The Application of Bilayer Artificial Dermis Combined with VSD Technology in Chronic Wounds

Background: Bilayer artificial dermis promotes wound healing and offers a treatment option for chronic wounds. Aim: Examine the clinical efficacy of bilayer artificial dermis combined with Vacuum Sealing Drainage (VSD) technology in the treatment of chronic wounds. Method: From June 2021 to December 2023, our hospital treated 24 patients with chronic skin tissue wounds on their limbs using a novel tissue engineering product, the bilayer artificial dermis, in combination with VSD technology to repair the wounds. The bilayer artificial dermis protects subcutaneous tissue, blood vessels, nerves, muscles, and tendons, and also promotes the growth of granulation tissue and blood vessels to aid in wound healing when used in conjunction with VSD technology for wound dressing changes in chronic wounds. Results: In this study, 24 cases of chronic wounds with exposed bone or tendon larger than 1.0 cm2 were treated with a bilayer artificial skin combined with VSD dressing after wound debridement. The wounds were not suitable for immediate skin grafting. At 2 - 3 weeks post-treatment, good granulation tissue growth was observed. Subsequent procedures included thick skin grafting or wound dressing changes until complete wound healing. Patients were followed up on average for 3 months (range: 1 - 12 months) post-surgery. Comparative analysis of the appearance, function, skin color, elasticity, and sensation of the healed chronic wounds revealed superior outcomes compared to traditional skin fl repairs, resulting in significantly higher satisfaction levels among patients and their families. Conclusion: The application of bilayer artificial dermis combined with VSD technology for the repair of chronic wounds proves to be a viable method, yielding satisfactory therapeutic effects compared to traditional skin flap procedures.

Artificial dermis combined with skin grafting for the treatment of hand skin and soft tissue defects and exposure of bone and tendon

BACKGROUND The recovery time of hand wounds is long,which can easily result in chronic and refractory wounds,making the wounds unable to be properly repaired.The treatment cycle is long,the cost is high,and it is prone to recurrence and disability.Double layer artificial dermis combined with autologous skin transplantation has been used to repair hypertrophic scars,deep burn wounds,exposed bone and tendon wounds,and post tumor wounds.AIM To investigate the therapeutic efficacy of autologous skin graft transplantation in conjunction with double-layer artificial dermis in treating finger skin wounds that are chronically refractory and soft tissue defects that expose bone and tendon.METHODS Sixty-eight chronic refractory patients with finger skin and soft tissue defects accompanied by bone and tendon exposure who were admitted from July 2021 to June 2022 were included in this study.The observation group was treated with double layer artificial dermis combined with autologous skin graft transplantation(n=49),while the control group was treated with pedicle skin flap transplantation(n=17).The treatment status of the two groups of patients was compared,including the time between surgeries and hospital stay.The survival rate of skin grafts/flaps and postoperative wound infections were evaluated using the Vancouver Scar Scale(VSS)for scar scoring at 6 mo after surgery,as well as the sensory injury grading method and two-point resolution test to assess the recovery of skin sensation at 6 mo.The satisfaction of the two groups of patients was also compared.RESULTS Wound healing time in the observation group was significantly longer than that in the control group(P<0.05,27.92±3.25 d vs 19.68±6.91 d);there was no significant difference in the survival rate of skin grafts/flaps between the two patient groups(P>0.05,95.1±5.0 vs 96.3±5.6).The interval between two surgeries(20.0±4.3 d)and hospital stay(21.0±10.1 d)in the observation group were both significantly shorter than those in the control group(27.5±9.3 d)and(28.4±17.7 d),respectively(P<0.05).In comparison to postoperative infection(23.5%)and subcutaneous hematoma(11.8%)in the control group,these were considerably lower at(10.2%)and(6.1%)in the observation group.When comparing the two patient groups at six months post-surgery,the excellent and good rate of sensory recovery(91.8%)was significantly higher in the observation group than in the control group(76.5%)(P<0.05).There was also no statistically significant difference in two point resolution(P>0.05).The VSS score in the observation group(2.91±1.36)was significantly lower than that in the control group(5.96±1.51),and group satisfaction was significantly higher(P<0.05,90.1±6.3 vs 76.3±5.2).CONCLUSION The combination of artificial dermis and autologous skin grafting for the treatment of hand tendon exposure wounds has a satisfactory therapeutic effect.It is a safe,effective,and easy to operate treatment method,which is worthy of clinical promotion.

