To investigate the changes of genetic damage in patients with arsenism caused by coal-burning in 9 years. To analyze the relationship between the changes of genetic damage and disease progression and provide a basis for condition monitoring.
Alcohol-induced liver disease (ALD) is a leading cause of non-accident-related deaths in the world. Identification of an early specific signature of ALD would aid in therapeutic intervention. Scoparone is an important constituent of Yinchenhao, and displayed bright prospects in hepatoprotective effect. However, its precise molecular mechanism has not been well explored. The present study was designed to assess the effects and possible mechanisms of scoparone against alcohol-induced liver injury. UPLC/ESI-Q-TOF/MS combined with pattern recognition approaches including PCA, and PLS-DA were integrated to get differentiating metabolites for the pathways and clarify mechanisms of disease, highlight insights into drug discovery. The results indicated four ions in the positive mode were characterized as potential differentiating metabolites which can be regulated by scoparone treatment, and suggested that therapeutic effect of scoparone could regulated the dysfunctions of citrate cycle, sphingolipid metabolism, taurine and hypotaurine.
Proteinuria serves as a direct causative factor of renal tubular cell injury and is highly associated with the progression of chronic kidney disease via uncertain mechanisms. Recently, evidence demonstrated that both NLRP3 inflammasome and mitochondria are involved in the chronic kidney disease progression. The present study was undertaken to examine the role of NLRP3 inflammasome/mitochondria axis in albumin-induced renal tubular injury. In patients with proteinuria, NLRP3 was significantly up-regulated in tubular epithelial cells and was positively correlated with the severity of proteinuria. In agreement with these results, albumin remarkably activated NLRP3 inflammasome in both in vitro renal tubular cells and in vivo kidneys in parallel with significant epithelial cell phenotypic alteration and cell apoptosis. Genetic disruption of NLRP3 inflammasome remarkably attenuated albumin-induced cell apoptosis and phenotypic changes under both in vitro and in vivo conditions. In addition, albumin treatment resulted in a significant mitochondrial abnormality as evidenced by the impaired function and morphology, which was markedly reversed by invalidation of NLRP3/caspase-1 signaling pathway. Interestingly, protection of mitochondria function by Mn(III)tetrakis (4-benzoic acid) porphyrin (MnTBAP) or cyclosporin A (CsA) robustly attenuated albumin-induced injury in mouse proximal tubular cells. Collectively, these findings demonstrated a pathogenic role of NLRP3 inflammasome/caspase-1/mitochondria axis in mediating albumin-induced renal tubular injury. The discovery of this novel axis provides some potential targets for the treatment of proteinuria-associated renal injury.
Mitochondrial abnormality has been shown in many kidney disease models. However, its role in the pathogenesis of chronic kidney diseases (CKDs) is still uncertain. In present study, a mitochondrial complex I inhibitor rotenone was applied to the mice subjected to unilateral ureteral obstruction (UUO). Following 7-days rotenone treatment, a remarkable attenuation of tubular injury was detected by PAS staining. In line with the improvement of kidney morphology, rotenone remarkably blunted fibrotic response as shown by downregulation of fibronectin (FN), plasminogen activator inhibitor-1 (PAI-1), collagen I, collagen III, and ?-SMA, paralleled with a substantial decrease of TGF-? 1. Meanwhile, the oxidative stress markers thiobarbituric acid-reactive substances (TBARS) and heme oxygenase 1 (HO-1) and inflammatory markers TNF-?, IL-1?, and ICAM-1 were markedly decreased. More importantly, the reduction of mitochondrial DNA copy number and mitochondrial NADH dehydrogenase subunit 1 (mtND1) expression in obstructed kidneys was moderately but significantly restored by rotenone, suggesting an amelioration of mitochondrial injury. Collectively, mitochondrial complex I inhibitor rotenone protected kidneys against obstructive injury possibly via inhibition of mitochondrial oxidative stress, inflammation, and fibrosis, suggesting an important role of mitochondrial dysfunction in the pathogenesis of obstructive kidney disease.
Induction of metallothionein (MT) expression is involved in metal homeostasis and detoxification. To identify the key pathways that regulate metal-induced cytotoxicity, we investigate how phosphorylated metal-responsive transcription factor-1 (MTF-1) contributed to induction of MT expression. Immortal human embryonic kidney cells (HEK cells) were treated with seven kinds of metals including cadmium chloride (CdCl2), zinc sulfate (ZnSO4), copper sulfate(CuSO4), lead acetate (PbAc), nickel sulfate (NiSO4), sodium arsenite (NaAsO2), and potassium bichromate (K2Cr2O7). The MT expression was induced in a dose-response and time-dependent manner upon various metal treatments. A cycle of phosphorylation and dephosphorylation was required for translocation of MTF-1 from cytoplasm to nucleus, leading to the up-regulation of MTs expression. Protein phosphatase 2A (PP2A) participated in regulating MT expression through dephosphorylation of MTF-1. A loss-of-function screen revealed that the specific PP2A complexes containing PR110 were involved in metal-induced MT expression. Suppression of PP2A PR110 in HEK cells resulted in the persistent MTF-1 phosphorylation and the disturbance of MTF-1 nuclear translocation, which was concomitant with a significant decrease of MT expression and enhanced cytotoxicity in HEK cells. Notably, MTF-1 was found in complex with specific PP2A complexes containing the PR110 subunit upon metal exposure. Furthermore, we identify that the dephosphorylation of MTF-1 at residue Thr-254 is directly regulated by PP2A PR110 complexes and responsible for MTF-1 activation. Taken together, these findings delineate a novel pathway that determines cytotoxicity in response to metal treatments and provide new insight into the role of PP2A in cellular stress response.
Metabolomics is a new approach based on the systematic study of the full complement of metabolites in a biological sample. Extracting biomedical information from large datasets is of considerable complexity. Furthermore, the traditional method of assessing metabolomics data is not only time-consuming but it is often subjective work. Here we used sensitive ultra-performance LC-ESI/Q-TOF high-definition mass spectrometry (UPLC-ESI-Q-TOF-MS) in positive ion mode coupled with a new developed software program TransOmics for widely untargeted metabolomics, which incorporates novel nonlinear alignment, deconvolution, matched filtration, peak detection, and peak matching to characterize metabolites as a case study. The TransOmics method can facilitate prioritization of the data and greatly increase the probability of identifying metabolites related to the phenotype of interest. By this means, 17 urinary differential metabolites were identified (less than 10 min) involving the key metabolic pathways including tyrosine metabolism, glutathione metabolism, phenylalanine metabolism, ascorbate and aldarate metabolism, arginine and proline metabolism, and so forth. Metabolite identification has also been significantly improved, using the correlation peak patterns in contrast to a reference metabolite panel. It can detect and identify metabolites automatically and remove background noise, and also provides a user-friendly graphical interface to apply principal component analyses, correlation analysis and compound statistics. This investigation illustrates that metabolomics combined with the proposed bioinformatic approach (based on TransOmics) is important to elucidate the developing biomarkers and the physiological mechanism of disease, and has opened the door for the development of a new genre of metabolite identification methods.
Abstract Trace elements play an important role in maintaining the normal metabolic and immune function. The onset of recurrent respiratory tract infection (RRI) is associated with the immune function, genetic factors and nutritional status. However, the association between the levels of trace elements and RRI remains inconclusive. We aimed to investigate the alterations of hair levels of zinc (Zn), copper (Cu) and iron (Fe) in Chinese children with RRI by performing a meta-analysis. A predefined electronic databases search was performed to identify eligible studies for the analysis of hair Zn, Cu or Fe levels in Chinese children with RRI. Thirteen studies were included. RRI patients displayed significantly lower levels of hair Zn (13 studies, random effects SMD: - 1.215, 95% CI: - 1.704 to - 0.725, p < 0.0001), Cu (11 studies, random effects SMD: - 0.384, 95% CI: - 0.717 to - 0.052, p = 0.023) and Fe (12 studies, random effects SMD: - 0.569, 95% CI: - 0.827 to - 0.312, p < 0.0001) compared with controls. No evidence of publication bias was observed. Sensitivity analysis did not change the results significantly. In conclusion, the deficiency of Zn, Cu and Fe may be contributing factors for the susceptibility of RRI in Chinese children. However, more studies in different ethnicities should be performed in the future.
