Hypertrophic Scar In Rabbit Ears Research Articles

Development of a bioactive silk fibroin bilayer scaffold for wound healing and scar inhibition

In cases of deep skin defects, spontaneous tissue regeneration and excessive collagen deposition lead to hyperplastic scars. Conventional remedial action after scar formation is limited with a high recurrence rate. In this study, we designed a new artificial skin bilayer using silk fibroin nanofibers films (SNF) as the epidermis, and silk fibroin (SF) / hyaluronic acid (HA) scaffold as the dermal layer. The regenerated SF film was used as a binder to form a functional SNF-SF-HA bilayer scaffold. The bilayer scaffold showed high porosity, hydrophilicity, and strength, and retained its shape over 30 days in PBS. In vitro, human umbilical vein endothelial cells were seeded into the bilayer scaffold and showed superior cell viability. In vivo analyses using the rabbit ear hypertrophic scar (HS) model indicated that the bilayer scaffold not only supported the reconstruction of new tissue, but also inhibited scar formation. The scaffold possibly achieved scar inhabitation by reducing wound contraction, weakening inflammatory reactions, and regulating collagen deposition and type conversion, which was partly observed through the downregulation of type I collagen, transforming growth factor-β, and α-smooth muscle actin. This study describes a new strategy to expand the application of silk-based biomaterials for the treatment of hyperplastic skin scars.

Gallic Acid Treats Hypertrophic Scar in Rabbit Ears via the TGF-β/Smad and TRPC3 Signaling Pathways.

Hypertrophic scars (HSs) develop due to excessive collagen deposition and abnormal fibroblast proliferation during wound healing, significantly impacting patient quality of life. Three dosages of GA ointments were administered to rabbit ear HS models to investigate the potential efficacy and mechanism of gallic acid (GA) on HS. Daily application of ointment was performed on the matrix group, the GA ointment groups, and the silicone gel group for 28 days. (No drug treatment was performed on the skin and model groups as a blank group and vehicle group, and silicone gel ointment was topically administered to the silicone gel group as a positive control group.) Scar specimens were collected for histopathology analysis, RNA sequencing analysis, real-time quantitative polymerase chain reaction, and Western blot analysis at the first, second, and fourth weeks after the treatment. Low-dose and medium-dose GA effectively suppressed HS formation and markedly decreased fibroblast infiltration levels and scar thickness. Moreover, decreased expression of TRPC3 mRNA and TGF-β1, p-Smad2/3, and Smad2/3 protein was observed in the low- and medium-dose GA groups and the silicone gel group. This study provides evidence for the efficacy of GA in treating HS and sheds light on its potential underlying pharmacological mechanisms.

The effects of Shikonin on the hypertrophic scar of rabbit ears via the TLR4/NF-κB signaling pathway.

As a traditional Chinese medicine, Zihuang Shengji Ointment has obvious effects on promoting postoperative wound healing and reducing scar formation in clinical application. Shikonin is the major phytochemical in Zihuang Shengji Ointment. As a kind of naphquinone compound with anti-tumor, anti-viral, anti-inflammatory, anti-bacterial and other biological activities extracted from Lithospermum erythrorhizon, shikonin exerts an important role in many diseases. Shikonin has impacts on the development of hypertrophic scars (HS), however, these effects are yet mostly unknown. As a result, we created the Newland white rabbit ear HS model, administered shikonin to it, and then assessed scar hypertrophy using HE and VG staining. The degree of scarring is assessed by HI, NA, as well as AA. The expression levels of collagen I, collagen III, as well as α-SMA as well as fibroblast proliferation, are also measured using real-time PCR, immunohistochemistry, and western blot. TUNEL tests are used to assess fibroblast apoptosis. In our work, HE staining and VG staining showed that the shikonin-treated group had normal bundles of collagen fibers and regular fibroblasts. Shikonin suppresses the production of HS, according to histopathological features, HI, NA, and AA measures. Shikonin also causes fibroblast apoptosis and lowers the production of α-SMA, collagen I, as well as collagen III in the HS rat. Notably, we discover that NF-κB activation and TLR4 activity are inhibited by shikonin. Overall, the results show that the signaling pathway of TLR4/NF-κB is modulated by shikonin's inhibitory effect on scar formation, which represses the levels of collagen I, collagen III, α-SMA, as well as fibroblasts.