Treatment of intractable orbital implant exposure with a large conjunctival defect by secondary insertion of the implant after preceding dermis fat graft

AIM: To report a procedure and results of a two-stage operation to manage intractable extensive orbital implant exposure with a large conjunctival defect which was difficult to treat with dermis fat grafts due to repeated graft necrosis. METHODS: A retrospective chart review of four patients who had extensive orbital implant exposures with large conjunctival defects and had past histories of repeated autologous or preserved dermis graft failures was done. As a first-stage operation, the problematic pre-existing orbital implants were removed and autologous dermis fat grafts alone were performed on the defect area. Four months later, new orbital implants were secondarily inserted after confirmation of graft survival. The size of the conjunctival defects and state of the extraocular muscles were checked preoperatively. Success of the operations and complications were investigated.RESULTS: The mean size of the conjuctival defects was 17.3mm×16.0mm, and the mean time from the initial diagnosis of orbital implant exposure to implant removal and autologous dermis fat graft was 20.8 months. After implant removal and autologous dermis fat graft, no graft necrosis was observed in any patients. Also, implant exposure or fornix shortening was not observed in any patients after new orbital implant insertion. CONCLUSION: The secondary insertion of a new orbital implant after pre-existing implant removal and preceding dermis fat graft is thought to be an another selective management of intractable orbital implant exposure in which dermis fat grafts persistently fail.

Temperature Distributions for Regional Hypothermia Based on Nonlinear Bioheat Equation of Pennes Type: Dermis and Subcutaneous Tissues

We have used a nonlinear one-dimensional heat transfer model based on temperature-dependent blood perfusion to predict temperature distribution in dermis and subcutaneous tissues subjected to point heating sources. By using Jacobi elliptic functions, we have first found the analytic solution corresponding to the steady-state temperature distribution in the tissue. With the obtained analytic steady-state temperature, the effects of the thermal conductivity, the blood perfusion, the metabolic heat generation, and the coefficient of heat transfer on the temperature distribution in living tissues are numerically analyzed. Our results show that the derived analytic steady-state temperature is useful to easily and accurately study the thermal behavior of the biological system, and can be extended to such applications as parameter measurement, temperature field reconstruction and clinical treatment.

Use of Artificial Dermis and Cultured Epithelial Autograft for Extensive Deep Dermal Burns —A Case Report

In the treatment of extensive burns, cultured epithelial autograph (CEA) became available commercially in Japan from 2009. Based on the 6 years multicenter surveillance data on using CEA for extensive burns, it is reported that using 6:1 split thickness skin graft together with CEA is successful after wound bed preparation for extensive deep dermal burn or patients with deep burn [1].

Effect of artificial dermis combined with jinfu ning on skin healing and bacterial detection rate of finger abdomen

Objective: To explore the effect of artificial dermis combined with rhGM-CSF(Jinfuning) on healing of soft tissue defect of finger ventral skin and the influence of bacterial detection rate. Methods: Totally 110 patients with finger injury admitted to the rehabilitation department of our department from January 2017 to June 2018 were collected and divided into control group and observation group according to the random number table method with 55 cases in each group. The control group received direct artificial derma lrepairing after thorough debridement, while the observation group received recombinant gm-csf gel coating on the wound surface before artificial dermal repairing, Wound healing, wound inflammation, bacterial detection rate, inflammatory factor expression, follow-up and adverse reactions were compared between the two groups. Results: The wound healing rate of the observation group at 7, 14, 21 and 28 days after treatment was significantly higher than that of the control group (t= 11.211, P =0.000).( T = 14.895, P =0.000;T = 25.346, P=0.000;T =8.247, P=0.000). The wound healing time of the observation group was (19.7±2.3) d, and that of the control group was (27.4±3.3) d. The average wound healing time of the observation group was significantly shorter than that of the control group, and the difference was statistically significant (t=14.197, P= 0.000). Observation group wound inflammation at each time point score was significantly lower than the control group, the group rooms, time points, ·point interaction effect between the comparison, the differences were statistically significant (P <0.05), the observation group wound bacteria detection rate of 7.27% (4 cases) : the control bacteria detection rate was 21.81% (12 cases), difference was statistically significant (chi-square = 4.68, P= 0.0305), the observation group of bacteria detection rate was significantly lower than the control group;The bacteria detected in the two groups were mainly e. coli, tetanus bacillus and fungi. There was no significant difference in the indicators between the two groups before treatment, and the values of inflammatory cytokines il-1 and TNF- IOD in the two groups were significantly decreased after treatment, and the observation group was significantly lower than the control group, with statistically significant differences (P < 0.05). No serious adverse reactions occurred in either group during the treatment. Conclusion: the application of artificial dermals combined with jinfuning can promote wound healing of skin and soft tissue defect of finger abdomen, effectively inhibit bacterial infection of wound surface, reduce inflammation and infection,reducing bacterial detection rate.

