Hisako Ishise

Calcium-Based Nanoparticles Accelerate Skin Wound Healing

Introduction Nanoparticles (NPs) are small entities that consist of a hydroxyapatite core, which can bind ions, proteins, and other organic molecules from the surrounding environment. These small conglomerations can influence environmental calcium levels and have the potential to modulate calcium homeostasis in vivo. Nanoparticles have been associated with various calcium-mediated disease processes, such as atherosclerosis and kidney stone formation. We hypothesized that nanoparticles could have an effect on other calcium-regulated processes, such as wound healing. In the present study, we synthesized pH-sensitive calcium-based nanoparticles and investigated their ability to enhance cutaneous wound repair. Methods Different populations of nanoparticles were synthesized on collagen-coated plates under various growth conditions. Bilateral dorsal cutaneous wounds were made on 8-week-old female Balb/c mice. Nanoparticles were then either administered intravenously or applied topically to the wound bed. The rate of wound closure was quantified. Intravenously injected nanoparticles were tracked using a FLAG detection system. The effect of nanoparticles on fibroblast contraction and proliferation was assessed. Results A population of pH-sensitive calcium-based nanoparticles was identified. When intravenously administered, these nanoparticles acutely increased the rate of wound healing. Intravenously administered nanoparticles were localized to the wound site, as evidenced by FLAG staining. Nanoparticles increased fibroblast calcium uptake in vitro and caused contracture of a fibroblast populated collagen lattice in a dose-dependent manner. Nanoparticles also increased the rate of fibroblast proliferation. Conclusion Intravenously administered, calcium-based nanoparticles can acutely decrease open wound size via contracture. We hypothesize that their contraction effect is mediated by the release of ionized calcium into the wound bed, which occurs when the pH-sensitive nanoparticles disintegrate in the acidic wound microenvironment. This is the first study to demonstrate that calcium-based nanoparticles can have a therapeutic benefit, which has important implications for the treatment of wounds.

Hypertrophic scar contracture is mediated by the TRPC3 mechanical force transducer via NFkB activation

Wound healing process is a complex and highly orchestrated process that ultimately results in the formation of scar tissue. Hypertrophic scar contracture is considered to be a pathologic and exaggerated wound healing response that is known to be triggered by repetitive mechanical forces. We now show that Transient Receptor Potential (TRP) C3 regulates the expression of fibronectin, a key regulatory molecule involved in the wound healing process, in response to mechanical strain via the NFkB pathway. TRPC3 is highly expressed in human hypertrophic scar tissue and mechanical stimuli are known to upregulate TRPC3 expression in human skin fibroblasts in vitro. TRPC3 overexpressing fibroblasts subjected to repetitive stretching forces showed robust expression levels of fibronectin. Furthermore, mechanical stretching of TRPC3 overexpressing fibroblasts induced the activation of nuclear factor-kappa B (NFκB), a regulator fibronectin expression, which was able to be attenuated by pharmacologic blockade of either TRPC3 or NFκB. Finally, transplantation of TRPC3 overexpressing fibroblasts into mice promoted wound contraction and increased fibronectin levels in vivo. These observations demonstrate that mechanical stretching drives fibronectin expression via the TRPC3-NFkB axis, leading to intractable wound contracture. This model explains how mechanical strain on cutaneous wounds might contribute to pathologic scarring.

A new mouse model of impaired wound healing after irradiation

Abstract Radiation has many benefits and is an important treatment for cancer therapy. However, it also has unfavourable side-effects. Among these side-effects, the impairment of wound healing in the skin is a major problem in clinics. Although many attempts have been made to overcome this shortcoming, there are few effective treatments for impaired wound healing after irradiation. One reason for this is that it is hard to obtain good animal models for researching this topic. In this study, two different models were created and investigated. In one model, rectangular flaps were created on the backs of mice and irradiated while the other parts of their bodies were covered with a lead board. In another model, the lower limbs were exposed to radiation. In each model, several doses of irradiation were tested. Skin ulcers were created in the irradiated area, and the wound healing process was observed. In order to verify the usefulness of the model, adipose derived stromal cells were injected into the wound and the healing rate was calculated. In the flap model, the flaps contracted and formed linear scars. On the other hand, in the thigh model, 15 Gy irradiation resulted in slow wound healing but no strong inflammation or necrosis. The transplantation of adipose tissue derived stromal cells into the irradiated thigh wound improved the wound healing. This study suggested that irradiation of the lower limb at ∼ 15 Gy might be an appropriate model for basic research into wound healing in irradiated skin.

Influence of continuous or intermittent negative pressure on bacterial proliferation potency in vitro.

