THE EFFECTS OF PRP-CE TREATMENT ON WOUND HEALING IN A MURINE SKIN INJURY MODEL. OBJECTIVE :

1. This study was done to estimate effect the biological activity of Proline Rich Polypeptide  Colostrum Extract  (PRP-CE) from BioPharma. We provided topical application on the wound healing process in mice after skin mechanical injury. Mice were anesthetized and two full-thickness excisional wounds were made on both sides on the dorsal area of mice using surgical scissors and forceps. The size of the wound was 5 mm diameter. The bottom of the wound was panniculus carnosus, with no visible bleeding. Wounds of the mice of first group were treated with PRP-CE ( - right side ) and with distilled water (- left side), respectively. The mice of the second group were treated as follows: with distilled water ( - right side wound) and no any treatment ( – left side ). Wounds were treated once a day for ten days. We found that topical application of PRP-CE improves wound healing process in mice about 22% better to control.We also provide assessment of PRP-CE anti-allergic properties. The two methods were used. Method of active sensibilization of guinea pigs by P. Anderson (1980) which is based on one time injection of ovalbumin and Histamine model of induced bronchial spasm. In case of  active sensibilization an anti-allergic effect was observed. Introduction of PRP-CE prior to ovalbumin resulted in death only in 28% of animals, compared to 100% in control group. Interestingly, PRP-CE did not have any noticeable effect on the development of system anaphylaxis induced in sensibilized guinea pigs by xenogenic histamine.   The effects of Proline Rich Polypeptide  Collostrum Extract treatment on wound healing in a murine skin injury model and assessment of it’s antiallergic properties on system anaphylaxis in guinea pigs.A. Krylov, E. Bogdanenko, T. Bogush, R. Zhdanov, Institute of General Pathology and Pathophisiology, Baltijskaja 8, 125315, Moscow, Russia,  E-mail: akrylo2@mail.ru, Fax: 7-495-151-17-56 Fig. 1. A typical image of mouse with dorsal wounds. Fig. 2. Representative image of mouse from Immulox treated group in 5 days after injury. Proline-rich polypeptides (PRPs) are small, very low-weight molecules (6,000 daltons) that have an active immune-modulating effect upon the thymus. They have been variously described as biological response modulators, infopeptides, transfer factors and colostrinin.9,10 Immunodeficiency (Th1-deficit states), as in HIV, EBV and herpes, may be thus counteracted, while immune hyperactivity (Th2 hyperactivity), as in autoimmune and allergic diathesis conditions, may be inhibited.11 PRPs isolated from colostrum and taken sublingually have shown great promise as an immune-equilibrating nutraceutical and as a potential therapy for Alzheimer’s disease!12 Table 1. Effect of PRP-CE on wounds healing in mice 5 days after skin injury THE EFFECTS OF PRP-CE TREATMENT ON WOUND HEALING IN A MURINE SKIN INJURY MODEL. OBJECTIVE: The purpose of the study is estimate effect of PRP-CE topical application on wound healing process in mice after skin mechanical injury. METHODS: Mice: Female Balb/c mice with average weight 34 g were used. Mice were anesthetized with an intraperitoneal injection of phenobarbital. Two full-thickness excisional wounds were made on both sides on the dorsal area of mice using surgical scissors and forceps. The size of the wound is about 5 mm diameter. The bottom of the wound was panniculus carnosus, with no visible bleeding. Two groups of mice were used in the experiment with 10 animals in each group. Wounds of the mice of first group were treated with PRP-CE- right side and with distilled water - left side respectively. The mice of the second group were treated as follows: with distilled water - right side wound and no any treatment – left side. Wounds were treated once a day for ten days. Treatment began immediately after injury. Mice were imaged immediately after injury and then daily. The size of the wounds was determined by the measurement of wound area obtained from the captured digital images. RESULTS: Initially all mice had a wounds of similar size which was accomplished by using stencil. The Figure 1 demonstrates a typical image of mouse after mechanical skin injury. The measurements of wound size after injury were presented in Table 1 after 5 days as difference in healing effect was most prominent compared to earlier periods. Healing process was practically completed after 10 days in all cases. The results of the measurements are presented in Table 1 and Table 2. Table 1. Effect of PRP-CE on wounds healing in mice 5 days after skin injury Size of the wounds in the images is presented in arbitrary units. Ratio between experimental and control wound areas (R/L) demonstrates the effect of Immulox compared to control side on wound healing. The value R/Ldetermined from the Tableis= 0.78 ± 0.19. Obtained result indicates that Immulox treatment is 22% more effective than control healing. Table 2 Comparison of water treated with dry wound healing process in mice 5 days after injury. Obtained from the Table 2 value R/L is 1.02 ± 0.06. It shows that the difference between dry and water treated healing rate is insignificant. • CONCLUSIONS • It has been shown that topical application of PRP-CE improves wound healing process in mice about 22% better compared to control. • REFERENCES • Wang RY, Abrams T, Monfils P, Salness R, Raymond RM. The effects of colchicine treatment on wound healing in a caustic murine injury model, J. Toxicol Clin Toxicol. 