Novel pre-vascularized tissue-engineered dermis based on stem cell sheet technique used for dermis-defect healing

Insufficient donor dermis and the shortage of three-dimensional vascular networks are the main limitations in the tissue-engineered dermis(TED).To solve these problems,we initially constructed pre-vascularized bone marrow mesenchymal stem cell sheet(PBMCS)and pre-vascularized fibroblasts cell sheet(PFCS)by cell sheet technology,and then superimposed or folded them together to construct a pre-vascularized TED(PTED),aiming to mimic the real dermis structure.The constructed PTED was implanted in nude mice dorsal dermis-defect wound and the wound-healing effect was quantified at Days 1,7 and 14 via the methods of histochemistry and immunohistochemistry.The results showed that PTED could rapidly promote the wound closure,especially at Day 14,and the wound-healing rate of three-layer PTED could reach 97.2%(P<0.01),which was faster than the blank control group(89.1%),PBMCS(92.4%),PFCS(93.8%)and six-layer PTED(92.3%).In addition,the vessel density in the PTED group was higher than the other groups on the 14th day.Taken together,it is proved that the PTED,especially three-layer PTED,is more conducive to the fullthickness dermis-defect repair and the construction of the three-dimensional vascular networks,indicating its potential application in dermis-defect repair.

Accelerated healing of diabetic wound using artificial dermis constructed with adipose stem cells and poly（L-glutamic acid）/ chitosan scaffold

Background Diabetic wound is one of the most serious complications of diabetes mellitus. There are no significantly effective therapies for chronic non-healing diabetes ulcer so far. This study aimed to explore the feasibility of healing impaired wound using artificial dermis constructed with human adipose derived stem cells （ASCs） and poly（L-glutamic acid）/chitosan （PLGA/CS） scaffold in streptozotocin-induced diabetic mice. Methods ASCs were isolated from fresh human lipoaspirates and expanded ex vivo for three passages, and then cells were seeded onto PLGNCS scaffold to form artificial dermis. Expression of VEGF and TGFI31 by ASCs presented in artificial dermis was determined. The artificial dermis was transplanted to treat the 20 mm ~ 20 mm full-thickness cutaneous wound created on the back of diabetic mice. Wound treated with scaffold alone and without treatment, and wound in normal non-diabetic mice served as control. Results Cells growing within scaffold showed great proliferation potential, depositing abundant collagen matrix. Meanwhile, expression of VEGF and TGF-131 by seeded ASCs maintained at a consistent high level. After treated with ASC based artificial dermis, diabetic wounds exhibited significantly higher healing rate compared with wounds treated with scaffold alone or without treatment. Histological examination also demonstrated an improvement in cutaneous restoration with matrix deposition and organization. Further quantitative analysis showed that there was a significant increase in dermis thickness and collagen content on artificial dermis treated wounds. Conclusion ASC/PLGA artificial dermis can effectively accelerate diabetic wound healing by promoting angiogenic growth factors and dermal collagen synthesis.

