Differential immunomodulating effects of pegylated liposomal doxorubicin nanoparticles on human macrophages.

PURPOSE: The pegylated liposomal doxorubicin (PLD) has been widely accepted in treatment of various cancers. However, the composition of two currently marketed PLD nanoparticles differs in structure and composition of lipids, and their differential effects remain unknown. Macrophages of the mononuclear phagocyte system are pivotal in determining PLD clearance in vivo. The aim of this study was to compare the effect of these two PLDs on drug uptake, cell viability, morphology and immune function of human macrophages. METHODS: Two PLD nanoparticles were used in this study. The major difference between PLD-D and PLD-H is that their phospholipid bilayers are composed of distearoyl phosphatidylcholine (DSPC) and hydrogenated soybean phosphatidylcholine (HSPC), respectively. Human CD14+ monocytes were isolated from peripheral blood to prepare macrophages. Comparative assays included: flow cytometry for detection of doxorubicin penetration into cells, MTT for cell viability, Trypan blue exclusion for cell membrane integrity, Liu's stain for morphologic evaluation, and inactivated yeast co-culture for phagocytosis. RESULTS: The uptake of PLD-H was rapidly detected at 10 min and kept increasing to 4 h followed by a decline thereafter, whereas that of PLD-D had similar profile with much less doxorubicin fluorescence detected, indicating a greater amount of doxorubicin retention of PLD-H. PLD-H, at higher concentration, decreased the viability and impaired cell membrane integrity of macrophages with an extent greater than PLD-D. The morphological observation showed a more extensive necrosis in PLD-H-treated macrophages. The phagocytosis function of macrophage was inhibited with a greater extent in PLD-H-treated macrophages. CONCLUSIONS: The PLD containing HSPC may cause retention of doxorubicin with greater amount and longer period in human macrophages than that containing DSPC. This effect was accompanied by greater toxicity and more profound dysfunction. The correlation of this differential effect to clinical outcome remains to be extensively investigated by performing in vivo experiments or conducting clinical trials.

Functional Analysis of the Collagen Binding Proteins of Streptococcus parasanguinis FW213.

Streptococcus parasanguinis is a dominant isolate of dental plaque and an opportunistic pathogen associated with subacute endocarditis. As the expression of collagen binding proteins (CBPs) could promote the establishment of S. parasanguinis in the host, the functions of three putative CBP-encoding loci, Spaf_0420, Spaf_1570, and Spaf_1573, were analyzed using isogenic mutant strains. It was revealed that S. parasanguinis FW213 bound effectively to fibronectin and type I collagen, but the strain's affinity for laminin and type IV collagen was quite low. By using various deletion derivatives, it was found that these three loci mediated the binding of S. parasanguinis to multiple extracellular matrix molecules, with type I collagen as the common substrate. Derivative strains with a deletion in any of the three loci expressed reduced binding to trypsin-treated swine heart valves. The deletion of these loci also reduced the viable count of S. parasanguinis bacteria within macrophages, especially the loss of Spaf_0420, but only strains with deletions in Spaf_0420 and Spaf_1570 expressed reduced virulence in the Galleria mellonella larva model. The deletion of Spaf_1570 and Spaf_1573 affected mainly the structure, but not the overall mass, of biofilm cultures in a flow cell system. Thus, CBPs are likely to be more critical for the initial colonization of S. parasanguinis on host tissues during the development of endocarditis.IMPORTANCE Bacteria generally can utilize multiple adhesins to establish themselves in the host. We found that Streptococcus parasanguinis, a dominant oral commensal and an opportunistic pathogen for subacute endocarditis, possesses at least three collagen-binding proteins that enable S. parasanguinis to successfully colonize damaged heart tissues and escape innate immune clearance. The binding specificities of these three proteins for extracellular matrix molecules differ, although all three proteins participate in biofilm formation by S. parasanguinis The "multiligand for multisubstrate" feature of these adhesins may explain the high adaptability of this microbe to different tissue sites.

Pegylated gold nanoparticles induce apoptosis in human chronic myeloid leukemia cells.

Gold nanoparticles (AuNPs) have several potential biological applications as well as excellent biocompatibility. AuNPs with surface modification using polyethylene glycol (PEG-AuNPs) can facilitate easy conjugation with various biological molecules of interest. To examine the anticancer bioactivity of PEG-AuNPs, we investigated their effect on human chronic myeloid leukemia K562 cells. The results indicated that PEG-AuNPs markedly inhibited the viability and impaired the cell membrane integrity of K562 cells. The particles caused morphological changes typical of cell death, and a marked increase in the sub-G1 population in DNA histogram, indicating apoptosis. In addition, PEG-AuNPs reduced the mitochondrial transmembrane potential, a hallmark of the involvement of intrinsic apoptotic pathway in K562 cells. Observation of ultrastructure under a transmission electron microscope revealed that the internalized PEG-AuNPs were distributed into cytoplasmic vacuoles and damaged mitochondria, and subsequently accumulated in areas surrounding the nuclear membrane. In conclusion, PEG-AuNPs may have the potential to inhibit growth and induce apoptosis in human chronic myeloid leukemia cells.

Complete genome and transcriptomes of Streptococcus parasanguinis FW213: phylogenic relations and potential virulence mechanisms.

Streptococcus parasanguinis, a primary colonizer of the tooth surface, is also an opportunistic pathogen for subacute endocarditis. The complete genome of strain FW213 was determined using the traditional shotgun sequencing approach and further refined by the transcriptomes of cells in early exponential and early stationary growth phases in this study. The transcriptomes also discovered 10 transcripts encoding known hypothetical proteins, one pseudogene, five transcripts matched to the Rfam and additional 87 putative small RNAs within the intergenic regions defined by the GLIMMER analysis. The genome contains five acquired genomic islands (GIs) encoding proteins which potentially contribute to the overall pathogenic capacity and fitness of this microbe. The differential expression of the GIs and various open reading frames outside the GIs at the two growth phases suggested that FW213 possess a range of mechanisms to avoid host immune clearance, to colonize host tissues, to survive within oral biofilms and to overcome various environmental insults. Furthermore, the comparative genome analysis of five S. parasanguinis strains indicates that albeit S. parasanguinis strains are highly conserved, variations in the genome content exist. These variations may reflect differences in pathogenic potential between the strains.