1999;37(7):845-53 • Padgett DA, Marucha PT, Sheridan JF Restraint stress slows cutaneous wound healing in mice, Brain Behav Immun. 1998 Mar;12(1):64-73 • Joseph HL, Anderson GL, Barker JH, Roisen FJ, Weiner LJ, Tobin GR, Inhibition of wound contraction with colchicine and D-penicillamine, J Surg Res. 1996 Feb 15;61(1):197-200 • Oberbaum M, Markovits R, Weisman Z, Kalinkevits A, Bentwich Z, Wound healing by homeopathic silica dilutions in mice, Harefuah. 1992 Aug;123(3-4):79-82, 156 • Bergman A, Yanai J, Weiss J, Bell D, David MP, Acceleration of wound healing by topical application of honey. An animal model., Am J Surg. 1983 Mar;145(3):374-6 / ASSESSMENT OF PRP-CE ANTIALLERGIC PROPERTIES ON SYSTEM ANAPHYLAXIS IN GUINEA PIGS. Material and methods Study was conducted on guinea pigs maintained in standard conditions. Ovalbumin (GIII) and Histamine dihydrochloride (Sigma) were used in the study. Ovalbumin and histamine were introduced into the animals with help of compressor nebulaiser (Pari). Method of active sensibilization of guinea pigs by P. Andersson (1980) Guinea pigs with weight 250-300 g were sensibilized by one time injection of ovalbumin (OA) 10 mg and aluminum hydroxide (Al(OH3)) 100 mg in 1 ml of water volume. Dosage was distributed by 0.2 ml intramuscularly in two hips and 0.6 ml intra abdominally. Observation of allergic reaction was done 5 weeks later, when enough amount of IgE antibodies formed and antigen introduction results in development bronchial spasms. Histamine model of induced bronchial spasm in guinea pigs. ненаркотизированных 0.2% histamine in 0.9% solution was injected with nebulaiser by inhalation into guinea pigs placed in special chamber. Duration of bronchial spasm reaction was measured in seconds. Two phases of spasm are observed: 1 – acute phase: animal rests in side position, frequency of breath 10 – 15 times a minute; 2 – sub acute phase: animal sits, increased frequency of breath up to 40-50 times a minute. Introduction of PRP-CE was done by the spray method 5 or 10 times into animal mouth 30 minutes before histamine injection. Experimental design: 1. Assessment of bronchial spasm duration after injection of histamine (internal control). 2. Introduction of PRP-CE 24 hours after histamine injection, and repeated histamine injection and assessment of duration of bronchial spasm reaction. Antigen-induced bronchial spasm model on immunized guinea pigs Experiment was conducted on actively-sensibilized by ovalbumin guinea pigs (in accordance with approach developed by P. Anderson (1980)). Ovalbumin in dosage of 2.5 mg/kg was used as inductor of bronchial spasm. Animals were divided in two groups. Ovabumin 2.5 mg/kg dissolved in 1 ml of physiological solution was introduced by inhalation with nebulaiser during 3 min into animal of first group – control. PRP-CE was introduced to the animals of experimental group Table 2. Effect of Immulox on the development Ovalbumin-induced bronchial spasm in sensibilized guinea pigs. Control Table 1. Effect of PRP-CE on the development histamine-induced bronchial spasm in sensibilized guinea pigs. Control Results PRP-CE in dosages 5 and 10 sprays did not inhibit development of bronchial spasm reaction in sensibilized guinea pigs followed by histamine inhalation, see Table 1. Duration of acute and sub acute phases were practically similar in control (histamine only) and experimental (PRP-CE and histamine) groups. Antiallergic effect has been observed in ovalbumin-induced bronchial spasm model in sensibilized guinea pigs, see Table 2. Injection of permissive dosage of Ovalbumin results in bronchial spasm reaction in 100% of animals (6 out 6), 3 animals (50%) died from difficulty of breathing. Average duration of acute phase of spasm reaction was about 300 sec in other 3 animals (50%) Intoduction PRP-CE prior to Ovalbumin resulted in death only in 2 animals out 7 (28.6%). The other animals of the same group 5 out 7 (71.4%) did not develop bronchial spasm reaction on permissive dosage of antigen (Ovalbumin), see Table 2. The difference in duration of latent period (time between introduction of permissive dosage of antigen and the beginning of acute phase of spasm) between experimental (with PRP-CE and control group was also statistically significant, (p<0.05 using Fisher’s criterion). CONCLUSIONS PRP-CE did not have any noticeable effect on the development of system anaphylaxis, induced in sensibilized guinea pigs by xenogenic histamine. PRP-CE has showed clear anti-allergic activity and inhibited the development of systemic anaphylaxis, induced in sensibilized guinea pigs by Ovalbumin. PRP-CE PRP-CE