Study on potential markers for diagnosis of renal cell carcinoma by serum untargeted metabolomics based on UPLC-MS/MS.

Objective: Renal cell carcinoma (RCC) is the most common malignancy of the kidney. However, there is no reliable biomarker with high sensitivity and specificity for diagnosis and differential diagnosis. This study aims to analyze serum metabolite profile of patients with RCC and screen for potential diagnostic biomarkers. Methods: Forty-five healthy controls (HC), 40 patients with benign kidney tumor (BKT) and 46 patients with RCC were enrolled in this study. Serum metabolites were detected by ultra-high performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), and then subjected to multivariate statistical analysis, metabolic pathway analysis and diagnostic performance evaluation. Results: The changes of glycerophospholipid metabolism, phosphatidylinositol signaling system, glycerolipid metabolism, d-glutamine and d-glutamate metabolism, galactose metabolism, and folate biosynthesis were observed in RCC group. Two hundred and forty differential metabolites were screened between RCC and HC groups, and 64 differential metabolites were screened between RCC and BKT groups. Among them, 4 differential metabolites, including 3-ß-D-Galactosyl-sn-glycerol, 7,8-Dihydroneopterin, lysophosphatidylcholine (LPC) 19:2, and γ-Aminobutyryl-lysine (an amino acid metabolite), were of high clinical value not only in the diagnosis of RCC (RCC group vs. HC group; AUC = 0.990, 0.916, 0.909, and 0.962; Sensitivity = 97.73%, 97.73%, 93.18%, and 86.36%; Specificity = 100.00%, 73.33%, 80.00%, and 95.56%), but also in the differential diagnosis of benign and malignant kidney tumors (RCC group vs. BKT group; AUC = 0.989, 0.941, 0.845 and 0.981; Sensitivity = 93.33%, 93.33%, 77.27% and 93.33%; Specificity = 100.00%, 84.21%, 78.38% and 92.11%). Conclusion: The occurrence of RCC may involve changes in multiple metabolic pathways. The 3-ß-D-Galactosyl-sn-glycerol, 7,8-Dihydroneopterin, LPC 19:2 and γ-Aminobutyryl-lysine may be potential biomarkers for the diagnosis or differential diagnosis of RCC.

A Review of Traditional Chinese Medicine on Treatment of Diabetic Nephropathy and the Involved Mechanisms.

Diabetic nephropathy (DN) is a common microvascular complication of diabetes mellitus (DM), which can lead to renal failure in diabetic patients. At present, the first-line drugs for DN are mainly the renin-angiotensin system (RAS) inhibitors or angiotensin receptor blockers, and the latest approved aldosterone receptor antagonist finerenone, which delay the progression of DN to end-stage renal disease (ESRD), but the therapeutic effect is still not ideal. With a history of thousands of years, traditional Chinese medicine (TCM) has rich experience in the treatment of DN. Based on the theory of TCM, the clinical treatment of DN mainly focuses on generating fluid and nourishing blood, nourishing Qi and Yin, detoxifying and detumescent. In recently years, the therapeutic effects and mechanisms of TCM prescription, Chinese herbal medicine, and its active components on DN have received extensive attention in new drug development. This paper reviews the research progress of the mechanism of TCM on DN.

Circ_000829 Plays an Anticancer Role in Renal Cell Carcinoma by Suppressing SRSF1-Mediated Alternative Splicing of SLC39A14.

Background: Covalently closed circular RNAs (circRNAs) play critical oncogenic or anticancer roles in various cancers including renal cell carcinoma (RCC), pointing to their regulation as a promising strategy against development of RCC. We, thus, studied the tumor-suppressive role of circ_000829 in RCC through in vitro and in vivo experiments. Methods: The expression of circ_000829 was validated in clinical RCC tissues and RCC cell lines. Based on ectopic expression and knockdown experiments, we examined the interactions among circ_000829, serine and arginine rich splicing factor 1 (SRSF1), and solute carrier family 39 member 14 (SLC39A14, zinc transporter). Then, the effects of circ_000829, SRSF1, and SLC39A14 on cell cycle distribution and proliferation in vitro and on tumor growth in vivo were evaluated in RCC cells. Results: Circ_000829 was poorly expressed in RCC tissues and cells, while SRSF1 was highly expressed. Restoration of circ_000829 reduced the levels of SRSF1 and SLC39A14B, thereby repressing the RCC cell proliferation in vitro and tumor growth in vivo. Meanwhile, overexpression of SRSF1 and SLC39A14B promoted the proliferation and cell cycle entry of RCC cells. Mechanistically, circ_000829 directly bound to SRSF1, and SRSF1 enhanced the expression of SLC39A14B by mediating the alternative splicing of SLC39A14. SLC39A14B upregulation negated the effect of SLC39A14 knockdown on RCC cell proliferation. Conclusion: Hence, this study suggests the antiproliferative role of circ_000829 in RCC growth and further elucidates the underlying mechanism involving the inhibited SRSF1-mediated alternative splicing of SLC39A14 mRNA.

