Hepatic Effect of 2,3,5,4'-tetrahydroxystilbene-2-O-ß-D-glucoside, the Signature Component of Traditional Chinese Medicine Heshouwu: Advances and Prospects.

Traditional Chinese medicine Heshouwu, named Polygoni Multiflori Radix in Pharmacopoeia of the People's Republic of China (PPRC, 2020), is derived from the root tuber of Polygonum multiflorum Thunb., Heshouwu or processed Heshouwu is well known for its function in reducing lipids and nourishing the liver. However, increasing cases of Heshouwu-induced hepatotoxicity were reported in recent years. Researchers have begun to study the paradoxical effects of Heshouwu on the liver. 2,3,5,4'-tetrahydroxystilbene-2-O-ß-D-glucoside (TSG), an abundant functional component of Heshouwu, shows various biological activities, among which its effect on the liver is worthy of attention. This paper reviews the current studies of TSG on hepatoprotection and hepatotoxicity, and summarizes the doses, experimental models, effects, and mechanisms of action involved in TSG's hepatoprotection and hepatotoxicity, aiming to provide insight for future study of TSG and understanding the effects of Heshouwu on the liver. Emerging evidence suggests that TSG ameliorates both pathological liver injury and chemical-induced liver injury by modulating lipid metabolism, inhibiting the inflammatory response and oxidative stress in the liver. However, with the reports of clinical cases of Heshouwu induced liver injury, it has been found that long-term exposure to a high dose of TSG cause hepatocyte or hepatic tissue damage. Moreover, TSG may cause hepatotoxicity by affecting the transport and metabolism of other possible hepatoxic compounds in Heshouwu. Studies indicate that trans-TSG can be isomerized into cis-TSG under illumination, and cis-TSG had a less detrimental dose to liver function than trans- TSG in LPS-treated rats. In brief, TSG has protective effects on the liver, but liver injury usually occurs under highdose TSG or is idiosyncratic TSG-induced liver injury.

Potential biomarkers screening of Polygonum multiflorum radix-induced liver injury based on metabonomics analysis of clinical samples.

ETHNOPHARMACOLOGICAL RELEVANCE: Polygonum multiflorum Radix (PMR) is the dried root tuber of Polygonum multiflorum Thunb., which has been used in the clinic for a variety of pharmacological activities. However, Polygonum multiflorum Radix-induced liver injury (PMR-ILI) has been reported in recent years, which has limited its clinical use to some extent. The occurrence of PMR-ILI is not universal, so finding the different metabolic characteristics between PMR-ILI and Polygonum multiflorum Radix-tolerance group (PMR-T) is very important for the PMR rational clinical application and PMR-ILI early clinical diagnosis. METHODS: In this study, 6 clinical plasma samples of PMR-ILI and 13 PMR-T were collected and analyzed by high-resolution liquid chromatography-mass spectrometry. Firstly, the differential metabolites of the two groups were screened by conventional screening methods such as multivariate statistical analysis. Secondly, the characteristic metabolites with greater contribution, correlation with liver injury and high sensitivity were screened by correlation analysis with clinical liver injury indicators, random forest analysis, and receiver operating characteristic curve (ROC). RESULTS: After multivariate statistical analysis and screening analysis, 29 differential metabolites were identified. Compared with PMR-T group, the metabolism of glycerol and phospholipid, glutamine and glutamate, phenylalanine, sphingolipid and tryptophan in PMR-ILI group were disturbed. After correlation analysis with liver injury indexes and random forest screening, 8 potential biomarkers closely related to clinical liver injury were obtained. Finally, 3 potential biomarkers with high expression in PMR-ILI, hypoxanthine, LysoPC(P-16:0/0:0) and taurochenodesoxycholic acid, were screened out through the analysis of ROC, which can provide a basis for the early clinical diagnosis. CONCLUSION: Based on the analysis of the PMR-ILI and PMR-T plasma samples by LC-MS, three biomarkers of clinical liver injury of Polygonum multiflorum Radix were selected: hypoxanthine, LysoPC(P-16:0/0:0) and taurochenodeoxycholic acid.