The Susceptibility of Primary Dermis Fibroblasts from the Chinese Tree Shrew to Human Cytomegalovirus Infection

As a universal pathogen leading to neonatal defects and transplant failure,human cytomegalovirus(HCMV)has strict species specificity and this has prevented the development of a suitable animal model for the pathogenesis study.The mechanism of cross-species barrier remains elusive and there are so far no non-human cell culture models that support HCMV replication.The Chinese tree shrew(Tupaia belangeri chinensis)is a small laboratory animal and evolutionary closely related with primates.We investigated the susceptibility of primary tree shrew dermis fibroblasts(TSDF)to HCMV infection.Infection with a GFP-expressing HCMV virus resulted in green fluorescence in infected cells with the expression of IE1,UL44 and pp28.The titers of cell-free viruses reached 10^3 PFU/mL at 96 hpi,compared to titers of 10^4 PFU/mL observed in primary human foreskin fibroblasts.Our results suggested that TSDF was semi-permissive for HCMV infection.The TSDF model could be further used to investigate key factors influencing cross-species multiplication of HCMV.

Saturation Effects on Immunopathogenic Mechanism of Psoriasis:A theoretical approach

Psoriasis vulgaris is a common chronic autoimmune inflammatory skin disease characterized by T-cell mediated hyper proliferation of keratinocytes.This type of skin disorder is characterized by macroscopic and also microscopic skin alterations.It was observed that,psoriatic lesions are sharply demarcated,red and slightly raised lesions along with silver whitish scales.The striking feature of T-cell mediated chronic psoriatic lesions are the epidermal infiltration of predominantly oligoclonal CD8^+T-cells and probably also of CD4^+T-cells in the dermis.In this research article we proposed a mathematical model for the chronic disease,psoriasis,consisting a set of differential equations, involving T-Lymphocytes cells,Dendritic cells and the epidermal Keratinocytes,incorporating half saturation constant through the T lymphocyte cells and also in that sense,it has been infiltrated in the activation process of keratinocytes in the course of the plaque of psoriasis.Cell biological research on psoriasis has been established the suppression of epidermal T-cell density.Here we are interested to know how the cell biological system changes for introducing the half saturation constant in presence of suppression taking place DCs on the model dynamics.Our analytical and numerical result reveal that due to control effect,the macrophages population density does not impede for the interaction between T-cell and keratinocytes.T-cell and keratinocyte population ultimately become stable after a certain period of time span.

How and when of eyelid reconstruction using autologous transplantation

Reconstructive surgery of the eyelid after tumor excision,trauma or other causes can be challenging,especially due to the complexities of the anatomic structures and to the necessity of both functional and aesthetic successful outcomes.The aim of this minireview was to investigate the use of tissue transplantation in eyelid reconstruction.Surgical procedures are various,based on the use of both flaps,pedicled or free,and grafts,in order to guarantee adequate tissue reconstruction and blood supply,which are necessary for correct healing.Common techniques normally include the use of local tissues,combining non-vascularized grafts with a vascularized flap for the two lamellae repair,to attempt a reconstruction similar to the original anatomy.When defects are too wide,vast,deep,and complex or when no adjacent healthy tissues are available,distant area tissues need to be recruited as free flaps or grafts and paired with mucosal layer reconstruction.With regards to the anterior lamella,full thickness skin grafts are commonly preferred.With regards to the reconstruction of posterior lamella,there are different graft options,which include conjunctival or tarsoconjunctival,mucosal or palatal or cartilaginous grafts usually combined with local flaps.Free flap transplantation,normally reserved for rare select cases,include the use of the radial forearm and anterolateral flaps combined with mucosal grafts,which are surgical options currently reported in the literature.

Reconstruction of massive skin avulsion of the scrota and penis by combined application of dermal regeneration template(Pelnac) and split-thickness skin graft with vacuum-assisted closure: A case report

BACKGROUND Although skin avulsions to male external genitalia are rare, they can be both physically and psychologically traumatic. Thus, the necessity for judicious management poses significant challenges to surgeons in order to avoid potential permanent disabilities. We report a case of massive penoscrotal skin avulsion and a composite graft was creatively applied to cover the defect which achieved good results. We believe that this case is of great reference value for fellow surgeons.CASE SUMMARY A 52-year-old male presented with massive traumatic avulsion of the penile and scrotal skin following mishandling of an electric drill. The avulsed skin was missing. The patient was diagnosed with massive skin avulsion of external genitalia. Following initial complete debridement of devitalized or infected tissues, Pelnac dermal substitute was secured to the defect with the assistance of negative-pressure wound closure. In the final step, the silicone layer of Pelnac was removed and a split-thickness skin graft was applied. The defect had healed at the two-month follow-up. The patient now has normal erections and satisfactory sexual function.CONCLUSION Our experience with this wound repair demonstrated that the combination of a dermal regeneration template and a split-thickness skin graft with vacuumassisted closure is a safe, well-tolerated and efficient solution for the reconstruction of massive penoscrotal skin defects.