Experimental Study on the Effect of Caffeine Hydrogel on the Expression of TGF -β1, α-SMA and Collagen in Hypertrophic Scar of Rabbit Ears.

This study evaluated the effects of topical use of caffeine hydrogel on hypertrophic scar in a rabbit ear wound model. Nine rabbits were randomly divided into three groups: control group, caffeine hydrogel group, and matrix group. Punched defects were established on each rabbit's ear which resulted in a hypertrophic scar. When the wound epithelialization and scar hyperplasia could be seen, control group did not do any treatment, while caffeine hydrogel group and matrix group were treated with caffeine hydrogel and hydrogel matrix, respectively. After 3 weeks of administration, the general morphological changes of scar were observed, and the scar tissue of rabbit ears was stained with HE and Masson. The relative expressions of TGF β-1, α-SMA, type I collagen, and type III collagen in scar tissue were detected by Western blot. In all three groups, findings showed that caffeine hydrogel can inhibit scar growth by reducing the expression of TGF β-1, reducing the proliferation of fibroblasts, improving collagen arrangement and reducing collagen deposition. The overall study shows efficacy and mechanism of caffeine. It concluded that caffeine could be an effective therapeutic agent for hypertrophicscars.

Effectiveness of fractional carbon dioxide laser combined with botulinum toxin type A in a rabbit ear model with the underlying mechanism.

Hypertrophic scar (HS) is a common disease in plastic and cosmetic surgery, with limited treatment options, and is a challenge for clinicians. This study aimed to evaluate the efficacy of fractional carbon dioxide (CO2 ) laser combined with botulinum toxin type A (BTXA) in treating HSs in rabbit ears and to provide new strategies for treating HS. Twenty-four New Zealand white rabbits with induced HSs were randomly divided into one control and three treatment groups. After 4 weeks of modeling, BTXA (2.0U) was injected into the HS of the BTXA and combination groups, whereas a fractional CO2 laser (combo mode, deep energy: 12.5mJ; super energy: 90 mJ) was used in the fractional CO2 laser and combination groups. The laser treatments were repeated after 2 weeks. The HSs in the rabbit ears were observed and photographed 5 weeks after the first treatment. The scar thickness in each group was measured and compared, and the scar elevation index (SEI) was determined using hematoxylin and eosin staining. Collagen content and alignment were observed using Masson's trichrome staining. Western blotting and immunohistochemistry were performed to analyze scar-related protein levels. Hypertrophic scars were reduced in all treatment groups compared with the control group. The combination group had lower scar thickness, SEI, and expression of scar-related proteins in HSs, with an appearance similar to that of normal rabbit ear skin. Furthermore, the fibroblast content and collagen deposition decreased significantly in the combination group (p < 0.001). Fractional CO2 laser combined with BTXA more effectively reduced HSs by inhibiting fibroblast proliferation, decreasing transforming growth factor-β1 and α- smooth muscle actin expression, and causing collagen remodeling.

Quantitative simulation of photothermal effect in laser therapy of hypertrophic scar.

Laser technology has been widely used in the treatment of hypertrophic scar (HPS). Due to the lack of effective quantitative relationship between laser doses and thermal effect of lesion tissue, the selection of laser doses in clinical laser treatment of HPS is blind, which cannot guarantee the best treatment effect. The photothermal model of HPS was established by using finite element method. The effects of laser dose parameters such as laser energy density, pulse width, and spot diameter on the thermal effects of laser treatment were analyzed. According to tissue temperature threshold and thermal damage degree of the simulation results, the optimal laser doses of HPS were selected for the laser treatment experiments of rabbit ear HPSs to verify the rationality of the quantitative photothermal model. The temperature rise and thermal damage degree of HPS following laser treatment were directly correlated to the laser doses, which grew with the increase of energy density and laser pulse width. For the different spot diameters, the temperature rise decreased with the increase of spot diameter, whereas the thermal damage degree worsened with the increase of spot diameter. Both simulation and experimental results show that the optimal treatment parameters of HPS were as follows: The laser energy density was 7.5J/cm3 , the pulse width was 4ms, and the spot diameter was 7mm. The laser dose parameters optimized by the photothermal model have achieved good therapeutic effects in the rabbit ear HPS, indicating that the model can be used for quantitative evaluation of laser doses before clinical treatment.

Intradermal delivery of an angiotensin II receptor blocker using a personalized microneedle patch for treatment of hypertrophic scars.

