Two Pre-Expanded Free Flaps Used for Auricular Reconstruction and Ipsilateral Facial Scar Release.

OBJECTIVES: Auricular reconstruction is a great challenge for surgeons to achieve good aesthetic outcomes when adjacent tissues were burned. Compared with pedicle flap therapies, there are some advantages of pre-expanded free flaps for ear rebuilding, such as thinner layer tissues for aesthetic requirements of delicate auricular structures and less donor site deformity. In this study, the authors introduced 6 sequential surgical procedures for total auricular reconstruction with severe ipsilateral facial scar. METHODS: Pre-expanded deltopectoral flap was used to release periauricular contracture and repair facial scar. The injured ear was restored by expanded forearm flap including autologous cartilage framework. The surgical procedures were lasted more than 2 years. An 8 and half year's follow-up was performed from November 2012 to April 2021. The clinical data and surgical techniques were recorded and analyzed. RESULTS: The patient was satisfied with the aesthetic outcomes of the new ear. The skin texture and color of the grafts were approximately matched to the recipient sites. Facial expression was not affected severely. Sensations of the transferred flap and new ear had partially recovered. The donor sites were recovered without severe complication. CONCLUSIONS: The pre-expanded free forearm flap is a feasible method for total ear reconstruction when local flap therapies could not be applied. Repair of ipsilateral facial scar is beneficial for auricular procedures.

Improving the Retention Rate of Fat Grafts in Recipient Areas via Botulinum Toxin A Treatment.

BACKGROUND: Improving the retention rate of transplanted fat is, currently, of great concern. Partial immobilization, angiogenesis, and adipose tissue-derived stem cells, all proven to be influenced by botulinum toxin A (BTX-A), are significant in fat graft retention. OBJECTIVES: The authors sought to determine the impact of BTX-A on fat grafts. METHODS: Our study included 12 Sprague Dawley rats and each rat's hind limbs were randomly designated as the BTX-A side and control side. We injected 0.2 mL of BTX-A-treated fat into the quadriceps femoris and subcutaneous space of the BTX-A sides. This was also done for the control sides but with untreated fat. We performed electroneuromyography of recipient muscles at 1 week post-operation. The rats were euthanized at 12 weeks post-operation and we observed the fat retention rate, the fat's histologic characteristics, and the density of vessels and mature adipocytes. RESULTS: The amplitudes of electroneuromyography were smaller for the BTX-A sides than the control sides. For intramuscularly injected fat, the BTX-A sides had better retention rates and histologic characteristics and a higher density of vessels and mature adipocytes than the control sides. For subcutaneously injected fat, the BTX-A sides had better histologic characteristics and a higher density of vessels and mature adipocytes than the control sides, but the retention rates were not significantly different between the 2 sides. CONCLUSIONS: Injecting BTX-A-treated fat grafts can immobilize the surrounding muscles. BTX-A can improve the density of vessels and mature adipocytes, histologic characteristics of fat grafts, and retention rate of fat grafts transplanted into muscles.

Usnic acid inhibits hypertrophic scarring in a rabbit ear model by suppressing scar tissue angiogenesis.

Hypertrophic scarring is a common condition in the Chinese population; however, there are currently no satisfactory drugs to treat the disorder. Previous studies showed that angiogenesis plays an important role in the early phase of hypertrophic scarring and inhibition of angiogenesis has been reported as an effective strategy for anti-hypertrophic scar therapy. A recent study showed that usnic acid (UA), an active compound found mainly in lichens, inhibited tumor angiogenesis both in vivo and in vitro. To investigate the therapeutic effects of UA on hypertrophic scarring and to explore the possible mechanism involved, a rabbit ear hypertrophic scar model was established. Scars were treated once a week for four weeks with UA, DMSO or triamcinolone acetonide acetate. Histological evaluation of hematoxylin and eosin staining indicated that UA significantly inhibited hypertrophic scar formation, with obvious reductions in scar height and coloration. The scar elevation index (SEI) was also evidently reduced. Masson's trichrome staining showed that UA significantly ameliorated accumulation of collagen tissue. Immunohistochemical analysis of CD31 expression showed that UA significantly inhibited scar angiogenesis. In vitro, UA inhibited endothelial cell migration and tube formation as well as the proliferation of both human umbilical vein endothelial cells and scar fibroblast cells. These results provide the first evidence of the therapeutic effectiveness of UA in hypertrophic scar formation in an animal model via a mechanism that involves suppression of scar angiogenesis.

