Investigation of relationships between metabolic chemical reporter structures and S-glyco-modification effects.

The approach of metabolic chemical reporters (MCRs) for labeling proteins has been widely used in the past several decades. Nevertheless, artificial side reaction generated with fully protected MCRs, termed S-glyco-modification, occurs with cysteine residues through base-promoted ß-elimination and Michael addition, leading to false positives in the proteomic identification. Therefore, next generation of MCRs, including partially protected strategy and modifications on the backbone of monosaccharides, have emerged to improve the labeling efficiency. In this paper, we prepared fifteen kinds of unnatural monosaccharides to investigate the relationships of structures and S-glyco-modification labeling. Our results demonstrated that Ac4GlcNAz and Ac4GalNAz exhibited the most remarkable labeling effects among the detected compounds. Of note, Ac4ManNAz, Ac46AzGlucose and Ac46AzGalactose containing similar structures but did not show similar robust signals as them. Moreover, other modifications on the 1-, 2-, 3-, 4- and 6-site indicated minimal side reactions of S-glyco-modification, raising a possibility that subtle modifications of monosaccharide substrate may alter its role in the process of biosynthesis, for example, by change of electronegativity or enhancement of steric hindrance effects. In conclusion, our discoveries provide a new avenue to choose appropriate probe for selective label proteins in vitro and in vivo without undesired S-glyco-modification.

GB1a Activates SIRT6 to Regulate Lipid Metabolism in Mouse Primary Hepatocytes.

Lipid accumulation, oxidative stress, and inflammation in hepatocytes are features of nonalcoholic fatty liver disease (NAFLD). Garcinia biflavonoid 1a (GB1a) is a natural product capable of hepatic protection. In this study, the effect of GB1a on anti-inflammatory, antioxidant, and regulation of the accumulation in HepG2 cells and mouse primary hepatocytes (MPHs) was investigated, and its regulatory mechanism was further explored. The result showed that GB1a reduced triglyceride (TG) content and lipid accumulation by regulating the expression of SREBP-1c and PPARα; GB1a reduced reactive oxygen species (ROS) and improved cellular oxidative stress to protect mitochondrial morphology by regulating genes Nrf2, HO-1, NQO1, and Keap1; and GB1a reduced the damage of hepatocytes by inhibiting the expression of inflammatory cytokines interleukin-6 (IL-6), interleukin-1ß (IL-1ß), tumor necrosis factor-alpha (TNF-α), and nuclear factor kappa B (NF-κB) p65. The activities of GB1a were lost in liver SIRT6-specific knockout mouse primary hepatocytes (SIRT6-LKO MPHs). This indicated that activating SIRT6 was critical for GB1a to perform its activity, and GB1a acted as an agonist of SIRT6. It was speculated that GB1a may be a potential drug for NAFLD treatment.

Long Circulating Cancer Cell-Targeted Bionic Nanocarriers Enable Synergistic Combinatorial Therapy in Colon Cancer.

Cancer nanomedicine treatment aims to achieve highly specific targeting and localization to cancer cells. Coating of nanoparticles with cell membranes endows them with homologous cellular mimicry, enabling nanoparticles to acquire new functions and properties, including homologous targeting and long circulation in vivo, and can enhance internalization by homologous cancer cells. Herein, we fused a human-derived HCT116 colon cancer cell membrane (cM) with a red blood cell membrane (rM) to fabricate an erythrocyte-cancer cell hybrid membrane (hM). Oxaliplatin and chlorin e6 (Ce6) co-encapsulated reactive oxygen species-responsive nanoparticles (NPOC) were camouflaged by hM and obtained a hybrid biomimetic nanomedicine (denoted as hNPOC) for colon cancer therapy. hNPOC exhibited prolonged circulation time and recognized homologous targeting ability in vivo since both rM and HCT116 cM proteins were maintained on the hNPOC surface. hNPOC showed enhanced homologous cell uptake in vitro and considerable homologous self-localization in vivo, producing effective synergistic chemophotodynamic therapy efficacy under irradiation with a homologous HCT116 tumor compared to that with a heterologous tumor. Together, the biomimetic hNPOC nanoparticles showed prolonged blood circulation and preferential cancer cell-targeted function in vivo to provide a bioinspired strategy for chemophotodynamic synergistic therapy of colon cancer.