High-quality postoperative rehabilitation is the focus of most patients currently, and hypertrophic scar (HS) greatly reduces the patient's quality of life due to the symptom of severe itching. Traditional HS therapies are associated with limitations, such as poor drug delivery efficiency for topical administration and severe pain for intralesional injection. In this study, we developed a personalized microneedle patch system for minimally invasive and effective treatment of HSs. The microneedle patches were personalized designed and fabricated with 3D printing in order to adapt to individual HS. The optimized microneedle patches were composed of dissolving gelatin and starch and loaded with losartan. Losartan, as a drug class of angiotensin II receptor blockers (ARBs), can effectively inhibit the proliferation and migration of hypertrophic scar fibroblasts (HSFs) and downregulate the gene expression related to scar formation in HSFs. The dissolving microneedle patches exhibited strong mechanical strength, effectively penetrated the stratum corneum of HSs and increased the losartan delivery into HSs upon dissolution of gelatin and starch. Together, the losartan-loaded microneedle patches effectively inhibited the formation of HSs in rabbit ears with reduced scar elevation index (SEI), and decreased fibrosis and collagen deposition in HSs. This personalized microneedle patch system increases the drug delivery efficiency into HSs with minimal invasion, and opens a new window for personalized management and treatment of skin diseases.

Effects of mechanical tension on the formation of hypertrophic scars in rabbit ears and transforming growth factor-β1/Smad signaling pathway

Objective: To explore the effects of mechanical tension on the formation of hypertrophic scars in rabbit ears and transforming growth factor-β1 (TGF-β1)/Smad signaling pathway. Methods: The experimental research method was adopted. Six New Zealand white rabbits, male or female, aged 3-5 months were used and 5 full-thickness skin defect wounds were made on the ventral surface of each rabbit ear. The appearance of all rabbit ear wounds was observed on post surgery day (PSD) 0 (immediately), 7, 14, 21, and 28. On PSD 28, the scar formation rate was calculated. Three mature scars in the left ear of each rabbit were included in tension group and the arch was continuously expanded with a spiral expander. Three mature scars in the right ear of each rabbit were included in sham tension group and only the spiral expander was sutured without expansion. There were 18 scars in each group. After mechanical tension treatment (hereinafter referred to as treatment) for 40 days, the color and texture of scar tissue in the two groups were observed. On treatment day 40, the scar elevation index (SEI) was observed and calculated; the histology was observed after hematoxylin eosin staining, and the collagen morphology was observed after Masson staining; mRNA expressions of TGF-β1, Smad3, collagen Ⅰ, collagen Ⅲ, and α-smooth muscle actin (α-SMA) in scar tissue were detected by real-time fluorescence quantitative reverse transcription polymerase chain reaction; and the protein expressions of TGF-β1, collagen Ⅰ, collagen Ⅲ, and α-SMA, and phosphorylation level of Smad3 in scar tissue were detected by Western blotting. The number of samples of each group in the experiments was 3. Data were statistically analyzed with independent sample t test. Results: On PSD 0, 5 fresh wounds were formed on all the rabbit ears; on PSD 7, the wounds were scabbed; on PSD 14, most of the wounds were epithelialized; on PSD 21, all the wounds were epithelialized; on PSD 28, obvious hypertrophic scars were formed. The scar formation rate was 75% (45/60) on PSD 28. On treatment day 40, the scar tissue of rabbit ears in tension group was more prominent than that in sham tension group, the scar tissue was harder and the color was more ruddy; the SEI of the scar tissue of rabbit ears in tension group (2.02±0.08) was significantly higher than 1.70±0.08 in sham tension group (t=5.07, P<0.01). On treatment day 40, compared with those in sham tension group, the stratum corneum of scar tissue became thicker, and a large number of new capillaries, inflammatory cells, and fibroblasts were observed in the dermis, and collagen was more disordered, with nodular or swirling distribution in the scar tissue of rabbit ears in tension group. On treatment day 40, the mRNA expressions of TGF-β1, Smad3, collagen Ⅰ, collagen Ⅲ, and α-SMA in the scar tissue of rabbit ears in tension group were respectively 1.81±0.25, 5.71±0.82, 7.86±0.56, 4.35±0.28, and 5.89±0.47, which were significantly higher than 1.00±0.08, 1.00±0.12, 1.00±0.13, 1.00±0.14, and 1.00±0.14 in sham tension group (with t values of 5.36, 9.82, 20.60, 18.26, and 17.13, respectively, all P<0.01); the protein expressions of TGF-β1, collagen Ⅰ, collagen Ⅲ, and α-SMA, and phosphorylation level of Smad3 in the scar tissue of rabbit ears in tension group were respectively 0.865±0.050, 0.895±0.042, 0.972±0.027, 1.012±0.057, and 0.968±0.087, which were significantly higher than 0.657±0.050, 0.271±0.029, 0.631±0.027, 0.418±0.023, and 0.511±0.035 in sham tension group (with t values of 5.08, 21.27, 15.55, 16.70, and 8.40, respectively, all P<0.01). Conclusions: Mechanical tension can inhibit the regression of hypertrophic scars in rabbit ears through stimulating the hyperplasia of scars, inhibiting the normal arrangement of dermal collagen fibers, and intensifying the deposition of collagen fibers, and the mechanism may be related to the activation of TGF-β1/Smad signaling pathway by mechanical tension.