Association of Autologous Fat Injection in Facial Artery With Ophthalmological Complications: An Experimental Animal Study.

IMPORTANCE: Complications caused by autologous fat filling have been reported. Comprehensive knowledge of the possible adverse effects of autologous fat filling is needed. OBJECTIVE: To determine the association of autologous fat filling with ophthalmic function complications. DESIGN, SETTING, AND PARTICIPANTS: Four adult New Zealand white rabbits were killed for a facial anatomy study. Sixty-four adult New Zealand white rabbits underwent fat harvest using the Coleman technique. Autologous fat was minced or digested with collagenase 1 and centrifuged to separate fat lipid and fat granules. Either 0.2 mL or 0.4 mL of minced fat, fat granules, fat lipid, or saline (control) was retrogradely injected into the facial artery of rabbit models. Electroretinography and ophthalmic fundoscopy were performed to measure the retina and fundus artery occlusions 2 weeks after surgery. MAIN OUTCOMES AND MEASURES: Visual impairment, blindness, and death. RESULTS: Injection of 0.2 mL of fat granules, fat lipid, and saline resulted in 100% (8 of 8), 62.5% (5 of 8), and 0 ophthalmic complications, respectively; and 0.4 mL resulted in 87.5% (7 of 8), 12.5% (1 of 8), and 0 ophthalmic complications, respectively. Injection of 0.2 mL and 0.4 mL minced fat led to 100% (8 of 8) ophthalmic complications and death, respectively. The mortality rates were 37.5% (3 of 8), 12.5% (1 of 8), and 0 for 0.2 mL emboli injection, and 100% (8 of 8), 50% (4 of 8), and 0 for 0.4 mL, respectively. CONCLUSIONS AND RELEVANCE: In this study, minced fat injection was associated with more ophthalmic complications than injection of fat granules and fat lipid. Increasing the injection volume of fat tissues could raise the incidence of morbidity and mortality. LEVEL OF EVIDENCE: NA.

TIGIT-Fc alleviates acute graft-versus-host disease by suppressing CTL activation via promoting the generation of immunoregulatory dendritic cells.

Graft-versus-host disease (GVHD) is the most common complication and major limitation of allogeneic hematopoietic stem cell transplantation. The CD226/TIGIT-CD155 signal is critical for the cross-talk between T cells and dendritic cells (DCs). Studies have shown that blockade of the CD226-CD155 interaction, using an anti-CD226 antibody, can significantly ameliorate GVHD. It has also been reported that a TIGIT-Fc fusion protein exerts immunosuppressive effects by binding to CD155 on DCs. Here, we used a mouse allogeneic acute GVHD model to explore the therapeutic potential and mechanism of action of TIGIT-Fc. C57/BL6 and Balb/c mice were used as hematopoietic cell graft donors and recipients, respectively. In the TIGIT-Fc-treated mice, GVHD symptom occurrence and mortality were delayed compared to that in isotype control group mice. Histopathological analyses revealed that following TIGIT-Fc treatment, liver and small intestine tissue damage was reduced with minimal lymphocytic infiltration. The percentage of CD8+IFN-γ+ and CD8+ granzyme B+ cells significantly decreased in the TIGIT-Fc group. Moreover, treatment with TIGIT-Fc, even after the onset of GVHD, ameliorated symptoms and prolonged survival. TIGIT-Fc also inhibited CD8+ T cell activation in vitro; this was dependent on the presence of CD155 on bone marrow-derived dendritic cells (BMDCs) and on IL-10 production. In addition, TIGIT-CD155 ligation triggered both Erk phosphorylation and STAT3 nuclear translocation. These data indicate that TIGIT plays an important role in the development of GVHD and is an ideal molecular target to treat acute GVHD.