Inhibition of CYP3A4 enhances aloe-emodin induced hepatocyte injury.

Aloe-emodin (AE) is a natural hydroxyanthraquinone derivative that was found in many medicinal plants and ethnic medicines. AE showed a wide array of pharmacological activities including anticancer, antifungal, laxative, antiviral, and antibacterial effects. However, increasing number of published studies have shown that AE may have some hepatotoxicity effects but the mechanism is not fully understood. Studies have shown that the liver injury induced by some free hydroxyanthraquinone compounds is associated with the inhibition of some metabolic enzymes. In this study, the CYP3A4 and CYP3A1 were found to be the main metabolic enzymes of AE in human and rat liver microsomes respectively. And AE was metabolized by liver microsomes to produce hydroxyl metabolites and rhein. When CYP3A4 was knocked down in L02 and HepaRG cells, the cytotoxicity of AE was increased significantly. Furthermore, AE increased the rates of apoptosis of L02 and HepaRG cells, accompanied by Ca2+ elevation, mitochondrial membrane potential (MMP) loss and reactive oxygen species (ROS) overproduction. The mRNA expression of heme oxygenase-1 in L02 and HepaRG cells increased significantly in the high-dose of AE (40 µmol/L) group, and the mRNA expression of quinone oxidoreductase-1 was activated by AE in all concentrations. Taken together, the inhibition of CYP3A4 enhances the hepatocyte injury of AE. AE can induce mitochondrial injury and the imbalance of oxidative stress of hepatocytes, which results in hepatocyte apoptosis.

Exploration of components and mechanisms of Polygoni Multiflori Radix-induced hepatotoxicity using siRNA -mediated CYP3A4 or UGT1A1 knockdown liver cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Polygoni Multiflori Radix, the dried root of Polygonum multiflorum Thunb., and its processed products have been used as restoratives for centuries in China. However, the reports of Polygoni Multiflori Radix-induced liver injury (PMR-ILI) have received wide attention in recent years, and the components and mechanism of PMR-ILI are not completely clear yet. Our previous studies found that the PMR-ILI was related to the down-regulation of some drug metabolism enzymes (DME). AIM OF THE STUDY: To explore the effect of the inhibition of CYP3A4 or UGT1A1 on PMR-ILI, screen the relevant hepatotoxic components and unveil its mechanism. METHODS: RT-qPCR was used to detect the effects of water extract of Polygoni Multiflori Radix (PMR) and its main components on the mRNA expression of CYP3A4 and UGT1A1 in human hepatic parenchyma cell line L02. High-performance liquid chromatography (HPLC) was employed to detect the content of major components in the PMR. And then, the stable CYP3A4 or UGT1A1 knockdown cells were generated using short hairpin RNAs (shRNA) in L02 and HepaRG cells. Hepatotoxic components were identified by cell viability assay when PMR and its four representative components, 2,3,5,4'-tetrahydroxy stilbene glycoside (TSG), emodin (EM), emodin-8-O-ß-D-glucoside (EG), and gallic acid (GA), acted on CYP3A4 or UGT1A1 knockdown cell lines. The PMR-ILI mechanism of oxidative stress injury and apoptosis in L02 and HepaRG cells were detected by flow cytometry. Finally, the network toxicology prediction analysis was employed to excavate the targets of its possible toxic components and the influence on the metabolic pathway. RESULTS: PMR and EM significantly inhibited the mRNA expression of CYP3A4 and UGT1A1 in L02 cells, while TSG and GA activated the mRNA expression of CYP3A4 and UGT1A1, and EG activated CYP3A4 expression while inhibited UGT1A1 expression. The contents of TSG, EG, EM and GA were 34.93 mg/g, 1.39 mg/g, 0.43 mg/g and 0.44 mg/g, respectively. The CYP3A4 or UGT1A1 knockdown cells were successfully constructed in both L02 and HepaRG cells. Low expression of CYP3A4 or UGT1A1 increased PMR cytotoxicity remarkably. Same as PMR, the toxicity of EM and GA increased in shCYP3A4 and shUGT1A1 cells, which suggested EM and GA may be the main components of hepatotoxicity in PMR. Besides, EM not only inhibited the expression of metabolic enzymes but also reduced the cytotoxicity threshold. EM and GA affected the level of ROS, mitochondrial membrane potential, Ca2+ concentration, and dose-dependent induced hepatocyte apoptosis in L02 and HepaRG cells. The network toxicology analysis showed that PMR-ILI was related to drug metabolism-cytochrome P450, glutathione metabolism, and steroid hormone biosynthesis. CONCLUSION: The inhibition of mRNA expression of CYP3A4 or UGT1A1 enhanced hepatotoxicity of PMR. EM and GA, especially EM, may be the main hepatotoxic components in PMR. The mechanism of PMR, EM and GA induced hepatotoxicity was proved to be related to elevated levels of ROS, mitochondrial membrane potential, Ca2+ concentration, and induction of apoptosis in liver cells.

