Intercellular adhesion molecule-1 and CD36 synergize to mediate adherence of Plasmodium falciparum-infected erythrocytes to cultured human microvascular endothelial cells.

We have compared the adhesion of Plasmodium falciparum-infected erythrocytes to human dermal microvascular endothelial cells (HDMEC) and human umbilical vein endothelial cells (HUVEC) and have assessed the relative roles of the receptors CD36 and intercellular adhesion molecule-1 (ICAM-1). HUVEC (a cell line that expresses high levels of ICAM-1 but no CD36) mediate low levels of adhesion, whereas HDMEC (which constitutively express CD36) mediate high levels of adhesion even before ICAM-1 induction ICAM-1 expression leads to yet greater levels of adhesion, which are inhibited both by anti-ICAM-1 and CD36 mAbs, despite no increase in the expression of CD36. The results indicate the presence of a substantial population of infected cells that require the presence of both receptors to establish adhesion. Synergy between these receptors could be demonstrated using a number of parasite lines, but it could not be predicted from the binding of these same parasite lines to purified ICAM-1 and CD36. This phenomenon could not be reproduced using either purified receptors presented on plastic, or formalin-fixed HDMEC, suggesting that receptor mobility is important. This is the first study to demonstrate receptor synergy in malaria cytoadherence to human endothelial cells, a phenomenon necessary for parasite survival and associated with disease severity.

Identification of heparin as a ligand for the A-domain of Plasmodium falciparum thrombospondin-related adhesion protein.

Thrombospondin-related adhesion protein (TRAP) is a Plasmodium falciparum transmembrane protein that is expressed within the micronemes of sporozoites, and is implicated in host cell invasion and motility. Contained within the extracellular region of TRAP is an A-domain, a module found in a number of membrane, plasma and matrix proteins, that is often involved in ligand recognition. In order to determine the role of the TRAP A-domain, it has been expressed as a glutathione S-transferase fusion protein and its ligand binding compared with that of other characterised glutathione S-transferase A-domain fusion proteins. Using a solid phase assay to screen for binding to known A-domain ligands, the TRAP A-domain was found to bind heparin. Binding to heparin appeared to be specific as it was saturable, and was inhibited by soluble heparin, fucoidan and dextran sulfate, but not by other negatively charged sulfated glycosaminoglycans such as chondroitin sulfates. Furthermore, unlike some A-domain ligand interactions, the A-domain of both TRAP and the leukocyte integrin, Mac-1, bound to heparin in the absence of divalent cations. It has been shown previously that another domain within TRAP, which is homologous to region II-plus of circumsporozoite protein, binds to sulfatide and to heparan sulfate on the immortalised hepatocyte line HepG2. The TRAP A-domain also bound to sulfatide and to HepG2 cells. Thus the A-domain shares certain binding properties already attributed to the region II-plus-like domain of TRAP, and may contribute to the binding of TRAP to heparan sulfate on hepatocytes.

A comparison of large block macrosectioning and conventional techniques in breast pathology.

Two techniques for the histological handling of breast specimens, namely conventional sampling using small blocks (SB) and a large block (LB) macrosectioning technique have been compared, with special emphasis on tumour size and in situ carcinoma, in an attempt to objectively demonstrate the advantages of the latter method. This is considered to be of particular importance in guiding the clinicians in their use of the many treatment modalities available for breast carcinoma. All cases were from the routine surgical caseload; 100 examined by the LB technique and 111 using conventional SB. The LB technique gave a reliable measurement of invasive carcinoma in 100% of cases compared to only 63% of SB cases. Ductal carcinoma in situ (DCIS), was found more frequently (80% versus 64%) and its extent was more easily and reliably measured in the large blocks. The extent of DCIS was significantly greater in all cases using large block techniques. Concurrent carcinomas were found more frequently in the LB series and these tumours were smaller than those in the SB series. Similar differences were noted with radial scars, and other proliferative lesions also had a higher incidence in the LB series. We conclude that the LB technique has sufficient advantages to recommend it as a standard technique in breast pathology.

Nucleotide sequence of transferrin cDNAs and tissue-specific of the transferrin gene in Atlantic cod (Gadus morhua).