Improvement in the Retention Rate of Transplanted Fat in Muscle by Denervation.

BACKGROUND: Improvement in the retention rate of transplanted fat is currently a topic of interest. The retention of transplanted fat relies heavily on the reestablishment of blood supply and the function of the adipose-derived stem cells (ADSCs), which may both be impeded by mechanical force. However, the effect of mechanical force on the retention of adipose implants remains unclear. OBJECTIVES: This study aimed to evaluate the effectiveness of immobilization on fat retention rate. METHODS: Immobilization was carried out by denervation of the hind limb of rats to reduce the mechanical force. Sprague-Dawley (SD) rats were used, and the two hind limbs were assigned at random to the immobilization side and the control side. On average, 0.4 mL of fat was injected into the bilateral muscle and subcutaneous space of the hind limb, and 6 rats were sacrificed at each time point. The outcome measures included the retention rate, the histologic evaluation, and the density of new vessels and proliferative ADSCs. RESULTS: For the muscle fat, the retention rate improved, and more proliferative ADSCs and new vessels were found in the immobilization group. The histologic evaluation between the two sides was of no statistical significance. For the fat in the subcutaneous space, no statistical difference was observed in all the outcome measures between the two sides. CONCLUSIONS: Regional immobilization of the recipient site by denervation can improve the retention of the fat graft in muscles owing to improved density of the new vessels and proliferative ADSCs.

Silibinin Induced Human Glioblastoma Cell Apoptosis Concomitant with Autophagy through Simultaneous Inhibition of mTOR and YAP.

Silibinin, also known as silybin, is the major flavonolignan isolated from Silybum marianum. Although previous reports demonstrated that silibinin exhibits significant tumor suppressor activities in various cancers by promoting cell apoptosis, it was also shown to trigger autophagy to counteract apoptosis induced by exogenous stresses in several types of cells. However, there is no report to address the role of silibinin induced autophagy in human A172 and SR glioblastoma cells. Our study showed that silibinin treatment not only inhibited the metabolic activities of glioblastoma cells but also promoted their apoptosis through the regulation of caspase 3 and PARP-1 in concentration- and time-dependent manners. Meanwhile, silibinin induced autophagy through upregulation of microtubule-associated protein a light chain 3- (LC3-) II. And autophagy inhibition with chloroquine, a lysosomotropic agent, significantly enhanced silibinin induced glioblastoma cell apoptosis. Moreover, silibinin dose-dependently downregulated the phosphorylation levels of mTOR at Ser-2448, p70S6K at Thr-389, and 4E-BP1 at Thr-37/46. Furthermore, the expression of YAP, the downstream effector of Hippo signal pathway, was also suppressed by silibinin. These results suggested that silibinin induced glioblastoma cell apoptosis concomitant with autophagy which might be due to simultaneous inhibition of mTOR and YAP and silibinin induced autophagy exerted a protective role against cell apoptosis in both A172 and SR cells.

Blocking CD226 Promotes Allogeneic Transplant Immune Tolerance and Improves Skin Graft Survival by Increasing the Frequency of Regulatory T Cells in a Murine Model.