The Relationship between Serum Trace Elements and Oxidative Stress of Patients with Different Types of Cancer.

OBJECTIVE: Many studies have identified causal and promotive roles of oxidative stress (OxS) and oxidative damage caused by OxS in the occurrence and progression of cancer. Many biomarkers in the blood circulation of patients may change correspondingly with the development of tumors. This study is aimed at investigating the correlation between OxS and serum trace element (TE) levels of patients with different types of cancer. METHODS: 1143 different types of cancer patients and 178 healthy controls from Mar. 2018 to Aug. 2020 in Mianyang Central Hospital were involved in this study. Their levels of OxS parameters (including total oxidant status (TOS), total antioxidant status (TAS), and oxidant stress index (OSI)) and the concentrations of serum TEs (including Cu, Zn, Fe, and Se) were determined. RESULTS: Compared with healthy controls, all types of cancer patients had higher TOS level (all P adj < 0.001) and OSI level (z = 6.228 ~ 9.909, all P adj < 0.001) and lower TAS level (all P adj < 0.001). Compared with healthy controls, the changes of four TE levels in serum were different in different types of cancer patients, among which Cu increased in all groups, but there was no statistical difference in gastric and brain cancer; Se decreased in all groups, but there was no statistical difference in gastric, colorectal, esophageal, and other cancer; Zn was significantly decreased in breast cancer patients (P adj < 0.001); there was no statistical difference in the change of Fe in liver, kidney, and other cancer. Spearman correlation showed that the change of Cu concentration was most closely related to the three OxS parameters and was strongly correlated in the observed several types of tumors (r s > 0.6). Multinomial logistic regression showed that the risks of different tumors are related to the level change of multiple TEs and OxS parameters (ORTOS = 1.19 ~ 2.82, OROSI = 2.56 ~ 4.70, ORTAS = 0.20 ~ 0.46, ORCu = 0.73 ~ 1.44, ORZn = 0.81 ~ 0.91, ORFe = 0.68 ~ 1.18, and ORSe = 0.22 ~ 0.45, all P < 0.006). CONCLUSIONS: The OxS exists in the occurrence and development of cancer, which may be related to the changes of certain trace elements. In order to evaluate OxS correctly, it is necessary to detect TAS and TOS and at the same time, their ratio OSI should be detected. Assessment of markers representing the overall level of OxS and TEs may guarantee improved the monitoring of disease occurrence and development risk in cancer patients.

Analgesic effect and related amino acids regulation of ginsenoside Rg3 in mouse pain models.

AIMS: The present study aims to evaluate the analgesic effect of ginsenoside Rg3 in different mouse pain models. MAIN METHODS: Formalin-, carrageenan- and S180 tumor cells induced mouse pain models were built in the study. The licking and biting time and PEG2 contents in the inflammatory sites were measured. The excitatory and inhibitory amino acids in the brains were determined by pre-column derivation FLD-HPLC method. KEY FINDING: We have found that ginsenoside Rg3 treated the pain phases and decreased the PGE2 in formalin and carrageenan induced models, respectively. It significantly increased the contents of EAAs (Asp and Glu) in the brains of S180 tumor inducing pain mice, meanwhile, the IAAs (Gly, Tau and GABA) decreased. SIGNIFICANCE: Our results revealed that ginsenoside Rg3 acted central and peripheral analgesic effect and regulated the inflammatory factors and pain-related amino acids. It could re-balance the abnormal EAAs/IAAs value when the pain occurred. The analgesic mechanism and the clinical application of ginsenoside Rg3 need be evaluated furtherly.

mTORC2 facilitates endothelial cell senescence by suppressing Nrf2 expression via the Akt/GSK-3ß/C/EBPα signaling pathway.

