Viewing Hyaluronan: Imaging Contributes to Imagining New Roles for This Amazing Matrix Polymer

Hyaluronan (HA) is an ubiquitous extracellular matrix polymer that plays many roles in health and disease. The ability to view the spatial and temporal expression of HA in tissues and on/in cells has provided researchers with insights into the tremendously diverse biological processes in which HA is involved. Biochemical extraction, quantity, and size measurement of HA can tell part of the story, but these techniques are incomplete in placing HA at the scene of a biological event and determining which other molecules are likely to be cooperating. HA, however, is not immunogenic, so preparing antibodies for histochemistry is problematic. Fortunately, a probe for HA was devised based on the HA binding region of aggrecan, and today this probe is commercially available and very useful for histochemistry. This article discusses the conditions and considerations that the authors’ lab and others have developed for optimal HA staining in many tissues and cell types

Rhop-3 Protein Conservation Among Plasmodium Species and Induced Protection Against Lethal P. yoelii and P. berghei Challenge

In the present study, Rhop-3 polymorphism among Plasmodium falciparum field and laboratory isolates and among rodent Plasmodium species was investigated and identified. The Rhop-3 gene was found in all Plasmodium species so far tested. The overall structure of the Rhop-3 protein was found conserved among P. falciparum, Plasmodium yoelii, and Plasmodium berghei. However, it was more conserved among rodent Plasmodium species than between P. falciparum and Plasmodium vivax. The most conserved regions of Rhop-3 are the second half of exon 6 (amino acid #548 to #665) and the beginning of exon 3 (amino acid #59 to #210). Recombinant C-terminal partial and full-length Rhop-3 proteins of P. yoelii and P. berghei were expressed in Escherichia coli and purified. Immunization-challenge experiments in mice using recombinant Rhop-3 proteins led to a delay in parasite development and protected mice from a homologous lethal challenge infection. In a group of eight outbred Carworth Farm White (CFW) mice immunized with P. yoelii C-terminal recombinant His-Y1412 protein, three mice (37.5%) were protected from a lethal P. yoelii challenge. In BALB/cJ mice one mouse (20%) survived the infection. Immunization of mice with P. berghei recombinant full-length Rhop-3 protein in BALB/cJ mice led to a 40% survival from lethal P. berghei challenge. CFW mice immunized with P. berghei recombinant full-length Rhop-3 protein showed a significant delay in parasite development with a heterologous P. yoelii challenge. The Rhop-3 protein is a promising candidate for an asexual stage malaria vaccine. © Springer-Verlag 2006

Trafficking and Association of Plasmodium falciparum Mc-2tm With the Maurer’s Clefts

In this study, we investigated stage specific expression, trafficking, solubility and topology of endogenous PfMC-2TM in P. falciparum (3D7) infected erythrocytes. Following Brefeldin A (BFA) treatment of parasites, PfMC-2TM traffic was evaluated using immunofluorescence with antibodies reactive with PfMC-2TM. PfMC-2TM is sensitive to BFA treatment and permeabilization of infected erythrocytes with streptolysin O (SLO) and saponin, showed that the N and C-termini of PfMC-2TM are exposed to the erythrocyte cytoplasm with the central portion of the protein protected in the MC membranes. PfMC-2TM was expressed as early as 4 h post invasion (hpi), was tightly colocalized with REX-1 and trafficked to the erythrocyte membrane without a change in solubility. PfMC-2TM associated with the MC and infected erythrocyte membrane and was resistant to extraction with alkaline sodium carbonate, suggestive of protein-lipid interactions with membranes of the MC and erythrocyte. PfMC-2TM is an additional marker of the nascent MCs