A sensitive high-performance liquid chromatography method coupled with diode array detection (HPLC-DAD) was developed for the quality control of Phellodendri amurensis cortex (PAC), the quality control included the simultaneous determination of seven major constituents, namely phellodendrine, magnoflorine, jatrorrhizine, palmatine, berberine, obaculactone and obacunone. The chromatographic separation was accomplished on a Diamonsil-C18 column (4.6 mm × 200 mm, 5 ?m) with acetonitrile and 0.1% phosphoric acid (0.02 mol sodium dihydrogen phosphate per liter) by linear gradient elution. The established method was successfully validated by acceptable linearity, limits of detection and quantitation, precision, repeatability, stability and accuracy. The HPLC-DAD fingerprint chromatograph under 220 nm consisting of 21 peaks was constructed for the evaluation of the 11 batches of PAC. The HPLC fingerprints were analyzed by similarity analysis, hierarchical clustering analysis and principal component analysis. The results indicated that the combination of multicomponent determination method and chromatographic fingerprint analysis could be employed for the quantitative analysis and identification of PAC, as well as pharmaceutical products containing this herbal material.
Angiotensin II (Ang II) stimulates (pro)renin receptor (PRR) expression in the renal collecting duct, triggering the local renin response in the distal nephron. Our recent study provided evidence for involvement of cyclooxygenase-2-prostaglandin E2 pathway in Ang II-dependent stimulation of PRR expression in the collecting duct. Here, we tested the role of E-prostanoid (EP) subtypes acting downstream of cyclooxygenase-2 in this phenomenon. In primary rat inner medullary collecting duct cells, Ang II treatment for 12 hours induced a 1.8-fold increase in the full-length PRR protein expression. To assess the contribution of EP receptor, the cell was pretreated with specific EP receptor antagonists: SC-51382 (for EP1), L-798106 (for EP3), L-161982 (for EP4), and ONO-AE3-208 (ONO, a structurally distinct EP4 antagonist). The upregulation of PRR expression by Ang II was consistently abolished by L-161982 and ONO and partially suppressed by SC-51382 but was unaffected by L-798106. The PRR expression was also significantly elevated by the EP4 agonist CAY10598 in the absence of Ang II. Sprague-Dawley rats were subsequently infused for 1 or 2 weeks with vehicle, Ang II alone, or in combination with ONO. Ang II infusion induced parallel increases in renal medullary PRR protein and renal medullary and urinary renin activity and total renin content, all of which were blunted by ONO. Both tail cuff plethysmography and telemetry demonstrated attenuation of Ang II hypertension by ONO. Overall, these results have established a crucial role of the EP4 receptor in mediating the upregulation of renal medullary PRR expression and renin activity during Ang II hypertension.
Abstract The uromodulin (UMOD) is a glycoprotein expressed exclusively by renal tubular cells lining the thick ascending limb of the loop of Henle. UMOD acts as a regulatory protein in health and in various conditions. For kidney diseases, its role remains elusive. On one hand, UMOD plays a role in binding and excretion of various potentially injurious products from the tubular fluid. On the other hand, chronic kidney disease is associated with higher serum levels of UMOD. Signaling pathways might be very important in the pathogenesis of kidney diseases. We performed this review to provide a relatively complete signaling pathway flowchart for UMOD to the investigators who were interested in the role of UMOD in the pathogenesis of kidney diseases. Here, we reviewed the signal transduction pathway of UMOD and its role in the pathogenesis of kidney diseases.
Prostate cancer is a leading cause of cancer deaths in men worldwide. Although prostate-specific antigen (PSA) has been extensively used as a serum biomarker to detect prostate cancer, this screening method has suffered from a lack of specificities and sensitivities. The successful prevention and treatment of prostate cancer relies on the early and accurate detection of the disease; therefore, more sensitive biomarkers are urgently needed. Prostate has long been known to exhibit unique metabolite profiles, fortunately, metabolomics, the study of all metabolites produced in the body, can be considered most closely related to a patient's phenotype. It may provide clinically useful biomarkers applied toward identifying metabolic alterations in prostate cancer and has introduced new insights into the pathology of prostate cancer. This advanced bioanalytic method may now open door for diagnostics. Metabolomics has a great and largely potential in the field of disease, and the analysis of the cancer metabolome to identify novel biomarkers and targets can now be undertaken in many research laboratories. In this review, we take a closer look at the metabolomics in the field of prostate cancer and highlight the interesting publications as references for the interested reader.
Adriamycin (ADR)-induced nephropathy in animals is an experimental analog of human focal segmental glomerulosclerosis, which presents as severe podocyte injury and massive proteinuria and has a poorly understood mechanism. The present study was designed to test the hypothesis that the peroxisome proliferator-activated receptor-? coactivator (PGC)-1?-mitochondria axis is involved in ADR-induced podocyte injury. Using MPC5 immortalized mouse podocytes, ADR dose dependently induced downregulation of nephrin and podocin, cell apoptosis, and mitochondrial dysfunction based on the increase in mitochondrial ROS production, decrease in mitochondrial DNA copy number, and reduction of mitochondrial membrane potential and ATP content. Moreover, ADR treatment also remarkably reduced the expression of PGC-1?, an important regulator of mitochondrial biogenesis and function, in podocytes. Strikingly, PGC-1? overexpression markedly attenuated mitochondrial dysfunction, the reduction of nephrin and podocin, and the apoptotic response in podocytes after ADR treatment. Moreover, downregulation of PGC-1? and mitochondria disruption in podocytes were also observed in rat kidneys with ADR administration, suggesting that the PGC-1?-mitochondria axis is relevant to in vivo ADR-induced podocyte damage. Taken together, these novel findings suggest that dysfunction of the PGC-1?-mitochondria axis is highly involved in ADR-induced podocyte injury. Targeting PGC-1? may be a novel strategy for the treatment of ADR nephropathy and human focal segmental glomerulosclerosis.
Ligand binding promotes conformational rearrangement of the epidermal growth factor receptor (EGFR) leading to receptor autophosphorylation and downstream signaling. However, transient interactions between unstimulated EGFR molecules on the cell surface are not fully understood. In this report, we describe the investigation of homodimer formation of EGFR by means of an SNAP-tag based selective crosslinking approach (S-CROSS). EGFR homodimers were selectively captured in living cells and utilized for analysis of protein receptor interactions on the plasma membrane and ligand-induced activation. We showed that EGFR forms homodimers in unstimulated cells with efficiencies similar to those seen in cells treated with the epidermal growth factor ligand (EGF) supporting the existence of constitutive transient receptor-receptor interactions. EGFR crosslinked homodimers displayed a substantially increase in kinase activation upon ligand stimulation. Interestingly, in unstimulated cells the levels of spontaneous phosphorylation were found to correlate with the yields of the crosslinked homodimers species. In addition, we demonstrated that this crosslinking approach can be applied to interrogate the effect of small molecule inhibitors on receptor dimerization and kinase activity. Our crosslinking assay provides a new tool to dissect ligand-independent dimerization and activation mechanisms of receptor tyrosine kinases, many of which are important anticancer drug targets.
Increasing numbers of people are suffering from allergic contact dermatitis. However, the immunosuppressive drug candidate with negligible toxicity is still deficient. In the present study, we identified a natural cyclodepsipeptide named trichomide A that effectively inhibited the proliferation of activated T cells and reduced the production of proinflammatory cytokines but had almost no toxic effect on naive T cells at 0.3-3??M. In addition, trichomide A caused G0/G1 phase arrest, suppressed the activation of AKT and STAT3, and increased the level of phosphorylated SHP2 in activated T cells in dose- and time-dependent manners. Furthermore, an in vivo experiment demonstrated that trichomide A significantly ameliorated picryl chloride (PCI)-induced contact hypersensitivity in mice. Such effects of trichomide A in the aforementioned experiments were significantly reversed by the inhibition of SHP2 activity using the SHP2-specific inhibitor PHPS1 or conditional SHP2 knockout mice in T cells, suggesting the SHP2-dependent action of trichomide A. Taken together, trichomide A showed an immunosuppressive activity against T cell-mediated immune responses both in vitro and in vivo, which has potential for the treatment of immune-related skin diseases.
It is well documented that oxidative stress is involved in the pathogenesis of idiopathic nephrotic syndrome (INS). Malondialdehyde (MDA) is a measurement of lipid oxidation; vitamin C and E are important components of antioxidants. However, the association between MDA, vitamin C or E levels and INS remains elusive. A meta-analysis was performed to investigate the alteration of serum levels of MDA, vitamin C and E in INS compared with controls. Eight studies were included in our meta-analysis according to predefined criteria. Active INS patients demonstrated significantly higher level of serum MDA (SMD: 2.13, 95% CI: 1.511 to 2.749, p < 10(-4)), markedly lower levels of serum vitamin C (SMD: -1.449, 95% CI: -2.616 to -0.281, p = 0.015) and E (SMD: -1.45, 95% CI: -2.544 to -0.356, p = 0.009) compared with those in controls. Active steroid-sensitive nephrotic syndrome (SSNS) patients showed comparable levels of serum vitamin C and E to those in controls. INS subjects in the remission stage demonstrated significantly higher level of serum MDA (SMD: 1.028, 95% CI: 0.438 to 1.617, p < 10(-4)), markedly lower level of serum vitamin C (SMD: -2.235, 95% CI: -3.048 to -1.421, p < 10(4)) and similar level of serum vitamin E compared with those in controls. No significant publication bias was observed. In conclusion, the disorder of MDA and vitamin C persists in the remission stage of INS. It seems that the serum levels of vitamin C and E is associated with the responsiveness of INS to steroids. However, more studies should be performed in the future.