BACKGROUND/AIMS: Regulatory T cells (Tregs) play key roles in maintaining peripheral tolerance and preventing autoimmune disease. Treg modulation could be helpful in treating malignancies, autoimmune disease, and allergies, as well as to facilitate organ transplantation. Signals transduced by co-stimulatory molecules are essential for Treg differentiation, homeostasis, and function. One well-known active receptor, CD226, also known as DNAM-1 or PTA1, is an adhesion molecule that interacts primarily with CD155 and is involved in Treg differentiation and immune tolerance to transplanted tissue. METHODS: Anti-CD226 monoclonal antibody (mAb) and truncated recombinant CD226 proteins were employed to manipulate the CD226 signal. Various T cell markers on freshly isolated splenocytes and T lymphocytes were characterized by flow cytometry Cell proliferation was measured by carboxyfluorescein succinimidyl ester dye, mRNA transcripts by q-RT PCR, and protein expression by western blotting. A BALB/c-to-C57BL/6 skin allograft model was used to determine the effects of CD226 blocking treatment. RESULTS: We observed that both intact extracellular domains of CD226 were necessary for functional interaction of the receptor with its ligand CD155, even though one domain was shown to bind CD155 with lower affinity in a solid binding assay. Importantly, CD226 mAb promoted Treg expansion in a mixed lymphocyte culture and inhibited the cytotoxicity of effector cells. In allogeneic skin transplant mice, administering CD226 mAb reduced inflammation and prolonged allogeneic graft survival, with an increase in the frequency of Tregs. CONCLUSIONS: Our results reveal the mechanism underlying CD226-CD155 interactions and indicate that CD226 signals can be manipulated to promote Treg expansion. Moreover, we provide new evidence that suggests the therapeutic potential of anti-CD226 with allogeneic transplantation.

Effect of the vascularized bone components on the survival of vascularized composite allografts.

BACKGROUND: Vascularized composite allograft (VCA), such as hand and face allograft, contains a vascularized bone component that may provide an immunologic benefit and induce tolerance for the simultaneous inclusion of marrow cells and a marrow microenvironment. We developed a chimeric groin cutaneous/femur flap to investigate the effect of vascularized bone marrow on VCA survival and its ability to induce chimerism. METHODS: Brown Norway and Lewis rats were used as donors and recipients, respectively. The experimental groups were as follows: groin flap transplantation alone, flap plus intravenous donor bone marrow cells and flap plus simultaneous femur transplantation. Animals received a nonmyeloablative conditioning regimen that consisted of 7-Gy thymic irradiation, 0.75-mL antilymphocyte serum, and 8-mg-1kg-1d cyclosporine A. The flap survival time, peripheral blood chimerism, and the bone marrow of transplanted femurs were analyzed and compared between groups. RESULTS: Our data showed that the conditioning regimen was effective in T cell ablation. Simultaneous femur transplantation significantly prolonged the median flap survival time (78.8 ± 13.0 d, n = 8) compared with the intravenous bone marrow infusion group (60.9 ± 2.2 d, n = 7) and the control group (58.6 ± 1.3 d, n = 5). Peripheral blood chimerism of 5.81% ± 1.98% was persistently detected for 60 d in recipients of femur transplants but not in the other two groups. Viable bone marrow was confirmed within the transplanted femur on postoperative d 60, but it was gradually replaced by recipient origin cells and eventually developed rejection and fibrosis. CONCLUSIONS: Vascularized bone component plays some protective roles on VCA survival but fails to provide a continuous source of donor cells.

Transfusion of ethylene carbodiimide-fixed donor splenocytes prolongs survival of vascularized skin allografts.

BACKGROUND: Allograft rejection is a major obstacle to the widespread clinical application of vascularized composite allotransplantation. Recent studies revealed a noncytoreductive strategy to protect allografts by the transfusion of ethylene carbodiimide-fixed donor splenocytes (ECDI-SPs). To determine whether this approach offers advantages in protecting skin allografts, we examined the immunological protection of infusing ECDI-SPs with a 30-d administration of rapamycin on the skin allografts of mice. MATERIALS AND METHODS: C57BL/6 recipient mice received BALB/c donor full-thickness skin or vascularized skin transplants at day 0, along with the infusion of donor ECDI-SPs 7 d before and 1 d after allotransplantation and a 30-d course of rapamycin. Recipients received ECDI-untreated splenocytes or C3H allografts as controls. In vitro allostimulatory activity of ECDI-SPs and donor-specific ex vivo hyporesponsiveness were tested. Production of related cytokines (TGF-ß, IL-10, IL-1ß, and TNF-α) and expression of CD4+Foxp3+ regulatory T cells (Tregs) were also examined. RESULTS: Transfusion of ECDI-SPs combined with rapamycin significantly prolonged survival of full-thickness skin (median survival time [MST]: 28 d) and full-thickness skin allografts (MST: 71 d) compared with untreated splenocytes (MSTs: 11 d and 30 d) or C3H allografts (MSTs: 11 d and 38 d). This effect was accompanied by increased production of IL-10 and TGF-ß, decreased production of IL-1ß and TNF-α, and expansion of Tregs in vitro and in vivo. CONCLUSIONS: ECDI-SP infusion combined with short-term rapamycin administration provides a promising approach to prolong the skin allograft survival.

