Identification of Protein Quality Markers in Toad Venom from Bufo gargarizans.

Toad venom is a traditional Chinese medicine with high medicinal value. The existing quality evaluation standards of toad venom have obvious limitations because of the lack of research on proteins. Thus, it is necessary to screen suitable quality markers and establish appropriate quality evaluation methods for toad venom proteins to guarantee their safety and efficacy in clinical applications. SDS-PAGE, HPLC, and cytotoxicity assays were used to analyze differences in protein components of toad venom from different areas. Functional proteins were screened as potential quality markers by proteomic and bioinformatic analyses. The protein components and small molecular components of toad venom were not correlated in content. Additionally, the protein component had strong cytotoxicity. Proteomics analysis showed that 13 antimicrobial proteins, four anti-inflammatory and analgesic proteins, and 20 antitumor proteins were differentially expressed extracellular proteins. A candidate list of functional proteins was coded as potential quality markers. Moreover, Lysozyme C-1, which has antimicrobial activity, and Neuropeptide B (NPB), which has anti-inflammatory and analgesic activity, were identified as potential quality markers for toad venom proteins. Quality markers can be used as the basis of quality studies of toad venom proteins and help to construct and improve safe, scientific, and comprehensive quality evaluation methods.

Effects of dietary inulin supplementation on the composition and dynamics of cecal microbiota and growth-related parameters in broiler chickens.

Inulin, a prebiotic, is an attractive alternative to antibiotic growth promoters in chickens. Dietary supplementation with inulin can improve growth performance, carcass yield, immune system activity, and serum biochemical parameters in chickens. A few studies investigated the impact of dietary inulin supplementation on chicken intestinal microbiota. In this study, we investigated how and why dietary supplementation with 1, 2, and 4% inulin can affect body weight gain, feed intake, food conversion rate, immunological parameters, serum biochemical parameters, and composition and dynamics of the cecal microbiota of Tegel broiler chickens using quantitative fluorescence in situ hybridization (qFISH). We showed that inulin inclusion has a negative effect on growth performance parameters before day 21 and a positive effect subsequently up to day 42. Quantitative FISH data revealed an age-dependent change in the cecal microbiota in the control broilers fed no inulin. Thus, relative abundances of Firmicutes and Actinobacteria decreased from 52.8 to 48.3% of total cells and from 8.7 to 1.4% at days 7 and 42, respectively. However, relative abundances of Bacteroidetes and Proteobacteria gradually increased from 9.3 to 26.9% of the total cells and from 10.7 to 21.1%, respectively, over the same periods. Inulin inclusion appeared to lower the relative abundances of Lactobacillus johnsonii and Bifidobacterium species at an early bird age, but it subsequently significantly (P < 0.05) increased their relative abundances. Such increases positively correlated with body weight gain of the birds, determined after day 21. Thus, dietary supplementation with inulin together with the addition of L. johnsonii and Bifidobacterium (B. gallinarum and B. pullorum) cultures at an early age may help overcome its early negative influence on growth performance. We believe that these findings can improve our knowledge on how inulin can change the intestinal microbiota of broiler chickens and help in developing an inulin feeding regime to optimize its beneficial role in chicken development.

Effects of dietary supplementation with lysozyme on the structure and function of the cecal microbiota in broiler chickens.

