Chitosan-microencapsulated rhEGF in promoting wound healing.

AIMS: Chitosan and epidermal growth factor (EGF) have been shown to improve wound healing. This study investigates the healing effects of a spray solution (NewEpi, JoyCom Bio-Chem Co. Ltd., Taiwan) containing recombinant human EGF (rhEGF) delivered via a newly patented technology-chitosan microencapsulated nanoparticles. METHODS: On Wistar rats, two full-thickness wounds on the dorsum bilateral of the spine were created. The rats were randomised to the following treatment groups: hydrogel, wet dressing, foam, rhEGF spray and rhEGF spray+foam. Sterile dressings were applied and changed daily. A total of 2µg of rhEGF was administered in two sprays during each dressing change. All animals were euthanised on day 14. Tissue samples were taken from the wound bed, including an area of 2cm surrounding the wound margin for histological evaluations. RESULTS: Wounds treated with the rhEGF spray achieved the greatest size reduction by day 14 compared with other types of conventional dressings. An overall significant difference in levels of collagen synthesis existed between groups (p<0.01). Pair-wise comparisons showed that the rhEGF spray treatment significantly promoted higher levels of mature Type I collagen than any other conventional dressings (p<0.01), whereas non-rhEGF treatments resulted in higher levels of Type III collagen. The regenerated tissue in rhEGF spray treatment groups was also in alignment with that of normal skin. Epidermis, dermis and hair follicles were easily observed in wounds treated with the rhEGF spray. CONCLUSION: The major challenge of topical application of rhEGF was overcome by using a new drug delivery technology: chitosan-rhEGF nanoparticles. The positive healing effects observed in this study suggest the therapeutic potentials of this novel rhEGF topical spray solution.

A mini-pig model for evaluating the efficacy of autologous platelet patches on induced acute full thickness wound healing.

BACKGROUND: Autologous platelet concentrates are currently widely used across different areas of regenerative medicine in order to enhance the wound healing process. Although several protocols for platelet concentrates are available, their application remains difficult due to different protocols leading to distinct products with vary potential biological uses. In this study, we attempted to make a platelet patch (PP) using mixtures of platelet rich plasma (PRP) injection and platelet rich fibrin (PRF) to promote wound repair and regeneration. RESULTS: Experiments were performed using a full-thickness wound model in mini-pigs. Autologous PRP, PRF and PP were prepared immediately before creating four full-thickness skin wounds in pigs. We quantified concentrations of platelets, thrombin and various growth factors to ensure that the desired effect can be produced. After surgery, hydrocolloid dressing, PRP injection, PRF and PP was applied to experimentally induced wounds. Application efficacy was evaluated by measurement of wound sizes and histological examination. The results indicated that all wounds showed a significant size reduction. Wound repair efficacy in response to PP treatment exhibited enhanced re-epithelialization compared to PRP and PRF (P < 0.05) and higher wound contraction than did PRF application (P < 0.05). Another aspect, experiment using DsRed transgenic pigs as blood donors demonstrated that leucocytes in PP were incorporated into the wound bed at the end of the study, suggesting that leucocytes activity is stimulated in response to PP application. Safety of the experimental processes was also confirmed by examination of organ biopsies. CONCLUSIONS: We used a mini-pig model to evaluate the efficacy of lab-made PP on induced full-thickness wound healing. Results demonstrated that application of one piece of PP was enough to obtain comparable efficacy versus general utilization of PRP or PRF for wound care. We also demonstrated that leucocytes in PP were incorporated into the wound bed and no safety concerns have been found in the whole experiment. This study provides a novel and feasible method for veterinary or clinical wound care.

Functional evaluation for adequacy of MDCK-lineage cells in influenza research.