Cyclooxygenase 2 (COX-2) has an established role in postnatal kidney development. 15-Hydroxyprostaglandin dehydrogenase (15-PGDH) is recently identified as an endogenous inhibitor of COX-2, limiting the production of COX-2-derived prostanoids in several pathological conditions. The present study was undertaken to examine the regulation of renal 15-PGDH expression during postnatal kidney development in rats compared with COX-2. qRT-PCR and immunoblotting demonstrated that 15-PGDH mRNA and protein in the kidney were present in neonates, peaked in the second postnatal week, and then declined sharply to very low level in adulthood. Immunostaining demonstrated that at the second postnatal week, renal 15-PGDH protein was predominantly found in the proximal tubules stained positive for Na/H exchanger 3 and brush borders (periodic acid-Schiff), whereas COX-2 protein was restricted to macular densa and adjacent thick ascending limbs. Interestingly, in the fourth postnatal week, 15-PGDH protein was redistributed to thick ascending limbs stained positive for the Na-K-2Cl cotransporter. After 6 wk of age, 15-PGDH protein was found in the granules in subsets of the proximal tubules. Overall, these results support a possibility that 15-PGDH may regulate postnatal kidney development through interaction with COX-2.
Urchin-like NiCo2S4 sub-micron spheres integrated with nano-sized and micro-sized structures, which were synthesized via a facile one-pot method, deliver efficient electrocatalytic activities for oxygen reduction and evolution reactions. The excellent electrocatalytic property of NiCo2S4 sub-micron spheres is originated from their unique urchin-like microstructure, composition and d-electronic configurations of the transition metal ions.
The association between megsin 2093C/T, 2180C/T and C25663G gene polymorphisms and IgA nephropathy (IgAN) risk remains unclear. We aimed to evaluate the association between megsin 2093C/T, 2180C/T and C25663G gene polymorphisms and IgAN risk by performing a meta-analysis. Eligible studies were searched according to predefined criteria by using electronic databases. Six articles were identified for the analysis of the association between megsin 2093C/T, 2180C/T and C25663G gene polymorphisms and IgAN risk. 2093C/T C allele was associated with IgAN risk in overall populations and Asians (overall populations: p?=?0.014, Asians: p?=?0.037). 2093C/T CC/TT genotype was not associated with IgAN risk in overall populations, Caucasians and Asians. 2180C/T C allele was correlated with IgAN risk in Caucasians (p?=?0.024). 2180C/T CC/TT genotype was not associated with IgAN risk in overall populations, Caucasians and Asians. C25663G gene polymorphism was not associated with IgAN onset in Asians. In conclusion, megsin 2093C/T C allele may be genetic marker for IgAN susceptibility in overall populations and Asians. 2180C/T C allele may be risk factor for IgAN onset in Caucasians. However, more studies should be performed in the future.
Hyperuricemia is associated with kidney complications including glomerulosclerosis and mesangial cell (MC) proliferation by poorly understood mechanisms. The present study investigated the underlying mechanisms that mediate uric acid (UA)-induced MC proliferation. A rat MC line, HBZY-1, was treated with various concentrations of UA in the presence or absence of a specific extracellular-regulated protein kinase 1/2 (ERK1/2) inhibitor (U0126), apocynin. UA dose dependently stimulated MC proliferation as shown by increased DNA synthesis and number of cells in the S and G2 phases in parallel with the upregulation of cyclin A2 and cyclin D1. In addition, UA time dependently promoted MC proliferation and significantly increased phosphorylation of ERK1/2 but not c-Jun NH2-terminal kinase and p38 MAPK in MCs as assessed by immunoblotting. Inhibition of ERK1/2 signaling via U0126 markedly blocked UA-induced MC proliferation. More importantly, UA induced intracellular reactive oxygen species (ROS) production of MCs dose dependently, which was completely blocked by apocynin, a specific NADPH oxidase inhibitor. Toll-like receptor (TLR)2 and TLR4 signaling had no effect on NADPH-derived ROS and UA-induced MC proliferation. Interestingly, pretreatment with apocynin inhibited ERK1/2 activation, the upregulation of cyclin A2 and cyclin D1, and MC proliferation. In conclusion, UA-induced MC proliferation was mediated by NADPH/ROS/ERK1/2 signaling pathway. This novel finding not only reveals the mechanism of UA-induced MC cell proliferation but also provides some potential targets for future treatment of UA-related glomerular injury.
Many studies have shown that LPS mainly activates four signal transduction pathways to induce inflammation, namely the p38, ERK1/2, JNK and IKK/NF-?B pathways. Studies have demonstrated that 5'-AMP-induced hypothermia (AIH) exhibits high anti-inflammatory capabilities. In this study, we explore that how AIH inhibits the inflammatory response. Wistar rats were divided into five groups: a control group, an LPS group, a 5'-AMP pre-treatment group, a 5'-AMP post-treatment group and a 5'-AMP group. For each group, plasma and lung were collected from the rats at 6h and 12h after LPS injection. ELISA assays were used to detect plasma levels of CD14, CRP and MCP-1. Inflammatory pathway activation and TLR4 expression were assayed separately by Western blot analysis and immunohistochemistry. Our results showed that rats treated with AIH either before or after an LPS-challenge had a significant decrease in plasma levels of CD14, CRP and TLR4 compared with rats that received LPS only. Western blot analysis showed that AIH inhibited the activation of extracellular signal-regulated kinases (ERK) 1/2, p38, c-Jun N-terminal kinase (JNK) and NF-?B in inflammatory rats. Our study concluded that AIH attenuated LPS-induced inflammation mainly by inhibiting activation on the ERK1/2, p38, JNK and NF-?B signaling pathways.
It is well documented that selenium (Se) is involved in the metabolism of glucose. However, whether Se supplementation could lower the risk of type 2 diabetes mellitus (T2DM) remains elusive. We aimed to evaluate the association between Se supplementation and the risk of T2DM by performing a meta-analysis. We searched the Pubmed, Embase, and Cochrane databases from January 1990 to November 2013 to identify randomized controlled trials (RCTs) that met pre-stated inclusion criteria. Reference lists of retrieved articles were also reviewed. Either a fixed-effects or, in the presence of heterogeneity, a random-effects model was used to calculate the pooled prevention effects. Four RCTs involving 20,294 participants were included in this meta-analysis. The combined relative risks (RRs) for subjects administered with Se compared with control groups were 1.09 (95 % CI: 0.99-1.20, p = 0.085). Omission of any single study did not change the overall risk estimates significantly. Meta- regression analyses showed almost no impact on the RRs of age and study length. No evidence of publication bias was observed. In conclusion, our findings do not support the routine application of Se supplementation for T2DM prevention in Caucasians. Larger studies are needed to investigate the effects of Se supplementation on T2DM prevention among various populations and further elucidate the impact of age and study length.
The chemical constituents of Simiao Wan (SW), a traditional Chinese medicine preparation, are difficult to determine and remain unclear. To more efficiently detect ions, a multiple data processing approach has been used in the characterization of the compounds. In this study, a rapid and sensitive method based on ultra high performance liquid chromatography with mass spectrometry and the multiple data processing approach was established to characterize the chemical constituents of SW. Ultra high performance liquid chromatography with mass spectrometry coupled with the multiple data processing approach could efficiently remove nonrelated ion signals from accurate mass data. We report the application of the multiple data processing approach for comprehensive detection and rapid identification of chemical constituents of SW. Of note, the total analysis time for separation was less than 20 min without losing any resolution. In the variable, importance in projection plot of orthogonal projection to latent structure-discriminant analysis, a total of 72 ions of interest (37 ions in positive mode, 38 ions in negative mode and three ions in both mode) were extracted or tentatively characterized based on their retention times, exact mass measurement for each molecular ion and subsequent fragment ions. In summary, the methodology proposed in this study could be valuable for the structural characterization and identification of the multiple constituents in the traditional Chinese medicine formula SW.
