Abstract: Fip-gts, an immunomodulatory protein purified from Ganoderma tsugae, has been reported to possess therapeutic effects in the treatment of cancer and autoimmune disease. For medicinal application, a recombinant Fip-gts was successfully expressed and purified in Sf21 insect cells by our previously work. It is important to evaluate the immunomodulatory activity of the rFip-gts. To assess the immunomodulatory potential of rFip-gts, the T lymphocytes of murine splenocytes were used in the present study. Results revealed that rFip-gts induced cellular aggregation formation. Additionally, the expression of IL-2 and IFN-r were up-regulated after the treatment of rFip-gts, and a corresponding increased production of IL-2 and IFN-r in a dose-dependent manner. The results showed that rFip-gts has an immunomodulatory activity in inducing Th1 lymphocytes from murine splenocytes released IL-2 and IFN-γ, thus suggest that rFip-gts may have therapeutic potential in vivo as an immune modulator.
Abstract: Characterization and evaluation of the activity of Vespa basalis DPP-IV, which expressed in Spodoptera frugiperda 21 cells. The expression of rDPP-IV was confirmed by SDS–PAGE, Western blot analyses, LC-MS/MS and measurement of its peptidase specificity. One-step purification by Ni-NTA affinity chromatography and the total amount of rDPP-IV recovered was approximately 6.4mg per liter from infected culture medium; an equivalent amount would be produced by 1x109 infected Sf21 insect cells. Through the affinity purification led to highly stable rDPP-IV enzyme was recovered and with significant peptidase activity. The rDPP-IV exhibited classical Michaelis–Menten kinetics, with kcat/Km in the range of 10-500 mM-1×S-1 for the five synthetic substrates and optimum substrate is Ala-Pro-pNA. As expected in inhibition assay, the enzymatic activity of rDPP-IV was significantly reduced by 80 or 60% in the presence of sitagliptin (a DPP-IV inhibitor) or PMSF (a serine protease inhibitor), but was not apparently affected by iodoacetamide (a cysteine protease inhibitor).
Abstract: Highly pathogenic avian influenza (HPAI) H5N1 viruses have created demand for a cost-effective vaccine to prevent a pandemic of the disease. Here, we report that Trichoplusia ni (T. ni) larvae can act as a cost-effective bioreactor to produce recombinant HA5 (rH5HA) proteins as an potential effective vaccine for chickens. To facilitate the recombinant virus identification, virus titer determination and access the infected larvae, we employed the internal ribosome entry site (IRES) derived from Perina nuda virus (PnV, belongs to insect picorna like Iflavirus genus) to construct a bi-cistronic baculovirus expression vector that can express the rH5HA protein and enhanced green fluorescent protein (EGFP) simultaneously. Western blot analysis revealed that the 70 kDa rH5HA protein and partially cleaved products (40 kDa H5HA1) were generated in T. ni larvae infected with recombinant baculovirus carrying the H5HA gene. These data suggest that the baculovirus-larvae recombinant protein expression system could be a cost-effective platform for H5N1 vaccine production.