Astragali radix total flavonoid synergizes cisplatin to inhibit proliferation and enhances the chemosensitivity of laryngeal squamous cell carcinoma.

Laryngeal squamous cell carcinoma (LSCC) is the most common head and neck cancer. Astragali radix extracts play crucial roles in the regulation of cancer progression. However, the role of Astragali radix extracts in LSCC and the related mechanisms remains unclear. Here, we evaluated the inhibitory effects of the combined use of Astragali radix total flavonoid (TFA) and cisplatin (CDDP) on an LSCC mouse model by pharmacodynamics. Ultra-high-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) was employed to define the prototype of TFA in vivo. The potential drug targets were identified through the integrative analysis of LSCC microarrays, RNA sequencing data and the main bioactive component of TFA. Furthermore, a protein-protein interaction network, compound-target network and target-pathway network were constructed based on the prototype and potential drug targets to identify the main targets and pathways. Animal experiments showed that TFA has significant synergistic antitumor activity with cisplatin and attenuates the nephrotoxicity caused by CDDP chemotherapy, improving the survival of LSCC-bearing mice. Using UPLC-MS/MS, we identified 8 constituents of TFA in experimental mice serum: formononetin, ononin, calycosin, calycosin-7-O-ß-D-glucoside, 7,2'-dihydroxy-3',4'-dimethoxyisoflavan, 7,2'-dihydroxy-3',4'-dimethoxyisoflavaneglucoside, 3-hydroxy-9,10-dimethoxypterocarpan and 9,10-dimethoxyptercarpan-3-O-ß-d-glucoside. Integrative analysis predicted 19 target genes for TFA constituents, and the target genes were mainly involved in the EGFR-related cancer signaling, metabolism and oxidative stress. Collectively, these findings highlight the role of TFA in the regulation of LSCC and provide potential targets for a high-efficiency and low-toxicity therapeutic strategy of LSCC.

Relationship Between HIV Infection, Antiretroviral Therapy, Inflammatory Markers, and Cerebrovascular Endothelial Function Among Adults in Urban China.