Diabetic nephropathy (DN) is a severe complication of diabetes and serves as the leading cause of chronic renal failure. In the past decades, angiotensin-converting enzyme inhibitors (ACEIs)/angiotensin II receptor blockers (ARBs) based first-line therapy can slow but cannot stop the progression of DN, which urgently requests the innovation of therapeutic strategies. Thiazolidinediones (TZDs), the synthetic exogenous ligands of nuclear receptor peroxisome proliferator-activated receptor- ? (PPAR ? ), had been thought to be a promising candidate for strengthening the therapy of DN. However, the severe adverse effects including fluid retention, cardiovascular complications, and bone loss greatly limited their use in clinic. Recently, numerous novel PPAR ? agonists involving the endogenous PPAR ? ligands and selective PPAR ? modulators (SPPARMs) are emerging as the promising candidates of the next generation of antidiabetic drugs instead of TZDs. Due to the higher selectivity of these novel PPAR ? agonists on the regulation of the antidiabetes-associated genes than that of the side effect-associated genes, they present fewer adverse effects than TZDs. The present review was undertaken to address the advancements and the therapeutic potential of these newly developed PPAR ? agonists in dealing with diabetic kidney disease. At the same time, the new insights into the therapeutic strategies of DN based on the PPAR ? agonists were fully addressed.
Asymmetric dimethylarginine (ADMA) is considered an independent mortality and cardiovascular risk factor in chronic kidney disease (CKD) patients, and contributes to the development of renal fibrosis. Quercetin (QC), a natural component of foods, protects against renal injury. Here, we explored the possible mechanisms that are responsible for ADMA-induced renal fibrosis and the protective effect of QC. We found that ADMA treatment activated the endoplasmic reticulum (ER) stress sensor proteins phosphorylated protein kinase RNA-activated-like ER kinase (PERK) and inositol requiring-1? (IRE1), which correspondingly induced C/EBP homologous protein (CHOP) expression and phosphorylated c-Jun N-terminal kinase (JNK) phosphorylation in glomerular endothelial cells (GEnCs). Following this, ADMA promoted ER stress-induced apoptosis and resulted in transforming growth factor ? (TGF-?) expression in GEnCs. SP600125, an inhibitor of JNK, and CHOP siRNA protected against ADMA-induced cell apoptosis and TGF-? expression. QC prevented ADMA-induced PERK and IRE1 apoptotic ER stress pathway activation. Also, ADMA-induced GEnCs apoptosis and TGF-? expression was reduced by QC. Overexpression of CHOP blocked QC-mediated protection from apoptosis in ER stressed cells. Overall, these observations indicate that ADMA may induce GEnCs apoptosis and TGF-? expression by targeting the PERK-CHOP and IRE1-JNK pathway. In addition, drugs such as QC targeting ER stress may hold great promise for the development of novel therapies against ADMA-induced renal fibrosis.
Exposure of humans to inorganic arsenic can cause skin cancer. The acquisition of cancer stem cell-like properties is involved in the initiation of some cancers, and there are changes in let-7 levels in some tumors. The mechanisms of action, however, remain obscure. Here, we report that there are decreased levels of let-7a, let-7b, and let-7c in human keratinocyte HaCaT cells during malignant transformation induced by a low concentration (1.0?M) of arsenite. The process by which arsenite reduces the level of let-7c apparently involves methylation, for 5-aza-2'-deoxycytidine, an inhibitor of methyltransferases, prevents arsenite-induced hypermethylation, decreases the level of let-7c, and thereby blocks arsenite-induced activation of the Ras/NF-?B signal pathway. Let-7c is an up-stream regulator of the Ras/NF-?B signal pathway and down-regulates activation of this pathway. In arsenite-transformed HaCaT cells, the acquisition of cancer stem cell-like properties is prevented by over-expression of let-7c, and over-expression of let-7c decreases the malignancy of transformed HaCaT cells. Thus, we conclude that epigenetic silencing of let-7c via Ras/NF-?B is involved in the acquisition of cancer stem cell-like properties and neoplastic transformation of HaCaT cells induced by arsenite, which contribute to the tumorigenesis of arsenite.
Metabolomics represents an emerging discipline concerned with comprehensive assessment of small molecule endogenous metabolites in biological systems and provides a powerful approach insight into the mechanisms of diseases. Type 2 diabetes (T2D), called the burden of the 21st century, is growing with an epidemic rate. However, its precise molecular mechanism has not been comprehensively explored. In this study, we applied urinary metabolomics based on the UPLC/MS integrated with pattern recognition approaches to discover differentiating metabolites, to characterize and explore metabolic pathway disruption in an experimental model for high-fat-diet induced T2D. Six differentiating urinary metabolites were found in the negative mode, and two (2-(4-hydroxy-3-methoxy-phenyl) acetaldehyde sulfate, 2-phenylethanol glucuronide) of which were identified involving the key metabolic pathways linked to pentose and glucuronate interconversions, starch, sucrose metabolism and tyrosine metabolism. Our study provides new insight into pathophysiologic mechanisms and may enhance the understanding of T2D pathogenesis.
Mitochondrial dysfunction has gained recognition as a contributing factor in many diseases. The kidney is kind of organ with high energy demand, rich in mitochondria. As such, mitochondrial dysfunction in the kidney plays a critical role in the pathogenesis of kidney diseases. Despite of the recognized importance mitochondria play in the pathogenesis of the diseases, there is limited understanding on various aspects of mitochondrial biology. This review examines the physiology and pathophysiology of mitochondria. It begins by discussing mitochondrial structure, mitochondrial DNA, mitochondrial reactive oxygen species production, mitochondrial dynamics and mitophagy, before turning to inherited mitochondrial cytopathies in kidneys (inherited or sporadic mitochondrial DNA or nuclear DNA mutations in genes that affect mitochondrial function). Glomerular diseases, tubular defects and other renal diseases are then discussed. Next, acquired mitochondrial dysfunction in kidney diseases is discussed, emphasizing the role of mitochondrial dysfunction in the pathogenesis of chronic kidney disease and acute kidney injury, as their prevalence is increasing. Finally, it summarizes the possible beneficial effects of mitochondrial-targeted therapeutic agents for treatment of mitochondrial dysfunction-mediated kidney injury-genetic therapies, antioxidants, thiazolidinediones, sirtuins and resveratrol-as mitochondrial-based drugs may offer potential treatments for renal diseases.
Colorectal cancer (CRC), a major public health concern, is the second leading cause of cancer death in developed countries. There is a need for better preventive strategies to improve the patient outcome that is substantially influenced by cancer stage at the time of diagnosis. Patients with early stage colorectal have a significant higher 5-year survival rates compared to patients diagnosed at late stage. Although traditional colonoscopy remains the effective means to diagnose CRC, this approach generally suffers from poor patient compliance. Thus, it is important to develop more effective methods for early diagnosis of this disease process, also there is an urgent need for biomarkers to diagnose CRC, assess disease severity, and prognosticate course. Increasing availability of high-throughput methodologies open up new possibilities for screening new potential candidates for identifying biomarkers. Fortunately, metabolomics, the study of all metabolites produced in the body, considered most closely related to a patients phenotype, can provide clinically useful biomarkers applied in CRC, and may now open new avenues for diagnostics. It has a largely untapped potential in the field of oncology, through the analysis of the cancer metabolome to identify marker metabolites defined here as surrogate indicators of physiological or pathophysiological states. In this review we take a closer look at the metabolomics used within the field of colorectal cancer. Further, we highlight the most interesting metabolomics publications and discuss these in detail; additional studies are mentioned as a reference for the interested reader.
Abstract Metabolomics technologies enable the examination and identification of endogenous biochemical reaction products, revealing information on the precise metabolic pathways and processes within a living cell. Metabolism is either directly or indirectly involved with every aspect of cell function, and metabolomics is thus believed to be a reflection of the phenotype of any cell. Metabolomics analysis of cells has many potential applications and advantages compared to currently used methods in the postgenomics era. Cell metabolomics is an emerging field that addresses fundamental biological questions and allows one to observe metabolic phenomena in cells. Cell metabolomics consists of four sequential steps: (a) sample preparation and extraction, (b) metabolic profiles of low-weight metabolites based on MS or NMR spectroscopy techniques, (c) pattern recognition approaches and bioinformatics data analysis, (d) metabolites identification resulting in putative biomarkers and molecular targets. The biomarkers are eventually placed in metabolic networks to provide insight on the cellular biochemical phenomena. This article analyzes the recent developments in use of metabolomics to characterize and interpret the cellular metabolome in a wide range of pathophysiological and clinical contexts, and the putative roles of the endogenous small molecule metabolites in this new frontier of postgenomics biology and systems medicine.
Metabolomics is comprehensive analysis of low-molecular-weight endogenous metabolites in a biological sample. It could enable mapping of perturbations of early biochemical changes in diseases and hence provide an opportunity to develop predictive biomarkers that could provide valuable insights into the mechanisms of diseases. The aim of this study was to elucidate the changes in endogenous metabolites and to phenotype the metabolic profiling of d-galactosamine (GalN)-inducing acute hepatitis in rats by UPLC-ESI MS.
