Huoxue Prescription Research Articles

Rapid identification of the compounds of Bushen Huoxue Prescription based on offline two-dimensional liquid chromatography with high-resolution mass spectrometry and molecular network technology.

The comprehensive and efficient characterization of components in traditional Chinese medicine is crucial for elucidating its active constituents and uncovering its mechanism. Identifying the compounds of the Bushen Huoxue Prescription (BHP) is difficult because of its complex composition and the large difference in concentration among its compounds. In this study, a hydrophilic interaction liquid chromatography coupled with reversed-phase LC (HILIC × RPLC) offline 2D-LC tandem high-resolution mass spectrometry method was established to analyze the total compounds of the BHP. Database screening and molecular networking were performed to identify the compounds. In contrast to conventional 1D chromatography, 2D chromatography increased peak capacity, enriched trace ingredients, and prevented the masking of high-abundance compounds. A total of 165 compounds were identified, and 14 potential compounds needed to be further identified. This study provided an effective method for comprehensively analyzing the complex system of traditional Chinese medicine compounds.

Exploring the mechanism of Bushen Huoxue prescription in the treatment of early diabetic retinal edema from the perspective of inner blood-retinal barrier injury

Diabetic retinopathy (DR) is one of the most severe complications of diabetes mellitus, diabetes belongs to the category of "emaciation-thirst disease" in traditional Chinese medicine (TCM). Bushen Huoxue Prescription (BHP) is composed of traditional Chinese materia medica, which has therapeutic effects on DR and early diabetic retinal edema (EDRE). However, the therapeutic mechanism is unclear. Exploring the mechanism of BHP against EDRE. Feeding Sprague Dawley (SD) rats a high-fat, high-sugar diet as well as providing intraperitoneal injections of streptozotocin (STZ) to promote inner blood-retinal barrier (iBRB) damage that can trigger EDRE, evaluating the therapeutic effect of BHP by the level of expressiveness of TJ proteins (ZO-1,Occludin) of the iBRB and the leakage of rhodamine B isothiocyanate (RITC) in the retina. The combination of network pharmacology and metabolomics was employed to study the mechanism of BHP in preventing of EDRE, then four proteins which were closely to the damage of iBRB were chosen for the validation by employing Western Blot (WB). Research of network pharmacology had shown that BHP had efficacy against EDRE by regulating targets such as AKT1, ALB, TNF, PPARG, etc, its potential pathways mainly involving signaling pathways such as HIF-1. In untargeted metabolomics analysis of serum, 15 differential metabolites were identified, with the metabolic pathways focusing on ketone body metabolism and synthesis, sphingolipid metabolism and phenylalanine metabolism. The conclusions of metabolomics and network pharmacology revealed that BHP can treat EDRE by alleviating hypoxia and oxidative stress and exerting protection of the iBRB. Finally, BHP's protection behavior of the iBRB was validated by WB experiments. Through integrating pharmacodynamics, network pharmacology and metabolomics, BHP was discovered to have a crucial function in EDRE therapy by preserving the integrity of iBRB. This comprehensive strategy also provided a reasonable way to reveal the multi-components, multi-targets, multi-pathways mechanism of TCM.

Integrated serum pharmacochemistry and serum pharmacology to investigate the active components and mechanism of Bushen Huoxue Prescription in the treatment of diabetic retinopathy

