Diabetic wounds, a prevalent chronic disease, are associated with older age. The hyperglycemic microenvironment in diabetic wounds significantly reduces the immune system, inducing bacterial invasion. The coupling of tissue repair and antibacterial treatment is critical for infected diabetic ulcer regeneration. In this study, a dual-layered sodium alginate/carboxymethyl chitosan (SA/CMCS) adhesive film cored with an SA-bFGF microsphere-loaded small intestine submucosa (SIS) hydrogel composite dressing with a graphene oxide (GO)-based antisense transformation system was developed to promote infected diabetic wound healing and bacterial eradication. Initially, our injectable SIS-based hydrogel composite stimulated angiogenesis, collagen deposition, and immunoregulation in diabetic wound repair. The GO-based transformation system subsequently inhibited bacterial viability in infected wounds by post-transformation regulation. Meanwhile, the SA/CMCS film provided stable adhesion covering the wound area to maintain a moist microenvironment, which promoted in situ tissue repair. Our findings provide a promising clinical translation strategy for promoting the healing of infected diabetic wounds.

Gene silencing is an important biological strategy for studying gene functions, exploring disease mechanisms and developing therapeutics. 8-17 DNAzyme is of great potential for gene silencing, due to its higher RNA-cleaving activity. However, it is not generally used in practice, due to its divalent cation dependence and poor understanding of its cellular mechanisms. To address these issues, we have explored its activity in vitro and in cells and found that it can cleave RNA substrates under the simulated physiological conditions, and its gene-silencing activity is additionally enhanced by its RNase H compatibility, offering both cleavage and antisense activities in cells. Further, chemical modifications can facilitate its stability, substrate binding affinity and gene-silencing activity. Our research results suggest that this DNAzyme can demonstrate high levels of activities for both actions in cells, making it a useful tool for exploring biomedical applications.

UNASSIGNED: Staphylococcus aureus (S. aureus) is the leading cause of various infective diseases, including topical soft tissue infections. The goals of this study were to investigate the roles of YycF and CodY in the regulation of biofilm formation and pathogenicity.
UNASSIGNED: Electrophoretic mobility shift assay (EMSA) was conducted to validate the bound promoter regions of YycF protein. We constructed the codY up-regulated or down-regulated S. aureus mutants. The biofilm biomass was determined by crystal violet microtiter assay and scanning electron microscopy (SEM). Quantitative RT-PCR analysis was used to detect the transcripts of biofilm-related genes. The live and dead cells of S. aureus biofilm were also investigated by confocal laser scanning microscopy (CLSM). We constructed an abscess infection in Sprague Dawley (SD) rat models to determine the effect of CodY on bacterial pathogenicity. We further used the RAW264.7, which were cocultured with S. aureus, to evaluate the effect of CodY on macrophages apoptosis.
UNASSIGNED: Quantitative RT-PCR analyses reveled that YycF negatively regulates codY expression. EMSA assays indicated that YycF protein directly binds to the promoter regions of codY gene. Quantitative RT-PCR confirmed the construction of dual- mutant stains codY + ASyycF and codY-ASyycF. The SEM results showed that the biofilm formation in the codY + ASyycF group was sparser than those in the other groups. The crystal violet assays indicated that the codY + ASyycF group formed less biofilms, which was consistent with the immunofluorescence results of the lowest live cell ration in the codY + ASyycF group. The expression levels of biofilm-associated icaA gene were significantly reduced in the codY + strain, indicating codY negatively regulates the biofilm formation. Furthermore, CodY impedes the pathogenicity in a rat-infection model. After cocultured with bacteria or 4-h in vitro, the apoptosis rates of macrophage cells were lowest in the codY + group.
UNASSIGNED: YycF negatively regulate the expression of codY. By interaction with codY, YycF could modulate S. aureus biofilm formation via both eDNA- dependent and PIA- dependent pathways, which can be a significant target for antibiofilm. CodY not only impedes the pathogenicity but also has a role on immunoregulation. Thus, the current evidence may provide a supplementary strategy for managing biofilm infections.