Vascular endothelial cell senescence is a leading cause of age-associated and vascular diseases. Mammalian target of rapamycin complex 2 (mTORC2) is a conserved serine/threonine (Ser/Thr) protein kinase that plays an important regulatory role in various cellular processes. However, its impact on endothelial senescence remains controversial. In this study we investigated the role and molecular mechanisms of mTORC2 in endothelial senescence. A replicative senescence model and H2O2-induced premature senescence model were established in primary cultured human umbilical vein endothelial cells (HUVECs). In these senescence models, the formation and activation of mTORC2 were significantly increased, evidenced by the increases in binding of Rictor (the essential component of mTORC2) to mTOR, phosphorylation of mTOR at Ser2481 and phosphorylation of Akt (the effector of mTORC2) at Ser473. Knockdown of Rictor or treatment with the Akt inhibitor MK-2206 attenuated senescence-associated ß-galactosidase (ß-gal) staining and expression of p53 and p21 proteins in the senescent endothelial cells, suggesting that mTORC2/Akt facilitates endothelial senescence. The effect of mTORC2/Akt on endothelial senescence was due to suppression of nuclear factor erythroid 2-related factor 2 (Nrf2) at the transcriptional level, since knockdown of Rictor reversed the reduction of Nrf2 mRNA expression in endothelial senescence. Furthermore, mTORC2 suppressed the expression of Nrf2 via the Akt/GSK-3ß/C/EBPα signaling pathway. These results suggest that the mTORC2/Akt/GSK-3ß/C/EBPα/Nrf2 signaling pathway is involved in both replicative and inducible endothelial senescence. The deleterious role of mTORC2 in endothelial cell senescence suggests therapeutic strategies (targeting mTORC2) for aging-associated diseases and vascular diseases.

Binding effect of fluorescence labeled glycyrrhetinic acid with GA receptors in hepatocellular carcinoma cells.

Glycyrrhetinic acid (GA) is a natural active component from licorice, which is broadly used in traditional Chinese medicine. Lots of glycyrrhetinic acid receptors (GA-R) are proved to locate on the surface of liver cells. Many reports about the hepatocellular carcinoma (HCC) treatment were dependent on GA modified carriers. However, the reality of GA-R in HCC cells was not clear. In this paper, 18ß-glycyrrhetinic acid (18ß-GA) was labeled with fluorescence (FITC) by chemical synthesis. Together with the binding effect of fluorescence labeled glycyrrhetinic acid (FITC-GA), the competitive action of 18ß-GA with GA-R was investigated in HCC cells. The results showed that in HepG2 cells, 18ß-GA and FITC-GA presented similar cytotoxicity. The specific binding saturation of GA showed the dissociation constant (Kd) was 7.457±2.122pmol/L and the maximum binding counts (Bmax) was 2.385±0.175pmol/2.5×106 cells, respectively. FITC-GA bound to cytomembrane specifically and 18ß-GA competed to bind the sites significantly in HepG2 cells. Therefore, there is binding effect between fluorescence labeled GA and GA-R. The GA-R on HCC cells is confirmed as expected, which provides a useful reference of active target modified by GA and a novel approach for receptors and ligands study.

[Biological assay in quality control of animal medicines].

Animal medicine is a unique part of traditional Chinese medicine. They have strong effects, but their effective compounds are not entirely known. The efficiency and safety of animal medicines can't be effectively controlled by current quality assurance system and evaluation method, which has deeply influenced the development of animal medicines. Biological assay does not focus on efficacy of single component, but directly reflects the pharmacodynamics and safety of animal medicines by biological effect. With the development of biotechnology, many new technologies have emerged, such as biochip and high content analysis. Based on the related targets, pathways and key biochemical factors, the field of biological assay has been expanded. With advantages of pharmacology andoverall controllability, as well as the characteristics of in line with the quality control of Chinese Medicine, biological assay will become one of the important development directionsfor quality standardization of animal medicines.

Analysis of the diagnostic efficiency of serum oxidative stress parameters in patients with breast cancer at various clinical stages.

BACKGROUND: Reactive oxygen species (ROS) are balanced through enzymatic mechanisms and exogenous antioxidants; imbalance results in oxidative stress (OxS). It is known that OxS plays an important role in the occurrence, development, and metastasis of breast cancer. The present study aimed to assess serum total oxidant status (TOS), total antioxidant status (TAS), and oxidant stress index (OSI) in patients at different clinical stages of breast cancer and to evaluate their diagnostic accuracy. METHODS: Serum TOS, TAS, and OSI were determined in 91 patients with breast cancer at different stages, 51 patients with benign breast tumors, and 35 healthy adults. RESULTS: Significant differences in serum TOS (F=104.384, p=0.000), TAS (F=18.247, p=0.000), and OSI (F=62.598, p=0.000) were observed among the 3 groups (benign breast tumor patients, breast cancer patients, and healthy women). Of the enrolled breast cancer patients, significant differences were also observed among different tumor stages, with TOS and OSI gradually increasing as the disease progressed, while TAS diminished. Receiver operating characteristic curve analysis revealed that the area under the ROC curve for OSI (AUCOSI) was significantly higher than AUCTAS (z=2.344, p=0.019) in distinguishing breast cancer from control groups (including disease control and the healthy control). The AUCOSI (z=4.700, p=0.001) or AUCTOS (z=4.700, p=0.001) was significantly higher than AUCTAS in distinguishing breast cancer from the healthy control. The AUCOSI (z=5.907, p=0.000) or AUCTOS (z=5.667, p=0.000) was significantly higher than AUCTAS in distinguishing benign breast tumors from the healthy control. CONCLUSION: Oxidative stress parameters might serve as important indexes for monitoring breast cancer occurrence and progression. The combined evaluation of TOS, TAS, and OSI could be more beneficial for clinical assessment.

