Green synthesis and characterization of gold nanoparticles and their application for the rapid detection of glycyrrhizin with immunochromatographic strips.

In this study, a facile and environmentally friendly synthesis process was proposed without regular chemical additives. We successfully synthesized spherical gold nanoparticles (AuNPs) coated with glycyrrhizin (GL) by using GL as both a reductant and a stabilizer to reduce chloroauric acid. The obtained NPs were approximately 35 nm in size. The formation of these GL-AuNPs was verified by the presence of a surface plasmon resonance band at 526 nm. We also experimentally determined that in terms of chemical structure, GL can be used as a reducing agent to obtain colloidal gold. The d-glucuronic acid structure, rather than glycyrrhetinic acid (GA), plays an important reducing role in colloidal gold production. From this, we hypothesized that other compounds with sugar structures in Glycyrrhiza may also have the ability to reduce chloroauric acid. To mitigate the high cost and low efficiency of current GL detection methods, we applied AuNPs to the immunochromatographic system. Then, a colloidal gold immunochromatographic test strip based on the indirect competition method was developed for the rapid detection of GL, and the detection limit of this strip was 25 ng mL-1. The cross-test showed that the strip has high specificity. The test results are consistent with the data obtained by high-performance liquid chromatography (HPLC) with a coincidence rate of up to 100%. The rapid test strip is simple, fast, highly efficient and inexpensive, making it suitable for large-scale, rapid glycyrrhizin content determination.

Effect of dietary oridonin supplementation on growth performance, gut health, and immune response of broilers infected with Salmonella pullorum.

BACKGROUND: The effects of dietary supplementation of oridonin (ORI) on growth performance, cecal microbiota, epithelium development and antioxidant and immune parameters of broilers infected with S. pullorum were studied. A total of 300 1-d-old male chicks were selected and divided into 5 trial groups (6 replicates of 10 chickens): 1) nonchallenge control chicks (CON), 2) chicks treated with Salmonella Challenged Control (SCC), 3) chicks treated with S. pullorum challenge and 50 mg/kg ORI (O1), 4) chicks treated with S. pullorum challenge and 80 mg/kg ORI (O2), and 5) chicks treated with S. pullorum challenge and 100 mg/kg ORI (O3). RESULTS: The results showed that S. pullorum had no effect on the feed intake (FI), body weight gain (BWG) or feed conversion ratio (FCR) of broilers compared with the values measured for the CON group (P > 0.05). However, compared with the characteristics of CON, S. pullorum showed effects on the counts of Salmonella and Lactobacillus at 7 d and at 14 d (P < 0.05), on jejunal development at 7 d (P < 0.05), and on jejunal immunoglobulin A (IgA) concentration at 7 d (P < 0.05). The addition of 100 mg/kg ORI had the greatest effect on the counts of Lactobacillus and Salmonella in cecal content (P < 0.05), malonaldehyde (MDA) content in the jejunum (P < 0.05), villi height of the small intestine, and IgA concentrations in the jejunum (P < 0.05). CONCLUSIONS: The results suggest that ORI can improve Salmonella-induced immune responses and protect intestinal health, not only through its immune inhibitory properties but also through its multi-protective effects on gut health.

Effects of oridonin on immune cells, Th1/Th2 balance and the expression of BLys in the spleens of broiler chickens challenged with Salmonella pullorum.

The aim of this study was to investigate the effects of oridonin (ORI) on the immune cells, Th1/Th2 balance and the expression of B lymphocyte stimulator (BLys) in the spleens of broilers infected with Salmonella pullorum. In a completely randomized design, 300 one-day-old AA male broilers were divided to 5 treatments. The groups included a noninfection control (CON) group receiveed a basal diet; a S. pullorum infect control group received the basal diet; and S. pullorum infect group receiveed the basal diet plus 50, 80, and 100 mg/kg ORI, respectively. The results showed that Salmonella challenge increased the relative weights of the spleen, white blood cell counts, lymphocyte and heterophil percentage, H/L ratio, the concentration and mRNA levels of spleen proinflammatory mediators, including tumor necrosis factor-α (TNF-α), interleukin-2 (IL-2), IL-4, IL-6, and IL-10, as well as the anti-inflammatory target Blys (P < .05), and modulated the Th1/Th2 balance (P < .05). ORI pretreatment decreased the relative weight of the spleen and inhibited the release and expression of these proinflammatory mediators and the anti-inflammatory target BLys. The results suggested that ORI supplementation may have immunosuppressive and multiple modulation effects on activated microglia through modulation of the Th1/Th2 balance and BLys expression.