Aspirin inhibits the growth of hypertrophic scar in rabbit ears via regulating Wnt/β-catenin signal pathway.

Steroidal anti-inflammatory drugs have certain side effects in the treatment of hypertrophic scar, and the scar recurrence is easy after withdrawal of steroid anti-inflammatory drugs. Finding reliable alternative drugs is an effective means to improve this defect. Aspirin, a traditional non-steroidal anti-inflammatory drug, is safe for topical use and has anti-inflammatory effects similar to those of steroidal anti-inflammatory drugs, which may have similar effects on the treatment of hypertrophic scar. This study aims to investigate the inhibitory effect of aspirin on the proliferation of hypertrophic scar in rabbit ears and the underlying mechanism. The rabbit ear hypertrophic scar models were prepared. The rabbits were randomly divided into a normal skin group (group A), a blank control group (group B), a 0.9% NaCl group (group C), a 0.2% aspirin group (group D), a 0.5% aspirin group (group E), a 2% aspirin group (group F), and a triamcinolone acetonide group (group G). Macroscopic observation of hyperplasia was performed 8 weeks after local injection of the scar, followed by collecting the scar tissue samples for HE staining, Masson staining, and immunohistochemistry, respectively to assess the proliferation of fibroblasts and collagen fibers, and calculate the hypertrophic index, microvessel density, and immunohistochemical score. All rabbit ear hypertrophic scar models were successfully constructed. In groups B and C, the hypertrophic scar edge was irregular, with reddish protruding epidermis, significant contracture and hard touch. In group D, E, and F, with the increase of aspirin administration concentration, the scar became thinner and gradually flat, the proliferation of fibrocytes and collagen fibers was weakened, and the hypertrophic index was gradually decreased (P<0.05). Immunohistochemistry showed that the expression of β-catenin was decreased in the group D, E and F in turn, and the immunohistochemical score was gradually decreased (P<0.05). There was no significant difference in hypertrophic index, microvessel density, and immunohistochemical score (all P>0.05). Local injection of aspirin can reduce the generation of hypertrophic scar in a dose-dependent manner within a certain concentration range; aspirin inhibits the growth of hypertrophic scar in rabbit ears by inhibiting Wnt/β-catenin signal pathway; 2% aspirin and 40 mg/mL triamcinolone acetonide have similar curative efficacy on hypertrophic scar.

Lycorine Inhibits Hypertrophic Scar Formation by Inducing ROS-Mediated Apoptosis.

Background: Hypertrophic scar (HS) is a fibrotic cutaneous disease with few effective therapies. Lycorine is a drug with pro-apoptotic ability and anti-fibrosis potential. This study aimed to test whether lycorine could trigger the apoptosis of hypertrophic scar fibroblasts (HSFs) to inhibit HS formation. Methods: The proapoptotic and anti-fibrosis effects of lycorine on the viability and apoptosis of human primary HSFs and their reactive oxygen species (ROS) production as well as a rabbit ear model of HS were determined by CCK-8, flow cytometry, Western blot, immunofluorescence, transwell migration, collagen gel contraction assays. Results: Lycorine treatment selectively decreased the viability of HSFs, and induced their apoptosis, but not normal fibroblasts (NFs). Lycorine treatment increased the relative levels of Bax and cleaved PARP expression, cytochrome C cytoplasm translocation, but decreased Bcl-2, caspase-3 and caspase-9 expression, the mitochondrial membrane potential (MMP) in HSFs. Lycorine inhibited the migration and contraction of HSFs, and reduced the expression of collagen I, collagen III and α-SMA. Mechanistically, lycorine treatment stimulated high levels of ROS production, leading to apoptosis of HSFs while treatment with NAC, a ROS inhibitor, significantly mitigated or abrogated the pro-apoptotic and antifibrotic activity of lycorine in HSFs. Moreover, lycorine treatment mitigated the severity of HS in rabbit ears by inducing fibroblast apoptosis. Conclusion: These results indicate that lycorine has a potent anti-fibrotic activity and is a potential drug for intervention of HS.

