MOESM1 of Chiral polymer modified nanoparticles selectively induce autophagy of cancer cells for tumor ablation

Additional file 1: Figure S1. Stability in different solutions as indicated of l-PAV-AuNPs and d-PAV-AuNPs. (a) Photos and (b) UV–Vis-NIR of l-PAV-AuNPs or d-PAV-AuNPs in different solutions including saline, PBS, cell medium, fetal bovine serum and dilution of whole blood of the mice for 3 days. Figure S2. The toxicity study of l/d-PAV-AuNPs. (a) Dose- and chirality-dependent cytotoxicity of l/d-PAV-AuNPs in MDA-MB-231 cells, 3T3 fibroblasts and HBL-100 cells respectively. (b) Apoptosis rates of the MDA-MB-231 cells, 3T3 fibroblasts and HBL-100 cells treated with PAV-AuNPs, respectively. FCM analysis was tested via Annexin V-FITC and PI as probes. (c) Expression levels of LC3 in MDA-MB-231 cells, 3T3 fibroblasts and HBL-100 cells with PAV-AuNPs treatment, separately. GAPDH was used as a loading control. Figure S3. Biodistribution of PAV-AuNPs in vivo. The in vivo biodistribution of PAV-AuNPs was analyzed by testing the Au content in main organs (liver, kidneys, spleen, heart, and lung) of mice at 1 and 30 days post intravenous injection, separately. * and ** present p < 0.05 and p < 0.01, respectively

Glutamine sustains energy metabolism and alleviates liver injury in burn sepsis by promoting the assembly of mitochondrial HSP60-HSP10 complex via SIRT4 dependent protein deacetylation

Burns and burn sepsis, characterized by persistent and profound hypercatabolism, cause energy metabolism dysfunction that worsens organ injury and systemic disorders. Glutamine (Gln) is a key nutrient that remarkably replenishes energy metabolism in burn and sepsis patients, but its exact roles beyond substrate supply is unclear. In this study, we demonstrated that Gln alleviated liver injury by sustaining energy supply and restoring redox balance. Meanwhile, Gln also rescued the dysfunctional mitochondrial electron transport chain (ETC) complexes, improved ATP production, reduced oxidative stress, and protected hepatocytes from burn sepsis injury. Mechanistically, we revealed that Gln could activate SIRT4 by upregulating its protein synthesis and increasing the level of Nicotinamide adenine dinucleotide (NAD+), a co-enzyme that sustains the activity of SIRT4. This, in turn, reduced the acetylation of shock protein (HSP) 60 to facilitate the assembly of the HSP60-HSP10 complex, which maintains the activity of ETC complex II and III and thus sustain ATP generation and reduce reactive oxygen species release. Overall, our study uncovers a previously unknown pharmacological mechanism involving the regulation of HSP60-HSP10 assembly by which Gln recovers mitochondrial complex activity, sustains cellular energy metabolism and exerts a hepato-protective role in burn sepsis.</p

Intracellular bacteriolysis contributes to pathogenicity of <i>Staphylococcus aureus</i> by exacerbating AIM2-mediated inflammation and necroptosis

Staphylococcus aureus can survive within phagocytes. Indeed, we confirm in this study that approximately 10% of population persists in macrophages during S. aureus infection, while the rest are eliminated due to bacteriolysis, which is of particular interest to us. Herein, we observe that the bacteriolysis is an early event accompanied by macrophage death during S. aureus infection. Furthermore, the cell death is significantly accelerated following increased intracellular bacteriolysis, indicating that intracellular bacteriolysis induces cell death. Subsequently, we establish that the cell death is not apoptosis or pyroptosis, but AIM2-mediated necroptosis, accompanied by AIM2 inflammasome activation. This finding challenges the classical model that the cell death that accompanies inflammasome activation is always pyroptosis. In addition, we observe that the apoptosis-associated genes are highly inhibited during S. aureus infection. Finally, we establish in vivo that increased bacteriolysis significantly enhances S. aureus pathogenicity by promoting its dissemination to kidney and leading to an inflammatory cytokine storm in AIM2-mediated manner. Collectively, our data demonstrate that bacteriolysis is detrimental when triggered in excess and its side effect is mediated by AIM2. Meanwhile, we propose a potential immune manipulation strategy by which S. aureus sacrifices the minority to trigger a limited necroptosis, thereby releasing signals from dead cells to inhibit apoptosis and other anti-inflammatory cascades of live cells, eventually surviving within host cells and establishing infection.</p

