging. All stretched samples had upregulated epidermal proliferation and angiogenesis. Real-time reverse transcriptase-polymerase chain reaction of epidermal growth factor, transforming growth factor beta1, and nerve growth factor demonstrated greater expression in cyclically stretched skin when compared to static stretch. Hypoxia-induced factor 1alpha was significantly upregulated in cyclically stretched skin, but poststretch analysis demonstrated well-oxygenated tissue, collectively suggesting the presence of transient hypoxia. Waveform-specific mechanical loads may accelerate tissue growth by mechanotransduction and as a result of repeated cycles of temporary hypoxia. Further analysis of mechanotransduction signaling pathways may provide additional insight to improve skin tissue engineering methods and optimize our device.

Quiescent platelets stimulate angiogenesis and diabetic wound repair.

INTRODUCTION: Platelets partake in hemostasis, wound healing, and tumor growth. Although platelet-rich-plasma (PRP) has been used in surgery for several years, its mechanism of action and application methods are still poorly characterized. MATERIALS AND METHODS: A single unit of human platelets obtained by plateletpheresis was diluted in plasma and divided into three equal volumes. One volume was stored at room temperature as fresh platelets (RT), another volume was frozen by storage at -80 degrees C (FZ), and the third volume was frozen at -80 degrees C with 6% DMSO (FZ6). Plasma (PL) was used as control. Using flow cytometry, platelets were tested for platelet glycoprotein GPIb and annexin V binding, as survival and activation markers, respectively. Hemostatic function was assessed by thromboelastometry. In vivo, platelets were topically applied on 1 cm,(2) full-thickness wounds on db/db mice (n = 10/group) and healing was staged microscopically and macroscopically. RESULTS: All platelet preparations showed hemostatic ability. RT platelets were GPIb positive (nonactivated-quiescent platelets) and stimulated angiogenesis by threefold, and cell proliferation by fourfold in vivo. FZ platelets were positive for annexin V, indicating activated platelets and, in vivo, increased only wound granulation. FZ6 platelets contained 30% nonactivated-quiescent and 50% activated platelets and stimulated granulation, angiogenesis, cell proliferation, and promoted re-epithelialization in vivo. CONCLUSIONS: Platelets showed distinct mechanisms to induce hemostasis and wound healing. Quiescent platelets are required to induce angiogenesis in vivo. Platelets stored at room temperature and frozen with 6% DMSO and stored at -80 degrees C achieved optimal wound healing in diabetic mice.

Poly-N-acetyl glucosamine nanofibers: a new bioactive material to enhance diabetic wound healing by cell migration and angiogenesis.

INTRODUCTION: In several fields of surgery, the treatment of complicated tissue defects is an unsolved clinical problem. In particular, the use of tissue scaffolds has been limited by poor revascularization and integration. In this study, we developed a polymer, poly-N-acetyl-glucosamine (sNAG), with bioactive properties that may be useful to overcome these limitations. OBJECTIVE: To develop a scaffold-like membrane with bioactive properties and test the biologic effects in vitro and in vivo in diabetic wound healing. METHODS: In vitro, cells-nanofibers interactions were tested by cell metabolism and migration assays. In vivo, full thickness wounds in diabetic mice (n = 15 per group) were treated either with sNAG scaffolds, with a cellulosic control material, or were left untreated. Wound healing kinetics, including wound reepithelialization and wound contraction as well as microscopic metrics such as tissue growth, cell proliferation (Ki67), angiogenesis (PECAM-1), cell migration (MAP-Kinase), and keratinocyte migration (p 63) were monitored over a period of 28 days. Messenger RNA levels related to migration (uPAR), angiogenesis (VEGF), inflammatory response (IL-1beta), and extracellular matrix remodeling (MMP3 and 9) were measured in wound tissues. RESULTS: sNAG fibers stimulated cell metabolism and the in vitro migratory activity of endothelial cells and fibroblasts. sNAG membranes profoundly accelerated wound closure mainly by reepithelialization and increased keratinocyte migration (7.5-fold), granulation tissue formation (2.8-fold), cell proliferation (4-fold), and vascularization (2.7-fold) compared with control wounds. Expression of markers of angiogenesis (VEGF), cell migration (uPAR) and ECM remodeling (MMP3, MMP9) were up-regulated in sNAG treated wounds compared with controls. CONCLUSIONS: The key mechanism of the bioactive membranes is the cell-nanofiber stimulatory interaction. Engineering of bioactive materials may represent the clinical solution for a number of complex tissue defects.

Tumors stimulate platelet delivery of angiogenic factors in vivo: an unexpected benefit.

The interaction between platelets and the tumor microenvironment results in the modulation of angiogenesis, although the mechanisms governing this regulation remain unclear. This study explores the differences in the communication between wounded tissues and healthy, tumor-conditioned, and frozen platelets. Platelet-rich plasma obtained from healthy (PRP) or tumor-bearing (TPRP) mice was applied to dorsal, full-thickness wounds on diabetic mice. Wound healing was evaluated using macroscopic criteria and a staging system based on angiogenesis and stromal cell proliferation. Proteomic analysis was used to compare the levels of angiogenic proteins contained in the platelet preparations. TPRP-treated wounds reached 90% wound closure 5.6 to 9.5 days earlier than PRP-treated and nontreated wounds, respectively. TPRP induced a fourfold increase in stromal cell proliferation compared with nontreated wounds, and a 2.5-fold increase compared with PRP-treated wounds. TPRP induced the highest stimulation of angiogenesis with a fourfold increase compared with nontreated controls. On day 21, wounds treated with TPRP showed a typical architecture with thick collagen bundles. Although the levels of angiogenesis regulators detected via SELDI-ToF were similar between the PRP and TPRP treatment regimens, the enhanced healing capacity of TPRP suggests improved platelet delivery as indicated by frozen TPRP preparations that had lost most of their pro-angiogenic drive. In conclusion, these results show that intact tumor-conditioned platelets display an improved ability to deliver angiogenesis regulators to wounded tissues.