Transcriptome profiling reveals CD73 and age-driven changes in neutrophil responses against Streptococcus pneumoniae

Neutrophils are required for host resistance against Streptococcus pneumoniae but their function declines with age. We previously found that CD73, an enzyme required for antimicrobial activity, is down-regulated in neutrophils from aged mice. This study explored transcriptional changes in neutrophils induced by S. pneumoniae to identify pathways controlled by CD73 and dysregulated with age. Pure bone marrow-derived neutrophils isolated from wild type (WT) young, old, and CD73KO young mice were mock-challenged or infected with S. pneumoniae ex vivo . RNA sequencing was performed to identify differentially expressed genes (DEGs). We found that infection triggered distinct global transcriptional changes across hosts, that were strongest in CD73KO neutrophils. Surprisingly, there were more down-regulated than up-regulated genes in all groups upon infection. Down-regulated DEGs indicated a dampening of immune responses in old and CD73KO hosts. Further analysis revealed that CD73KO neutrophils expressed higher numbers of long non-coding RNAs (lncRNAs) compared to WT controls. Predicted network analysis indicated that CD73KO specific lncRNAs control several signaling pathways. We found that genes in the JNK-MAPK-pathway were up-regulated upon infection in CD73KO and WT old but not in young mice. This corresponded to functional differences, as phosphorylation of the downstream AP-1 transcription factor component c-Jun was significantly higher in infected CD73KO and old mice neutrophils. Importantly, inhibiting JNK/AP-1 rescued the ability of these neutrophils to kill S. pneumoniae . Altogether, our findings revealed that the ability of neutrophils to modify their gene expression to better adapt to bacterial infection is in part regulated by CD73 and declines with age.

The DNA-to-cytoplasm ratio broadly activates zygotic gene expression in Xenopus

In multicellular animals, the first major event after fertilization is the switch from maternal to zygotic control of development. During this transition, zygotic gene transcription is broadly activated in an otherwise quiescent genome in a process known as zygotic genome activation (ZGA). In fast developing embryos, ZGA often overlaps with the slowing of initially synchronous cell divisions at the mid-blastula transition (MBT). Initial studies of the MBT led to the nuclear-to-cytoplasmic ratio model where MBT timing is regulated by the exponentially increasing amounts of some nuclear component N titrated against a fixed cytoplasmic component C. However, more recent experiments have been interpreted to suggest that ZGA is independent of the N/C ratio. To determine the role of the N/C ratio in ZGA, we generated Xenopus frog embryos with ~3-fold differences in genomic DNA (i.e., N) by using X. tropicalis sperm to fertilize X. laevis eggs with or without their maternal genome. Resulting embryos have otherwise identical X. tropicalis genome template amounts, embryo sizes, and X. laevis maternal environments. We used the X. tropicalis paternally derived mRNA to identify a high confidence set of exclusively zygotic transcripts. Both ZGA and the increase in cell cycle duration are delayed in embryos with ~3-fold less DNA per cell. Thus, DNA is an important component of the N/C ratio, which is indeed a critical regulator of zygotic genome activation in Xenopus embryos.

Transcriptome profiling reveals CD73 and age-driven changes in neutrophil responses against Streptococcus pneumoniae