This study aimed to investigate the active components of Bushen Huoxue Prescriptions (BHP), and further clarify the mechanism by the integration of serum pharmacochemistry and serum pharmacology based on spectrum-effect relationship in vivo. In this paper, the components absorbed into serum were analyzed by ultra-high performance liquid chromatography-quadrupole-Exactive Orbitrap-high resolution mass spectrometry (UPLC-Q-Exactive Orbitrap-HRMS). And Müller cells were chosen as target cells to further investigate the mechanism. After cell purification, the well-grown cells were identified by Hematoxylin-eosin staining (HE) staining and immunofluorescence assay, such as glial fibrillary acidic protein (GFAP) and glutamine synthetase (GS). The logarithmic phase cells were divided into normal group, model group and 12 BHP groups. The hyperglycemic and hypoxic model was induced by 50 mmol/L glucose and 1 mmol/L sodium disulfite. Enzyme-linked immunesorbnent assay (ELISA) was used to detect the expressions of five factors closely related to DR, named vascular endothelial growth factor (VEGF), hypoxia-inducible factor1-alpha (HIF-1α), protein kinase C-β (PKC-β), angiopoietin-2 (ANG-2) and transforming growth factor-β (TGF-β). Finally, the spectrum-effect relationship was investigated to screen the active components of BHP by partial least squares regression (PLSR). The results showed that 83 metabolic components, containing 30 prototypes and 53 metabolites were found in BHP serum. 12 characteristic common peaks were chosen to establish spectrum-effect relationship. Significantly, all the 12 BHP serum exhibited stronger inhibition on the expression of VEGF, PKC-β, and ANG-2, and the expression of VEGF, PKC-β, ANG-2 was chosen to establish the spectrum-effect relationship in vivo. The results of PLSR revealed that the content of methylation and sulfuration of caffeic acid, dehydroxylation and sulfation of Danshensu, daidzein, O-demethylangolanolin, cryptotanshinone, tanshinone IIA and protopanaxatriol were inversely correlated with VEGF expression of Müller cells; the areas of dihydrocaffeic acid, methylation and sulfuration of caffeic acid, dehydroxylation and sulfation of Danshensu, daidzein, cryptotanshinone, tanshinone IIA were negative correlation with the expression of PKC-β; while the coefficient of hydroxytyrosol sulfation, R-equol, O-demethylangolanolin, dihydrotanshinone IIA, hydrated cryptotanshinone, protopanaxatriol showed negative correlation with the expression of ANG-2. The above results indicated that cryptotanshinone, tanshinone IIA, daidzein and protopanaxatriol need be focused in our future research. In addition, this research idea provides feasible ways to investigate and determine pharmacodyamic material basis and screen the Q-markers of TCM and its formulas.

Effects of modified Bushen Huoxue Prescription combined with active immunotherapy on immunomodulatory functions and pregnancy outcomes of patients with unexplained recurrent spontaneous abortion

Effects of modified Bushen Huoxue Prescription combined with active immunotherapy on immunomodulatory functions and pregnancy outcomes of patients with unexplained recurrent spontaneous abortion

Exploring the quality markers and mechanism of Bushen Huoxue Prescription in prevention and treatment of diabetic retinopathy based on Chinmedomics strategy.

Chinese herbal medicines have complex chemical composition; therefore, revealing the effective substances of Chinese herbal medicine becomes a prerequisite for scientific elucidation of the mechanism of action of Bushen Huoxue Prescription (BHP) against diabetic retinopathy (DR) and the development of new drugs. The Chinmedomics technique was used to evaluate the pharmacodynamic ingredients and mechanism of action of BHP against DR rats. The overall physiological condition of the rats, including body weight, blood glucose, inflammatory factor levels, histological staining, and urine metabolic profile were examined to evaluate the model and its effects. The chemical composition of BHP in vivo and ex vivo was fully analyzed utilizing UPLC-Q-Exactive Orbitrap MS in conjunction with TCM serum pharmacochemistry. Finally, correlation analysis between biomarkers, and serum migration components was used to identify Quality markers (Q-markers) that were significantly associated with effectiveness. The UPLC-Q-Exactive Orbitrap MS platform was used to identify a total of 29 chemicals in serum, 17 of which were highly linked with effectiveness and can be potentially employed as pharmacodynamic substances for BHP against DR. In addition, 14 biomarkers related to galactose metabolism, starch and sucrose metabolism, pantothenate and CoA biosynthesis, glycine, serine, and threonine metabolism were identified. These pathways reveal that DR may be inextricably linked to levels of oxidative stress and inflammation in the organism. Finally, five active ingredients were identified as potential Q-markers of BHP against DR, namely ajugol, protocatechuic acid, tanshinone IIA, panaxatriol and puerarin. This study successfully clarified the efficacy and Q-markers of BHP through the Chinmedomics strategy, which is of great significance in determining the quality standards of BHP.

Integrating network analysis and experimental validation to reveal the mitophagy-associated mechanism of Yiqi Huoxue (YQHX) prescription in the treatment of myocardial ischemia/reperfusion injury

Myocardial ischemia/reperfusion (I/R) injury is the main cause of increasing postischemic heart failure and currently there is no definite treatment for myocardial I/R injury. It has been suggested that oxidative stress-induced mitochondrial dysfunction plays an important role in the pathological development of myocardial I/R. In this study, Yiqi Huoxue (YQHX) prescription, as a kind of Chinese herbal formula, was developed and shown to alleviate I/R injury. Network analysis combined with ultrahigh-performance liquid chromatography-high resolution mass spectrometry expounded the active components of YQHX and revealed the mitophagy-regulation mechanism of YQHX treating I/R injury. In vivo experiments confirmed YQHX significantly alleviated I/R myocardial injury and relieved oxidative stress. In vitro experiments validated that YQHX could relieve hypoxia/reoxygenation injury and attenuate oxidative stress via improving the structure and function of mitochondria, which was strongly related to regulating mitophagy. In summary, this study demonstrated that YQHX, which could alleviate I/R injury via targeting mitophagy, might be a potential therapeutic strategy for myocardial I/R injury.

