Polarization of Macrophages, Cellular Adhesion, and Spreading on Bacterially Contaminated Gold Nanoparticle-Coatings in Vitro.

Biomaterial-associated infections often arise from contaminating bacteria adhering to an implant surface that are introduced during surgical implantation and not effectively eradicated by antibiotic treatment. Whether or not infection develops from contaminating bacteria depends on an interplay between bacteria contaminating the biomaterial surface and tissue cells trying to integrate the surface with the aid of immune cells. The biomaterial surface plays a crucial role in defining the outcome of this race for the surface. Tissue integration is considered the best protection of a biomaterial implant against infectious bacteria. This paper aims to determine whether and how macrophages aid osteoblasts and human mesenchymal stem cells to adhere and spread over gold nanoparticle (GNP)-coatings with different hydrophilicity and roughness in the absence or presence of contaminating, adhering bacteria. All GNP-coatings had identical chemical surface composition, and water contact angles decreased with increasing roughness. Upon increasing the roughness of the GNP-coatings, the presence of contaminating Staphylococcus epidermidis in biculture with cells gradually decreased surface coverage by adhering and spreading cells, as in the absence of staphylococci. More virulent Staphylococcus aureus fully impeded cellular adhesion and spreading on smooth gold- or GNP-coatings, while Escherichia coli allowed minor cellular interaction. Murine macrophages in monoculture tended toward their pro-inflammatory "fighting" M1-phenotype on all coatings to combat the biomaterial, but in bicultures with contaminating, adhering bacteria, macrophages demonstrated Ym1 expression, indicative of polarization toward their anti-inflammatory "fix-and-repair" M2-phenotype. Damage repair of cells by macrophages improved cellular interactions on intermediately hydrophilic/rough (water contact angle 30 deg/surface roughness 118 nm) GNP-coatings in the presence of contaminating, adhering Gram-positive staphylococci but provided little aid in the presence of Gram-negative E. coli. Thus, the merits on GNP-coatings to influence the race for the surface and prevent biomaterial-associated infection critically depend on their hydrophilicity/roughness and the bacterial strain involved in contaminating the biomaterial surface.

Gene disruption of Plasmodium falciparum p52 results in attenuation of malaria liver stage development in cultured primary human hepatocytes.

Difficulties with inducing sterile and long lasting protective immunity against malaria with subunit vaccines has renewed interest in vaccinations with attenuated Plasmodium parasites. Immunizations with sporozoites that are attenuated by radiation (RAS) can induce strong protective immunity both in humans and rodent models of malaria. Recently, in rodent parasites it has been shown that through the deletion of a single gene, sporozoites can also become attenuated in liver stage development and, importantly, immunization with these sporozoites results in immune responses identical to RAS. The promise of vaccination using these genetically attenuated sporozoites (GAS) depends on translating the results in rodent malaria models to human malaria. In this study, we perform the first essential step in this transition by disrupting, p52, in P. falciparum an ortholog of the rodent parasite gene, p36p, which we had previously shown can confer long lasting protective immunity in mice. These P. falciparum P52 deficient sporozoites demonstrate gliding motility, cell traversal and an invasion rate into primary human hepatocytes in vitro that is comparable to wild type sporozoites. However, inside the host hepatocyte development is arrested very soon after invasion. This study reveals, for the first time, that disrupting the equivalent gene in both P. falciparum and rodent malaria Plasmodium species generates parasites that become similarly arrested during liver stage development and these results pave the way for further development of GAS for human use.

Pfs47, paralog of the male fertility factor Pfs48/45, is a female specific surface protein in Plasmodium falciparum.

The genome of Plasmodium falciparum contains a small gene family that expresses proteins characterized by the presence of 6-cysteine domains. Most of these proteins are expressed on the surface of the parasite and some are known to play a role in cell-cell interactions. Two members of this family, Pfs48/45 and Pfs230, form a complex localized on the surface of gametes and are recognized as important targets for transmission-blocking vaccines. In this study we report the analysis of an additional member of this family, Pfs47 the closest paralog of Pfs48/45. We demonstrate that Pfs47 is expressed only in female gametocytes and is located on the surface of female gametes following emergence from red blood cells. In contrast to the critical function of P48/45 for male fertility, Pfs47 does not appear crucial for female fertility. Parasites lacking Pfs47 through targeted gene disruption, produce normal numbers of oocysts when included in the blood meal of the mosquito vector. In addition, three monoclonal antibodies against Pfs47 were unable to inhibit oocyst development when present in a blood meal containing wild type parasites. These results show redundancy in protein function for Pfs47 and reduce the support for candidacy of Pfs47 as a transmission-blocking vaccine target.