Abstract Negative pressure wound therapy is helpful and effective in the treatment of intractable skin ulcers and defects, not only acute wounds. However, application of negative pressure wound therapy for an infected wound is still controversial. The authors developed an in-vitro model of negative pressure wound therapy and investigated the influence of various types of negative pressure environment on the proliferation potency of non-pathogenic Escherichia coli. E. coli in Luria-Bertani liquid media was cultured at 37°C under different environments, which were normal atmosphere in group 1, continuous negative pressure of 75 mmHg in group 2, intermittent negative pressure of 75 mmHg with cycle time of 1 minute aspiration and 1 minute abeyance in group 3, with the one of 3 minutes aspiration and 3 minutes abeyance in group 4. The relative amounts of E. coli in each group were investigated at different times. The proliferation potency of E. coli was higher under negative pressure than under normal atmosphere; higher under intermittent negative pressure than under continuous negative pressure; and higher under intermittent negative pressure with a short cycle than with a long cycle. It is important to consider the possibility that the intermittent and continuous mode of negative pressure wound therapy may promote proliferation of bacteria in an infected wound with no blood flow like necrotic tissue.

Comparative study of the antibacterial penetrating effects of wound dressings

Abstract In an infected wound, bacteria are present not only on the surface but also within the granulation tissue. The presence of bacteria inside the granulation tissue is one of the reasons for our inability to control infection. We have developed an in vitro model of an infected wound using Luria-Bertani (LB) agar media and non-pathogenic Escherichia coli, and compared the differences in the antibacterial effects of various types of antibacterial wound dressings. The results have shown that the penetrating antibacterial effects differed according to the type of wound dressing selected. As the thickness of the layer of the LB agar media increased, the potency of antibacterial effects caused by penetration decreased. In conclusion, it was suggested that antibacterial wound dressings can have antibacterial effects against bacteria not only on the surface of an infected wound but inside, so it is necessary to use different wound dressings according to the conditions of each infected wound.

Fibrous dysplasia and Klippel–Trenaunay syndrome: a rare association

Abstract Fibrous dysplasia is a rare congenital disorder, with abnormal hypertrophy of affected bone. A 17-year-old girl with a protrusion on her right forehead presented and was diagnosed as craniofacial fibrous dysplasia. Although she had no gait problem, her right leg was longer than the other. She had vascular malformation on the right leg. The condition was diagnosed as Klippel–Trénaunay syndrome, which also is a rare disorder. As the prevalence of these disorders is scarce, the probability of coincidental association of them is extremely low. No previous report about the association of fibrous dysplasia and Klippel–Trénaunay syndrome could be found. We concluded that, those two rare congenital disorders arose coincidentally in this patient.

Usage of Skin Tumor Images on the Internet for Personal Computer Based Automated Cognition

Visual diagnosis of skin tumors is one of the most important steps in dealing with them. Nowadays, artificial intelligence has been booming and technology of automated cognition by computer has been improved. One of the bottlenecks in building an efficient cognition system is providing adequate amount of data to base on. The internet may be a mine of the data. Feasibility of using macroscopic skin tumor images on the internet for automated cognition was studied by a personal computer. Skin tumor images were collected with scraping software. The quality of images varied. The most time-consumed process was to select images visually. After the selection, 600 macroscopic images consisting of 5 categories were qualified. The images were plotted with cluster analysis algorithms. Unsupervised data clustering: k-means clustering, principal component analysis and t-distributed stochastic neighbor embedding could not cluster them in human comprehensible fashion. Three-dimensional plotting of a supervised data clustering: Linear discrimination analysis, showed relatively clear clustering. Convolution neural networks were trained and tested for categorical accuracy. Time, consumed for training networks on a personal computer, was satisfactory. Categorical accuracy depended on what network it employed. Fine-tuning of a pre-trained network scored best categorical accuracy. Augmenting training data statistically increased categorical accuracy. Despite variation of image quality, using skin tumor images on the internet is a feasible approach for automated cognition.

Abstract 174: TRPC3 Channels Regulate Expression of Fibronectin Via NFκB Signling in Response to Cyclic Strain And Contribute to Hypertrophic Scar Formation

PURPOSE: In the cutaneous wound healing process, fibronectin is robustly secreted from fibroblasts. Fibronectin serves a variety of important functions in the wound healing process beyond just providing as a supporting scaffold. Fibronectin induces cell differentiation, migration, coagulation and formation of ECM, which ultimately serves to promote wound contraction. The expression of fibronectin is under complex control. Among the transcriptional factors interacting with the fibronectin promoter, Nuclear factor-kappa B (NFκB) also regulates the transcriptional activity of fibronectin.

Abstract 169: Calcium Entry Via TRPC3 Channels Transduce Mechanical Force And Accerelate Wound Contraction

118 type mice. Doppler analysis of the wounded area showed increased blood flow in both CD47 and thrombospondin-1 null mice. Accelerated wound closure in the CD47 null mice was associated with increased fibrosis as demonstrated by a 4-fold increase in collagen fraction. Wound tissue of CD47 null mice showed increased thrombospondin-1 mRNA and protein expression and TGF-β1 mRNA levels. Activation of latent TGF-β1 was increased in thermally injured CD47null tissue as assessed by phosphorylation of the TGF-β1 receptor-regulated transcription factors SMAD-2 and -3.