Effect of Huatan Qushi Huoxue prescription on the ultrastructure of hepatocyte mitochondria in rats with nonalcoholic steatohepatitis

Effect of Huatan Qushi Huoxue prescription on the ultrastructure of hepatocyte mitochondria in rats with nonalcoholic steatohepatitis

Simultaneous optimization of the extraction process of Yangyin Yiqi Huoxue prescription with natural deep eutectic solvents for optimal extraction yield and antioxidant activity: A comparative study of two models

IntroductionNatural deep eutectic solvents (NaDESs) are green and effective solvents that are used to extract 3 flavonoids from Yangyin Yiqi Huoxue prescription, a traditional Chinese prescription. MethodsA total of 6 types of NaDESs were systematically screened and evaluated for the total extraction yield of puerarin, calycosin, and formononetin by high-performance liquid chromatography. Then, a 4-factor-three-level experimental scheme designed by the Box-Benhnken Design was applied on the basis of a single experiment to determine the extraction yield and the antioxidant property. Finally, the extraction process was optimized through response surface methodology (RSM) and the genetic neural network (GNN), respectively. ResultsThe use of betaine-lactic acid as an extractant displayed significant advantages in the screening process. The optimum extraction parameters provided by GNN were as follows: water content 25% (v/v), liquid to material ratio 190 mg/ml, extraction time 37 min, and extraction temperature 63 °C. Under this condition, the average experimental comprehensive evaluation values of the extraction yield and antioxidant properties were 3.12 mg/g and 86.27%, and the relative deviations to the predicted values were 0.30% and 1.44%, respectively. In addition, the experimental results of GNN were better than those of RSM (p < 0.01). ConclusionsWe found the application of GNN to be effective and credible for bi-objective optimization of extraction yields and antioxidant activity in this study. Moreover, our results provide a reference and a theoretical basis for experimental and future industrial extraction for multi-objective situations.

Clinical Effect of Bushen Huoxue Method Combined with Platelet-Rich Plasma in the Treatment of Knee Osteoarthritis and Its Effect on IL-1, IL-6, VEGF, and PGE-2.

Objective To observe the clinical efficacy of the Bushen Huoxue method combined with platelet-rich plasma (PRP) in the treatment of knee osteoarthritis (KOA) and its effect on serum and joint fluid interleukin-1 (IL-1), interleukin-6 (IL-6), vascular endothelial growth factor (VEGF), and prostaglandin E2 (PGE-2). Methods A total of 64 cases of KOA diagnosed and treated from January 2020 to January 2021 were randomly divided into research group and control group, with 32 cases in each group. The control group was treated with PRP, and the research group took the prescription of the Bushen Huoxue method on the basis of the control group. The clinical efficacy was assessed according to the criteria in “the diagnosis and Treatment of osteoarthritis,” osteoarthritis index score and pain visual analogue score (VAS). Serum and articular fluid VAS, IL-1, IL-6, VEGF, and PGE-2 levels were detected by the enzyme-linked immunosorbent assay (ELISA). Results The clinical effective rate of the research group was 93.8%, which was significantly higher than that of the control group (90.6%). There was no significant difference in the scores of osteoarthritis index between the two groups before treatment, but the scores of both groups decreased after treatment and was lower in the research group than those in the control group. VAS was significantly decreased in two groups after treatment and it was lower in the research group than that in the control group. After treatment, the levels of IL-1, IL-6, and PGE-2 in serum and articular fluid all indexes were decreased, and the levels in the research group were lower than those in the control group. Conclusions PRP joint cavity injection combined with oral administration of Bushen Huoxue prescription, and PRP joint cavity injection alone can improve the efficacy of KOA, relieve knee pain, and promote the recovery of knee function. The mechanism may be related to the reduction of IL-1, IL-6, VEGF levels, and PGE-2 levels in the serum and joint fluid. However, the efficacy of combination therapy was superior to PRP alone.