Staphylococcus aureus (S. aureus) is an opportunistic bacterium that causes several infections in humans. However, chronic biofilms remain a major challenge associated with recalcitrance toward traditional treatments. Herein, an antibacterial hydrogel composed of antisense DNA oligonucleotides, graphene oxide and alginate is construed for biofilm management and infection care. The hydrogel is established through noncovalent binding and possesses injectability and degradability properties. Furthermore, hydrogels present controllable release of cargoes, genetic targeting antibacterial effects and stem cell supporting capabilities. Our in vivo results reveal a high antibiofilm performance and good biocompatibility, which significantly improve tissue regeneration. The hydrogel inhibits biofilm formation by decreasing the expression of YycFG with antisense and viability of strains by graphene oxide. Thus, antisense hydrogels can be a promising antibacterial bioactive material for potential therapeutic S. aureus infection.

Immune cell invasion after the transplantation of solid organs is directed by chemokines binding to glycosaminoglycans (GAGs), creating gradients that guide immune cell infiltration. Renal transplant is the preferred treatment for end stage renal failure, but organ supply is limited and allografts are often injured during transport, surgery or by cytokine storm in deceased donors. While treatment for adaptive immune responses during rejection is excellent, treatment for early inflammatory damage is less effective. Viruses have developed highly active chemokine inhibitors as a means to evade host responses. The myxoma virus-derived M-T7 protein blocks chemokine: GAG binding. We have investigated M-T7 and also antisense (ASO) as pre-treatments to modify chemokine: GAG interactions to reduce donor organ damage. Immediate pre-treatment of donor kidneys with M-T7 to block chemokine: GAG binding significantly reduced the inflammation and scarring in subcapsular and subcutaneous allografts. Antisense to N-deacetylase N-sulfotransferase1 (ASONdst1) that modifies heparan sulfate, was less effective with immediate pre-treatment, but reduced scarring and C4d staining with donor pre-treatment for 7 days before transplantation. Grafts with conditional Ndst1 deficiency had reduced inflammation. Local inhibition of chemokine: GAG binding in donor organs immediately prior to transplant provides a new approach to reduce transplant damage and graft loss.

BACKGROUND: Outbreaks of infection due to multidrug-resistant (MDR) bacteria, especially Gram-negative bacteria, have become a global health issue in both hospitals and communities. Antisense oligonucleotides (ASOs) based therapeutics hold a great promise for treating infections caused by MDR bacteria. However, ASOs therapeutics are strangled because of its low cell penetration efficiency caused by the high molecular weight and hydrophilicity.
RESULTS: Here, we designed a series of dendritic poly-peptides (DPP1 to DPP12) to encapsulate ASOs to form DSPE-mPEG2000 decorated ASOs/DPP nanoparticles (DP-AD1 to DP-AD12) and observed that amphipathic DP-AD2, 3, 7 or 8 with a positive charge ≥ 8 showed great efficiency to deliver ASOs into bacteria, but only the two histidine residues contained DP-AD7 and DP-AD8 significantly inhibited the bacterial growth and the targeted gene expression of tested bacteria in vitro. DP-AD7anti-acpP remarkably increased the survival rate of septic mice infected by ESBLs-E. coli, exhibiting strong antibacterial effects in vivo.
CONCLUSIONS: For the first time, we designed DPP as a potent carrier to deliver ASOs for combating MDR bacteria and demonstrated the essential features, namely, amphipathicity, 8-10 positive charges, and 2 histidine residues, that are required for efficient DPP based delivery, and provide a novel approach for the development and research of the antisense antibacterial strategy.

Circular RNAs (circRNAs) represent a large group of non-coding RNAs that are widely detected in mammalian cells. Although most circRNAs are generated in a sense orientation, there is a group of circRNAs that are synthesized in an antisense orientation. High-throughput analysis of breast cancer specimens revealed a significant enrichment of 209 antisense circRNAs. The tumor suppressor SCRIB was shown to potentially produce thirteen circRNAs, three of which are in an antisense orientation. Among these three circRNAs, circSCRIB (hsa_circ_0001831) was the most enriched in the breast cancer panel. This antisense SCRIB circRNA was shown to span one intron and two exons. We hypothesized that this circRNA could decrease pre-mRNA splicing and mRNA translation. To test this, we generated a hsa_circ_0001831 expression construct. We found that there was decreased SCRIB mRNA production but increased cancer cell proliferation, migration, and invasion. In comparison, an exonic sequence construct did not affect mRNA splicing but decreased protein translation, leading to increased E-cadherin expression and decreased expression of N-cadherin and vimentin. Thus, there was increased cell migration, invasion, proliferation, colony formation, and tumorigenesis. Our study suggests a novel modulatory role of antisense circRNAs on their parental transcripts. This may represent a promising approach for developing circRNA-directed therapy.