Zhibai dihuang pill is a famous traditional Chinese medicine formula containing the herbal medicine Phellodendri amurensis cortex. In this work, an integrative pattern recognition approach including principal component analysis and orthogonal partial least squared discriminant analysis based on ultra-performance liquid chromatography coupled to MS was successfully applied for the rapid discovery of natural compounds from herbal medicines. In a 24 min analysis, 93 compounds were identified or tentatively characterized from Zhibai dihuang pill based on their fragmentation behaviors, 23 of which were from Phellodendri amurensis cortex. A total of 26 metabolites in plasma were identified from Phellodendri amurensis cortex in Zhibai dihuang pill formula, among them, 12 prototypes and 14 metabolites through metabolic pathways of demethylation, methylation, hydrolysis, sulfate conjugation, and glucuronide conjugation, which were seven metabolites more than that of the single drug, suggesting the importance of the compatibility of traditional Chinese medicine. The present study provided important structural information on the metabolism of Zhibai dihuang pill. Furthermore, the results of this work have demonstrated the feasibility of ultra-performance liquid chromatography coupled to MS for the rapid and reliable characterization of metabolites from herbal medicines. Based on these results, this method could be a novel approach for identifying potentially bioactive components in other traditional Chinese medicines.
Pediatric primary nephrotic syndrome (PNS) is a chronic disease promoted by metabolic and immune dysfunctions. Peroxisome proliferator-activated receptor (PPAR) polymorphisms have been associated with a variety of metabolic and kidney disorders. We therefore hypothesized that PPAR polymorphisms might be involved in the pathophysiology of PNS. We compared the distributions of the PPAR-? Pro12Ala and Val290Met, PPAR-? coactivator-? (PGC-1?) Gly482Ser, and PPAR-? Leu162Val single nucleotide polymorphisms (SNPs) between children with PNS and normal controls and analyzed their correlations with clinical and metabolic indicators and steroid responsiveness. There were no significant differences in distributions of any of the polymorphisms between PNS cases and controls. However, PNS patients with the PPAR-? (Pro12Ala) PP genotype had significantly higher fasting serum insulin, IgA, and HOMA-IR levels and lower insulin sensitivity than did patients with PA and AA genotypes. Additionally, the PGC-1? (Gly482Ser) A allele was associated with lower CD8+ T-cell counts and higher triglyceride and complement C3 levels compared with the G allele. No polymorphisms were related to hormone sensitivity. These results suggest that the PPAR-? (Pro12Ala) and PGC-1? (Gly482Ser) SNPs may influence insulin and triglyceride metabolism in children with PNS and may thus be relevant to the prognosis of this chronic condition.
We previously showed that mitochondrial dysfunction (MtD) is involved in an aldosterone (Aldo)-induced podocyte injury. Here, the potential role of MtD in the initiation of podocyte damage was investigated. We detected the dynamic changes of urinary protein, urinary F2-isoprostane and renal malondialdehyde levels, kidney ultrastructure morphology, mitochondrial DNA (mtDNA) copy number, mitochondrial membrane potential (??m), and nephrin and podocin expressions in Aldo-infused mice. Aldo infusion first induced renal oxidative stress, as evidenced by increased levels of urinary F2-isoprostane and renal malondialdehyde, and MtD, as demonstrated by reduced mtDNA, ??m, and ATP production. Later, at 5 days after Aldo infusion, proteinuria and podocyte injury began to appear. In cultured podocytes, Aldo or hydrogen peroxide (H2O2) induced MtD after 2-8 h of treatment, whereas the podocyte damage, as shown by decreased nephrin and podocin expressions, occurred later after 12 h of treatment. Thus Aldo treatment both in vitro and in vivo indicated that MtD occurred before podocyte damage. Additionally, MtDNA depletion by ethidium bromide or mitochondrial transcription factor A (TFAM) RNAi induced MtD, further promoting podocyte damage. TFAM expression was found to be reduced in Aldo-infused mice and Aldo-treated podocytes. Adenoviral vector-mediated overexpression of TFAM prevented Aldo-induced MtD and protected against podocyte injury. Together, these findings support MtD as an early event in podocyte injury, and manipulation of TFAM may be a novel strategy for treatment of glomerular diseases such as podocytopathy.
Kidney diseases (KD), a major public health problem that affects about 10 % of the general population, manifest in progressive loss of renal function, which ultimately leads to complete kidney failure. However, current approaches based on renal histopathological results and clinical parameters lack sensitivity and are not sufficient to characterize the category and progression of nephrology or to predict nephrology progression risk reliably or to guide preventive interventions. The high incidence and financial burden of KD make it imperative to diagnose KD at early stages when therapeutic interventions are far more effective. Nowadays, the appearance of metabolomics (the high-throughput measurement and analysis of metabolites) has provided the framework for a comprehensive analysis of KD and serves as a starting point for generating novel molecular diagnostic tools for use in nephrology. Changes in the concentration profiles of a number of small-molecule metabolites found in either blood or urine can be used to localize kidney damage or assess kidneys suffering from injury. The power of metabolomics allows unparalleled opportunity to query the molecular mechanisms of KD. Novel metabolomics technologies have the ability to provide a deeper understanding of the disease beyond classical histopathology, redefine the characteristics of the disease state, and identify novel approaches to reduce renal failure. This review gives an overview of its application to important areas in clinical nephrology, with a particular focus on biomarker discovery. Great strides forward are being made in breaking down important barriers to the successful prevention and treatment of this devastating disorder.
Acupuncture has a history of over 3000 years and is a traditional Chinese medical therapy that uses hair-thin metal needles to puncture the skin at specific points on the body to promote wellbeing, while its molecular mechanism and ideal biological pathways are still not clear. High-throughput metabolomics is the global assessment of endogenous metabolites within a biologic system and can potentially provide a more accurate snap shot of the actual physiological state. We hypothesize that acupuncture-treated human would produce unique characterization of metabolic phenotypes. In this study, UPLC/ESI-HDMS coupled with pattern recognition methods and system analysis were carried out to investigate the mechanism and metabolite biomarkers for acupuncture treatment at "Zusanli" acupoint (ST-36) as a case study. The top 5 canonical pathways including alpha-linolenic acid metabolism, d-glutamine and d-glutamate metabolism, citrate cycle, alanine, aspartate, and glutamate metabolism, and vitamin B6 metabolism pathways were acutely perturbed, and 53 differential metabolites were identified by chemical profiling and may be useful to clarify the physiological basis and mechanism of ST-36. More importantly, network construction has led to the integration of metabolites associated with the multiple perturbation pathways. Urine metabolic profiling might be a promising method to investigate the molecular mechanism of acupuncture.
Traditional Chinese medicine (TCM) formula has been playing a very important role in health protection and disease control for thousands of years. Guided by TCM syndrome theories, formula are designed to contain a combination of various kinds of crude drugs that, when combined, will achieve synergistic efficacy. However, the precise mechanism of synergistic action remains poorly understood. One example is a famous TCM formula Yinchenhao Tang (YCHT), whose efficacy in treating hepatic injury (HI) and Jaundice syndrome, has recently been well established as a case study. We also conducted a systematic analysis of synergistic effects of the principal compound using biochemistry, pharmacokinetics and systems biology, to explore the key molecular mechanisms. We had found that the three component (6,7-dimethylesculetin (D), geniposide (G), and rhein (R)) combination exerts a more robust synergistic effect than any one or two of the three individual compounds by hitting multiple targets. They can regulate molecular networks through activating both intrinsic and extrinsic pathways to synergistically cause intensified therapeutic effects. This paper provides an overview of the recent and potential developments of chemical fingerprinting coupled with systems biology advancing drug discovery towards more agile development of targeted combination therapies for the YCHT.
