RNA quality is of utmost importance to perform gene expression quantification by qPCR. The classical methods used to determine RNA quality are based on electrophoresis and spectrophotometer assessment, namely A(260)/A(280) and A(260)/A(230) ratios. It was previously shown that due to the complex nature of Staphylococcus epidermidis biofilms, RNA extraction procedures could impact mRNA quality and thus accurate quantification. Herein, we contaminated and degraded RNA extracted from S. epidermidis biofilms, and assessed the effect on gene expression by qPCR. As expected, thermal degradation of RNA had a significant impact on gene expression on two out of the three tested genes. On the other hand, the contamination of the extracted RNA yielded an interesting result: while most contaminants did not changed the purity indicators or the integrity of RNA, significant changes on gene expression levels were found. This work confirms that poor RNA extraction has an important impact in qPCR quantification, emphasizing the consequences of carry-over contaminants on gene expression studies. Additionally, our results show that the parameters commonly used to assess the quality of extracted RNA from bacterial cultures seem to be insufficient to ensure reliable gene expression determination.
Implant integration is a complex process mediated by the interaction of the implant surface with the surrounding ions, proteins, bacteria, and tissue cells. Although most implants achieve long-term bone-tissue integration, preventing pervasive implant-centered infections demands further advances, particularly in surfaces design. In this work, we analyzed classical microrough implant surfaces (only acid etched, AE; sandblasted then acid etching, SB 1 AE) and a new calcium-ion-modified implant surface (AE 1 Ca) in terms of soft-and hard-tissue integration, bacterial adhesion, and biofilm formation. We cultured on the surfaces primary oral cells from gingiva and alveolar bone, and three representative bacterial strains of the oral cavity, emulating oral conditions of natural saliva and blood plasma. With respect to gingiva and bone cells and in the presence of platelets and plasma proteins, AE 1 Ca surfaces yielded in average 86% higher adhesion, 44% more proliferation, and triggered 246% more synthesis of extracellular matrix biomolecules than AE-unmodified controls. Concomitantly, AE 1 Ca surfaces regardless of conditioning with saliva and/or blood plasma showed significantly less bacterial adhesion (67% reduction in average) and biofilm formation (40% reduction in average) than unmodified surfaces. These results highlight the importance of a calcium-rich hydrated interface to favor mammalian cell functions over microbial colonization at implant surfaces.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.