BACKGROUND: Cerebrovascular risk is increased in people living with HIV infection compared with age-matched uninfected individuals. Cerebrovascular endothelial dysfunction related to antiretroviral therapy (ART) and inflammation may contribute to higher stroke risk in HIV infection. METHODS: We compared cerebral vasoreactivity-a measure of cerebrovascular endothelial function assessed by the breath-holding index (BHI) using transcranial Doppler ultrasound-between virologically suppressed Chinese HIV-infected individuals followed in an HIV clinic in Beijing, China, and uninfected controls. We constructed mixed-effects models to evaluate the association of HIV, ART, and inflammatory markers with cerebral vasoreactivity. RESULTS: In an unadjusted model, HIV infection was associated with a trend toward lower cerebral vasoreactivity (BHI 1.08 versus 1.26, P = 0.079). In multivariable analyses, cholesterol modified the association between HIV infection and cerebral vasoreactivity (P = 0.015 for interaction). At a lower total cholesterol of 4.15 mmol/L, HIV was associated with lower cerebral vasoreactivity (BHI -0.28, P = 0.019), whereas at a cholesterol of 5.15 mmol/L, the reduction in cerebral vasoreactivity associated with HIV was no longer statistically significant (BHI -0.05, P = 0.64). Among HIV-infected individuals, use of lopinavir/ritonavir compared with efavirenz was associated with lower cerebral vasoreactivity (BHI -0.24, P = 0.040). We did not find a significant association between inflammatory markers and cerebral vasoreactivity. CONCLUSIONS: Cerebrovascular endothelial dysfunction associated with HIV infection may be most relevant for individuals with less traditional vascular risk, such as those with lower cholesterol. Further study of the impact of ART on cerebrovascular endothelial function is warranted to aid with ART selection in individuals at high cerebrovascular risk.

NMR-based metabonomics and correlation analysis reveal potential biomarkers associated with chronic atrophic gastritis.

Chronic atrophic gastritis (CAG) is one of the most important pre-cancerous states with a high prevalence. Exploring of the underlying mechanism and potential biomarkers is of significant importance for CAG. In the present work, 1H NMR-based metabonomics with correlative analysis was performed to analyze the metabolic features of CAG. 19 plasma metabolites and 18 urine metabolites were enrolled to construct the circulatory and excretory metabolome of CAG, which was in response to alterations of energy metabolism, inflammation, immune dysfunction, as well as oxidative stress. 7 plasma biomarkers and 7 urine biomarkers were screened to elucidate the pathogenesis of CAG based on the further correlation analysis with biochemical indexes. Finally, 3 plasma biomarkers (arginine, succinate and 3-hydroxybutyrate) and 2 urine biomarkers (α-ketoglutarate and valine) highlighted the potential to indicate risks of CAG in virtue of correlation with pepsin activity and ROC analysis. Here, our results paved a way for elucidating the underlying mechanisms in the development of CAG, and provided new avenues for the diagnosis of CAG and presented potential drug targets for treatment of CAG.

[Risk analysis for cerebral hyperperfusion syndrome after carotid endarterectomy].

OBJECTIVE: To analyze risk factors for cerebral hyperperfusion syndrome (CHS) after carotid endarterectomy (CEA). METHODS: From September 2010 to September 2012, 183 consecutive patients with carotid artery stenosis who had indications for CEA entered the study. There were 149 male and 34 female patients, aged from 38 to 83 years with an average of (66 ± 9) years. Intracranial blood flow changes were monitored through transcranial Doppler routinely. Pre- and post-operative middle cerebral artery velocity (VMCA) were recorded. CHS was diagnosed by the combination of hyperperfusion syndrome and 100% increase of VMCA after operation compared with pre-operative baseline values. The patients who had CHS during hospitalization were recorded. Pre-operative and operative related factors were analyzed by univariate analysis, followed by Logistic regression model, to identify the risk factors of CHS. RESULTS: Overall, CHS occurred in 15 patients (8.2%) after CEA. The average onset time was (2.6 ± 0.2) days after surgery. By decreasing blood pressure and using dehydration medicine, all the patients with CHS recovered before discharge. None of them developed to intracranial hemorrhage. On univariate analysis, significant risk factors for CHS were history of stoke, symptomatic carotid artery stenosis and shunting during operation. On Logistic regression model, independent risk factor was symptomatic carotid artery stenosis (OR = 6.733, 95%CI: 1.455-31.155, P = 0.015), while shunting during operation (OR = 0.252, 95%CI: 0.067-0.945, P = 0.041) was a protective factor. CONCLUSIONS: Symptomatic carotid artery stenosis is an independent risk factor for CHS after CEA and shunting during operation is a protective factor. Using shunt may be an effective method of preventing CHS after CEA.