Additional file 1: Figure S1. of NLRP3 inflammasome activation in murine macrophages caused by Neospora caninum infection

Effect of inhibitors on N. caninum replication in Vero cells. To evaluate whether the inhibitors used in this study could influence N. caninum replication, the inhibitors zVAD-fmk (z), Ac-YVAD-CHO (y) and glyburide (g) were added to Vero cells for 45 min, and the Vero cells were then infected with N. caninum tachyzoites (MOI = 3:1; parasite:cell) for 12 h. Total DNA from the infected cells was extracted and was used for a quantitative analysis by qPCR described in this study. The results showed that only glyburide could slightly inhibit N. caninum replication (P = 0.0393), while zVAD-fmk and Ac-YVAD-CHO could not influence parasite replication compared with the Nc group. Meanwhile, parasitophorous vacuoles can be observed in each group (data not shown), indicating that these inhibitors did not influence the viability of N. caninum. (*P <0.05; **P < 0.01; ***P <0.001 vs the Nc group). (TIF 4400 kb

Morphology of phages as revealed by TEM.

<p>Three phages displayed different morphology. GH-K2 displayed a short and contractile tail. The tails of GH-K1 and GH-K3 were long, and the tail of GH-K3 appeared more flexible than that of GH-K1. The bars represent 200 nm.</p

Delayed treatment with phage cocktail.

<p>Mice were inoculated intraperitoneally with K7 at the dose of 2.5×10⁸ cfu. Cocktail phages at the dose of 3.0×10⁴ pfu or a buffer were administered into the peritoneal cavities of mice at the indicated time intervals after challenging with K7. Phage cocktail was given at 1 h (black squares), 2 h (black diamond), or 3 h (black triangles) after the K7 challenge. Infected mice treated with buffer (white squares) under the same conditions were used as control. Each symbol represents the average of three experiments.</p

Survival rate of bacteremic mice treated with different doses of phages.

<p>Mice were inoculated intraperitoneally with K7 at the dose of 2.5×10⁸ cfu. Thirty minutes later, different doses of GH-K1 (○), GH-K2 (□), GH-K3 (Δ), or phage cocktail (◊) were injected into the peritoneal cavity of the mice. Every group contained five mice and each symbol represents the average of three experiments. The error bars indicate SD.</p

Electron micrographs of <i>K. pneumoniae</i> strains.

<p>A, B, and C show K7, and D shows K7R¹. B and C are higher magnification views of A. C and D are at the same magnification. The images show that K7 adsorbed to the thick jelly and formed large bacterial aggregates.</p

Comparison of the mice states at 12 h after treated with different single phage and the phage cocktail.

<p>Four groups of five mice were treated with different single phage and phage cocktail at the MPD, while the control group was treated with PBS. The mice were scored for their state of health on a scale of 5 to 0, based on progressive disease states. A score of 5 indicates normal health and unremarkable condition. Slight illness, defined as decreased physical activity and ruffled fur, was scored as 4. Moderate illness, defined as lethargy and a hunched back, was scored as 3. Severe illness, with the aforementioned signs, plus exudative accumulation around partially closed eyes, was scored as 2. A moribund state was scored as 1. Death was scored as 0. Each bar indicates the state of health of a single BALB/c mouse.</p

Colony counts of bacteria and titers of phages in blood samples obtained at regular intervals.

<p>The mice were challenged with K7 at the dose of 2.5×10⁸ cfu. At the indicated times, bacterial counts (•) and phage titers (▪) in three mice treated with either the MPD of (A)GH-K1, (B)GH-K2, (C)GH-K3, or (D) the phage cocktail were determined from peripheral blood samples taken from the caudal vein. The arrow indicates the moment at which the phage was injected (30 min after challenge). Each symbol represents the average of three experiments.</p