Neutrophils are required for host resistance against Streptococcus pneumoniae but their function declines with age. We previously found that CD73, an enzyme required for antimicrobial activity, is down-regulated in neutrophils from aged mice. This study explored transcriptional changes in neutrophils induced by S. pneumoniae to identify pathways controlled by CD73 and dysregulated with age. Ultrapure bone marrow-derived neutrophils isolated from wild type (WT) young, old, and CD73KO young mice were mock-challenged or infected with S. pneumoniae ex vivo. RNA sequencing was performed to identify differentially expressed genes (DEGs). We found that infection triggered distinct global transcriptional changes across hosts, that were strongest in CD73KO neutrophils. Surprisingly, there were more down-regulated than up-regulated genes in all groups upon infection. Down-regulated DEGs indicated a dampening of immune responses in old and CD73KO hosts. Further analysis revealed that CD73KO neutrophils expressed higher numbers of long non-coding RNAs (lncRNAs) compared to WT controls. Predicted network analysis indicated that CD73KO specific lncRNAs control several signaling pathways. We found that genes in the JNK-MAPK-pathway were up-regulated upon infection in CD73KO and WT old but not in young mice. This corresponded to functional differences, as phosphorylation of the downstream AP-1 transcription factor component c-Jun was significantly higher in infected CD73KO and old mice neutrophils. Importantly, inhibiting JNK/AP-1 rescued the ability of these neutrophils to kill S. pneumoniae. Altogether, our findings revealed that neutrophils modify their gene expression to better adapt to bacterial infection and that this capacity declines with age and is regulated by CD73.

Evaluation of the Antibacterial Activities of Isolated Bioactive Components from the plant Adenodolichos paniculatus

The three bioactive components isolated included: component A (major phytochemicals were Bis (2-ethylhexyl) phthalate (16.36 %), 9,12-Octadecadienoic acid, ethyl ether (14.77 %) and 9.cis., 11.trans.-octadecadie noate (14.77 %), component B (major phytochemicals were 9,12-Octadecadienal (Linoleic acid) (40.98 %), Octadecanoic acid (Stearic acid) (9.26 %), Undecanoic acid, 10-bromo- (10-bromoudecanoic acid) (9.26 %) and n-Hexadecanoic acid (Palmitic acid) and component C (cis-9-octadecenoic acid (Oleic Acid) (30.45 %), Octadecanoic acid (Stearic acid) (17.33 %)). These components isolated from the chloroform fraction of Adenodolichos paniculatus are used by traditional medicinal practitioners for the management of mouth and throat infections. The antibacterial activities against Streptococcus pyogenes, Staphylococcus aureus and Pseudomonas aeruginosa were evaluated using bioautography and agar-well diffusion methods. The bioautogram result showed that component A had inhibited spots against S. pyogenes (17.50 mm) and P. aeruginosa (16.00 mm), corresponding to the TLC spots with Rf values of 0.594, 0.55 and 0.26, respectively. Component B showed inhibition spots against Streptococcus pyogenes (36.50 mm), Staphylococcus aureus (16.00 mm) and Pseudomonas aeruginosa (11.00 mm), corresponding to the TLC spots with Rf values 0.891, 0.87, 0.85 and 0.25, respectively. Component C showed inhibition spots against Streptococcus pyogenes (16.50 mm), Staphylococcus aureus (15.00 mm) and Pseudomonas aeruginosa (10.50 mm), corresponding to the TLC spots Rf values of 0.938, 0.44, 0.21 and 0.90, respectively. For the agar-well diffusion method, component A at 1 mg/ml inhibited Streptococcus pyogenes, Staphylococcus aureus and Pseudomonas aeruginosa growths with zones of inhibition 23.0, 19.5 and 17.50 mm, respectively. MIC and MBC of component A were 125, 250 and 250 and 250, 500 and 500 μg/ml, respectively. Component B at 1 mg/ml inhibited Streptococcus pyogenes, Staphylococcus aureus and Pseudomonas aeruginosa growth with zones of inhibition 30.0, 28.0 and 18.5 mm, respectively. MIC and MBC of the compound B were 31, 62 and 125 and 62, 125 and 250 μg/ml, respectively. Component C at 1 mg/ml inhibited Streptococcus pyogenes, Staphylococcus aureus and Pseudomonas aeruginosa growth with zones of inhibition 24.5, 20.5 and 17.0 mm, respectively. MIC and MBC of the component C were 62, 125 and 250 and 125, 250 and 500 μg/ml, respectively. This study confirmed that bioactive compounds of A. paniculatus root have antibacterial properties and support the use of this part of the plant as a traditional remedy for mouth and throat infections possibly caused by the test bacteria.