Meta-Analysis of Bushen Huoxue Method in the Treatment of Recurrent Spontaneous Abortion Due to Prethrombotic State

Objective: To evaluate the effectiveness and safety of Bushen Huoxue prescription in the treatment of recurrent spontaneous abortion (RSA) due to prethrombotic state (PTS). Methods: Databases such as CNKI, WanFang, VIP, CBM, PubMed, Embase, and Cochrane Library were searched for randomized controlled trials on Bushen Huoxue prescription in treating RSA due to PTS from inception to March 2021; meta-analysis was performed by RevMan Version 5.3.0 following quality evaluation. Results: Seven trials were included, with 496 patients; the meta-analysis indicated that Bushen Huoxue prescription has advantages on the improvement of total clinical effective rate [RR = 1.22, 95% CI (1.10, 1.35), Z = 3.80 (P = 0.0001)], embryo survival rate at pregnancy of 12 weeks [RR = 1.25, 95% CI (1.10, 1.41), Z = 3.53 (P = 0.0004)], D-dimer levels [SMD = -1.59, 95% CI (-2.20, -0.97), Z = 5.07 (P &lt; 0.00001)], and fibrinogen levels [MD = -1.00, 95% CI (-1.29, -0.70), Z = 6.61 (P &lt; 0.00001)], but the statistical heterogeneity was significant; in terms of incidence of adverse reactions, there was no statistical difference between Bushen Huoxue prescription and western medicine. Conclusion: Compared with western medicine alone, Bushen Huoxue prescription alone or in combination with western medicine showed significant advantages in improving the overall clinical efficiency, embryonic survival rate at 12 weeks of pregnancy, and reducing D-dimer values as well as fibrinogen levels, without any significant difference in the incidence of adverse effects; however, the number of included studies is small and there are drawbacks, such as small sample size and low quality; therefore, high-quality clinical studies with large sample size and rigorous trial designs are needed in the future to provide a reliable basis for the effectiveness and safety of TCM in reducing the incidence of RSA due to prethrombotic state.

A Study of the Protective Effect of Bushen Huoxue Prescription on Cerebral Microvascular Endothelia Based on Proteomics and Bioinformatics.

Diabetic cognitive dysfunction is a serious complication of type 2 diabetes mellitus (T2DM), which can cause neurological and microvascular damage in the brain. At present, there is no effective treatment for this complication. Bushen Huoxue prescription (BSHX) is a newly formulated compound Chinese medicine containing 7 components. Previous research indicated that BSHX was neuroprotective against advanced glycosylation end product (AGE)-induced PC12 cell insult; however, the effect of BSHX on AGE-induced cerebral microvascular endothelia injury has not been studied. In the current research, we investigated the protective effects of BSHX on AGE-induced injury in bEnd.3 cells. Our findings revealed that BSHX could effectively protect bEnd.3 cells from apoptosis. Moreover, we analyzed the network regulation effect of BSHX on AGE-induced bEnd.3 cells injury at the proteomic level. The LC-MS/MS-based shotgun proteomics analysis showed BSHX negatively regulated multiple AGE-elicited proteins. Bioinformatics analysis revealed these differential proteins were involved in multiple processes, such as Foxo signaling pathway. Further molecular biology analysis confirmed that BSHX could downregulate the expression of FoxO1/3 protein and inhibit its nuclear transfer and inhibit the expression of downstream apoptotic protein Bim and the activation of caspase, so as to play a protective role in AGE-induced bEnd.3 injury. Taken together, these findings demonstrated the role of BSHX in the management of diabetic cerebral microangiopathy and provide some insights into the proteomics-guided pharmacological mechanism study of traditional Chinese Medicine.

Investigation on the Q-markers of Bushen Huoxue Prescriptions for DR treatment based on chemometric methods and spectrum-effect relationship

Ethnopharmacological relevanceDiabetic retinopathy (DR) is a kind of complex complication of late diabetes mellitus with high incidence and risk of blindness. Bushen Huoxue Prescription (BHP), which consists of Rehmanniae radix (RR), Salviae miltiorrhizae radix et rhizoma (SMRR), Ginseng radix et rhizome (GRR) and Puerariae lobatae radix (PLR), has an active effect on the treatment of DR. However, the quality markers (Q-markers) of BHP are not entirely clear. PurposeThis study aimed to screen the Q-markers of BHP for DR treatment based on the establishment of spectrum-effect relationship and verified experiment. Materials and methodsIn this study, 12 BHP samples (S1–S12) for fingerprint analysis and pharmacological evaluation were prepared according to a four-factor and twelve-level uniform design. High performance liquid chromatography-ultraviolet detector-evaporative light scattering detector (HPLC-UV-ELSD) was employed to analyze the fingerprint on the basis of the characteristics of BHP components. The evaluation of sample similarity was carried out by similarity analysis (SA) and hierarchical cluster analysis (HCA). The pharmacological indicators, including expression of vascular endothelial growth factor (VEGF) and hypoxia-inducible factor-1α (HIF-1α) in the retina of Sprague Dawley (SD) rats induced by streptozotocin (STZ), were detected by enzyme-linked immunosorbent assay (ELISA). Besides, the spectrum-effect relationship between common peaks of fingerprints and the pharmacological results was investigated by partial least squares regression (PLSR) and canonical correlation analysis (CCA). The results of spectrum-effect relationship were verified by the expression of VEGF and HIF-1α on primary culture retinal Müller cells induced by hyperglycemia and hypoxia. ResultsIn the HPLC-UV-ELSD fingerprint, 23 common peaks in UV and 14 common peaks in ELSD were identified. The pharmacological results indicated that the expression of VEGF and HIF-1α in the retina of SD rats was inhibited by 12 BHP samples to varying degrees compared with the model group. Based on SA and heatmap of HCA, S4 and S8 were clearly distinguished from other samples. The results of PLSR and CCA revealed that the contents of puerarin, daidzin, salvianolic acid B and ginsenoside Rb1 were inversely correlated with the expression of VEGF and HIF-1α. Hence, the four compounds may be the main active components to prevent and treat DR. The results of intervention on primary culture retinal Müller cells showed that puerarin, daidzin, salvianolic acid B, and ginsenoside Rb1 can significantly inhibit the expression of VEGF and HIF-1α. ConclusionsThe spectrum-effect relationship of BHP was successfully established, and the Q-markers of BHP for the prevention and treatment of DR were preliminarily confirmed. It provides a feasible method for the research of quality control.