RNA-seq-based transcriptome analysis of the anti-inflammatory effect of artesunate in the early treatment of the mouse cerebral malaria model.

BACKGROUND: cerebral malaria (CM) is an important complication of malaria with a high mortality rate. Artesunate is recommended as the first-line artemisinin compound treatment for severe malaria. Due to the difficulty of obtaining brain tissue samples clinically, the use of animals to research host responses to CM parasite infections is necessary. Rodent malaria models allow for detailed time series studies of host responses in multiple organs. To date, studies on the transcriptome of severe malaria are only limited to the parasites in the peripheral blood of patients, and there is little data on the transcriptional changes in brain tissue in mice with CM treated with artesunate. METHOD AND RESULT: in this study, fresh tissue samples (three biological replicates per mouse) from the same area of the brain in each animal were collected from the uninfected, Plasmodium berghei ANKA-infected and artesunate-treated C57BL/6 mice, and then transcriptome research was performed by the RNA-seq technique. Differentially expressed genes (DEGs) included Il-21, Tnf, Il-6, Il-1ß, Il-10, Ifng, and Icam-1. Among which, Il-6, Il-10, Tnf-α and Il-1ß were further verified and validated via qRT-PCR and ELISA. This revealed that Il-1ß (p < 0.0001), Il-10 (p < 0.05) and Tnf-α (p < 0.05) were significantly up-regulated in the Pb ANKA-infected versus uninfected group, while Il-1ß (p < 0.0001) and Tnf-α (p < 0.05) were significantly down-regulated after artesunate treatment. All DEGs were closely related to the top 3 artesunate treatment pathways, including the JAK-STAT signaling pathway, apoptosis, and Toll-like receptor signaling pathway. CONCLUSION: the mechanism of improving the prognosis of cerebral malaria by artesunate may not only involve the killing of plasmodium but also the inhibition of a cytokine storm in the host. This study provides new insights into the molecular mechanism by which artesunate improves the prognosis of cerebral malaria.

Garcinia Biflavonoid 1 Improves Lipid Metabolism in HepG2 Cells via Regulating PPARα.

Garcinia biflavonoid 1 (GB1) is one of the active chemical components of Garcinia kola and is reported to be capable of reducing the intracellular lipid deposition, which is the most significant characteristic of non-alcoholic fatty liver disease. However, its bioactive mechanism remains elusive. In the current study, the lipid deposition was induced in HepG2 cells by exposure to oleic acid and palmitic acid (OA&PA), then the effect of GB1 on lipid metabolism and oxidative stress and the role of regulating PPARα in these cells was investigated. We found that GB1 could ameliorate the lipid deposition by reducing triglycerides (TGs) and upregulate the expression of PPARα and SIRT6, suppressing the cell apoptosis by reducing the oxidative stress and the inflammatory factors of ROS, IL10, and TNFα. The mechanism study showed that GB1 had bioactivity in a PPARα-dependent manner based on its failing to improve the lipid deposition and oxidative stress in PPARα-deficient cells. The result revealed that GB1 had significant bioactivity on improving the lipid metabolism, and its potential primary action mechanism suggested that GB1 could be a potential candidate for management of non-alcoholic fatty liver disease.

Comparison of non-anthocyanin polyphenol accumulation in the berry skins of muscadine and European grapes during ripening in China.