Inhibition of Lymphatic Drainage With a Self-Designed Surgical Approach Prolongs the Vascularized Skin Allograft Survival in Rats.

Vascularized composite allotransplantation (VCA) is an emerging treatment for significant tissue defects. However, VCAs usually consist of multiple highly antigenic skin tissues. Previous studies have shown that the lymphatic system in skin plays important roles in the initiation of immune responses during acute rejection, by transporting T cells and antigen-presenting dendritic cells to regional lymph nodes. Therefore, we designed a new surgical treatment to inhibit lymphatic drainage of skin allografts and investigated whether this approach could promote the survival of allografts and suppress immunological events after transplantation. This procedure was achieved by connecting the vascularized allografts to recipient tissues with only an annular plastic holder, allowing the minimum of allograft contact with recipients. Our results showed that the self-designed treatment for inhibiting lymphatic drainage promoted the survival of allografts, reduced the serum concentration of IL-2, and decreased the percentage of CD4CD25 and CD8CD25 from the lymphatic nodes draining the transplantation region. In conclusion, these data suggest that self-designed surgical approach is effective in inhibiting lymphatic drainage of skin allografts, and the lymphatic system may be new therapeutic targets for developing techniques or drugs against acute rejection after VCAs.

Lipoamide Inhibits NF1 Deficiency-induced Epithelial-Mesenchymal Transition in Murine Schwann Cells.

BACKGROUND AND AIMS: Neurofibromatosis type I (NF1) is one of the most common neurocutaneous syndromes characterized by development of adult neurofibromas which is mainly made up of Schwann cells. The disease is generally accepted to be caused by inactivation mutation of Nf1 gene. And Nf1 deficiency had been reported to lead to ROS overproduction and epithelial-mesenchymal transition (EMT) phenotype. This study was designed to investigate whether excessive ROS conferred to Nf1 deficiency-induced EMT in Schwann cells. METHODS: Colony formation, wound healing assay and transwell assay was used to evaluate the effects of stable Nf1 knockdown in SW10 Schwann cells. Western blot and ROS assay was conducted to explore the molecular mechanisms of Nf1 inactivation in tumorigenesis. Animal experiments were performed to assess the inhibitory effects of lipoamide, which is the neutral amide of α-lipoic acid and functions as a potent antioxidant to scavenge ROS, on Nf1-deficiency tumor growth in vivo. RESULTS: Nf1 knockdown enhanced the cellular capacities of proliferation, migration and invasion, promoted ROS generation, decreased the expression of epithelial surface marker E-cadherin, and up-regulated several EMT-associated molecules in Schwann cells. Moreover, lipoamide dose-dependently inhibited not only Nf1 deficiency-induced EMT but also spontaneous EMT. Furthermore, lipoamide markedly suppresses tumor growth in a mouse model of NF1-associated neurofibroma. CONCLUSIONS: Our results clearly reveal that ROS overproduction is responsible for Nf1 deficiency-induced EMT and plays a crucial role in NF1 tumor growth. The findings presented herein shed light on the potential of antioxidant therapy to prevent the progression of NF1-associated neurofibroma.

Targeting of angiopoietin 2-small interfering RNA plasmid/chitosan magnetic nanoparticles in a mouse model of malignant melanoma in vivo.