Experimental comparative study of the effect of fractional CO2 laser combined with pulsed dye laser versus each laser alone on the treatment of hypertrophic scar of rabbit ears.

The use of fractional CO2 laser and pulsed dye laser (PDL) therapy to treat and/or prevent scarring following burn injury is becoming more widespread with a number of studies reporting reduction in scar erythema and pruritus following treatment with lasers. The objective of this study was to directly compare the efficacy of PDL, fractional CO2 , and PDL and fractional CO2 laser therapy in immature red hypertrophic scars in a standardized animal model. Ten New Zealand big-eared rabbits were used to make rabbit ear hypertrophic scar models. A total of 80hypertrophic scar models were obtained and randomly divided into groups: control (Group A), Fractional CO2 laser (Group B), pulse dye laser (Group C), combined (Group D), 20 in each group. Wound sites were treated with PDL, Fractional CO2 laser, or both at 28days post-grafting. Grafts receiving no laser therapy served as controls. Hypertrophic scar appearance, morphology, size, and erythema were assessed, and punch biopsies were collected. At days 7 and 28, additional tissue was collected for biomechanical analyses and markers for HE staining, Masson staining, immunohistochemical method to determine the CD31 content in the scar; Western blot to detect the expression of VEGF protein in scars. After the day 7 and 28, the scars were collected. Histomorphological change in scars was observed by hematoxylin-eosin staining, Masson staining. The expression of CD31, VEGF protein, and the cell apoptosis rate was detected by immunohistochemical method. (i) In morphological observation, HE staining and Masson staining, both the number of fibroblasts and amount of collagen fibrils in the experimental group were significantly reduced compared with those in control group. (ii) Micro-vessel Density (MVD) value can be found much smaller in the experimental groups than the control (p<0.05). Among the four experimental groups, there was a significant difference among 14d, 21d, and 28d groups (p<0.05). (iii) On days 7 and 28 after treatment, expression of VEGF could be regulated in experimental group (p<0.05). Among the three experimental groups, the decrease of VEGF in the combined group was significantly stronger than that of the other two. The effect of Fractional CO2 laser combined with pulsed dye laser treatment was shown to be better than that of Fractional CO2 laser or pulsed dye laser alone and inhibits the early hypertrophic scar in rabbit ears.

Treatment of hypertrophic scar with injection of triamcinolone leads to increased microRNA-26a in a rabbit ear model

Objectives: Differential expression of microRNAs (miRNAs) has been confirmed to promote hypertrophic scar (HS) formation. However, this mechanism remains to be further elucidated. This paper investigated the effects of microRNA26a on HS of rabbit ears or its mechanism. Methods: The rabbit HS models were established and randomly assigned to either the experimental group (20 rabbits with treatment through triamcinolone acetonide) or the scar group (20 rabbits without treatment). In addition, 10 unmodeled rabbits were served as control group. The expression of microRNA-26a in HS tissues was detected via a stem-loop real-time polymerase chain reaction (RT-PCR). Results: According to RT-PCR, we showed the decreased expression of microRNA-26a in the scar group compared with that in the experimental group, and in the experimental group compared with that in the control group ( p &lt; .01). In addition, the expression of microRNA-26a was negatively correlated with scar thickness (STs), number of fibroblasts (NFs), Collagen I (Col I) level, Collagen III (Col III) level, Interleukin-6 (IL-6) level, and Tumor necrosis factor-alpha (TNF-α) level (all p &lt; .01). Conclusions: Our findings revealed that the increase of microRNA-26a expression might alleviate excessive inflammation during the HS formation, inhibit fibroblast proliferation and collagen deposition in HS, and promote the treatment of HS.

Nanomicelle-generating Microneedles Loaded With Tranilast for Treatment of Hypertrophic Scars in a Rabbit Model.