The non-anthocyanin polyphenol accumulation in the berry skins of muscadine grapes from South China during ripening was investigated, in comparison to European grapes, for two consecutive years, using ultra-performance liquid chromatography tandem triple quadrupole time-of-flight mass spectrometry. A total of 121 components were tentatively identified, consisting of 48 ellagic acids and precursors (EP), 38 flavonols, 20 hydroxybenzoic acids, 6 hydroxycinnamic acids, 5 flavan-3-ols, and 4 stilbenes. EP (mostly ellagic acid derivatives) and flavonols (mainly isorhamnetin and quercetin derivatives) were, respectively, accumulated abundantly in the muscadine and European grape skins. Additionally, their variation had considerable differences during berry maturation. EP in muscadine cv. "Noble" skins showed a small decrease before veraison and a slight increase post-veraison, yet the flavonols significantly increased as berry maturation in European skins, with a few exceptions. This is the interactional result between genotype and environmental factors, which obviously influences the polyphenol composition and concentration. PRACTICAL APPLICATIONS: Muscadine grapes, indigenous to the southeast of United States, have been introduced to South China in recent years. These grapes have unique polyphenol composition that differs from non-muscadine grapes. Their polyphenol properties have been studied extensively in many but there are a few studies in our knowledge about the variation of these compounds at different berry developmental stages. This article evaluates the accumulation of non-anthocyanin polyphenols in muscadine skins during berry maturation, in comparison to European grapes. The findings exhibit the polyphenol characteristics of these grapes cultivated in new locations across South China, and make further efforts to increase breeding quality and agronomic production, through improving preharvest treatments and agronomic performance during berry developmental stages, that lead to a high increase of polyphenol.

HBD Inhibits the Development of Colitis-Associated Cancer in Mice via the IL-6R/STAT3 Signaling Pathway.

Colitis-associated cancer (CAC) is a malignant disease of the colon that is caused by recurrent episodes of chronic intestinal inflammation. Huangqi Baizhu decoction (HBD) is a classic prescription comprised of Radix Astragali and Rhizoma Atractylodis, which are usually used to treat digestive conditions, such as peptic ulcers, colitis, or colorectal carcinoma in clinics. HBD is well known for "tonifying qi and spleen" based on the theories of traditional Chinese medicine, and has the preponderant effect of alleviating chronic intestinal mucosa damage associated with disease. However, the underlying mechanism behind this is still unknown. In the current study, we employed the AOM/DSS mouse model to analyze the effects of HBD on the development of inflammation in colonic carcinoma. The in vivo study showed that HBD could significantly reduce the mortality of mice and control the incidence and size of colonic tumors by inhibiting the IL-6/STAT3 signaling pathway. In vitro, Astragaloside and Atractylenolide (CAA), the main components of HBD, inhibited the proliferation of HCT-116 cells as determined by an MTT assay. Furthermore, CAA notably suppressed the protein expression of IL-6R, STAT3, Survivin, and Cyclin D1 induced by IL-6 in HCT-116 and RAW264.7 cells. These results suggested that HBD exhibits anti-inflammatory and anti-proliferative effects, inhibiting the development of CAC in mice.

Identification of NPAC as a novel biomarker and regulator for hepatocellular carcinoma.

OBJECTIVE: Hepatocellular carcinoma (HCC) has a high morbidity and mortality around the world, yet the effective therapeutic option for HCC is still limited. NPAC, also known as glyoxylate reductase 1 homolog, is a new nuclear protein recently implicated in tumor biology. However, the role of NPAC in HCC remains unclear. The present study aimed to evaluate the clinical significance and potential role of NPAC in HCC. METHODS: The NPAC expression in HCC tissues and matched adjacent normal tissues was detected by real-time polymerase chain reaction, immunohistochemistry (IHC), and Western blot analysis. The clinical significance of the expression of NPAC in HCC was assessed by the Kaplan-Meier survival curve and the Cox regression model. In addition, we established a doxiline-induced overexpression of the NPAC system. The effects of NPAC on HCC cell proliferation, migration, and apoptosis were checked by CCK-8 proliferation assays, transwell, and flow cytometry, respectively. RESULTS: The NPAC expression was significantly downregulated in HCC tissues and HCC cell lines. NPAC reduction was significantly correlated with poorer survival among patients with HCC, and the multivariate analysis confirmed its independent prognostic value. Furthermore, overexpression of NPAC dramatically suppressed the proliferation of HCC cells and promoted HCC cells apoptosis. Besides, the levels of phosphorylation of janus kinase 2 (JAK2) and signal transduction and activator 3 (STAT3) were significantly reduced after overexpression of NPAC in HCC cell lines. CONCLUSIONS: These results suggest that NPAC may play an important role in the development and progression of HCC, and can act as a novel potential prognostic biomarker and therapeutic target for HCC.