Gene microarray assessment of multiple genes and signal pathways involved in androgen-dependent prostate cancer becoming androgen independent.

To study the gene expression change and possible signal pathway during androgen-dependent prostate cancer (ADPC) becoming androgen-independent prostate cancer (AIPC), an LNCaP cell model of AIPC was established using flutamide in combination with androgen-free environment inducement, and differential expression genes were screened by microarray. Then the biological process, molecular function and KEGG pathway of differential expression genes are analyzed by Molecule Annotation System (MAS). By comparison of 12,207 expression genes, 347 expression genes were acquired, of which 156 were up-ragulated and 191 down-regulated. After analyzing the biological process and molecule function of differential expression genes, these genes are found to play crucial roles in cell proliferation, differntiation, cell cycle control, protein metabolism and modification and other biological process, serve as signal molecules, enzymes, peptide hormones, cytokines, cytoskeletal proteins and adhesion molecules. The analysis of KEGG show that the relevant genes of AIPC transformation participate in glutathione metabolism, cell cycle, P53 signal pathway, cytochrome P450 metabolism, Hedgehog signal pathway, MAPK signal pathway, adipocytokines signal pathway, PPAR signal pathway, TGF-ß signal pathway and JAK-STAT signal pathway. In conclusion, during the process of ADPC becoming AIPC, it is not only one specific gene or pathway, but multiple genes and pathways that change. The findings above lay the foundation for study of AIPC mechanism and development of AIPC targeting drugs.

The diarrhoeogenic and antidiarrhoeal bidirectional effects of rhubarb and its potential mechanism.

AIM OF THE STUDY: The present study investigated the pharmacological effects of different extracts of rhubarb on intestinal function of mice, further to explore possible reasons for the adverse effects of long-term use of rhubarb as a purgative. MATERIALS AND METHODS: The total extract of rhubarb (TR) was extracted with 60% ethanol and the total anthraquinones extract (TA), total tannins extract (TT) and remaining components extract (RC) of rhubarb were separated from TR using macroporous resin. The pharmacological effects of each extract on the intestinal function of mice were evaluated by defecation test and the antidiarrhoeal activity of rhubarb tannins as well as its mechanism was studied by different animal models and histopathological examination. RESULTS: Both TR and TA produced purgative activities, but the purgative activity of TA was stronger than that of TR. Successive administration of TT produced an antidiarrhoeal activity in a time- and dose-dependent manner. Besides, successive administration of RC showed no significant effect on the intestinal function of mice. The antidiarrhoeal activity of rhubarb tannins was confirmed directly for the first time and its mechanism was probable that rhubarb tannins generated protein-precipitating reaction to the gastrointestinal mucosa due to its protein-precipitating action. CONCLUSIONS: The results confirmed that rhubarb had the diarrhoeogenic and antidiarrhoeal bidirectional effects due to the coexistence of anthraquinones and tannins. The bidirectional effects might be the reason or one of the reasons for the adverse effects of long-term use of rhubarb as a purgative.

Toxic effects caused by rhubarb (Rheum palmatum L.) are reversed on immature and aged rats.

AIM OF THE STUDY: Rhubarb is generally used to people of broad age, but diverse responses of people at different age to rhubarb have been little clarified. In this study, an attempt was made to access the safety of rhubarb to both immature and aged rats to provide some references for its clinical usage. MATERIALS AND METHODS: The total extract of rhubarb was administered intragastricly to both immature and aged rats once a day and lasted for 5 weeks. Then histopathologic and biochemical examinations were performed. RESULTS: No death was observed in immature rat groups, while 23.3% (21/90) subjects in aged rat groups died and most of the death cases were observed in the high-dosage (40 gkg(-1) of body weight per day od, counted on the quantity of crude material) group. The death rate between aged and immature rats was found of significantly statistical difference. Dosage-dependent histopathologic changes in kidney were observed in all the rhubarb-treated rats, principally involving the proximal tubules. Kidney changes in aged rats were severer than those observed in immature ones. Hepatic cells necrosis was occasionally observed in the middle- and high-dosage aged rat groups and minimal biliary hyperplasia was found in all the rhubarb-treated aged rats. Increased incidences of activated Kupffer cells and lymphocytic infiltration were found in all the rhubarb-treated rats. And dosage-dependent increase of interleukin 6 (IL-6) and notable increase of IL-8 was found in aged rat groups. CONCLUSIONS: The immature and aged rats showed reversed responses to the toxic potential of rhubarb extract. Elderly subjects were susceptible to the toxicity of high-dosage rhubarb, which drove rigorous consideration on rational use of rhubarb to aged people.