Hypertrophic scars (HS) are the result of pathological wound healing characterized by a red, raised scar formation. The goal of this research was development of a new method for treatment of HS formation. A tranilast-loaded microneedle (TMN) was developed and applied in a rabbit ear model to treat an induced HS. Scar elevation index, the thickness of dorsal skin by hematoxylin and eosin staining, collagen deposition by Masson trichrome staining and expression of myofibroblast biomarker proteins were evaluated. The 12×12 array of the TMN containing 2.9 μg tranilast per needle released more than 80% of the drug within 30 min. During the procedure, control, non-loaded MN and TMN loaded with three different doses of tranilast (low: 2.5-3, medium: 25-30, and high: 100-150 μg) were applied to the HS in rabbit ears. High-level TMN led to a clear and natural appearance of skin, a decrease in scar elevation index by 47% and decline in the thickness of the epidermis from 69.27 to 15.92 μm when compared to the control group. Moreover, the collagen density also decreased in groups treated with medium- or high-level TMNs, by 10.2% and 9.06%, respectively. Furthermore, the expression of transforming growth factor-β, collagen-1, and α-smooth muscle actin proteins was reduced in TMN-treated HSs compared to the control. The findings show the overall efficacy of TMNs in inhibiting HS. Thus, use of TMN is a simple and cosmetic remedy for HS, with good protection and reliability.

CO2 fractional laser-assisted transdermal delivery of silk nanofiber carriers in a rabbit ear hypertrophic scar model.

Hypertrophic scars are skin fibrotic diseases, characterized by fibroblast hyperproliferation and excessive accumulation of extracellular matrix. However, topical drug application for hypertrophic scars are unsatisfactory. The purpose of this study was to explore the permeability of silk nanofiber hydrogels (SNFs) loaded with rhodamine 6G (R6G) and rhodamine 110 (R110) mediated by CO2 fractional laser irradiation into hypertrophic scar tissues. In this work, R6G and R110 were chosen as hydrophilic and hydrophobic model molecules. They were loaded inside SNFs. In vivo rabbit ear hypertrophic scars were treated with CO2 fractional laser irradiation and then R6G/R110-laden SNFs were applied to the scars to evaluate their synergetic effect on drug penetration efficiency. Their permeability was quantified by fluorescence intensity and measured by confocal laser scanning microscopy on days 1, 3, 5 and 7. More specifically, the thermal coagulation zone (CZ) and its surrounding area (peri-CZ) caused by the thermal coagulation of the laser were discussed separately. Our data indicated that the SNFs promoted the penetration of R6G but not that of R110 in the peri-CZ on day 1 when combined with laser irradiation. Interestingly, both R6G and R110 were abundant in the CZ and remained stable on days 1, 3 and 5. Moreover, rapid re-epithelialization hindered the long-term permeability of both drugs. Combining CO2 fractional laser irradiation with SNF drug delivery could improve the efficiency of hydrophilic drug delivery within 24h before total re-epithelialization.

Fractional laser-assisted administration of human umbilical cord mesenchymal stem cells to reduce hypertrophic scars in rabbit ears.

The formation of hypertrophic scar is due to the abnormal accumulation and remodeling of the extracellular matrix, especially collagen tissue. Our research was designed to investigate the treatment effect of different administrations of human umbilical cord-derived stem cells and to hypertrophic scars on rabbit ears. Thirty New Zealand female white rabbits were treated as hypertrophic scar models. PBS was injected into the scars on the right ear of each group as control, while human umbilical cord-derived stem cells or condition medium of human umbilical cord-derived stem cells were administrated into the left ear through subcutaneous injection or fractional laser-assisted administration. Gross examination, scar elevation index (SEI) calculation and sampling were executed 5 weeks after administration. Then H&E and Masson staining analysis and the expression levels detections of α-SMA, Collagen I, TGF-β1, IL-1β, and IL-6 were performed. Our results demonstrated that the severity of hyperplasia was lower than the model group after stem cells and conditioned medium treatment. H&E and Masson staining results showed that the inflammation in scars was greatly alleviated and the degree of fibrosis was reduced after treatment. There was no significant difference in the therapeutic effect between subcutaneous injection or fractional laser-assisted administration. Both stem cells and conditioned medium can down-regulate SEI and factors expression levels in all groups. However, compared with the stem cells, the therapeutic effects of the conditioned medium were lower. The results confirmed that stem cells had an available treatment effect on hypertrophic scars of rabbit ears. In addition to the paracrine pathway, stem cells may have other ways to treat hypertrophic scars. Fractional laser-assisted administration may become a potential administration of stem cell clinical application in the future.