Atlantic cod (Gadus morhua) transferrin cDNAs were isolated from a liver cDNA library using a cod transferrin-derived polymerase chain reaction product as a hybridization probe. The composite nucleotide sequence of two overlapping clones was 2223 bp in length excluding the poly(A) sequence and was equivalent to 87% of the 3' end of the Atlantic salmon transferrin cDNA sequence. Comparison of the deduced amino acid sequence of cod, salmon, Xenopus and several mammalian transferrins revealed that the two fish sequences are more similar with respect to their amino acid sequence and the position of additions/deletions than to other vertebrate transferrins. Conservation of the iron-binding domains and cysteine residues involved in disulphide bridges indicates that all transferrins share similar tertiary structure and support the hypothesis that extant vertebrate transferrin genes were derived from a gene duplication before the divergence of fish, frogs and mammals. Cod transferrin mRNA was detected in both brain and liver RNA and to a much lesser extent in RNA isolated from kidney and heart in contrast to salmon and several other vertebrates in which the transferrin gene is not expressed in brain.

Sulfated glycoconjugates enhance CD36-dependent adhesion of Plasmodium falciparum-infected erythrocytes to human microvascular endothelial cells.

A novel adhesive pathway that enhances the adhesion of Plasmodium falciparum-infected erythrocytes (IEs) to endothelial cells has been identified. The sulfated glycoconjugates heparin, fucoidan, dextran sulfate 5000, and dextran sulfate 500 000 caused a dramatic increase in adhesion of IEs to human dermal microvascular endothelial cells. The same sulfated glycoconjugates had little effect on IE adhesion to human umbilical vein endothelial cells, a CD36-negative cell line. The effect was abolished by a monoclonal antibody directed against CD36, suggesting that enhanced adhesion to endothelium is dependent on CD36. No effect was observed on adhesion to purified platelet CD36 cells immobilized on plastic. The same sulfated glycoconjugates enhanced adhesion of infected erythrocytes to COS cells transfected with CD36, and this was inhibited by the CD36 monoclonal antibody. These findings demonstrate a role for sulfated glycoconjugates in endothelial adherence that may be important in determining the location and magnitude of sequestration through endogenous carbohydrates. In addition, they highlight possible difficulties that may be encountered from the proposed use of sulfated glycoconjugates as antiadhesive agents in patients with severe malaria.

The value of PE-10, a monoclonal antibody against pulmonary surfactant, in distinguishing primary and metastatic lung tumours.

A new monoclonal antibody (PE-10) raised against components of pulmonary surfactant has been assessed for its ability to distinguish primary from secondary carcinomas in the lung. We applied this antibody to a series of 107 primary lung carcinomas, 40 adenocarcinomas of other sites, and 26 cases of adenocarcinoma metastatic to lung and pleura. Of the primary lung carcinomas, all the non-mucinous bronchiolo-alveolar carcinomas were positive whereas all the mucinous cases were negative; 60% of other types of adenocarcinoma were positive and 10% of large cell undifferentiated carcinomas, 20% of small cell carcinomas and 40% of atypical carcinoids also showed focal positivity. Squamous cell carcinomas were all negative. Adenocarcinomas of the breast, kidney, large bowel and ovaries were all negative, as were all 26 cases of adenocarcinoma metastatic to the lung and pleura. We conclude that this antibody is highly specific and moderately sensitive for primary tumours of the lung.

Biology of malarial liver stages: implications for vaccine design.

The molecular events controlling sporozoite invasion and exo-erythrocytic (EE) development within hepatocytes are largely not understood, and EE parasites are probably better defined immunologically than biologically. The observation that the Plasmodium falciparum sporozoite antigen TRAP (thrombospondin-related anonymous protein) contains multiple adhesive domains that recognize endothelial and hepatocyte receptors indicates that, like leucocyte passage across capillaries, sporozoite invasion probably involves a co-ordinated interaction between sporozoite and hepatic molecules. The parallel with leucocyte extravasation is strengthened by the finding that TRAP contains a functional, integrin-like, I domain. EE parasites are an important target of immunity elicited by irradiated sporozoites, and much current effort is focused on developing malaria vaccines targeting EE parasites. Only one EE-specific antigen, liver-stage antigen 1 (LSA-1), is known to be expressed during EE development and may contribute to protective immunity elicited by irradiated P. falciparum sporozoites. In a study in Papua New Guinea, resistance to P. falciparum infection correlated with CD8+ T-cell interferon-gamma responses to an LSA-1 epitope that contains an HLA A11-restricted sequence. Since A11 is > 40% frequent in this population it is reasonable to suggest that, as with B53 responses to LSA-1 in The Gambia, P. falciparum has driven genetic selection of certain HLA haplotypes, as proposed by Haldane nearly 50 years ago. LSA-1 is thus an important vaccine candidate, and is being expressed in bacterial and phage vectors.