An isologous porcine promoter permits high level expression of human hemoglobin in transgenic swine.

We describe isologous promoter replacement as an approach to permit high level expression of human hemoglobin in transgenic swine. We linked the human beta globin genomic coding region to the porcine beta globin promoter and used this fusion gene in an expression construct containing the human beta locus control region and the human alpha and epsilon genes to produce transgenic pigs. The highest level of expression was 24% human (32g/liter) and 30% human alpha/pig beta hybrid (40g/liter) hemoglobin in one transgenic pig. This pig was bred to a non-transgenic animal resulting in the transmission of high level human hemoglobin expression to 5 of 12 progeny.

Polymorphonuclear neutrophil iodination response as an estimate of defective yeast opsonization.

Polymorphonuclear neutrophil iodide uptake can be used as a measure of yeast opsonization and optimal assay conditions are described for the identification of sera defective in this function. The use of standard normal and defective sera permits correction of inter-assay variations resulting from the use of different cell donors. The iodination assay correlated well (r = 0.74, P less than 0.001) with a direct assay of yeast opsonization when both were used to measure this function in a panel of 72 sera. Differences in results between the two assay systems for some sera may be explained by a requirement for two different surface-associated opsonic molecules in the initiation of neutrophil metabolic activity.

High-efficiency synthesis of human alpha-endorphin and magainin in the erythrocytes of transgenic mice: a production system for therapeutic peptides.

Chemical synthesis of peptides, though feasible, is hindered by considerations of cost, purity, and efficiency of synthesizing longer chains. Here we describe a transgenic system for producing peptides of therapeutic interest as fusion proteins at low cost and high purity. Transgenic hemoglobin expression technology using the locus control region was employed to produce fusion hemoglobins in the erythrocytes of mice. The fusion hemoglobin contains the desired peptides as an extension at the C end of human alpha-globin. A protein cleavage site is inserted between the C end of the alpha-globin chain and the N-terminal residue of the desired peptide. The peptide is recovered after cleavage of the fusion protein with enzymes that recognize this cleavage signal as their substrate. Due to the selective compartmentalization of hemoglobin in the erythrocytes, purification of the fusion hemoglobin is easy and efficient. Because of its compact and highly ordered structure, the internal sites of hemoglobin are resistant to protease digestion and the desired peptide is efficiently released and recovered. The applicability of this approach was established by producing a 16-mer alpha-endorphin peptide and a 26-mer magainin peptide in transgenic mice. Transgenic animals and their progeny expressing these fusion proteins remain health, even when the fusion protein is expressed at > 25% of the total hemoglobin in the erythrocytes. Additional applications and potential improvements of this methodology are discussed.

A prospective study of human polyomavirus infection in pregnancy.

Urine samples from 1,235 pregnant women were examined by light microscopy for cytologic evidence of virus infection. Smears of urine sediment from 40 women (3.2%) were observed to contain inclusion-bearing cells; polyomavirus infection was confirmed by virologic methods in 24 (60%). A polyomavirus was isolated from 12 women. Five isolates were identified as JC virus and one as BK virus. Another isolate designated AS virus appeared to be unique. Serologic studies on the 40 women were consistent with a high frequency of reactivation of JC virus, and virus excretion was related to gestation. The evidence suggests that selective excretion of JC virus may occur in pregnancy. Among 390 pregnant women without inclusion-bearing cells in their urine, 78 (20%) had a high or rising titer of serum antibody to JC or BK virus or both, a result suggesting virus reactivation, but virus excretion was not detected. In contrast to other reports, no evidence was found for transmission of BK virus to the fetus.