[Bacteriostasis of Platelet-Rich Plasma and Its Derivatives in Vitro and Its Relationship with PF4].

OBJECTIVE: To investigate the bacteriostatic effect of platelet-rich plasma (PRP) and its derivative platelet gel (PG) supernatant on Escherichia coli in vitro and its relationship with platelet factor 4 (PF4). METHODS: Apheresis platelets donated by healthy volunteers were obtained from the Blood station of Lu an Blood Center as the source of PRP. The counts of platelet, white blood cell and red blood cell in PRP and its derivative PG supernatant were detected by automatic hematology analyzer. Bacterial growth of PRP and PG supernatants co-cultured with bacteria for different time was observed by plate coating culture method, and the contents of PF4 in PRP and PG supernatants were detected by ELISA. RESULTS: Apheresis platelets were collected from 28 healthy volunteers with a median age of 33 (21-56) years old. PRP can inhibit the growth of escherichia coli, but there were individual differences in antibacterial effect within 24 hours. PRP of 13 healthy volunteers had strong antibacterial effect at 24 hours, 7 cases had weak antibacterial effect at 24 hours, and 8 cases had no antibacterial effect at 24 hours. PG supernatant showed no significant individual difference, and all of them had bacteriostatic effect within 12 hours, but no bacteriostatic effect after 12 hours. There was no statistical difference in the bacteriostatic effect of PRP at 24 hours between healthy volunteers aged ≤30 years and >30 years (P>0.05), and there was no statistical difference between the white blood cell count ≤0.1×109/L and (0.1-1) ×109/L groups (P>0.05). There was significant difference in the bacteriostatic effect of PRP between the two groups with platelet content ≤1 000×109/L and >1 000×109/L (P<0.05). The platelet count in PRP was higher than that in PG supernatant ï¼»(911.57±160.52) ×109/L vs 0ï¼½. The PF4 level in PRP was higher than that in PG supernatant (23623.34±9822.14 vs 6664.74±4065.83, P<0.05). CONCLUSION: Both PRP and PG supernatant have antibacterial effects in Escherichia coli. The bacteriostatic effect of PRP was better than that of PG supernatant, and the platelet and PF4 contents in PRP were higher than those in PG supernatant, suggesting that the platelet and PF4 levels play an important role in bacteriostasis.

[Preparation and bioactivity evaluation of SA-mIL15 fusing protein].

AIM: To prepare and characterize streptavidin-tagged murine interleukin-15 fusion proteins. METHODS: pET24a-SA-L-mIL15 and pET21a-mIL15-L-SA plasmids were constructed and expressed in Rosetta (DE3) host bacteria to generate SA/mIL15 fusion proteins. SA-mIL15 fusion protein was purified through the Ni-NTA affinity chromatography, and mIL15-SA fusion protein through anion exchange chromatography, followed by refolding. The efficiency of surface modification of the fusion proteins on the biotinylated RM-1 tumor cells was evaluated by a flow cytometer. MTT method was used to evaluate the proliferating effect of SA/mIL15 fusion proteins on mouse spleen lymphocytes stimulated by ConA. RESULTS: Both SA-mIL15 and mIL15-SA fusion proteins were highly expressed in Rosetta (DE3) at about 20% of total bacterial proteins. They exhibited the bi-functionality: proliferation-promoting activity of mIL15 on mouse spleen lymphocytes with the specific activity of 1x10(6); IU/mg for SA-mIL15 or 2 x 10(5); IU/mg for mIL15-SA, and SA-mediated high-affinity binding to the biotinylated surfaces of RM-1 tumor cells with about 95% surface modification efficiency. CONCLUSION: SA/mIL15 bi-functional fusion proteins were generated, which made feasible the development of mIL15-surface-modified cancer cell vaccine.