Dynamic changes of autophagy during hypertrophic scar formation and the role of autophagy intervention

BackgroundThe role of autophagy in the formation of hypertrophic scars (HS) remains unclear. This study aimed to explore the role and potential mechanism of autophagy during the development of HS. MethodsRNA and protein expression levels of Beclin-1, p62, and LC3II in normal skin tissues and HS specimens from different patients were examined. Autophagy inducers and inhibitors were used to cure established HS in rabbit ears, and the expression of Beclin-1, p62, and LC3II at the RNA and protein level was determined. Lastly, the effects of autophagy inducers and inhibitors on HS development were analyzed. ResultsCompared to normal skin tissues, the expression of LC3Ⅱ and Beclin-1 was higher (P<0.05), while that of p62 was lower (P<0.05) in HS tissues. In addition, the LC3II/LC3I ratio was increased during HS formation, and the altered expression of the three proteins stabilized after one year. Administration of autophagy inducers enhanced the formation of HS as well as the expression levels of LC3II and Beclin-1 but decreased p62 expression. Meanwhile, administration of autophagy inhibitors increased the expression of LC3II, Beclin-1, and p62, along with reduced HS formation. ConclusionAutophagic activity increased during HS initiation and subsequent stabilization. In addition, autophagy inhibitors were able to inhibit HS formation by suppressing autophagy, whereas autophagy inducers promoted scar hyperplasia by enhancing autophagy.

The therapeutic effect of Interleukin-18 on hypertrophic scar through inducing Fas ligand expression

ObjectiveAmong downstream interleukin-18 (IL-18) targets, Fas ligand (FasL) in particular, has been strongly implicated in many conditions. Our study aims to explore the role of IL-18 in hypertrophic scar through enhancing FasL expression. MethodsIL-18 expression in hypertrophic scar tissues and normal tissues were explored by immunohistochemistry, qRT-PCR and Western blotting, and the expression of IL-18 in normal skin fibroblasts and hypertrophic scar fibroblasts by immunofluorescence. Hypertrophic scar fibroblasts treated with recombinant human IL-18 (rhIL-18) were assessed with MTT, Annexin V-FITC/PI, qRT-PCR, ELISA and western blotting. In the hypertrophic scar of rabbit ears, rhIL-18 was injected to determine histological changes with HE and Masson staining. Additionally, the scars were rated based on contour and overall severity using a visual analog scale scores (VAS). ResultsIL-18 was decreased in hypertrophic scar tissues and fibroblasts compared to normal skin tissues and fibroblasts, respectively. Decreased proliferation and increased apoptosis of hypertrophic scar fibroblasts were found after rhIL-18 treatment with enhanced expression of FasL, sFasL FADD, Caspase-8, Caspase-9 and Caspase-3 in a dose-dependent manner. The VAS and thickness of scars in rabbit ears was decreased as time went on after rhIL-18 treatment, with decreases in scar elevation index (SEI) and the increases in FasL expression. ConclusionIL-18 curbs proliferation and promotes apoptosis of hypertrophic scar fibroblasts by enhancing FasL expression. IL-18is a potential target for treatment of hypertrophic scar.

Pigment epithelium-derived factor attenuates angiogenesis and collagen deposition in hypertrophic scars.

Scar forming wounds are often characterized by higher levels of vascularity than non-scarring wounds and normal skin, and inhibition of angiogenesis has been shown to inhibit scar formation in some model systems. The rabbit ear hypertrophic scar (HS) model has been widely used to study the mechanisms that underlie the development of HS as well as the effectiveness of various treatments. Although the rabbit ear HS model is well characterized in terms of scar formation, the rate and level of angiogenesis has not been investigated in this model, and the cause-effect relationship between angiogenesis and rabbit HSs has not been examined. In the current study, full-thickness excisional wounds were created on the ventral side of New Zealand White rabbit ears to induce HS formation, and the dynamic pattern of angiogenesis and the expression of angiogenic regulatory factors were examined over time. Blood vessel density was found to peak at 2.7% on day 14 post-wounding, decreasing to 1.7% by day 28. mRNA levels of the proangiogenic factor VEGF-A peaked at day 14, while the expression of the antiangiogenic factors pigment epithelium-derived factor (PEDF) and thrombospondin 1 (TSP1) peaked at day 28 post-wounding. To examine whether inhibition of angiogenesis influences HS formation in this model, wounds were treated with exogenous soluble antiangiogenic agents including recombinant PEDF (rPEDF) and a functional PEDF peptide (PEDF-335). rPEDF and PEDF-335 were administered intradermally from day 4 post-wounding every 3 days until day 19. Intradermal injection of rPEDF or PEDF-335 both led to decreased angiogenesis and decreased collagen deposition at the wound site. The results support the utility of antiangiogenic therapies, including rPEDF/PEDF-335, as a potential new strategy for the prevention or treatment of HSs.

