Toxic effects of Tripterygium glycoside tablets on the reproductive system of male rats by metabolomics, cytotoxicity, and molecular docking.

BACKGROUND: Tripterygium glycoside tablets (TGT) is the most common preparation from Tripterygium wilfordii Hook F, which is widely used in clinical for treating rheumatoid arthritis (RA) and other autoimmune diseases. However, its serious reproductive toxicity limits its application. PURPOSE: This study aimed to elucidate the toxic effects of TGT on the reproductive system of male RA rats and its potential toxic components and mechanism. METHODS: Collagen-induced arthritis (CIA) rat model was established, and TGT suspension was given at low, medium, and high doses. Gonadal index, pathological changes, and the number of spermatogenic cells were used to evaluate the toxic effects of TGT on the reproductive system. Non-targeted metabolomics of testicular tissue was conducted by UHPLC-QTOF/MS. Combined with network toxicology, the key targets of TGT-induced reproductive toxicity were screened and RT-qPCR was used to validation. In vitro toxicity of 19 components of TGT was evaluated using TM3 and TM4 cell lines. Molecular docking was used to predict the interaction between toxic components and key targets. RESULTS: TGT reduced testicular and epididymis weight. Pathology analysis showed a lot of deformed and atrophic spermatogenic tubules. The number of spermatogenic cells decreased significantly (P<0.0001). A total of 58 different metabolites including platelet-activating factor (PAF), lysophosphatidylcholine (Lyso PC), phosphatidylinositol (PI), glutathione (GSH), and adenosine monophosphate (AMP) were identified by testicular metabolomics. Glycerophospholipid metabolism, ether lipid metabolism, and glutathione metabolism were key pathways responsible for the reproductive toxicity of TGT. Ten key reproductive toxicity targets were screened by network toxicology. The cytotoxicity test showed that triptolide, triptonide, celastrol, and demethylzeylasteral could significantly reduce the viability of TM3 and TM4 cells. Alkaloids had no apparent toxic effects. Molecular docking showed that the four toxic components had a good affinity with 10 key targets. All binding energies were less than -7 kcal/mol. The RT-qPCR results showed the Cyp19a1 level was significantly up-regulated. Pik3ca and Pik3cg levels were significantly down-regulated. CONCLUSION: Through testicular metabolomics, we found that TGT may cause reproductive toxicity through CYP19A1, PIK3CA, and PIK3CG three target, which was preliminarily revealed. This study laid the foundation for elucidating the toxicity mechanism of TGT and evaluating its safety and quality.

Large-scale preparation of five polar polyphenols including three isomers from Phyllanthus emblica Linn. by preparative high-speed counter-current chromatography.

The separation of polar compounds is challenging work due to poor retention and insufficient selectivity. In the present study, an efficient strategy for large-scale preparation of five polar polyphenols including three isomers from Phyllanthus emblica Linn has been established by preparative high-speed counter-current chromatography. Macroporous resin column chromatography was used for the enrichment of the polar polyphenols. However, sugar and other ultra-polar impurities were co-washed out with the targets. Liquid-liquid extraction with ethyl acetate/water (1/1, v/v) solvent system was developed to remove the ultra-polar impurities with a clearance rate of 95%. Finally, the targets were introduced to preparative high-speed counter-current chromatography for separation using ethyl acetate/n-butanol/acetic acid/water (2/7/1/10, v/v/v/v) solvent system. As a result, 191 mg of Mucic acid 1,4-lactone 5-O-gallate, 370 mg of ß-Glucogallin, 301 mg of Gallic acid, 195 mg of Mucic acid 1,4-lactone 3-O-gallate and 176 mg of Mucic acid 1,4-lactone 2-O-gallate with purity higher than 98% were obtained from 1.5 g of sample. Mucic acid 1,4-lactone 3-O-gallate, Mucic acid 1,4-lactone 3-O-gallate, and Mucic acid 1,4-lactone 2-O-gallate are isomers. The results showed that high-speed counter-current chromatography could be well developed for the separation of polar compounds from natural products.

Effect of Low Frequency Repetitive Transcranial Magnetic Stimulation (rTMS) Combined with Hyperbaric Oxygen (HBO) on Awakening of Coma Patients with Traumatic Brain Injury.

Coma caused by craniocerebral injury is a common condition of neurosurgical acute injury. There is no specific method to promote awakening in a clinic. Early comprehensive treatment may be helpful to patients. The common methods are hyperbaric oxygen (HBO) and low-frequency repetitive transcranial magnetic stimulation (rTMS). However, the application effect and mechanism of rTMS combined with HBO on coma patients with traumatic brain injury need to be further studied. The brain stem auditory evoked potential (BAEP) is examined by the Kennedy coma recovery scale (CRS-R), the recovery of brain function and the state of consciousness are evaluated, and the therapeutic effect is evaluated by the Glasgow Coma Scale (GCS). Cerebrospinal fluid NE level, MCA blood flow velocity, and left brainstem and right brainstem auditory evoked potential are used to evaluate brain rehabilitation. RTMS combined with HBO could shorten the wake-up time, improve the wake-up rate, improve the GCS score and CRS-R score, shorten the brain wave latency time of the left and right brainstem, increase the NE level of cerebrospinal fluid, and decrease the blood flow velocity of MCA. RTMS combines with HBO can improve the nerve excitability of brain cells, reduce the disturbance of consciousness, promote the functional recovery of brain injury, and has a certain role in promoting the awakening of patients with traumatic brain injury coma.

Separation of five flavonoids with similar polarity from Caragana korshinskii Kom. by preparative high speed counter-current chromatography with recycling and heart cut mode.

Five flavonoids with similar polarity were obtained from Caragana korshinskii Kom. by preparative high speed counter-current chromatography. Due to their similar polarity, it is difficult to select a suitable solvent system for one-step separation. Three fractions including one pure compound were only obtained with a relatively suitable solvent system of ethyl acetate: n-butanol:water (0.5% glacial acetic acid) (4:6:10, v:v:v). In order to improve the separation efficiency, recycling and heart cut modes were introduced. As a result, two flavonoids with separation factor value at 1.13 and the other two with separation factor value at 1.20 were successfully separated. Finally, five flavonoids with purity higher than 98% were obtained. Fortunately, a new compound named myricetin 3'-methyl ether 3-O-glucoside-7-O-rhamnoside was obtained. The results indicated that the recycling combined with heart-cut mode could be effective for the preparation of natural compounds with similar polarity.