OBJECTIVE: Influenza is an acute respiratory disease caused by the influenza virus which circulates annually in populations of different species. Madin-Darby Canine Kidney (MDCK) is the most widely utilized cell-line for conducting influenza research. However, the infectivity of various influenza strains in MDCK cells is not equivalent and the productivity of viral propagation is also limited. RESULTS: We tested the functional adequacy of two MDCK-lineage cell lines, conventional MDCK and MDCK/London, were evaluated by assessing their infectivity of different influenza viral strains with focus forming assays and the cellular toxicity caused by influenza infections by lactate dehydrogenase assay. Moreover, the sensitivity of cells in the presence of the antiviral agent ribavirin was assessed by MTT assay. Our results showed that MDCK/London cells efficiently propagate virus across all influenza viruses tested, are comparable to the utility of Mv1Lu cells, and are superior to conventional MDCK cells in replicating virus as indicated by an increase in virus of three to four logs, particularly in H3N2 infection. Also, the MDCK/London cells were more sensitive to the presence of antiviral drug than conventional MDCK cells. In conclusion, MDCK/London cell line could be a better platform for influenza studies and vaccine development.

Use of platelet-rich plasma and platelet-derived patches to treat chronic wounds.

OBJECTIVE: To compare the healing results between platelet-rich plasma (PRP) and platelet-derived patches versus traditional advanced wound dressings in patients with chronic wounds. METHOD: Patients with and without diabetes were divided into two groups, each of which received either PRP patch treatments or the advanced wound dressings. All wounds were cleaned, debrided and assessed by physicians. The data were analysed and represented as mean ± standard deviation (SD). Student's t-test was used to calculate the significance of differences between both groups. Values of p<0.05 were considered statistically significant. RESULTS: Patients with and without diabetes receiving PRP patch treatments saw improvement in wound healing in two weeks (p=0.0083). Patients with diabetes who received platelet-derived patch treatment and PRP injection experienced wound size reduction to <25% of the original area by the fourth week of treatment, and >90% of the subjects had wounds of <10% their original size in the last three weeks of the trial. Conversely, the wound area in the control subjects receiving traditional advanced wound dressings remained at 25-50% of their original size from the fourth week of treatment to the end of the trial. The healing process of the PRP patch experimental group was statistically significant compared with the control group (p<0.0001). CONCLUSION: Combining treatments of PRP injections and platelet-derived patches significantly improved the healing outcomes of patients with chronic wounds, most notably in patients with diabetes, when compared with a traditional treatment of advanced wound dressings.

Anti-inflammatory effects of Antrodia camphorata, a herbal medicine, in a mouse skin ischemia model.

ETHNOPHARMACOLOGICAL EVIDENCE: Antrodia camphorata, a highly valued polypore mushroom native only to Taiwan, has been traditionally used as a medicine for anti-inflammation. AIM OF THE STUDY: In this study, anti-inflammatory effects of Antrodia camphorata (AC) and its active compound, ergostatrien-3ß-ol (ST1), were investigated in a mouse skin ischemia model induced by skin flap surgery on the dorsal skin. MATERIALS AND METHODS: A U-shaped flap was elevated on the dorsal skin of the nine-week-old male mice. Mice were randomly assigned to six groups for treatment (n=6) including normal skin/propylene glycol (PG), surgical skin flap/PG, solid-state-cultured AC (S/AC), wood-cultured AC (W/AC), high-dose ST1 (H-ST1), low-dose ST1 (L-ST1). Antrodia camphorata was dissolved in 25µL PG and smeared on the skin flap every six hours for 24h. At the end of the experiment, each mouse was anesthetized, and skin tissues were collected from their back for histopathological analysis, extracting RNA and protein according to our previous reports. RESULTS: Skin-flap-induced ischemia damage significantly increased the expression of the iNOS, COX2, and IL-6 proteins and decreased the expression of IκB protein. In addition, focal, moderate coagulative necrosis with inflammatory cell infiltration was found in the epidermis, and moderate inflammatory cells and necrosis with slight edema was noted in the sub-dermis at 24h after skin flap surgery. However, treatment with solid-state-cultured or wood-cultured AC, or with its derived ST1 active compound, significantly reduced the necrosis and inflammatory cell infiltration in both the epidermis and sub-dermis of the skin flap. The treatments also reduced the inflammatory response by decreasing the expression of inflammation-related genes including iNOS, IL-6, TNF-α, and NF-κB, as shown by changes in RNA and protein expression, when compared with the surgical skin flap procedure alone. CONCLUSIONS: These results demonstrated that methanolic extracts of wood-cultured fruiting bodies and solid-state-cultured mycelia from Antrodia camphorata have excellent anti-inflammatory activities and thus have great potential as an addition for hydrocolloid dressings.

