Chemical fingerprint analysis and metabolic profiling of 50% ethanol fraction of Lomatogonium rotatum by ultra-performance liquid chromatography/quadrupole-time of flight mass spectrometry.

Lomatogonium rotatum (L.) Fries ex Nym (L. rotatum), a member of Gentianaceae, is an important mongolian medicine in China used to treat febrile diseases in liver and gallbladder. The aim of present study was to investigate the chemical constituents and metabolites of the 50% ethanol fraction of L. rotatum (50EtLR). Firstly, the extract of L. rotatum was partitioned by macroporous resin to obtain the target fraction (50EtLR), then several compounds were isolated from 50EtLR to obtained the standards for further analysis of chemical constituents of 50EtLR. Secondly, the chemical constituents of 50EtLR were characterized using the ultra-high performance liquid chromatography coupled with quadrupole-time-of-flight mass spectrometry (UHPLC-Q-TOF-MS/MS). Finally, prototype constituents and related metabolites were analyzed after orally administerng 50EtLR to rats. As a result, a new compound, 6-O-[ß-d-xylopyranosyl-(1 → 6)-O-ß-d-glucopyranosyl]-1,4,8-trimethoxyxanthone (6) along with seven known compounds (1-5, 7 and 8) were isolated from the 50EtLR, 92 components were either unambiguously or tentatively identified. Additionally, 34 prototype constituents and 112 metabolites in rat plasma along with 32 prototype constituents and 53 metabolites in rat liver were tentatively identified. Therefore, xanthones and flavonoids were the main chemical constituents of 50EtLR and sulfation and glucuronidation are the main enzyme-induced metabolic pathways involved post-administration.

Gain of function in the mouse model of a recurrent mutation p53N236S promotes the formation of double minute chromosomes and the oncogenic potential of p19ARF.

The mutation p53N236S (p53S) has been identified as one of the recurrent mutations in human cancers by TCGA database. Our in vitro data revealed the oncogenic gain of function of p53S. To understand the function of p53S in vivo, we generated the p53S knock-in mouse. The p53S/S mice manifested highly invasive lymphomas and metastatic sarcomas with dramatically increased double minute chromosomes. The survival curve, the incidence of tumors and the tumor spectrum of p53S/S mice is very similar to the p53R172H mouse model. The p53S/+ mice showed delayed onset of tumorigenesis and a high metastasis rate (40%) and low loss of heterozygosity rate (2/16). The activation of CDKN2A pathway in p53S/S MEF and tumors, and the accumulation of p19ARF protein in tumor tissues suggested p19ARF might contribute to the accumulation of mutant p53S protein in the tumor and promote tumorigenesis. The high expression of p19ARF correlated with mutant p53 accumulation and tumor progression, suggesting a dual role of p19ARF in tumor promotion or suppression that might depend on the p53 mutation status in tumor cells. The oncogenic gain of function of this recurrent mutation p53S prompts the reconsideration of p53 mutations function that occurs at a low frequency.

Efficient and Stable Proton Exchange Membrane Water Electrolysis Enabled by Stress Optimization.

Proton exchange membrane water electrolysis (PEMWE) is a promising solution for the conversion and storage of fluctuating renewable energy sources. Although tremendously efficient materials have been developed, commercial PEMWE products still cannot fulfill industrial demands regarding efficiency and stability. In this work, we demonstrate that the stress distribution, a purely mechanical parameter in electrolyzer assembly, plays a critical role in overall efficiency and stability. The conventional cell structure, which usually adopts a serpentine flow channel (S-FC) to deliver and distribute reactants and products, resulted in highly uneven stress distribution. Consequently, the anode catalyst layer (ACL) under the high stress region was severely deformed, whereas the low stress region was not as active due to poor electrical contact. To address these issues, we proposed a Ti mesh flow channel (TM-FC) with gradient pores to reduce the stress inhomogeneity. Consequently, the ACL with TM-FC exhibited 27 mV lower voltage initially and an 8-fold reduction in voltage degradation rate compared to that with S-FC at 2.0 A/cm2. Additionally, the applicability of the TM-FC was demonstrated in cross-scale electrolyzers up to 100 kW, showing a voltage increase of only 20 mV (accounting for less than 2% of overall voltage) after three orders of magnitude scaleup.

Pu-erh tea inhibits tumor cell growth by down-regulating mutant p53.

