Elastic Plasma-Protein-Film Blended with Platelet-Releasate Accelerates Healing of Diabetic Mouse Skin Wounds.
Authors: Tanaka Ryuhei , Ichioka Shigeru , Sekiya Naomi , Ohura Norihiko , Uchino Satomi , Ishikawa Ikue , Ojima Aki , Itoh Yoshihiro , Ishihara Osamu , Nakatsuka Takashi , Ikebuchi Kenji 
Published: October 2019

Numerous plasma proteins and cytokines derived from platelets mediate wound healing process that is characterized by the migration and differentiation of several cell populations that give rise to angiogenesis, collagen synthesis, wound contraction and re-epithelialization. To evaluate the efficacy of the blood derived factors in wound healing, we have exploited a novel wound dressing consisting of human plasma proteins and platelet-releasate. Human blood unit was drawn into a bag containing ACD after obtaining informed consent from healthy donors. Platelet-rich plasma was separated and the plasma proteins were concentrated together with platelets using cold ethanol precipitation method. The thrombin activity from the same unit was added to the concentrate, thereby leading to emergence of elastic soft layer consisting of the concentrated plasma proteins with platelet-relaseate (CPPP). The CPPP has enough strength to dress cutaneous wounds and contains considerable amount of cytokines and fibronectin (Table). We applied the CPPP to excisional skin wound in db/db mouse: a well-known animal model of impaired wound healing. Full-thickness skin defects were created on the mice symmetrically on both sides. One side of the wound was covered with the CPPP and the fresh clot from the same blood donor was applied to the other side as a control. Both of the wounds were then covered with polyurethane film. These wounds were evaluated 10 days after wounding.

Bioactive factors for wound healing in the Serum and CPPP Serum CPPP (Day 0) CPPP (Day 7) ND = non-detectable, NE = not evaluated bFGF (pg/mL) ND 106 (75-150) 43 (21-72) VEGF (pg/mL) 172 (20-405) 870 (500-1125) 706 (550-1100) PDGF-AB (ng/mL) 30 (20-40) 68 (25-1670) 116 (100-140) TGF β1 (ng/mL) 43 (34-48) 218 (110-415) >250 Fibronectin (mg/mL) 0.6 (0.2-0.8) 13 (8-20) NE

CPPP sustains these cytokines at substantial level on Day 7 (Table), suggesting that these growth factors are delivered continuously to the wound. Every wound edge was traced and the area was measured. Skin sections were stained with anti-CD31 antibody to visualize the blood vessels and determined its density (i.e., number of blood vessels/mm2 in the section). The area of CPPP treated wounds significantly decreased compared with that of the control (65% vs. 94% of the original size, respectively, p=.032). The immunostained section revealed the striking effect of CPPP on vasculalization. The mean vascular density of CPPP treated wounds was 13.2/mm2. In contrast, that of the control wounds was 2.7/mm2 (p=.013). Our results suggest that CPPP is a promising bioactive dressing for the treatment of full-thickness skin wound. Every biological component of CPPP is from a unit of collected blood; namely, CPPP can be an entirely autologous biological dressing, suggesting that CPPP is free from the risk of transmission of pathogens through blood products. Taken together, we may be able to design and conduct a clinical trial to evaluate the efficacy of CPPP for patients with non-healing wound.