Previous work in our laboratory demonstrated that passive transfer of porcine reproductive and respiratory syndrome virus (PRRSV)-neutralizing antibodies (NA) protected pregnant sows against reproductive failure and conferred sterilizing immunity in sows and offspring. We report here on the dose requirement for protection by passive transfer with NA in young weaned pigs. The presence of a 1:8 titer of PRRSV-NA in serum consistently protected pigs against viremia. Nevertheless, their lungs, tonsils, buffy coat cells, and peripheral lymph nodes contained replicating PRRSV similar to the infected control group. Likewise, these animals excreted infectious virus to sentinels similar to the infectivity control animals. In an attempt to reach complete protective immunity equivalent to that previously observed in sows, the pigs were transferred with a higher titer of PRRSV-NA (1:32), and even then apparent sterilizing immunity was attained in only 50% of the animals. In conclusion, the presence of anti-PRRSV-NA in serum with a titer of 1:8 is enough to block viremia but not peripheral tissue seeding and transmission to contact animals. While a relatively low level of NA in blood is capable of conferring sterilizing immunity against PRRSV in sows, the amount of NA necessary to obtain full protection of a young weaned pig would be significantly higher, suggesting that differences exist in the PRRSV pathogenesis between both age groups. In addition, the titer of NA could be a helpful parameter of protection in the assessment of PRRSV vaccines.
Cell migration is a prerequisite for cancer invasion and metastasis, suggesting cell motility as a potential therapeutic target for cancer treatment. A synthetic library was screened to identify inhibitors of tumor cell migration. From this, we discovered that CAC-1098 (aurintricarboxylic acid) and CBI-0997 (5-(2,4-dimethoxy-5-ethylphenyl)-4-(4-bromophenyl) isoxazole) inhibited migration of MDA-MB-231 cells with IC 50 ؍ 5 and 50 nM, respectively. We synthesized KRIBB3 (5-(5-ethyl-2-hydroxy-4-methoxyphenyl)-4-(4-methoxyphenyl) isoxazole) by replacing the bromide group of CBI-0997 with a methoxyl group. Like CBI-0997, KRIBB3 has antimigratory and anti-invasive activities in MDA-MB-231 cells. Because KRIBB3 has a better drug-like structure, we focused our effort on further understanding its anti-migratory mechanism. Biotinyl-KRIBB3 was synthesized as an affinity probe for identification of KRIBB3-binding proteins. Using affinity chromatography, we identified Hsp27 as a target protein of KRIBB3 in vitro. Treatment of MDA-MB-231 cells with phorbol 12-myristate 13-acetate induced protein kinase C-dependent phosphorylation of Hsp27 and tumor cell migration. In contrast, treatment of MDA-MB-231 cells with KRIBB3 blocked phorbol 12-myristate 13-acetate-induced phosphorylation of Hsp27 and tumor cell migration. Furthermore, overexpression of Hsp27 antagonized the inhibitory effect of KRIBB3 on tumor cell invasion, and knockdown of Hsp27 using small interfering RNA inhibited tumor cell migration. Overall, our results demonstrate that KRIBB3 inhibits tumor cell migration and invasion by blocking protein kinase C-dependent phosphorylation of Hsp27 through its direct binding to Hsp27.Traditionally, genetic mutagenesis has proven to be a useful tool in solving the function of a wide range of genes in biological process. Recently, a chemical genetic approach has been developed to elucidate the principles of a wide range of biological processes (for review, see Refs. 1-3). In chemical genetics, instead of site-specific mutation as in traditional genetics, the function of a protein is inhibited or activated using small chemicals. Therefore, chemical genetics seeks to identify novel small molecules that afford functional dissection of cell biological pathways. Such chemicals are useful as bioactive molecular probes and allow further analysis of the relationship between target processes or proteins within cells and their cellular function.Metastasis plays a major role in morbidity and mortality from breast cancer (4). The metastatic potential of cancer cells is related to the ability to digest the extracellular matrix, migrate, cross blood vessel walls, and reach the blood circulation (for review, see Refs. 5-8). Cell movement is a complex process involving a number of steps, including the disruption of cell-cell junctions, cytoskeletal rearrangements, and the constant remodeling of adhesive contacts with the extracellular matrix (for review, see Refs. 9 -11). Cell migration contributes to several other important pathological pro...
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