Therapeutic effects of Yiqi Huoxue prescription on diabetic nephropathy: a meta-analysis and systematic review

Traditional Chinese medicine (TCM) has demonstrated excellent effects in treating diabetic nephropathy, and Yiqi Huoxue prescription has been widely used clinically. In the study, its effects on the kidney function and blood glucose of patients were explored.Chinese and English databases including PubMed, Medline, Embase, and Web of Sciences were used to retrieve articles comparing the treatment of diabetic nephropathy using Yiqi Huoxue prescription on the basis of conventional Western medicine treatment (experimental group) and conventional Western medicine treatment alone (control group) published from January 2000 to December 2020. The risk of bias assessment tool of the Cochrane System Review Manual 5.2.2 and the Jadad scale were used to evaluate the quality of the included literature. The outcome indexes were extracted, and the Review Manager 5.3 software was used for meta-analysis.A total of 13 articles that satisfied the inclusion/exclusion criteria were included in this study. After treatment, compared to the control group, the experimental group exhibited lower urine microalbumin excretion rate (UAER) [mean difference (MD) =-33.94, 95% confidence interval (CI), -42.60 to -25.28, P<0.00001], serum creatinine (SCr) (MD =-7.43, 95% CI, -11.50 to -3.36, P=0.0004), blood urea nitrogen (BUN) (SMD =-1.23, 95% CI, -2.49 to 0.03, P=0.04), blood glucose-related indexes [fasting blood glucose (FBG)] (MD =-0.43, 95% CI, -0.87 to 0.01, P=0.03), glycosylated hemoglobin (HbA1c) (MD =-0.38, 95% CI, -0.68 to -0.08, P=0.01), blood lipid-related indexes [triglycerides (TG)] (MD =-0.44, 95% CI, -0.76 to -0.13, P=0.006), and serum total cholesterol (TC) (MD =-0.37, 95% CI, -0.57 to -0.18, P=0.0002). Furthermore, the experimental group also showed higher effectiveness rate (odds ratio =3.81, 95% CI, 2.71 to 5.35, P<0.00001) after treatment.The included literature had low bias risk. Yiqi Huoxue prescription on the basis of conventional Western medicine can significantly improve the renal function and reduce the levels of blood glucose and blood lipids of patients with diabetic nephropathy.

Effects of Qingre Huatan Huoxue prescription on biological characteristics and cell cycle proteins of esophageal cancer stem cells.