To discover and screen the constituents or metabolites absorbed into blood after oral administration of herbal medicines tends to be more and more difficult. In this work, an integrative pattern recognition approach of principal component analysis (PCA) and orthogonal partial least squared discriminant analysis (OPLS-DA) was successfully applied for rapid discovery of natural compounds from herbal medicines. A rapid, sensitive, and reliable ultra performance liquid chromatography coupled with electrospray ionization/quadrupole-time-of-flight mass spectrometry (UPLC-ESI-Q-TOF-MS) method with Masslynx™ software was established to characterize the chemical constituents and rats metabolites of Phellodendri amurensis cortex (Guan Huangbai, GHB). The analysis was performed on a Waters UPLC HSS T3 column (2.1 × 100 mm, 1.8 ?m) using gradient elution system. A hyphenated electrospray ionization and quadrupole-time-of-flight analyzer was used for the determination of accurate mass of the protonated or deprotonated molecule and fragment ion in both negative and positive modes. A total of 46 peaks were obtained, 41 of which were tentatively characterized from GHB. In the S-plot of OPLS-DA, 24 interested ions (17 ions in positive mode and 6 ions in negative mode) were extracted, among them, 12 absorbed prototype components of GHB and 12 metabolites were identified in vivo. Major metabolic reactions of GHB were demethylation, methylation and glucuronidation. This is the first report on systematic analysis of chemical constituents and in vivo metabolites of GHB. It is concluded that UPLC-MS coupled with pattern recognition approach for the identification of herbal constituents in biological samples has been successfully developed. The method can also be applied to rapid discovery and global characterization of the constituents in rat serum after oral administration of other herbal medicines.
Schisandra chinensis Baill grows wild in Russia, China, Korea and Japan, and its fruit has been found to be effective in amnesia and insomnia. It is enriched in schisandra lignans (SL) that are major components responsible for therapeutic action. However, there are no reports on the biotransformation analysis of SL. An ultra-performance liquid chromatography/electrospray-ionization high-definition mass spectrometry (UPLC-Q-TOF-HDMS) method was developed to investigate the metabolism of SL in vivo. MS was performed on a Waters Micromass high-definition system with an electrospray ionization source in positive ion mode and automated MetaboLynx software analysis with excellent MS accuracy and enhanced MS data acquisition. An improved mass defect filter (MDF) method employing both drug and core structure filter templates was applied to the processing of UPLC-Q-TOF-HDMS data for the detection and structural characterization of metabolites. In this study, 30 metabolites were detected and identified in vivo, and demethylation and hydroxylation were confirmed as the primacy metabolic pathway for SL in rat plasma. In conclusion, the presently developed methodology was suitable for biotransformation research of SL and will find wide use in metabolic studies for other herbal medicines.
Denys-Drash syndrome (DDS) is a rare disorder characterized by nephropathy, male pseudohermaphroditism, and wilms tumor. Cases are thought to arise sporadically through a de novo mutation in the wilms tumor suppressor gene (WT1), which encodes a zinc finger protein that not only acts as a tumor suppressor but is essential for normal gonadogenesis, nephrogenesis, and development of the urogenital tract. In this report, we describe a family with the well-known missense mutation in exon 9 of the WT1 gene, 1180C>T (R394W), causing incomplete DDS and no symptoms in their father. The proband, a boy with 46, XY karyotype, was born with ambiguous genitalia, penoscrotal hypospadias, and bilateral inguinal hernias. At 2 years of age, he has proteinuria and diffuse mesangial sclerosis, but no wilms tumor has been detected. The elder sister of the proband, at 3 years of age, has normal genitalia, proteinuria, focal mesangial sclerosis but no wilms tumor. The WT1 mutation was detected in both patients, who have suspected DDS, and their father, who is phenotypically normal.
Abstract Scoparone is an active ingredient of Yinchenhao (Artemisia annua L.), a well-known Chinese medicinal plant, and has been utilized in prevention and therapy of liver damage. However, the molecular drug targets associated with the pharmacological effects of scoparone are largely unknown. In the present article, we extend the previous research on Yinchenhao through a study of its active ingredient and thus the putative targets of scoparone. We employed two-dimensional gel electrophoresis, and all proteins expressed were identified by MALDI-TOF/TOF MS and database research. Protein-interacting networks and pathways were also mapped and evaluated. The possible protein network associated with scoparone was constructed, and contribution of these proteins to the protective effect of scoparone against the carbon tetrachloride-induced acute liver injury in rats are discussed herein. Hepatoprotective effects of scoparone on liver injury in rats were associated with regulated expression of six proteins which were closely related in our protein-protein interaction network, and appear to be involved in antioxidation and signal transduction, energy production, immunity, metabolism, and chaperoning. These observations collectively provide new insights on the molecular mechanisms of scoparone action against hepatic damage in rats.
Adriamycin (ADR) administration in susceptible rodents such as the BALB/c mouse strain produces injury to the glomerulus mimicking human focal glomerular sclerosis. The goal of the present study was to use this model to investigate antiproteinuric action of nitro-oleic acid (OA-NO2), a nitric oxide-derived endogenous lipid product, which has exhibited multiple attractive signaling properties particularly in the kidney. BALB/c mice were pretreated for 2 days with OA-NO2 at 5 mg·kg(-1)·day(-1) via an osmotic minipump, followed by a single injection of vehicle or adriamycin (10 mg/kg) via the tail vein. Albuminuria and renal function were analyzed at 1 wk post-ADR treatment. ADR mice developed prominent albuminuria, hypoalbuminemia, hyperlipidemia, and severe ascites. In contrast, the symptoms of nephrotic syndrome were greatly improved by OA-NO2 treatment. In parallel, plasma creatinine and plasma urea nitrogen were elevated in the ADR group, and the severity was less in the ADR+OA-NO2 group. OA-NO2 attenuates ADR-induced glomerulosclerosis, podocyte loss, and tubulointerstitial fibrosis. Indices of oxidative stress, including plasma and urinary thiobarbituric acid-reactive substances and renal expression of NAD(P)H oxidase p47(phox) and gp91(phox), and inflammation, including renal expression of TNF-?, IL-1?, and MCP-1 in response to ADR, were all similarly suppressed. Together, these findings suggest that OA-NO2 exerts renoprotective action against ADR nephropathy likely via its anti-inflammatory and antioxidant properties.
Liver disease is any condition that may cause liver inflammation or tissue damage and affects liver function. Natural products that are found in vegetables, fruits, plant extracts, herbs, insects, and animals, have been traditionally used for treating liver diseases. They are chemical compounds that usually have biological activities for use in drug discovery and design. Many natural products have been clinically available as potent hepatoprotective agents against commonly occurring liver diseases. This review summarizes the current progress in the basic, clinical, and translational research on natural products in treatment of various liver diseases. Furthermore, we will focus on the discovery and biological evaluation of the natural products, which shows potential as a new therapeutic agent of liver diseases.
Hepatocellular carcinoma (HCC) is the commonest primary hepatic malignancy and the third most common cause of cancer-related death worldwide. Incidence remains highest in the developing world and is steadily increasing across the developed world. Current diagnostic modalities, of ultrasound and ?-fetoprotein, are expensive and lack sensitivity in tumor detection. Because of its asymptomatic nature, HCC is usually diagnosed at late and advanced stages, for which there are no effective therapies. Thus, biomarkers for early detection and molecular targets for treating HCC are urgently needed. Emerging high-throughput metabolomics technologies have been widely applied, aiming at the discovery of candidate biomarkers for cancer staging, prediction of recurrence and prognosis, and treatment selection. Metabolic profiles, which are affected by many physiological and pathological processes, may provide further insight into the metabolic consequences of this severe liver disease. Small-molecule metabolites have an important role in biological systems and represent attractive candidates to understand HCC phenotypes. The power of metabolomics allows an unparalleled opportunity to query the molecular mechanisms of HCC. This technique-driven review aims to demystify the metabolomics pathway, while also illustrating the potential of this technique, with recent examples of its application in HCC.
To enhance the therapeutic efficacy and reduce the adverse effects of traditional Chinese medicine, practitioners often prescribe combinations of plant species and/or minerals, called formulae. Unfortunately, the working mechanisms of most of these compounds are difficult to determine and thus remain unknown. In an attempt to address the benefits of formulae based on current biomedical approaches, we analyzed the components of Yinchenhao Tang, a classical formula that has been shown to be clinically effective for treating hepatic injury syndrome. The three principal components of Yinchenhao Tang are Artemisia annua L., Gardenia jasminoids Ellis, and Rheum Palmatum L., whose major active ingredients are 6,7-dimethylesculetin (D), geniposide (G), and rhein (R), respectively. To determine the mechanisms underlying the efficacy of this formula, we conducted a systematic analysis of the therapeutic effects of the DGR compound using immunohistochemistry, biochemistry, metabolomics, and proteomics. Here, we report that the DGR combination exerts a more robust therapeutic effect than any one or two of the three individual compounds by hitting multiple targets in a rat model of hepatic injury. Thus, DGR synergistically causes intensified dynamic changes in metabolic biomarkers, regulates molecular networks through target proteins, has a synergistic/additive effect, and activates both intrinsic and extrinsic pathways.