Evaluation of Mast Cells in Dermal versus Subcutaneous Hemangiomas and Hemangiosarcomas in Dogs

This study investigated the role of mast cells in canine cutaneous vascular tumors, and is the first such study to distinguish between tumors arising in the dermis versus the subcutis. Mast cell numbers in canine cutaneous hemangiomas (HA) and hemangiosarcomas (HSA) were evaluated to identify a relationship between mast cells, tumor type (HA, HSA), histologic location (dermis, subcutis) and tumor recurrence. One hundred and sixty-seven biopsies from 148 dogs were evaluated. Using only one biopsy from each dog, mast cell counts (MCC) for each tumor (n = 148) were obtained by averaging the number of mast cells counted in ten 400× fields. A significant difference in mean MCC was found only between tumor types, with HA having more mast cells than HSA (4.2 ± 4.2 vs. 2.2 ± 2.6;p < 0.001). No significant difference in mean MCC existed between tumors that recurred and those that did not. There was no difference in recurrence rate between tumor type or histologic location. Our results indicate that benign HA contain more mast cells than malignant endothelial cell tumors, regardless of histologic location;whether this is a cause or effect relationship remains to be determined.

Clinical Assessment of a Mesotherapy Formulation for Skin Rejuvenation in Healthy Volunteers

Mesotherapy is a mildly invasive procedure involving several subcutaneous drug injections, which are performed simultaneously. The concept is to inject a very small amount of active compounds (hyaluronic acid, vitamins, antioxidants, enzymes, humectants, …) and the aim is to stimulate the biosynthetic capacity of fibroblasts, increase collagen and elastin production and to give active essential molecules for a fibroblasts environment. Numerous studies suggest that the mesotherapy procedure improves skin hydration and mechanical properties;however, some results have not been statistically significant. In this study, we investigated an antiaging treatment as a complementary approach to a non-surgical rejuvenating procedure after 5 sessions of mesotherapy. Our main objective was to assess the improvement of face, neck and neckline skin quality, after intradermal microinjections of NCTF 135HA&reg. This mixture, manufactured by FILORGA Laboratories, includes a non-cross-linked hyaluronic acid, vitamins, antioxidants, mineral salts and enzymes. A statistically significant decrease in the depth of crow’s-feet wrinkles (43.3%) and in pore size (58.5%) was observed. Also, dermatological scores showed a significant decrease, and data suggest that skin tone was improved with a brighter tone. The injected mixture could then have an action on the skin pigmentation process with a photoprotective effect. Evolution of Dermis density was also investigated and an increase of dermal density and thickness were highlighted, showing an improvement of skin firmness with significant increases of dermis thickness (20%) and dermis density (24%), indicating a neocollagen activity.

Use of a selective enzymatic debridement agent(Nexobrid■)for wound management:Learning curve

AIM To evaluate the efficacy of Nexobrid~? in the initial management of burns and lessons learned with the procedure.METHODS From January 27^(th) 2015 until January 25^(th) 2016, 25 patients aged between 18-94 years old with deep partial and full thickness burns were treated with Nexobrid~? covering 1%-30% of their total body surface area(TBSA). The debridement was applied in the first 96 h post-injury following the protocol suggested for Nexobrid~?. In patients with burns of more than 15% TBSA a second application of Nexobrid~? was performed. After the removal of the product-4 h post application and after a 2 h period of wet dressing-we used several products to cover the wound like Suprathel~?, Biobrane~?, Mepitel~? with wet dressing, silver sulphadiazine 1% cream, and in some cases even autografts. We treated patients with inhalation injury as well. All the procedures were done under deep sedation, regional blocks in extremities or general anaesthesia in the intensive care unit room or in the operating theatre.RESULTS After these first 25 cases, we have observed that patients with partial thickness burns treated with Nexobrid~?, experienced great benefits in the reduction of the need for autografting compared with the standard of care. This isbecause after selective enzymatic debriding of the burn scar we can distinguish different wound beds, which can coexist in the same patient, and we also managed to associate each one to its ability to epithelize. In major burns, besides the improvement in wound healing, we observed an important improvement in their general state. This may be because SIRS significantly improved through a bloodless debridement of necrotic tissue, decreasing the requirements of vasoactive drugs and fluid resuscitation. Circumferential burns also benefited from enzymatic debridement, observing a decrease in the number of compartment syndromes and the need for escharotomies. At present, we have not observed a positive effect in the evolution and outcome of major burns with inhalation injury.CONCLUSION The introduction of Nexobrid~? shows significant improvement in burn treatment. Cumulative experiences are necessary to adapt its application in all Burns Centres.