e16052 Background: Esophageal cancer (EC) stem cells characterized with immature differentiation, high invasion, high tumorigenesis and other biological characteristics, which are the root of the occurrence, development, recurrence, metastasis of EC. In this study, its purpose is to select and identify EC stem cells from serum-free cultured EC9706 cells, explore the effect of Chinese herbal prescription and its separated prescriptions on the biological characteristics of EC9706 stem cells, and to reveal the cytological and molecular mechanisms of Chinese herbal prescription anti-cancer. Methods: Firstly, MTT method was applied to explore the serum-free culture conditions of EC9706 stem cells in diameter of 60 mm low adhesion culture dish, at the same time, applying flow cytometry to identify the p75NTR mark ratio of each passage of cell to screenEC9706 stem cells. Next, Plotting the growth curve of EC9706 stem cells, detecting the proliferation inhibition rate EC9706 stem cells by cisplatin, soft agar clone formation, cell cycle to identify the characteristics of EC9706 stem cells by MTT method, soft agar assay, flow cytometry. Then applying MTT method and orthogonal test to screen and optimize Qingre Huatan Huoxue prescription (QHHP), using flow cytometry and western blot to test the effects of QHHP and its separated prescriptions on EC9706 stem cells appraisal indicators and Cell Cycle Proteins. Results: EC9706 cells with concentration of 8×104 cells/dish inoculateed on diameter of 60mm low adhesion culture dish, with adding 5 ml DMEM/F12 medium, a daily supplement of 20ng EGF, 10ng bFGF, per 10 to 14 days to passage one generation. After 5 generations of cell passage, the expression rate of p75NTR was 4-5%, was selected as the experimental object.EC9706 stem cells with the biological characteristics of slower adhesive growth, stronger cisplatin resistance, higher soft agar clone formation and p75NTR expression rate than that of EC9706 cells, and The cell cycle distribution was concentrated in the G0/G1 phase. 6 Chinese herbs were screened out ( Coptis coptidis, Rhizoma officinalis, Cucurbita pedicle, Toona bark, Cantharidopsis cantharidis, Augustia japonica) and compose QHHP. QHHP and its separated prescriptions could change the morphology, colony formation ability and cycle distribution of EC9706 stem cells in serum-free culture condition, which were related to the BMI1/CyclinB1 signaling pathway. Conclusions: Serum-free cell culture is a method to enrich stem cell-like cells in EC9706 cells. Qingre Huatan Huoxue prescription and its separated prescriptions inhibited the proliferation of EC9706 stem cells and change esophageal cancer stem cells characters. The mechanism of QHHP and its separated prescriptions on stem cells in EC9706 cells was related to BMI1 /CyclinB1 signaling pathway.

Simultaneous determination of six main components in Bushen Huoxue prescription by HPLC-CAD

BackgroundThe Bushen Huoxue prescription is a traditional Chinese medicine formula treating diabetic retinopathy, which was developed by our research group. Catalpol, puerarin, salvianolic acid B, ginsenoside Rg1, ginsenoside Rb1 and ginsenoside Rd are six of main effective components, which could be partly representative of this prescription. The corona charged aerosol detector (corona CAD) is one kind of universal detectors equipped with the high performance liquid chromatography (HPLC). The CAD has many advantages for the analysis of complex mixtures, but too few applications in traditional Chinese medicine compounds. ObjectiveThe aims of this study are to establish a method for the determination of six components in Bushen Huoxue prescription, and to increase the use of the CAD in traditional Chinese medicine compounds. MethodsHPLC-CAD analysis was performed on an Inertsil ODS-SP (4.6 mm × 250 mm, 5 μm) with a mobile phase consisting of 0.5 % formic acid solution(A)-acetonitrile(B) at a flow rate of 1 mL/min (0−7 min, 1 % B; 7−12 min, 1 %–12 % B; 12−22 min, 12 %–19 % B; 22-40 min, 19 %–28 % B; 40−43 min, 28 %–33 % B; 43−50 min, 33 % B; 50−65 min, 33 %–42 % B). The column temperature maintained at 30 ℃, the injection volume was 20 μL, the atomization temperature mode was LOW, the filtration constant (filter) was 3.6 and data collection rate was 10 Hz. The methodology was examined and the linearity of regression of different functions was compared. Sixteen batches of samples were prepared and their contents were determined. ResultsThe six compounds showed a better linearity (R2 > 0.9990) in their concentration ranges when using the linear function. The average recoveries were 99.18 %–101.30 %. Although the RSD value of puerarin and ginsenoside Rg1 was slightly out of 3 % during the average recovery investigation, all the other methodological investigations of the six components were within 3 %. The precision, stability and repeatability of the method were good. In sixteen batches of Bushen Huoxue prescription samples, the contents of six components were 0.3138 %–0.6042 % for catalpol, 0.8095 %–1.2917 % for puerarin, 0.7416 %–1.1189 % for salvianolic acid B, 0.0231 %–0.0418 % for ginsenoside Rg1, 0.0702 %–0.1724 % for ginsenoside Rb1, 0.0384 %–0.1196 % for ginsenoside Rd. ConclusionIn this experiment, a method for the determination of six components in Bushen Huoxue prescription based on HPLC-CAD was established with high accuracy, good repeatability and simple operation, and it can provide references for the improvement of quality standard of the Bushen Huoxue prescription. It is reasonable and accessible for the CAD application in the determination of traditional Chinese medicine compound prescriptions.

Effect of Bushen Huoxue Prescription on Cognitive Dysfunction of KK-Ay Type 2 Diabetic Mice.