Effects of dietary supplementation with oridonin on the growth performance, relative organ weight, lymphocyte proliferation, and cytokine concentration in broiler chickens.

BACKGROUND: The present study was conducted to investigate the effects of oridonin (ORI) on growth performance, relative organ weight, lymphocyte proliferation, phagocytic function of neutrophils, and cytokine concentration in broiler chickens. A total of 240 one-day-old Arbor Acres male broilers were randomly assigned to four treatments with six replicate pens of 10 broiler chickens per pen. Broiler chickens were fed diets based on four levels of dietary ORI (0, 50, 80 and 100 mg/kg) for a 42-d feeding trial. The experimental diets were fed in three phases: 1 to 14 d, 15 to 28 d and 29 to 42 d. RESULTS: The results indicated that ORI has no influence on the growth performance (P > 0.05). However, ORI increased the relative weights of spleen and bursa, the number of proliferation peripheral blood T and B lymphocytes, the phagocytic rate of neutrophils, as well as the Interleukin-2 (IL-2), Interleukin-4 (IL-4) and tumor necrosis factor-a (TNF-a) serum concentrations in serum in broilers at days 14, 28 and 42 (P < 0.05). CONCLUSIONS: In conclusion, ORI can enhance immune function and resistance to disease in broiler chickens by stimulating T and B lymphocyte formation, division, and proliferation, as well as the modulation of Th1/Th2 cytokine secretion profiles.

Natural Killer Group 2A Expressed on Both Peripheral CD3-CD56+NK Cells and CD3+CD8+T Cells Plays a Pivotal Negative Regulatory Role in the Progression of Hepatitis B Virus-Related Acute-on-Chronic Liver Failure.

To explore the role of surface receptors natural killer group 2A (NKG2A) and natural killer group 2D (NKG2D) on CD3+CD8+T cells and CD3-CD56+NK cells in the progression of hepatitis B virus (HBV)-related acute-on-chronic liver failure (ACLF), we measured the expression of NKG2A and NKG2D on the surface of these 2 types of circulating cells by flow cytometry in 3 groups. One group consists of 36 patients with chronic hepatitis B (CHB), another one consists of 22 patients with HBV-related ACLF, and the last one has 12 normal controls (NC). The experimental result indicated that there was no significant difference in the proportion of CD3+CD8+T cells in total lymphocytes between the 3 groups. However, the percentage of CD3-CD56+NK cells in ACLF group was evidently higher than that in the CHB group (P < 0.05). In addition, the expression of NKG2D on CD3+CD8+T cells in the ACLF group was significantly lower than that in the CHB group (P < 0.05), but there were no statistically significant differences in its percentages on CD3-CD56+NK cells between the 3 groups. The expression of NKG2A on CD3+CD8+T cells in the ACLF group was significantly higher than that in the NC group (P < 0.05), and on NK cells was significantly higher than that in the CHB group (P < 0.05) and NC group (P < 0.01). The increase in ratios of NKG2A to NKG2D on CD3+CD8+T cells and CD3-CD56+NK cells in the ACLF group was significantly more than that in the CHB group and NC group. The results indicate that the imbalance between NKG2A and NKG2D may contribute to the progression of HBV-related ACLF mediated by CD3-CD56+NK cells and CD3+CD8+T cells. Compared with NKG2D, NKG2A expressed on both peripheral CD3-CD56+NK cells and CD3+CD8+T cells plays a more pivotal negative regulatory role in the progression of HBV-related ACLF.