The aim of the present study was to observe the in vivo targeting characteristic of angiopoietin 2-small interfering RNA (Ang2-siRNA) plasmid/chitosan magnetic nanoparticles in an established nude mouse model of malignant melanoma (MM) under an external magnetic field. The nude mouse MM model was first established, then divided into 3 groups, including the control group, the non-targeting group and the target group, the control group was given normal saline and the non-targeting and targeting groups were administrated particles through the tail vein; the non-targeting group was not under external magnetic field and the control group and the targeting group were under external magnetic field for 60 min. The mice were then sacrificed and the tumor tissues were stained with hematoxylin and eosin and Prussian blue in order to verify the particle distributions in the tumor tissues. The control group exhibited negative Prussian blue staining in the tumor tissues, the non-targeting group demonstrated weakly positive Prussian blue staining in tumor tissues and the targeting group revealed strongly positive Prussian blue staining in tumor tissues. Ang2-siRNA plasmid vector/chitosan magnetic nanoparticles directly moved towards tumor tissues under the action of external magnetic field, thus it demonstrated good targeting characteristic.

Comparison of Negative Pressure Wound Therapy and Conventional Therapy for Cranial Bone-exposed Wounds in Rabbits.

BACKGROUND: Bone-exposed wounds with intact or defected periosteum are difficult to heal. To provide relevant experimental evidence for guidance of clinical therapy, we established a rabbit model to compare the efficacies of negative pressure wound therapy (NPWT) and conventional guaze dressing therapy on the healing of cranial bone-exposed wounds. METHODS: Full-thickness excisional circular wounds of 2.0 cm in diameter with exposed bones covered with or without periosteum were created at the parietal regions in 88 rabbits that were further randomly divided into the following treatment groups: periosteum-intact wounds treated with conventional vaseline gauze dressings (P + Control group), periosteum-intact wounds treated with NPWT (P + NPWT group), periosteum-lacking wounds treated with conventional vaseline gauze dressings (P-Control group), and periosteum-lacking wounds treated with NPWT (P-NPWT group). The wounds of NPWT groups were treated using a negative pressure therapy assembly that was set at a continuous pressure of -125 mm Hg for 7 days, then covered with vaseline gauze. The wound healing rates, wound infection rates, hydroxyproline content, and wound tissue histology were determined and evaluated. RESULTS: The NPWT shortened the wound healing time by approximately 5 days when compared with the conventional gauze therapy. The histological characterization of wound tissues showed that NPWT decreased the inflammatory cells infiltration, accelerated reepithelialization and facilitated the organization of collagen fibers into neat layers on postoperative day (POD) 10. The NPWT enhanced bacterial clearances, reduced infection rates and increased the hydroxyproline contents in both types of wounds on PODs 10 and 15. The immunohistochemical staining of CD31 showed the NPWT treatment resulted in a significantly increased and persistent angiogenesis, and the wounds treated with NPWT showed well developed and more functional vessels at POD 7 compared with control. CONCLUSIONS: The NPWT is a more effective therapy for bone-exposed wounds than conventional guaze dressing therapy. The NPWT can promote bone-exposed wounds healing by increasing collagen contents and vessels densities while reducing inflammatory cells infiltration, reducing wound infection rates, and inducing an ordered collagen arrangement.

A Randomized, Controlled, Multicenter Study of Juvéderm Voluma for Enhancement of Malar Volume in Chinese Subjects.

BACKGROUND: Hyaluronic acid gels are used to restore volume to the midface, but there are few data published on this use in Asian subjects. METHODS: This study evaluated the safety and effectiveness in Chinese subjects of Juvéderm Voluma, a 20-mg/ml hyaluronic acid gel formulated for midface volumizing. This prospective, multicenter study randomized 119 subjects aged 18 years or older to a treatment group and 27 subjects to a no-treatment control group. The primary effectiveness endpoint was the objectively measured magnitude of change from baseline in volume of the midface area (right and left combined) calculated by digital analysis at month 6 using three-dimensional images for all subjects in both groups. Effectiveness was protocol-defined as a mean change for the treatment group that was significantly greater than that for the control group at month 6 using a one-side two-group t test performed at the 5 percent level. RESULTS: With a median volume of 2 ml of Voluma injected, the primary effectiveness endpoint was met, with the mean change from baseline to 6 months in malar volume for the treatment group (1.83 ml) being significantly greater than that for the control group (0.11 ml; p < 0.001). The secondary effectiveness endpoints of responder rate (malar volumization rated improved or much improved) using the Global Aesthetic Improvement Scale as assessed at month 6 by the investigator and by the subject were 98.2 and 93.8 percent, respectively. The most common treatment-related adverse events were mild injection-site swelling and bruising. CONCLUSION: Juvéderm Voluma is effective and well tolerated for midface augmentation in Chinese subjects. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, II.