Pu-erh tea is a kind of fermented tea with the incorporation of microorganisms' metabolites. Unlike green tea, the chemical characteristics and bioactivities of Pu-erh tea are still not well understood. Using water extracts of Pu-erh tea, we analyzed the tumor cell growth inhibition activities on several genetically engineered mouse tumor cell lines. We found that at the concentration that did not affect wild type mouse embryo fibroblasts (MEFs) growth, Pu-erh tea extracts could inhibit tumor cell growth by down-regulated S phase and cause G1 or G2 arrest. Further study showed that Pu-erh tea extracts down-regulated the expression of mutant p53 in tumor cells at the protein level as well as mRNA level. The same concentration of Pu-erh tea solution did not cause p53 stabilization or activation of its downstream pathways in wild type cells. We also found that Pu-erh tea treatment could slightly down-regulate both HSP70 and HSP90 protein levels in tumor cells. These data revealed the action of Pu-erh tea on tumor cells and provided the possible mechanism for Pu-erh tea action, which explained its selectivity in inhibiting tumor cells without affecting wild type cells. Our data sheds light on the application of Pu-erh tea as an anti-tumor agent with low side effects.

Lomatogonium Rotatum for Treatment of Acute Liver Injury in Mice: A Metabolomics Study.

Lomatogonium rotatum (L.) Fries ex Nym (LR) is used as a traditional Mongolian medicine to treat liver and bile diseases. This study aimed to investigate the hepatoprotective effect of LR on mice with CCl4-induced acute liver injury through conventional assays and metabolomics analysis. This study consisted of male mice (n = 23) in four groups (i.e., control, model, positive control, and LR). The extract of whole plant of LR was used to treat mice in the LR group. Biochemical and histological assays (i.e., serum levels of alanine transaminase (ALT) and aspartate transaminase (AST), and histological changes of liver tissue) were used to evaluate LR efficacy, and metabolomics analysis based on GC-MS and LC-MS was conducted to reveal metabolic changes. The conventional analysis and metabolomic profiles both suggested that LR treatment could protect mice against CCl4-induced acute liver injury. The affected metabolic pathways included linoleic acid metabolism, α-linolenic acid metabolism, arachidonic acid metabolism, CoA biosynthesis, glycerophospholipid metabolism, the TCA cycle, and purine metabolism. This study identified eight metabolites, including phosphopantothenic acid, succinic acid, AMP, choline, glycerol 3-phosphate, linoleic acid, arachidonic acid, and DHA, as potential biomarkers for evaluating hepatoprotective effect of LR. This metabolomics study may shed light on possible mechanisms of hepatoprotective effect of LR.

[Transfection Efficiency of Ad5F11p-GFP on CIK and NK-92 Cells and Its Influence on Biological Characteristics].

OBJECTIVE: To study transfection efficiency of Ad5F11p-GFP and its influence on biological characteristics of CIK and NK-92 cells in order to predict the application of Ad5F11p vector in immunotherapy. METHODS: Two kinds of immune cells, cytokine-induced killer (CIK) cells and natural-killer (NK) cell line NK-92 cells, were transfected by Ad5F11p-GFP at different multiplicity of transfection (MOI), and untransfected immune cells were used as negative control. GFP expression was determined by flow cytometry, the cell morphology was observed with microscope, the cell proliferation was analyzed by trypan blue staining, specific cytotoxicity of NK-92 cells was determined by LDH assay. RESULTS: About 90% of transfection efficiency for NK-92 cells could be achieved at a MOI of 25, while the transfection efficiency for CIK was less than 40% at a MOI of 200. In addition, the transfection efficiency basically unchanged at the same MOI for 48 h and 96 h, and the immune cells transfected with the virus trended to form agglomeration, displaying slower proliferation, increase of IFN-γ release and enhancement of tumor killing activity. CONCLUSION: Ad5F11p- modified NK-92 shows a good prospect for adoptive immunotherapy.

Adipose-Derived Mesenchymal Stem Cells Ameliorate Lipid Metabolic Disturbance in Mice.

UNLABELLED: : Adipose-derived mesenchymal stem cells (AD-MSCs) have been shown to ameliorate hyperglycemia in diabetic animals and individuals. However, little is known about whether AD-MSCs affect lipid metabolism. Here we have demonstrated for the first time that AD-MSC infusion can significantly suppress the increase in body weight and remarkably improve dyslipidemia in db/db obese mice and diet-induced obesity mice. Induction of white fat tissue "browning" and activation of adenosine monophosphate-activated protein kinase and its downstream hormone-sensitive lipase in adipose tissue contribute to the antiobesity and lipid-lowering effects. Thus, AD-MSC infusion holds great therapeutic potential for dyslipidemia and associated cardiovascular diseases. SIGNIFICANCE: Mesenchymal stem cells (MSCs) are considered one of the most promising types of stem cells for translational application because of their rich tissue sources, multilineage differentiation capacity, and easy amplification in vitro and unique immunobiological properties. This study demonstrated that adipose-derived MSCs (AD-MSCs) infusion can significantly suppress the increase in body weight and remarkably improve dyslipidemia in obese mice. Induction of white fat tissue "browning" and activation of adenosine monophosphate-activated protein kinase and its downstream hormone-sensitive lipase in adipose tissue were demonstrated to contribute to the antiobesity and lipid-lowering effects. Thus, AD-MSC infusion holds great therapeutic potential for dyslipidemia.