Staphylococcus aureus (S. aureus) is one of most common opportunistic pathogens and is attributed to several human infections. The increasing incidence of methicillin-resistant S. aureus (MRSA) is a serious clinical threat for osteomyelitis crisis. The YycFG two-component system of S. aureus regulates genes associated with biofilm formation. To investigate the potential role of an antisense yycF RNA in the regulation of transcription levels of yycF and associated effects on biofilm formation and pathogenicity, antisense yycF (ASyycF) RNA was detected by RT-PCR and 5\' RACE assays. ASyycF overexpression mutants were constructed, and the biofilm biomass was determined by crystal violet microtiter assay and scanning electron microscopy (SEM). Quantitative RT-PCR and Western blotting analyses were used to detect whether ASyycF overexpression inhibited the transcription and translation of biofilm-related genes. Then, a rat tibial infective model was used to evaluate the pathogenicity of ASyycF overexpression in vivo. ASyycF transcription led to reductions in YycF production and biofilm formation. Overexpression of ASyycF inhibited the transcription and translation of biofilm-related genes. The sensitivity to vancomycin was improved in ASyycF-overexpressing MRSA. Furthermore, ASyycF inhibited MRSA invasion in a rat tibial infection model. From this study, the expression of the YycF protein was found to be inversely correlated with different levels of ASyycF transcription. The biofilm biomass and pathogenicity decreased in the ASyycF-overexpressing mutant. Thus, the current evidence may support ASyycF as a supplementary strategy for managing S. aureus and MRSA infections.

The infectious pathogen Staphylococcus aureus (S. aureus) is primarily associated with osteomyelitis. Hydrogen peroxide drainage is an effective antimicrobial treatment that has been adopted to combat S. aureus infections. Previous investigations have indicated that the antisense RNA (asRNA) strategy negatively modulates S. aureus YycFG TCS, and it significantly disrupts biofilm formation. However, the effects of the antisense yycG RNA (ASyycG) strategy on the susceptibility of biofilm-producing S. aureus to hydrogen peroxide and the mechanisms underlying this effect have not been elucidated to date.
Overexpression of ASyycG inhibited the transcription of biofilm formation-related genes, including sarA and icaA. Additionally, the CFU counts and the live bacterial ratios of ASyycG biofilm-producing S. aureus treated with H2O2 were notably reduced across the groups. Notably, the predicted promoter regions of the sarA and icaA genes were directly regulated by YycF.
ASyycG was observed to sensitize biofilm-producing S. aureus to H2O2 intervention synergistically via the sarA and thus may represent a supplementary strategy for managing osteomyelitis. However, future in-depth studies should attempt to replicate our findings in animal models, such as the rat osteomyelitis model.

Anti-sense transcription is increasingly being recognized as an important regulator of gene expression. But the transcriptome complementation of anti-sense RNA in hybrid relative to their inbred parents was largely unknown. In this study, we profiled strand-specific RNA sequencing (RNA-seq) in a maize hybrid and its inbred parents (B73 and Mo17) in two tissues. More anti-sense transcripts were present in the hybrid compared with the parental lines. We detected 293 and 242 single-parent expression of anti-sense (SPEA) transcripts in maize immature ear and leaf tissues, respectively. There was little overlap of the SPEA transcripts between the two maize tissues. These results suggested that SPEA is a general mechanism that drives extensive complementation in maize hybrids. More importantly, extremely high-level expression of anti-sense transcripts was associated with low-level expression of the cognate sense transcript by reducing the level of histone H3 lysine 36 methylation (H3K36me3). In summary, these SPEA transcripts increased our knowledge about the transcriptomic complementation in hybrid.