Abstract
We have previously demonstrated that the human placenta contains a uniquely low sulfated extracellular aggrecan family chondroitin sulfate proteoglycan (CSPG). This CSPG is a major receptor for the adherence of Plasmodium falciparum-infected red blood cells (IRBCs) in placentas, causing pregnancy-specific malaria. However, it is not known whether such low sulfated CSPGs occur in placentas of other animals and, if so, whether IRBCs bind to those CSPGs. In this study, we show that rat placenta contains a uniquely low sulfated extracellular CSPG bearing chondroitin sulfate (CS) chains, which comprise only approximately 2% 4-sulfated and the remainder nonsulfated disaccharides. Surprisingly, the core protein of the rat placental CSPG, unlike that of the human placental CSPG, is a spongiotrophoblast-specific protein (SSP), which is expressed in a pregnancy stage-dependent manner. The majority of rat placental SSP is present in the CSPG form, and only approximately 10% occurs without CS chain substitution. Of the total SSP-CSPG in rat placenta, approximately 57% is modified with a single CS chain, and approximately 43% carries two CS chains. These data together with the previous finding on human placental CSPG suggest that the expression of low sulfated CSPG is a common feature of animal placentas. Our data also show that the unique species-specific difference in the biology of the rat and human placentas is reflected in the occurrence of completely different CSPG core protein types. Furthermore, the rat SSP-CSPG binds P. falciparum IRBCs in a CS chain-dependent manner. Since IRBCs have been reported to accumulate in the placentas of malaria parasite-infected rodents, our results have important implications for exploiting pregnant rats as a model for studying chondroitin 4-sulfate-based therapeutics for human placental malaria.
Highlights
We have previously shown that the low sulfated chondroitin sulfate proteoglycan (CSPG) is the receptor for the adherence of Plasmodium falciparum infected red blood cells (IRBCs) in the placentas of pregnant women and that the sulfate groupclustered regions of the C4S chains are the IRBC-binding sites [7,8,9]
As a part of our effort to determine whether pregnant rats can be useful as a model for studying pregnancy-specific malaria, we studied, in detail, the extracellular CSPG of the rat placenta
The CSPG was isolated by DEAE-Sephacel chromatography of isotonic buffer extracts of the pooled rat placental tissue and purified by CsBr density gradient centrifugation followed by gel filtration on columns of Sepharose CL-6B
Summary
Materials—Proteus vulgaris chondroitinase ABC, protease-free P. vulgaris chondroitinase ABC (120 units/mg), Arthrobacter aurescens chondroitinase AC II (87 units/mg), and super special grade C6S (shark cartilage) were purchased from Seikagaku America (Falmouth, MA). Purification of the CSPG by Size Exclusion Chromatography— The proteoglycan partially purified by CsBr density centrifugation was chromatographed on a column of Sepharose CL-6B (2 ϫ 65 cm) in 50 mM NaOAc, 150 mM NaCl, pH 6.0, containing 4 M GdnHCl. Fractions (2 ml) were collected and monitored for proteins by measuring absorption at 280 nm, and aliquots were analyzed for uronic acid contents [16]; yield was 4.9 mg. Analysis of Nonglycosylated and CSPG Form of SSP in Rat Placenta—The isotonic buffer extracts of rat placentas were analyzed on 15% polyacrylamide minigels before and after treatment with protease-free chondroitinase ABC, and the protein bands in the gels were transferred onto PVDF membranes. After washing the unbound cells, the bound cells were fixed, stained, and counted by light microscopy
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