Diabetic cognitive impairment is one of the common complications of type 2 diabetes, which can cause neurological and microvascular damage in the brain. Bushen Huoxue prescription (BSHX), a compound Chinese medicine, has been used clinically to treat diabetes-induced cognitive impairment. However, its underlying mechanisms remain unclear. In this study, KK-Ay diabetic model mouse was administered BSHX daily for 12 weeks. Bodyweight, random blood glucose (RBG), and fasting blood glucose (FBG) were measured every 4 weeks. Triglycerides (TG), cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), fasting serum insulin (FINS), and Morris water maze were tested after 12 weeks of administration. On the day of sacrifice, the hippocampus was collected for pathological staining and advanced glycation end products (AGEs) analysis to evaluate the neuroprotective effect of BSHX. Our results showed that BSHX treatment significantly ameliorated the T2DM related insults, including the increased bodyweight, blood glucose, TG, insulin levels, AGEs, the reduced HDL-C, the impaired spatial memory, and the neurological impairment. Moreover, Western blot analysis showed that increased expression of receptors of AGEs (RAGEs), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and activation of nuclear factor-κB (NF-κB) in the hippocampus were significantly inhibited by BSHX treatment. These results indicate that BSHX can significantly ameliorate glucose and lipid metabolism dysfunction, reduce the morphological changes in hippocampus tissues, and improve the cognitive function of KK-Ay mice. These protective effects of BSHX may involve regulation of the AGEs/RAGE/NF-κB signaling pathway.

The effects of Bushen Yiqi Huoxue prescription and its disassembled prescriptions on a diabetic retinopathy model in Sprague Dawley rats

BackgroundDiabetic retinopathy (DR) is one of the most serious complications in the late stages of diabetes, with a complex mechanism. As a complication affecting local lesions, few studies have compared differences of cytokine expression in the serum and retina. Owing to the specific value of traditional Chinese medicine (TCM) to complex diseases, TCM research has recently boomed in the prevention and treatment of diabetes. Bushen Yiqi Huoxue (BYH) prescription is a Chinese herbal compound that has been independently developed by our research group and has been proved to have a positive effect on DR; however, its specific mechanism and compatibility rule remain to be further explored. ObjectiveTo construct a DR model of Sprague Dawley (SD) rats, simultaneously detect multiple factor expression in the serum and retina of rats, explore the effect of BYH prescription and its disassembled prescriptions on DR, and discuss the influence of various compatibility combinations. MethodsBYH prescription was disassembled into two new compatibilities in the absence of Rehmanniae Radix (Yiqi Huoxue prescription, YH prescription) or Ginseng Radix et Rhizoma (Bushen Huoxue prescription, BH prescription). Male SD rats were induced using streptozotocin + high-fat and high-sugar diet to establish DR models and were divided into groups, then the intragastric administration and sampling. The body weight and fasting blood glucose of rats were continuously recorded during feeding; pathophysiological status observation of the retina by haematoxylin and eosin (HE) staining; advanced glycation end products (AGEs) and haemoglobin A1c (HbA1c) level detection in the rat serum by enzyme-linked immunosorbent assay; and the Luminex technique was used to detect the ICAM-1, IL-1β, IL-6, TNF-α and vascular endothelial growth factor (VEGF) expression concentrations in the retinal tissue and serum. ResultsThe results of blood glucose, body weight and HE staining proved that the model was successfully constructed, and the three combinations could reduce the retinal injury in DR rats. Serum AGEs and HbA1c levels of the model group increased compared with the control group (CG). Compared with the DR model group, only AGEs decreased in the BYH group, while the AGEs and HbA1c levels were significantly inhibited in the YH and BH groups, showing a significant correlation between the expression of AGEs and HbA1c in the serum of DR rats. In the serum of rats, IL-1β, IL-6, TNF-α and VEGF concentrations in the DR model group increased, although no statistical difference was observed in the ICAM-1 data compared with the CG. Compared with the DR model group, the IL-1β, IL-6 and TNF-α expression decreased in the BYH group. Moreover, the IL-6 and TNF-α expression decreased in the YH group and only the IL-6 expression decreased in the BH group. In the retina tissue, the model group had higher ICAM-1, IL-1β, IL-6, TNF-α and VEGF levels than the CG. Compared with the DR model group, TNF-α in the BYH group rats decreased, and the ICAM-1, IL-6 and TNF-α concentrations decreased in the YH and BH groups. Furthermore, differences in the ICAM-1 and VEGF expression in the serum and retina existed. ConclusionBYH compound and its disassembled prescriptions could improve the DR model rats induced with streptozotocin + high-fat and high-sugar diet, respectively, by inhibiting chronic blood glucose, AGEs, or inflammation response. The expression level and location of each factor are different, confirming that the effect of TCM prescriptions is not the simple addition of each single drug or its chemical components, but the rationality of its internal compatibility combination. Further, ICAM-1 and VEGF have exactly different expression levels, suggesting more attention to be paid by other researchers or doctors in future studies.