Association of p53 Arg72Pro polymorphism with esophageal cancer: a meta-analysis based on 14 case-control studies.

BACKGROUND: The p53 tumor suppressor gene Arg72Pro polymorphism has been associated with esophageal cancer. However, the results were not consistent. Herein, this meta-analysis was performed to estimate the association between p53 Arg72Pro polymorphism and esophageal cancer. METHODS: Electronic search of PubMed was conducted to select studies. Studies containing available genotype frequencies of Arg72Pro were chosen, and pooled odds ratio (OR) with 95% confidence interval (CI) was used to assess the association. RESULTS: The final meta-analysis included 14 published studies with 4184 esophageal cancer cases and 7308 controls. The results suggested that the variant genotype was associated with the esophageal cancer risk in additive model (Pro vs. Arg: OR=1.146, 95% CI: 1.016-1.293, p=0.027) and in recessive model (Pro/Pro vs. Arg/Arg+Arg/Pro: OR=1.258, 95% CI: 1.021-1.551, p=0.031). In the stratified analysis by ethnicity, the data suggested that the increased esophageal cancer risk associated with p53 Arg72Pro polymorphism was more evident in the Asian group. The symmetric funnel plot, the Egger's test (p>0.05) and the Begg's test (p>0.05) suggested the lack of publication bias. CONCLUSION: This meta-analysis suggests that p53 codon 72 polymorphism contributes to esophageal cancer risk, especially in Asians. To validate this association, further studies with more participants worldwide are needed to examine association between this polymorphism and esophageal cancer.

The gain of function of p53 mutant p53S in promoting tumorigenesis by cross-talking with H-RasV12.

The loss of wild type p53 tumor suppressive function and oncogenic gain-of-function of p53 mutants have been showing important implications in tumorigenesis. The p53(N236S) (p53(N239S) in human, p53S) mutation has been shown to lose wild type p53 function by yeast assay. However, its gain of function is still not clear. By gel shift assay, we showed that mutant p53S had lost its DNA binding ability to its target promoters. Further real-time PCR data confirmed that p53S had lost the function of regulating the transcription of p21( Cip1/Waf1), cyclin G, PUMA, and Bax in response to 10Gy irradiation. These data confirmed the loss of function of p53S in mammalian cells. By xenograft assay, we showed that the p53S per se was not oncogenic enough to form tumor, however, cooperating with H-RasV12, p53S could dramatically promote tumorigenesis in p53 null MEFs. Further study showed that co-expression of p53S and H-RasV12 could increase the expression level of H-RasV12 and partially eliminate the elevation of stress response proteins such as Chk2, γ-H2AX, Hsp70, Rb, p16(Ink4a) caused by either p53S or H-RasV12. These data suggested that p53S cross-talked with H-RasV12 and reduced the cellular stress response to oncogenic signals, which facilitated the cell growth and tumorigenesis. Together these data provided the molecular basis for the cooperation of p53S and H-RasV12 and revealed the gain of function of p53S in cross-talking with H-RasV12. This study revealed an important aspect of gain of function for p53 mutant, therefore might shed light on the clinical strategy in targeting p53 mutant.

Association between the MTHFR C677T polymorphism and recurrent pregnancy loss: a meta-analysis.

BACKGROUND: C677T polymorphism of the methylenetetrahydrofolate reductase (MTHFR) has been associated with recurrent pregnancy loss (RPL). However, results were conflicting. The aim of this study was to quantitatively summarize the evidence for MTHFR C677T polymorphism and RPL risk. METHODS: Electronic search of PubMed and the Chinese Biomedicine database was conducted to select studies. Case-control studies containing available genotype frequencies of C677T were chosen, and odds ratio (OR) with 95% confidence interval (CI) was used to assess the strength of this association. RESULTS: The case-control studies including 2427 cases and 3118 controls were identified. The meta-analysis stratified by ethnicity showed that individuals with the homozygous TT genotype had increased risk of RPL (OR=1.574, 95% CI: 1.163-2.13, p=0.003), in Asians (OR=1.663, 95% CI: 1.012-2.731, p=0.045). Results among Caucasians did not suggest an association (OR=1.269, 95% CI: 0.914-1.761, p=0.155). A symmetric funnel plot, the Egger's test (p=0.285), and the Begg's test (p=0.529) were all suggestive of the lack of publication bias. The studies conducted in each of the defined number of pregnancy losses-two or more pregnancy losses, and three or more pregnancy losses-showed no effect of the C677T polymorphism on RPL except for the TT versus CT+CT genotype comparison for the three or more pregnancy loss subgroup (OR=1.792, 95% CI: 1.187-2.704, p=0.005). CONCLUSION: This meta-analysis supports the idea that MTHFR C677T genotype is associated with increased risk of RPL, except for Caucasians. To draw comprehensive and true conclusions, further prospective studies with larger numbers of participants worldwide are needed to examine associations between MTHFR C677T polymorphism and RPL.