Lysozyme is known to eliminate intestinal pathogens in poultry and improve their growth performance. However, whether it can replace antibiotic growth promoters without the associated risk of the emergence of antibiotic-resistant bacterial strains is not known, and the effects of lysozyme supplementation on the composition, biodiversity, and function of the chicken gut microbiota remain unclear. Here, we used the 16S rRNA gene and ITS fragment Illumina sequencing combined with transcriptomic analysis to address this issue. A total of 400 1-d-old Di Gao chicks were allocated randomly to five groups, each consisting of four replicates (20 birds/group). The chicks were fed a starter (1-21 d) and a grower (22-42 d) diet supplemented with 0 (control), 40 (LYS40), 100 (LYS100), or 200 ppm (LYS200) lysozyme, or 400 ppm flavomycin as an antibiotic control for 6 weeks. Lysozyme administration did not contribute significantly (P > 0.05) to the growth of the broiler chickens. No significant (P > 0.05) differences in the diversity and composition of the bacterial and fungal communities in the cecal microbiota of chickens in the different diet groups were found. However, lysozyme supplementation led to a significant (P < 0.05) enrichment of genes involved in the synthesis/degradation of bacterial outer membranes and cell walls, cross-cell substrate transport, and carbohydrate metabolic processes, thus possibly promoting the cecal microbiota carbon and energy metabolism. Bacteroides contributed 31.9% of glycoside hydrolase genes (17,681-24,590), 26.1% of polysaccharide lyase genes (479-675), 20.7% of carbohydrate esterase genes (3,509-4,101), 8.8% of auxiliary activity genes (705-1,000), 16.2% of glycosyltransferase genes (5,301-6,844), and 13.9% of carbohydrate-binding module genes (8838-15,172) identified in the cecal samples. Thus, they were the main players in the breakdown of non-starch polysaccharides in the cecum, although Parabacteroides, Alistipes, Prevotella, Clostridium, Blastocystis, Barnesiella, Blautia, Faecalibacterium, Subdoligranulum, Megamonas, Eubacterium, Ruminococcus, Paenibacillus, Bifidobacterium, Akkermansia, and other bacteria also participated.

Identification of angiogenesis-inhibiting peptides from Chan Su.

Chan Su is a traditional medicine prepared from toxic secretions from the auricular and skin glands of Chinese toads. Previous studies show that active components in Chan Su can inhibit the proliferation of tumor cells. To study the effect of Chan Su peptides on angiogenesis, fresh Chan Su was collected and its component peptides were isolated by an extraction and precipitation method. A high-performance liquid chromatography (HPLC) fingerprint of the Chan Su component peptides revealed that there were more than 18 peptide component peaks. We demonstrate that Chan Su peptides inhibit angiogenesis in vitro by inhibiting human umbilical vein endothelial cell (HUVEC) proliferation and tube formation in a dose-dependent manner. Western blots indicated that Chan Su peptides inhibited the protein expression of VEGF165 and Ras, leading us to conclude that Chan Su peptide components exert anti-angiogenic effects by suppressing the VEGF165-VEGFR2-Ras signalling pathway. Finally, we identified the partial amino acid sequences of seven Chan Su peptides using the shotgun proteomics method.

Semiconducting polymer-based nanoparticles with strong absorbance in NIR-II window for in vivo photothermal therapy and photoacoustic imaging.

Near-infrared (NIR) light-induced photothermal therapy (PTT) has attracted much interest in recent years. In the NIR region, tissue penetration ability of the second biological near-infrared window (1000-1350 nm) is recognized to be stronger than that of the first window (650-950 nm). However, NIR light absorbers in the second NIR region (NIR-II) have been scant even though various NIR light absorbers in the first NIR region (NIR-I) have been widely explored. In this work, a thieno-isoindigo derivative-based semiconducting polymer, PBTPBF-BT, were formulated into PEGylated nanoparticles. The obtained nanoparticle NPPBTPBF-BT exhibited strong absorption in NIR-II region, inherent high photothermal conversion efficacy, and excellent photostability. The in vitro and in vivo PTT study employing 1064 nm laser in NIR-II window revealed that NPPBTPBF-BT could efficiently ablate tumor cell at a power density of 0.42 W/cm2 (the skin tolerance threshold value). Moreover, NPPBTPBF-BT with excellent photostability exhibited enhanced photoacoustic (PA) imaging of tumor in living mice, suggesting the great probability of using NPPBTPBF-BT for in vivo PA imaging-guided PTT in the NIR-II window.