For World Health Organization proposed Health for All, we should be to promote the use of traditional medicine. Traditional Chinese acupuncture has a history over 3000 years and is effective in the treatment of many conditions with few side effects. Acupuncture, an intrinsic part of traditional Chinese medicine (TCM), which utilizes fine needles to pierce through specific anatomical points (called "acupoints"), has been extensively used and has emerged as an important modality of complementary and alternative therapy to Western medicine. It is vital and necessary to explore the underlying biological mechanisms of acupuncture. Systems biology has become practically available and resembles acupuncture in many aspects and is current key technology that serves as the major driving force for translation of acupuncture medicine revolution into practice, will advance acupuncture therapy into healthcare for individuals. High-throughput genomics, proteomics and metabolomics in the context of systems biology have been able to identify potential candidates for the effects of acupuncture and provide valuable information toward understanding mechanisms of the therapy. To realize the full potential of TCM acupuncture, we describe the current status of principles and practice of acupuncture integrated with systems biology platform in the post-genomic era. Some characteristic examples are presented to highlight the application of this platform in omics and systems biology approaches to acupuncture research and some of the necessary milestones for moving acupuncture into mainstream health care.
The SNAP-tag labeling technology provides a simple, robust, and versatile approach to the imaging of fusion proteins for a wide range of experimental applications. Owing to the specific and covalent nature of the labeling reaction, SNAP-tag is well suited for the analysis and quantification of fused target protein using fluorescence microscopy techniques. In this report, we present our most recent findings on the labeling of SNAP-tag fusion proteins both in vitro and in cell culture with SNAP-tag substrates derived from single regioisomers of carboxyrhodamine dyes. Carboxyrhodamines are invaluable fluorescent dyes for biotechnology applications including DNA sequencing, detection on microarrays, and fluorescence in situ hybridization. We found that SNAP-tag reacts preferentially with the 6-positional regioisomer of carboxyrhodamine fluorescent dyes, whereas the 5-regioisomer predominantly contributes to background fluorescence. Our experimental study also indicates that benzylchloropyrimidine (CP) conjugates of 6-carboxyrhodamines exhibit a dramatic increase in the signal-to-noise ratio of fluorescently labeled cellular proteins compared to the benzylguanine (BG) conjugates, presumably due to higher cell permeability. These new SNAP-tag substrates based on pure 6-regioisomers can significantly improve fluorescence labeling in live cells and should become powerful tools for bioimaging applications.
Shaoyao-Gancao decoction (SGD), a traditional Chinese formulae containing Paeoniae Radix and Glycyrrhizae Radix, is commonly used to relieve abdominal pain. It has attracted increasingly much attention as one of the most popular and valuable herbal medicine in clinic. However, the systematic analysis of chemical constituents of SGD are difficult to determine and thus remain unclear. In this paper, a rapid, sensitive, and reliable ultra-performance LC-ESI/quadrupole-TOF high-definition MS (UPLC-ESI-Q-TOF-MS) with automated MetaboLynx analysis in negative ion mode were established to characterize the chemical constituents of SGD. The analysis was performed on a Waters UPLC(TM) HSS T3 (2.1 × 100 mm, 1.8 ?m) using gradient elution system. MS/MS fragmentation behavior was proposed for aiding the structural identification of the components. With the optimized conditions, a total of 58 peaks were tentatively characterized by comparing the retention time and mass spectrometry data and retrieving the reference literatures. Of note, 44 ingredients were identified from Glycyrrhizae Radix, and 14 were from Paeoniae Radix. It is concluded that a rapid and robust platform based on UPLC-ESI-Q-TOF-MS was successfully developed for globally identifying multiple-constituent of traditional Chinese medicine prescriptions. This is the first report on systematic analysis of chemical constituents and in vivo metabolites of SGD.
Shengmaisan (SMS) is a traditional Chinese medicine prescription widely used for the treatment of cardiovascular diseases in Asia. Its lignans are major components responsible for therapeutic action. A rapid and specific UPLC-Q-TOF/MS has been developed and validated for simultaneous quantification of the five main bioactive components, i.e. schisandrin, schisandrol B, schisantherin A, deoxyschisandrin, and schisandrin B, in rat plasma after oral administration of SMS. All calibration curves showed excellent linearity within the test ranges. Validation proved the repeatability of the method was good and recovery was satisfactory. The separation of these compounds was carried out on a Waters ACQUITY HSS T(3) column (2.1 × 100 mm, 1.8 ?m) by linear gradient elution using a mobile phase consisting of 0.01% formic acid in water and ACN containing 0.01% formic acid. In this work, plasma pharmacokinetic characteristics of lignans components after oral administration SMS were investigated using UPLC-Q-TOF/MS method. MS was performed on a Waters Micromass high-definition technology with an ESI source. Data were analyzed and estimated by compartmental methods and pharmacokinetic parameters calculated using WinNonlin Professional version 6.1. Results demonstrated that the proposed UPLC-Q-TOF/MS method was successfully applied to pharmacokinetic study of all components in rat plasma after oral administration of the SMS.
Geniposide is an important constituent of Gardenia jasminoides Ellis, a famous Chinese medicinal plant, and has displayed bright prospects in prevention and therapy of hepatic injury (HI). Unfortunately, the working mechanisms of this compound are difficult to determine and thus remain unknown. To determine the mechanisms that underlie this compound, we conducted a systematic analysis of the therapeutic effects of geniposide using biochemistry, metabolomics and proteomics. Geniposide significantly intensified the therapeutic efficacy as indicated by our modern biochemical analysis. Metabolomics results indicate 9 ions in the positive mode as differentiating metabolites which were associated with perturbations in primary bile acid biosynthesis, butanoate metabolism, citrate cycle (TCA cycle), alanine, aspartate and glutamate metabolism. Of note, geniposide has potential pharmacological effect through regulating multiple perturbed pathways to normal state. In an attempt to address the benefits of geniposide based on the proteomics approaches, the protein-interacting networks were constructed to aid identifying the drug targets of geniposide. Six identified differential proteins appear to be involved in antioxidation and signal transduction, energy production, immunity, metabolism, chaperoning. These proteins were closely related in the protein-protein interaction network and the modulation of multiple vital physiological pathways. These data will help to understand the molecular therapeutic mechanisms of geniposide on hepatic damage rats. We also conclude that metabolomics and proteomics are powerful and versatile tools for both biomarker discovery and exploring the complex relationships between biological pathways and drug response, highlighting insights into drug discovery.
Hepatitis B virus (HBV) is the fatal consequence of chronic hepatitis, and lack of biomarkers has been a long standing bottleneck in the clinical diagnosis. Metabolomics concerns with comprehensive analysis of small molecules and provides a powerful approach to discover biomarkers in biological systems. Here, we present metabolomics analysis applying ultra-performance liquid chromatography/electrospray ionization quadruple time-of-flight mass spectrometry. (UPLC-Q-TOF-HDMS) to determine metabolite alterations in HBV patients. Most important permutations are elaborated using multivariate statistical analysis and network analysis that was used to select the metabolites for the noninvasive diagnosis of HBV. In this study, the total 11 urinary differential metabolites were identified and contributed to HBV progress involving several key metabolic pathways by using pathway analysis with MetPA, which are promising biomarker candidates for diagnostic research. More importantly, of 11 altered metabolites, 4 metabolite markers were effective for the diagnosis of human HBV, achieved a satisfactory accuracy, sensitivity and specificity, respectively. It demonstrates that metabolomics has the potential as a non-invasive tool to evaluate the potential of these metabolites in the early diagnosis of HBV patients. These findings may be promising to yield a valuable insight into the pathophysiology of HBV and to advance the approaches of diagnosis, treatment, and prevention.
Metabolomics represents an emerging and powerful discipline that provides an accurate and dynamic picture of the phenotype of biosystems through the study of potential metabolites that could be used for therapeutic targets and discovery of new drugs. Metabolomic network construction has led to the integration of metabolites associated with the caused perturbation of multiple pathways. Herein, we present a method for the construction of efficient networks with regard to that Jujuboside B (JuB) protects against insomnia as a case study. UPLC/ESI-SYNAPT-HDMS coupled with pattern recognition methods including PCA, PLS-DA, OPLS-DA, and computational systems analysis were integrated to obtain comprehensive metabolomic profiling and pathways of the large biological data sets. Among the regulated pathways, twelve biomarkers were identified and tryptophan metabolism, phenylalanine, tyrosine, tryptophan biosynthesis, arachidonic acid metabolism, and phenylalanine metabolism related network were acutely perturbed. Results not only supplied a systematic view of the development and progression of insomnia but also were used to analyze the therapeutic effects of JuB, a widely used anti-insomina medicine in clinics. The results showed that JuB administration could provide satisfactory effects on insomnia through partially regulating the perturbed pathway. We have constructed a metabolomic feature network of JuB to protect against insomnia. The most promising use in the near future would be to clarify pathways for the drugs and get biomarkers for these pathways, to help guide testable predictions, provide insights into drug action mechanisms, and enable us to increase research productivity toward metabolomic drug discovery.