Highly efficient expression and purification system of small-size protein domains in Escherichia coli for biochemical characterization.

It is often essential to focus the study on the small-size domains of large proteins in eukaryotic cells in the post-genomic era, but the low expression level, insolubility, and instability of the domains have been continuing to hinder the massive purification of domain peptides for structural and biological investigation. In this work, a highly efficient expression and purification system based on a small-size fusion partner GB1 and histidine tag was utilized to solve these problems. Two vectors, namely pGBTNH and pGBH, were constructed to improve expression and facilitate purification. The linker and thrombin cleavage site have been optimized for minimal degradation during purification process. This system has been tested for eight domain peptides varying in size, linker, hydrophobicity, and predicted secondary structure. The results indicate that this system is achievable to produce these domain peptides with high solubility and stability for further biochemical characterization. Moreover, the fusion protein without the linker and thrombin cleavage site is also suitable for spectroscopic studies especially for NMR structural elucidation, if the target peptide is prone to precipitation or easily degraded during purification. This system will be beneficial to the research field of structure and function of small domain and peptide fragment.

Effects of segment substitution on the structure and stability of immunoglobulin G binding domain of streptococcal protein G.

Structural formation of segments plays pivotal roles in protein folding and stability, but how the segment influences the structural ensemble remains elusive. We engineered two hybrid proteins by replacing the central helical segment of immunoglobulin G binding domain of streptococcal protein G with an alpha-helix or beta(2)-strand element of a structural homologue, the immunoglobulin G binding domain of streptococcal protein L. The results show that substitution by the alpha-helical sequence retains a folded structure predominantly with a three-stranded beta-sheet but slightly destabilizes the compact ensemble, while substitution by the beta(2)-strand sequence completely destroys the structural formation. The finding implies that the local segment may influence the tertiary structure and overall stability, and the tertiary interactions may modulate structural formation of the segment, which might be considered when studying protein folding, prediction, design, and engineering.

[Gene mapping of a pedigree with autosomal dominant congenital cataract].

OBJECTIVE: To map the gene for autosomal dominant congenital cataract (ADCC) in a Chinese family. METHODS: Blood samples were collected from 14 members of this family. Linkage analysis was carried out using short tandem repeat polymorphism (STRP) in close proximity to genes and loci previously reported involving in human cataract. Two-point linkage analysis lod scores were calculated. RESULTS: The mutation gene locus in this pedigree was mapped to 17q, an 11.78-cM interval between markers D17S1288 and D17S933. Significant positive maximum LOD scores (Z(max)) at recombination fraction (theta) 0, were obtained for markers D17S805 (Z(max) = 2.03), D17S1294 (Z(max) = 2.49), and D17S1293 (Z(max) = 2.22). CONCLUSIONS: The mutation gene in this ADCC pedigree is located at chromosome 17q. This is the first report of an autosomal dominant congenital nuclear cataracts located at this locus. This result will be helpful for further studying of the pathogenesis of cataract.

[Autosomal dominant congenital nuclear cataract caused by a deletion mutation in the beta A1-crystallin gene].

OBJECTIVE: To identify the genetic defect causing automosal dominant congenital cataracts (ADCC) with nuclear opacities in a Chinese pedigree. METHODS: Linkage analysis was carried out with the short tandem repeat polymorphisms flanking the candidate genes. Mutation analysis of the candidate gene in the critical region was performed to detect the potential mutation. RESULTS: The cataract locus in this pedigree was mapped to 17q11.1-12, an 11.78 cM interval between markers D17S933 and D17S 1288. By means of sequencing the candiate gene, betaA1-crystallin (CRYBA1), a deletion mutation DeltaG91 in exon 4 was detected. This change cosegregated with the patients in the family but was not found in 50 normal unrelated individuals. CONCLUSION: It is a deletion mutation DeltaG91 of CRYBA1 gene that causes autosomal dominant congenital nuclear cataract. This is the first report of an autosomal dominant congenital nuclear cataract caused by the mutation in this gene.