Reversible Alopecia with Localized Scalp Necrosis After Accidental Embolization of the Parietal Artery with Hyaluronic Acid.

Hyaluronic acid (HA) filler injection is widely used for soft-tissue augmentation. Complications associated with HA filling are not uncommon; however, HA-induced alopecia is a rarely reported complication that could result in severe secondary psychological trauma. The etiology, clinical traits, treatment strategies, outcomes, and possible reversibility of HA-induced alopecia have not been characterized. Here, we report a case in which bilateral temple injections of 6.5 mL of HA led to persistent pain over the left scalp for several days. Although the pain was relieved at day 9 after 600 U of hyaluronidase were injected in the left temple, the patient developed localized alopecia at the left temporoparietal region with central skin necrosis at day 15. After topical applications of recombinant bovine basic fibroblast growth factor gel and 2% minoxidil spay, the necrotic skin wound was healed at day 42. Hair regrowth and normal hair density were restored at day 74. Analyses of Doppler ultrasound examinations and histopathology of the skin biopsy suggested that mild ischemia of the left temporoparietal region led to reversible alopecia, while the permanent hair loss in the left parietal area was associated with severe skin ischemia. Therefore, the key to treatment would be to focus on the effective correction of severe ischemia-induced skin necrosis to prevent permanent hair loss. Level of Evidence V 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 online Instructions to Authors www.springer.com/00266 .

Effects of miR-145-5p through NRAS on the cell proliferation, apoptosis, migration, and invasion in melanoma by inhibiting MAPK and PI3K/AKT pathways.

We aimed to detect the effects of miR-145-5p on the cell proliferation, apoptosis, migration, and invasion in NRAS-mutant, BRAF-mutant, and wild-type melanoma cells, in order to figure out the potential mechanisms and provide a novel therapeutic target of melanoma. RT-qPCR and western blot were used to detect the expression of miR-145-5p and NRAS in melanoma tumor tissues and cells, respectively. Luciferase assay was performed to determine whether miR-145-5p directly targeted NRAS. After transfecting miR-145-5p mimics, miR-145-5p inhibitors, NRAS cDNA and NRAS siRNA into CHL-1, VMM917 and SK-mel-28 cells, functional assays were used to detect the proliferation, apoptosis, invasion and migration, including MTT, flow cytometry, Transwell and wound healing assays. In addition, xenograft models in nude mice were also conducted to verify the role of miR-145-5p in vivo. MiR-145-5p was able to suppress proliferation, invasion, and migration of VMM917 and CHL-1 cells and induce apoptosis by inhibiting MAPK and PI3K/AKT pathways. However, aberrant expression of miR-145-5p and NRAS has little impact on the viability and metastasis of BRAF-mutant melanoma. The higher expression of miR-145-5p in xenograft models repressed the VMM917-induced and CHL-1-induced tumor growth observably and has little effect on SK-mel-28-induced tumor growth which was consistent with the results in vitro. Through targeting NRAS, miR-145-5p could suppress cell proliferation, invasion, and migration and induce apoptosis of CHL-1 and VMM917 melanoma cells by inhibiting MAPK and PI3K/AKT pathways.

A Mouse Model of Vascularized Skin Transplantation.