The mother and lateral root of Aconitum carmichaelii Debx, named "Chuanwu" (CW) and "Fuzi", respectively, has been used to relieve joint pain and treat rheumatic diseases for over 2000 years. However, it has a very narrow therapeutic range, and the toxicological risk of its usage remains very high. The traditional Chinese processing approach, Paozhi (detoxifying measure),can decompose poisonous Aconitum alkaloids into less or nontoxic derivatives and plays an important role in detoxification. The difference in metabolomic characters among the crude and processed preparations is still unclear, limited by the lack of sensitive and reliable biomarkers. Therefore, this paper was designed to investigate comprehensive metabolomic characters of the crude and its processed products by UPLC-Q-TOF-HDMS combined with pattern recognition methods and ingenuity pathway analysis (IPA). The significant difference in metabolic profiles and changes of metabolite biomarkers of interest between the crude and processed preparations were well observed. The underlying regulations of Paozhi-perturbed metabolic pathways are discussed according to the identified metabolites, and four metabolic pathways are identified using IPA. The present study demonstrates that metabolomic analysis could greatly facilitate and provide useful information to further comprehensively understand the pharmacological activity and potential toxicity of processed Aconite roots in the clinic.
A visual display terminal (VDT) mental fatigue task was designed to search for the objective indicator to diagnose and evaluate the VDT mental fatigue status. The signals of temperature and electrocardiograph (ECG) are collected from the subjects. The temperature, heart rate (HR) and heart rate variability (HRV) are extracted as the objective parameters after analyzing the temperature signals in time domain and the ECG signals in time domain and frequency domain. Compared with mental fatigue pre-experiment, the temperature increased significantly (P<0.001), and the RR-PNN50 increased obviously (P<0.05), while the S wavelet of ECG signals decreased obviously (P<0.05) after mental fatigue experiment. Compared with the end of mental fatigue experiment, the temperature increased, the changes of RR-PNN50 were not obvious, the S wavelet increased significantly (P<0.01), and the mental fatigue was somewhat remittance after natural recovery. The results showed that the objective indicator to evaluate the VDT mental fatigue status would be hopeful to be found out by analyzing the signals of temperature and ECG.
Metabolomics is the comprehensive assessment of endogenous metabolites and attempts to systematically identify and quantify metabolites from a biological sample. Small-molecule metabolites have an important role in biological systems and represent attractive candidates to understand disease phenotypes. Metabolites represent a diverse group of low-molecular-weight structures including lipids, amino acids, peptides, nucleic acids, organic acids, vitamins, thiols and carbohydrates, which makes global analysis a difficult challenge. The recent rapid development of a range of analytical platforms, including GC, HPLC, UPLC, CE coupled to MS and NMR spectroscopy, could enable separation, detection, characterization and quantification of such metabolites and related metabolic pathways. Owing to the complexity of the metabolome and the diverse properties of metabolites, no single analytical platform can be applied to detect all metabolites in a biological sample. The combined use of modern instrumental analytical approaches has unravelled the ideal outcomes in metabolomics, and is beneficial to increase the coverage of detected metabolites that can not be achieved by single-analysis techniques. Integrated platforms have been frequently used to provide sensitive and reliable detection of thousands of metabolites in a biofluid sample. Continued development of these analytical platforms will accelerate widespread use and integration of metabolomics into systems biology. Here, the application of each hyphenated technique is discussed and its strengths and limitations are discussed with selected illustrative examples; furthermore, this review comprehensively highlights the role of integrated tools in metabolomic research.
The "open" and "closed" isomers of the diarylethene molecule that can be converted between each other upon photo-excitation are found to have drastically different current-voltage characteristics when sandwiched between two graphene nanoribbons (GNRs). More importantly, when one GNR is metallic and another one is semiconducting, strong rectification behavior of the "closed" diarylethene isomer with the rectification ratio >10(3) is observed. The surprisingly high rectification ratio originates from the band gap of GNR and the bias-dependent variation of the lowest unoccupied molecular orbital of the diarylethene molecule, the combination of which completely shuts off the current at positive biases. Results presented in this paper may form the basis for a new class of molecular electronic devices.
The lateral root of Aconitum carmichaelii Debx is named "Fuzi" which is widely distributed across Asia and North America and has been used to relieve joint pain and treat rheumatic diseases for over two thousand years. However, it has very narrow therapeutic ranges and despite the toxicological risk, its usage remains very high. A traditional Chinese processing approach (Paozhi, detoxifying measure) is necessary to remove the poisonous Aconitum alkaloids mainly deriving from the diester diterpene alkaloids (DDAs) including aconitine, mesaconitine and hypaconitine. They can be decomposed into less or non-toxic derivatives through Paozhi that plays an essential role in detoxification. Processed Fuzi is mainly focused on the three main forms of Yanfuzi (YFZ), Heishunpian (HSP) and Baifupian (BFP) which are highly desirable in order to guarantee the clinical safety and their low toxicity in decoctions. The difference in metabolomic characters between Fuzi and its processed preparations is still completely unclear. Therefore, this paper was designed to investigate a comprehensive metabolome of Fuzi and its processed products by ultra-performance liquid-chromatography/electrospray-ionization synapt high-definition mass spectrometry (UPLC-Q-TOF-HDMS) combined with pattern recognition methods. The difference in metabolic profiles between Fuzi and its processed preparations was well observed by the principal component analysis (PCA) of the MS spectra. Significant changes of 19 metabolite biomarkers were detected in the Fuzi samples and three preparations. The underlying regulations of Paozhi-perturbed metabolic pathways were also discussed according to the identified metabolites. The present study proves that UPLC-Q-TOF-HDMS based metabolomic analysis greatly contributes to the investigation of Fuzi metabolism through Paozhi techniques, and provides useful information to further comprehensively understand the pharmacological activity and potential toxicity of processed Fuzi in a clinical environment.
Traditional Chinese medicine (TCM) has been widely used in many oriental countries for thousands of years and played an indispensable role in the prevention and treatment of diseases, especially the complicated and chronic ones. It is a very complex mixture containing hundreds or thousands of different components. Pharmacokinetic study on active constituents in TCM preparations is a good way for us to explain and predict a variety of events related to the efficacy and toxicity of TCM. A selective and sensitive method of ultra performance liquid chromatography coupled with electrospray ionization/quadrupole-time-of-flight mass spectrometry (UPLC-ESI-Q-TOF-MS/MS) was first developed to screen the potentially bioactive components in vivo, using the semi-quantitative determination of multicomponents in the rat plasma after a single oral administration of Yin-Chen-Hao-Tang (YCHT), a famous TCM formula for liver disorders. Hierarchical cluster analysis (HCA) and principal component analysis (PCA) were built to evaluate the differences of pharmacokinetic behaviors (time-course) of the absorbed components of YCHT. Here, we report that the developed method was successfully applied to monitoring the pharmacokinetic time-course of 21 compounds in rat plasma, and were grouped in 3 separate clusters using pattern recognition approaches (both HCA and PCA). Comparing the body dynamics of each composition, the initial choice of the following 9 compounds as the candidate components was: 7-methoxycoumarin-6-hydroxyl sulfate, genipingentiobioside, geniposide, 6,7-dimethylesculetin, peak 16, chimaphylin, 6-dementhoxycapillarisin, capillarisin, rhein. Pharmacokinetics based-UPLC-ESI-Q-TOF-MS/MS combined with HCA and PCA approaches can provide a reliable and suitable means of identifying and screening potentially bioactive components contributing to pharmacological effects of TCM, further prospecting natural products in the search for new leads in drug discovery.
Potential metabolites from the metabolic pathways could be therapeutic targets and useful for the discovery of broad spectrum drugs. UPLC/ESI-SYNAPT-HDMS coupled with pattern recognition methods including PCA, PLS-DA, OPLS-DA and Heatmap were integrated to examine the global metabolic signature of insomnia and intervention effects of Jujuboside A (JuA). Six unique pathways of the insomnia were identified using Ingenuity Pathway Analysis (IPA) software. The VIP-value threshold cutoff of the metabolites was set to 10, above this threshold, were filtered out as potential target biomarkers. Sixteen distinct metabolites were identified from these pathways, and 6 of them can be considered for rational drug design. It was further experimental validation that the changes in metabolic profiling were restored to their baseline values after JuA treatment according to the multivariate data analysis. Potential metabolite network of the insomnia was preliminarily predicted JuA-target interaction networks, and could be further explored for in silico docking studies with suitable drugs. Thus, our method is an efficient procedure for drug target identification through metabolic analysis. It can guide testable predictions, provide insights into drug action mechanisms and enable us to increase research productivity toward metabolomic drug discovery.
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