Analysis of Mechanical Behavior of Composite Tissues Using Vibrational Optical Coherence Tomography

Extracellular matrices (ECMs) found in vertebrate tissues are fiber reinforced composite materials that prevent premature mechanical failure, store, transmit, and dissipate mechanical energy generated by the musculoskeletal system. We have developed a new method using optical cohesion tomography and vibrational analysis to non-destructively and non-invasively measure the mechanical properties of composite tissues and polymeric materials. In addition, this method can be used to measure the moduli of individual components of composite materials and perform “mechanical spectroscopy” on materials. In addition, we propose that measurement of the resonant frequency of a material minimizes the viscoelastic behavior of a composite material. This approach simplifies the analysis of mechanical behavior of polymers and others materials that demonstrate time-dependence to their properties.

Biochemical, Biophysical and Mechanical Characterization of Decellularized Dermal Implants

Allografts have been used in a variety of applications to augment as well as replace tissues throughout the body. A number of steps are involved in selection, harvesting, processing and testing of dermal allografts. Grafts can be obtained that are: free of antibodies to viruses and low in viral titers. Cellular material can be eliminated from the tissue and the product becomes almost exclusively a collagen fiber network. The purpose of this paper is to examine the changes in collagen structure and properties that occur during processing of decellularized dermis. The results indicate that collagen fiber swelling occurs during processing although the product preserves the native collagen banding pattern at the fibrillar structural level. Fiber swelling and decreased collagen deformability of processed dermis, may lead to stress concentration at the implant-tissue interface and up-regulation of mechanotransduction. This may lead to premature mechanical failure due to creation of a chronic inflammatory condition at the implant-tissue interface. It is suggested that all dermal allografts be oriented such that Langer’s lines of the implant match those of the host tissue, and that wound closure by suturing be done under conditions that preserve the normal tension in skin in order to minimize implant-interfacial failure.

Full-thickness tissue engineered skin constructed with autogenic bone marrow mesenchymal stem cells

To explore the feasibility of repairing clinical cutaneous deficiency, autogenic bone marrow mesen-chymal stem cells (BMSCs) were isolated and differentiated into epidermal cells and fibroblasts in vitro supplemented with different inducing factors and biomaterials to construct functional tissue- engineered skin. The results showed that after 72 h induction, BMSCs displayed morphologic changes such as typical epidermal cell arrangement, from spindle shape to round or oval; tonofibrils, melano-somes and keratohyaline granules were observed under a transmission electronic microscope. The differentiated cells expressed epidermal stem cell surface marker CK19 (59.66% ± 4.2%) and epidermal cells differentiation marker CK10. In addition, the induced epidermal cells acquired the anti-radiation capacity featured by lowered apoptosis following exposure to UVB. On the other hand, the collagen microfibrils deposition was noticed under a transmission electronic microscope after differentiating into dermis fibroblasts; RT-PCR identified collagen type I mRNA expression in differentiated cells; radioimmunoassay detected the secretion of interleukin-6 (IL-6) and interleukin-8 (IL-8) (up to 115.06 pg/mL and 0.84 ng/mL, respectively). Further in vivo implanting BMSCs with scaffold material short-ened skin wound repair significantly. In one word, autogenic BMSCs have the potential to differentiate into epidermal cells and fibroblasts in vitro, and show clinical feasibility acting as epidermis-like and dermis-like seed cells in skin engineering.