BACKGROUND: Allogeneic skin tissues are highly antigenic and induce intensive immune rejection after composite tissue allotransplantation. Mouse models have advantages in mechanistic studies of immune rejection. However, due to technical challenge in vascular anastomosis with suture technique, mouse vascularized skin allotransplantation models are not widely used in studies of immune rejection. Therefore, the authors propose vascular anastomosis through cuff technique during allotransplantation of mouse donor free groin skin flaps to either recipient inguinal or cervical site. METHODS: Free groin skin flaps from BALB/c or C57BL/6 donor mice were transplanted to the groin (inguinal vascularized skin transplantation [IVST]) or to the neck of recipient sites (cervical vascularized skin transplantation [CVST]) of C57BL/6 mice. A nonsuture cuff technique was utilized to anastomose the donor vessels with either femoral vessels in IVST recipients or common carotid arteries and external jugular veins in CVST mice. Immunosuppressant drugs were used in the allogeneic skin group. RESULTS: The overall success rate was higher in the CVST (88.5%) when compared with the IVST (78.9%). Total operation time in CVST mice lasted 96 minutes (95% confidence interval, 92-101 minutes) that was shorter than that for IVST mice (136 minutes, 95% confidence interval, 127-176 minutes; P < 0.01). Complications, such as hindlimb necrosis and self-mutilation, were observed in IVST mice. Rapamycin (3 mg/kg, daily) significantly prolonged vascularized skin allografts survival with median survival time of 80 days. All syngeneic grafts survived for more than 80 days. CONCLUSIONS: We developed a novel mouse vascularized skin transplantation model that is feasible for the study of clinically relevant skin rejection and tolerance.

[Thinning of the free muscle flaps for the treatment of hand and foot defects].

Objective: To investigate the feasibility of one-stage thinning of latissimus dorsi muscle, rectus abdominis muscle and gracilis flap in reconstruction of the hand and foot defects. Methods: From June 2009 to April 2015,24 free muscle flaps were thinned during transfer operation by removing their superficial muscle layers on the basis of their vessel pedicles running in the deep surface of flaps. The surface of the muscle flaps were covered by split-thickness skin grafts. There were 16 latissimus dorsi muscle flaps,4 gracilis flaps and 4 rectus abdominis muscle flaps. Flap size ranged from 6 cm × 4 cm to 20 cm × 12 cm. Thinned muscle flaps were used to resurface 7 hand defects and 17 foot defects. Results: All muscle flaps survived the thinning procedures without any circulation problems except for one case which suffered total flap necrosis due to venous thrombosis. Skins grafts on muscle flaps also took well. One patient experienced partial skin loss. Two patients underwent secondary debridement and thinning procedure for infection and bulkiness. During the 0.3-20 months follow-up, the contour of thinned muscle flaps matched well with the recipient areas. Reconstructed feet are able to wear regular shoes. Conclusions: Intraoperative immediate thinning of free muscle flaps can be safely accomplished during the primary reconstruction procedure. This procedure prevents the limitations of muscle flap bulkiness and may provide an alternative for surface coverage.

Pharmacological interventions for melanoma: Comparative analysis using bayesian meta-analysis.

We conducted a network meta-analysis in order to compare different strategies for managing melanoma patients. Electronic databases were searched for eligible randomized trials that compared different strategies in efficacy and tolerability. Five interventions were associated with a significant improvement in PFS over chemotherapy (all HR < 1): Ipilimumab, Tremelimumab, Nivolumab, Pembrolizumab 2 mg/kg and Ipilimumab + Nivolumab. Three interventions exhibited significantly improved OS results over chemotherapy (all HR < 1): Ipilimumab, Nivolumab and Ipilimumab + Chemotherapy. Four interventions were superior to chemotherapy in CR and PR (all OR > 1): Nivolumab, Pembrolizumab 10 mg/kg, Pembrolizumab 2 mg/kg and Ipilimumab + Nivolumab. However, the other seven interventions were associated with an increased risk of pruritus compared to chemotherapy (all OR > 1). Ipilimumab, Tremelimumab, Ipilimumab + Nivolumab and Ipilimumab + Chemotherapy might result in a higher risk of diarrhea compared to chemotherapy (all OR > 1). Immune checkpoint therapy or combined interventions might be more effective than chemotherapy for managing melanoma patients. However, chemotherapy appears to be more tolerable than these combined strategies with respect to adverse events.