Binding of berberine to bovine serum albumin: spectroscopic approach.

Fluorescence spectroscopy in combination with UV-vis absorption spectroscopy was employed to investigate the binding of an important traditional medicinal herb berberine to bovine serum albumin (BSA) under the physiological conditions. In the mechanism discussion, it was proved that the fluorescence quenching of BSA by berberine is a result of the formation of berberine-BSA complex. Fluorescence quenching constants were determined using the Stern-Volmer equation and Scatchard equation to provide a measure of the binding affinity between berberine and BSA. The results of thermodynamic parameters ΔG, ΔH, ΔS at different temperatures indicate that the electrostatic interactions play a major role for berberine-BSA association. Site marker competitive experiments indicated that the binding of berberine to BSA primarily took place in site II. Furthermore, the Effect of supramolecules to berberine-BSA system, and the distance r between donor (BSA) and acceptor (berberine) was obtained according to fluorescence resonance energy transfer (FRET).

Site-selective binding of human serum albumin by palmatine: spectroscopic approach.

In this study, fluorescence spectroscopy in combination with UV-vis absorption spectroscopy and circular dichroism (CD) spectroscopy was employed to investigate the high affinity binding of palmatine to human serum albumin (HSA) under the physiological conditions. In the mechanism discussion it was proved that the fluorescence quenching of HSA by palmatine is a result of the formation of palmatine/HSA complex. Binding parameters calculating from Stern-Volmer method and Scatchard method showed that palmatine bind to HSA with the binding affinities of the order 10(4) L.mol(-1). The thermodynamic parameters studies revealed that the binding was characterized by negative enthalpy and positive entropy changes and the electrostatic interactions play a major role for palmatine-HSA association. Site marker competitive displacement experiments demonstrating that palmatine bind with high affinity to site I (subdomain IIA) of HSA. The specific binding distance r (2.91 nm) between donor (Trp-214) and acceptor (palmatine) was obtained according to fluorescence resonance energy transfer (FRET). Furthermore, the CD spectral result indicates that the secondary structure of HSA was changed in the presence of palmatine.

[Distribution of pathogen and resistance of nosocomial infections in the intensive care units].

OBJECTIVE: To determine the distribution of pathogens and their characteristics of drug susceptibility originating from nosocomial infections in the intensive care units (ICU), and to provide evidence for clinical anti-infection treatments. METHODS: Retrospective analysis to the pathogens and their drug susceptibility characteristics was carried out. These pathogens were isolated from the samples that came from patients infected in the ICU from 2002 to 2004. RESULTS: The main nosocomial infective pathogens in the ICU were gram negative bacilli (48.2%), and the next ones were gram positive bacteria (43.3%) and fungus (8.5%). The most common gram negative bacilli were Pseudomonas aeruginosa and Escherichia coli; while for gram positive bacteria, the main bacterin were Staphylococcus aureus. The gram negative bacilli could resist 4 or more than 4 antibiotics, and the rate for resistance exceeded 40%. Similarly, oxacillin resistance staphylococcus could resist 7 antibiotics, and the rate was over 50%. The detective rates of ESBLs and AmpC enzymes produced by Escherichia coli and K. peumoniae were 34.0% & 30.7% and 13.2% & 23.1%, respectively. The rate for oxacillin resistance staphylococcus was 66.3%, and there was relative high resistance rate ( > 55%) for most antibiotics: There was statistical difference, compared with that of non-resistant strains. CONCLUSION: Though gram positive coccus still play an important role, most infections are caused by gram negative bacilli of nosocomial infections in the ICU. The antibiotics resistant rate of all bacteria has been rising gradually. It shows strong resistance and multi-drug resistance. The most importment cause for resistance of gram negative bacilli is that the bacteria can produce ESBLs and AmpC enzymes. The antibiotic resistant rate for oxacillin resistance staphylococcus is really high.