Supine-to-standing transcranial Doppler test in patients with multiple system atrophy.

BACKGROUND: Supine-to-standing test, a transcranial Doppler (TCD) based technique, has been recently developed to evaluate cardiovascular dysautonomia. We explored the value of supine-to-standing TCD test in predicting the course of multiple system atrophy (MSA) with orthostatic hypotension (OH). METHODS: By monitoring the signals of middle cerebral artery during supine-to-standing posture changes, the trend curves of cerebral blood flow velocities, pulsatility index and resistance index were obtained from 38 MSA patients with OH and 31 healthy subjects. The correlation between TCD findings and the clinical outcome of the patients, which was determined by follow-up structured phone interview, was analyzed. Adverse outcome was defined if a patient died, was in bed-ridden state or had recurrent syncope (>1 per month). RESULTS: Two characteristic TCD findings were revealed in the MSA patients but not in the controls, i.e. a blunted cerebral blood flow velocity rebound after standing and/or sustained higher pulsatility index upon standing than supine baseline. Structured phone interview was completed in 31 of the 38 patients (mean follow-up time, 20 ± 11 months). While no subject had recurrent syncope before enrollment, 12 patients developed an adverse outcome during follow-up. The coexistence of two characteristic TCD findings predicted adverse outcomes with positive predictive value 66.7% and negative predictive value 87.5%. CONCLUSIONS: Supine-to-standing TCD test is valuable in predicting the course of MSA with OH at early stage. We hypothesize baroreflex failure effects and paradoxical cerebral vasoconstriction in response to OH may account for the TCD findings in MSA patients.

[The prevalence of combined extra- and intra-cranial artery stenosis in patients with extracranial internal carotid artery occlusive disease.].

OBJECTIVE: The intra-cranial arteries are more affected in Asian ischemic stroke patients. We thought that tandem or diffuse arterial occlusive disease may be more common among patients with extracranial internal carotid artery (ICAex) occlusive disease. In the current study, the frequency of other co-existing extracranial artery (EA) or intracranial artery (IA) stenosis in patients with ICAex occlusive disease and its risk factors were studied. METHOD: One hundred and ninety-eight consecutive patients with ischemic stroke or TIA who were admitted to our hospital from Jan 2001 to May 2008 and who had stenosis (70%) or occlusion of ICAex were enrolled in the study. All the patients were examined with carotid duplex and transcranial Doppler (TCD). Among them, 75 patients were also examined with DSA, 52 patients with intracranial MRA and 25 patients with extra- and intra-cranial CTA. Extracranial artery stenosis or occlusion were diagnosed with DSA/CTA in 93 patients, and with carotid duplex only in 105 patients. Intracranial artery stenosis was diagnosed with DSA/MRA/CTA in 118 patients, and with TCD only in 80 patients. Patients with presumably cardioembolism such as atrial fibrillation were excluded. RESULTS: Among the 198 patients, 172 (86.8%) had other combined EA or IA stenosis. One hundred and thirty-five patients (135/198, 68.2%) had other combined EA stenosis, which were more frequently found in the contralateral ICAex (71/198, 35.9%). One hundred and twenty-three patients (123/198, 62.1%) had combined IA stenosis, which were more frequently found in MCA (75/198, 37.9%). 52 (52/198, 26.3%) had ipsilateral terminal ICA and/or middle cerebral artery stenosis. CONCLUSION: Tandem or diffuse arterial occlusive disease is common among patients with steno-occlusion of ICAex. Therefore, it is important to evaluate other EA and IA stenosis before CEA or CAS in Chinese patients.