Progress in various crosslinking modification for acellular matrix

Objective To review the current crosslinking strategies for acelluar matrix scaffold,laying the foundation for subsequent experiment.Data sources Data were mainly obtained from recent papers published in PubMed or indexed by Web of Science,with keyword like crosslinking.Results Various crosslinking strategies,including chemical,physical and biological methods,have been introduced to facilitate the performance of fresh acellular matrix.Chemical crosslinking reagents,involved in synthetic and naturally derived agents,need to be eliminated before implantation in case of their potential biotoxicity,although several crosslinking agents with less toxicity and specific characteristics have been developed.Physical crosslinking methods present to be safe,additive-free and relatively controllable for rapid surface functionalization with no consideration of remaining radioactivity.Biological crosslinking strategies have attracted great interest,and have been demonstrated to enhance collagen-based crosslinking since their preparations do not need toxic or potentially biologically contaminated substances and can be carried out under physiological conditions.Conclusions Kinds of crosslinking methods with its potential advantages have been developed to modify raw acelluar matrix,of which the performance are promising after being crosslinked by several crosslinking treatments.Further preclinical and clinical evaluations should be taken to vertify their safety and efficacy for the tissues and organs substitutes in tissue and regenerative medicine.

Exosomes of adult human fibroblasts cultured on 3D silk fibroin nonwovens intensely stimulate neoangiogenesis

Background:Bombyx mori silk fibroin is a biomacromolecule that allows the assembly of scaffolds for tissue engineering and regeneration purposes due to its cellular adhesiveness,high biocompatibility and low immunogenicity.Earlier work showed that two types of 3D silk fibroin nonwovens(3D-SFnws)implanted into mouse subcutaneous tissue were promptly vascularized via undefined molecular mechanisms.The present study used nontumorigenic adult human dermal fibroblasts(HDFs)adhering to a third type of 3D-SFnws to assess whether HDFs release exosomes whose contents promote neoangiogenesis.Methods:Electron microscopy imaging and physical tests defined the features of the novel carded/hydroentangled 3D-SFnws.HDFs were cultured on 3D-SFnws and polystyrene plates in an exosome-depleted medium.DNA amounts and D-glucose consumption revealed the growth and metabolic activities of HDFs on 3D-SFnws.CD9-expressing total exosome fractions were from conditioned media of 3D-SFnws and 2D polystyrene plates HDF cultures.Angiogenic growth factors(AGFs)in equal amounts of the two groups of exosomal proteins were analysed via doubleantibody arrays.A tube formation assay using human dermal microvascular endothelial cells(HDMVECs)was used to evaluate the exosomes’angiogenic power.Results:The novel features of the 3D-SFnws met the biomechanical requirements typical of human soft tissues.By experimental day 15,3D-SFnws-adhering HDFs had increased 4.5-fold in numbers and metabolized 5.4-fold more D-glucose than at day 3 in vitro.Compared to polystyrene-stuck HDFs,exosomes from 3D-SFnws-adhering HDFs carried significantly higher amounts of AGFs,such as interleukin(IL)-1α,IL-4 and IL-8;angiopoietin-1 and angiopoietin-2;angiopoietin-1 receptor(or Tie-2);growth-regulated oncogene(GRO)-α,GRO-βand GRO-γ;matrix metalloproteinase-1;tissue inhibitor metalloproteinase-1;and urokinase-type plasminogen activator surface receptor,but lesser amounts of anti-angiogenic tissue inhibitor metalloproteinase-2 and pro-inflammatory monocyte chemoattractant protein-1.At concentrations from 0.62 to 10μg/ml,the exosomes from 3D-SFnws-cultured HDFs proved their angiogenic power by inducing HDMVECs to form significant amounts of tubes in vitro.Conclusions:The structural and mechanical properties of carded/hydroentangled 3D-SFnws proved their suitability for tissue engineering and regeneration applications.Consistent with our hypothesis,3D-SFnws-adhering HDFs released exosomes carrying several AGFs that induced HDMVECs to promptly assemble vascular tubes in vitro.Hence,we posit that once implanted in vivo,the 3DSFnws/HDFs interactions could promote the vascularization and repair of extended skin wounds due to burns or other noxious agents in human and veterinary clinical settings.