Therapeutic Potential of Andrographolide Isolated from the Leaves of Andrographis paniculata Nees for Treating Lung Adenocarcinomas.

Andrographolide is one of the major diterpene lactones found in Andrographis paniculata Nees and exhibits remarkable inhibitory effects on various cancers. In this study, the antipulmonary cancer effects of andrographolide were studied in a lung tumor mouse model induced by human vascular endothelial growth factor A 165 (hVEGF-A165). These results demonstrated that andrographolide significantly reduced the expression of hVEGF-A165 compared with a mock group in the Clara cells of the lungs. In addition, andrographolide also decreased tumor formation by reducing VEGF, EGFR, Cyclin A, and Cyclin B expression on the transcriptional and translational levels. These results indicated that andrographolide treatment on the overexpression of VEGF can arrest the cell cycle, which induced pulmonary tumors in transgenic mice. In conclusion, the antiangiogenesis and chemotherapeutic potential of andrographolide may provide a cure for pulmonary tumors in the future.

Active Component of Danshen (Salvia miltiorrhiza Bunge), Tanshinone I, Attenuates Lung Tumorigenesis via Inhibitions of VEGF, Cyclin A, and Cyclin B Expressions.

Tanshinone I (T1) and tanshinone II (T2) are the major diterpenes isolated from Danshen (Salvia miltiorrhiza Bunge). Three human lung adenocarcinoma cell lines, A549, CL1-0, and CL1-5, were treated with T1 and T2 for the in vitro antitumor test. Results showed that T1 was more effective than T2 in inhibiting the growth of lung cancer cells via suppressing the expression of VEGF, Cyclin A, and Cyclin B proteins in a dose-dependent manner. Moreover, a transgenic mice model of the human vascular endothelial growth factor-A165 (hVEGF-A 165) gene-induced pulmonary tumor was further treated with T1 for the in vivo lung cancer therapy test. T1 significantly attenuated hVEGF-A165 overexpression to normal levels of the transgenic mice (Tg) that were pretreated with human monocytic leukemia THP-1 cell-derived conditioned medium (CM). It also suppressed the formation of lung adenocarcinoma tumors (16.7%) compared with two placebo groups (50% for Tg/Placebo and 83.3% for Tg/CM/Placebo; P < 0.01). This antitumor effect is likely to slow the progression of cells through the S and G2/M phases of the cell cycle. Blocking of the tumor-activated cell cycle pathway may be a critical mechanism for the observed antitumorigenic effects of T1 treatment on vasculogenesis and angiogenesis.

Bovine lactoferrin inhibits lung cancer growth through suppression of both inflammation and expression of vascular endothelial growth factor.

Lung cancers are among the most common cancers in the world, and the search for effective and safe drugs for the chemoprevention and therapy of pulmonary cancer has become important. In this study, bovine lactoferrin (bLF) was used in both in vitro and in vivo approaches to investigate its activity against lung cancer. A human lung cancer cell line, A549, which expresses a high level of vascular endothelial growth factor (VEGF) under hypoxia, was used as an in vitro system for bLF treatment. A strain of transgenic mice carrying the human VEGF-A165 (hVEGF-A165) gene, which induces pulmonary tumors, was used as an in vivo lung cancer therapy model. We found that bLF significantly decreased proliferation of A549 cells by decreasing the expression of VEGF protein in a dose-dependent manner. Furthermore, oral administration of bLF at 300 mg/kg of body weight 3 times a week for 1.5 mo to the transgenic mice overexpressing hVEGF-A165 significantly eliminated expression of hVEGF-A165 and suppressed the formation of tumors. Additionally, treatment with bLF significantly decreased the levels of proinflammatory cytokines, such as tumor necrosis factor-α, and antiinflammatory cytokines, such as IL-4 and IL-10. Levels of IL-6, which is both a proinflammatory and an antiinflammatory cytokine, were also reduced. Treatment with bLF decreased levels of tumor necrosis factor-α, IL-4, IL-6, and IL-10 cytokines, resulting in limited inflammation, which then restricted growth of the lung cancer. Our results revealed that bLF is an inhibitor of angiogenesis and blocks lung cell inflammation; as such, it has considerable potential for therapeutic use in the treatment of lung cancer.