Liposome-encapsulated statins reduce hypertrophic scarring through topical application.

Hypertrophic scar is an important clinical problem with limited therapeutic options. Aside from their roles as 3-hydroxy-3-methylglutaryl-coenzyme A reductase inhibitors, statins have also been demonstrated to decrease scarring by reducing connective tissue growth factor (CTGF) expression. However, poor penetrative ability limits their utility as topical treatments for hypertrophic scar. Here, we aim to develop novel statin formulations using liposomes to enhance dermal penetrative ability and to evaluate their efficacy against formation of hypertrophic scar utilizing our validated rabbit ear hypertrophic scar model. Liposomal simvastatin or pravastatin were compounded using a novel, flexible liposomal formulation and applied topically to rabbit ear hypertrophic scars daily from postoperation day (POD) 14 until POD 25. Scar color, including erythema and melanin, was measured using reflectance spectrophotometry on POD 28, and scar tissue was harvested for evaluation of scar elevation index as well as gene and protein expression. Human foreskin fibroblasts were also treated with statin formulations and CCN2 expression was determined by quantitative PCR. Both simvastatin and pravastatin were efficiently encapsulated in liposomes, forming nanometer-scale particles possessing highly negative charges. Topical treatment with liposomal simvastatin and pravastatin at 6.5% concentration significantly reduced scar elevation index and decreased type I/III collagen content and myofibroblast persistence in the wound. The erythema/vascularity of scars was reduced by liposomal statin treatment, with concomitant decrease of CD31 expression as measured histologically. Expression levels of transcripts encoding CTGF, collagen I, and collagen III collagen in scar tissue were also decreased by liposomal pravastatin treatment, as were myofibroblast persistence and the type I/III collagen ratio as assessed by immunofluorescence and picrosirus red staining, respectively. Treatment of human foreskin fibroblasts with simvastatin or with liposome-encapsulated pravastatin resulted in decreased expression of transcript encoding CTGF. Overall, our novel statin formulations encapsulated in liposomes were successfully delivered through topical application, significantly reducing hypertrophic scarring in a rabbit ear model.

Anti-inflammatory effects of ginsenoside Rg3 on the hypertrophic scar formation via the NF-κB/IκB signaling pathway in rabbit ears.

The anti-inflammatory effects of Rg3 on the hypertrophic scar (HS) formation remain relatively obscure. Hence, this study aimed to explore the anti-inflammatory effects of Rg3 on the HS formation using a rabbit ear model and we assessed the involvement of the NF-κ B/IκB signaling pathway in this process. We constructed the Newland white rabbit ear HS model and treated it with Rg3. Using histological analyses, we evaluated scar hypertrophy based on the hematoxylin and eosin staining. The degree of scarring was evaluated using the scar elevation index (SEI). In addition, collagen I and collagen III expression levels were assessed by immunohistochemistry, while fibroblast apoptosis was examined using TUNEL assays. While MPO, IL-1β, IL-6, and TNF-α concentrations were quantified using ELISA, NF-κB and p-IκB activities were respectively measured using electrophoretic mobility shift assays (EMSAs) and western blots. SEI measurements and histological characteristics revealed that Rg3 could suppress the HS formation. Moreover, Rg3 could inhibit the HS formation by decreasing collagen I and collagen III synthesis and inducing fibroblast apoptosis. Besides, Rg3 treatment markedly inhibited the inflammatory cytokine production and ameliorated neutrophil infiltration. Notably, this study revealed that Rg3 inhibited NF-κB activation and the activity of p-IκB. Furthermore, this study suggested that the ability of Rg3 to decrease the scar formation might result from its ability to inhibit inflammation by modulating the NF-κB/IκB signaling. Overall, the findings of this study could support the use of Rg3 to prevent the HS formation.