Chinese Herbs Medicine Huatan Huoxue Prescription for obstructive sleep apnea hypopnea syndrome as complementary therapy: A protocol for a systematic review and meta-analysis.

Objective:The aim of this systematic review and meta-analysis is to assess effectiveness and safety of Chinese Herbs Medicine Huatan Huoxue Prescription (HTHXP) as complementary therapy in treating bronchiectasis.Methods:The following databases will be searched: Embase, Cochrane, PubMed, China National Knowledge Infrastructure, Wan Fang, and VIP database from their inception to April 1, 2020. We performed and completed meta-analysis and methodologic evaluation by Review Manager 5.3.3 and statas 12.0 software. Study selection, data extraction, quality assessment, and assessment of risk bias will be performed by 2 reviewers independently. Odds ratios and correlative 95% confidence intervals will be calculated to present the association between the HTHXP and western medicine treatment using Review Manager version 5.3 when there is sufficient available data.Results:The results will be disseminated through a peer-reviewed journal publication.Conclusion:These systematic review findings will summarize up-to-date evidence for that HTHXP is more effective and safe as adjunctive treatment for patients with bronchiectasis.Ethics and dissemination:Ethics approval and patient consent are not required as this study is a systematic review based on published articles.PROSPERO registration number:INPLASY202050079.

Urine metabolomics study of Bushen Huoxue Prescription on diabetic retinopathy rats by UPLC-Q-exactive Orbitrap-MS.

Diabetic retinopathy (DR) is a common microvascular complication of diabetes and remains one of the leading causes of blindness worldwide. Previous studies have shown that Bushen Huoxue Prescription (BP) possesses an effect on preventing and treating DR, but the mechanisms of action are not entirely clear. In order to clarify the mechanisms, the pharmacodynamical assessments of BP were investigated. Combining the pharmacodynamical studies, it was found that BP could inhibit high expression of VEGF and HIF-1α, indicating significant therapeutic effects on DR. In order to further investigate the mechanism of BP in treating DR, a urine metabolomics method based on ultrahigh-performance liquid chromatography coupled with Q-exactive quadrupole-electrostatic field Orbitrap mass spectrometry was established to observe the metabolic variations in DR rats and investigate the therapeutic effect of BP on DR. As a result, nine potential biomarkers associated with DR were found. The metabolic pathways related to these compounds were explored, and the results showed that these biomarkers were mainly associated with gut microbial metabolism, lipid metabolism and tryptophan metabolism. The results for the pharmacodynamics and metabolomics provided a theoretical basis for clarifying the mechanism of BP in the treatment of DR.

Yangyin Qingre Huoxue Method in Traditional Chinese Medicine Ameliorates Atherosclerosis in ApoE-/- Mice Suffering from High-Fat Diet and HSP65 Aggression.

Atherosclerosis (AS) is a complicated arterial disease resulting from abnormal lipid deposition and inflammatory injury, which is attributed to Yin deficiency, accumulation of heat materials, and stasis of blood flow in Traditional Chinese Medicine (TCM) theory. Thus, according to TCM theory, the method of nourishing Yin (Yangyin), clearing away heat (Qingre), and promoting blood circulation (Huoxue) is a reasonable strategy, which has achieved remarkable clinical efficacy in the treatment of AS, but the mechanisms remain to be known. In this study, we evaluated the effects of Yangyin Qingre Huoxue Prescription (YQHP) on AS in ApoE−/− mice suffering from a high-fat diet and heat shock protein (HSP65) attack. YQHP regulated levels of blood lipids and inflammation-linked cytokines as well as Th17/Treg ratio in peripheral blood. Suppressed IL-6-p-STAT3 signaling and restored IL-2-p-STAT5 signaling in the presence of YQHP may partake in the regulation of Th17 and Treg differentiation. Moreover, YQHP modulated transcriptional levels of costimulator CD80 in aortas as well corresponding to the downregulation of GM-CSF in serum and CD3 expression in CD4+ T cells, which might indicate the potential of YQHP to regulate antigen presenting cells. All these effects eventually promoted the improvement of atherosclerotic lesions. In addition, YQHP promoted less monocyte infiltration in the liver and lower levels of AST, ALT, and AKP production than simvastatin. Conclusively, lipid-regulating and anti-inflammatory functions mediated by YQHP with lower hepatotoxicity than simvastatin hindered the progression of HSP65 aggravated AS in ApoE−/− mice, indicating the effectiveness of Yangyin Qingre Huoxue Method in the treatment of AS.