Profile of Polyphenol Compounds of Five Muscadine Grapes Cultivated in the United States and in Newly Adapted Locations in China.

Polyphenol compositions and concentrations in skins and seeds of five muscadine grapes (cv. "Noble", "Alachua", "Carlos", "Fry", and "Granny Val") cultivated in the United States (Tallahassee-Florida, TA-FL) and South China (Nanning-Guangxi, NN-GX and Pu'er-Yunnan, PE-YN) were investigated, using ultra performance liquid chromatography tandem triple quadrupole time-of-flight mass spectrometry (UPLC Triple TOF MS/MS). Fourteen ellagitannins were newly identified in these muscadine grapes. The grapes grown in NN-GX accumulated higher levels of ellagic acid, methyl brevifolin carboxylate, and ellagic acid glucoside in skins, and penta-O-galloyl-glucose in seeds. In PE-YN, more flavonols were detected in skins, and higher contents of flavan-3-ols, ellagic acid, and methyl gallate were identified in seeds. Abundant seed gallic acid and flavonols were found among the grapes grown in TA-FL. Based on principal component analysis (PCA) of 54 evaluation parameters, various cultivars grown in different locations could be grouped together and vice versa for the same cultivar cultivated in different regions. This is the result of the interaction between genotype and environmental conditions, which apparently influences the polyphenol synthesis and accumulation.

Effects of parafibromin expression on the phenotypes and relevant mechanisms in the DLD-1 colon carcinoma cell line.

BACKGROUND: Parafibromin is a protein encoded by the HRPT2 (hyperparathyroidism 2) oncosuppressor gene and its down-regulated expression is involved in pathogenesis of parathyroid, breast, gastric and colorectal carcinomas. This study aimed to clarify the effects of parafibromin expression on the phenotypes and relevant mechanisms of DLD-1 colon carcinoma cells. METHODS: DLD-1 cells transfected with a parafibromin-expressing plasmid were subjected to examination of phenotype, including proliferation, differentiation, apoptosis, migration and invasion. Phenotype-related proteins were measured by Western blot. Parafibromin and ki-67 expression was detected by immunohistochemistry on tissue microarrays. RESULTS: The transfectants showed higher proliferation by CCK-8, better differentiation by electron microscopy and ALP activity and more apoptotic resistance to cisplatin by DNA fragmentation than controls. There was no difference in early apoptosis by annexin V, capase-3 activity, migration and invasion between DLD-1 cells and their transfectants. Ectopic parafibromin expression resulted in down-regulated expression of smad4, MEKK, GRP94, GRP78, GSK3ß-ser9, and Caspase-9. However, no difference was detectable in caspase-12 and -8 expression. A positive relationship was noted between parafibromin and ki-67 expression in colorectal carcinoma. CONCLUSIONS: Parafibromin overexpression could promote cell proliferation, apoptotic resistance, and differentiation of DLD-1 cells.

[Effects of Sijunzi Decoction on urine's xylose excretion rate and ATP in mucosa of spleen deficiency rats].

OBJECTIVE: To observe the effects of Sijunzi Decoction on D-xylose excretion rate and ATP content in the mucosa membranes of small intestines of rats with spleen deficiency. METHODS: Spleen deficiency model rats were made by reserpine injection. D-xylose excretion rate was measured with p-bromoaniline method, and the ATP content of small intestines mucosa was detected with bioluminescence method. The correlation between D-xylose excretion rate and ATP content of mucosa was also analyzed. RESULTS: Rats' body weight and D-xylose excretion rate decreased after reserpine injection (P < 0.01, vs control group), but increased after treated with Sijunzi Decoction (P < 0.05, vs model group). The ATP content of mucosa showed no significant difference between model group and control group. There was obviously positive correlation between the change of urine's D-xylose excretion rate and mucosa ATP content. CONCLUSION: Sijunzi Decoction has the activity of improving xylose absorption in spleen deficiency rats, but no obvious effect on their mucosa ATP content. The reducing of urine's D-xylose excretion rate in spleen deficiency rats is accompanied with the decrease of mucosa ATP content.