[Establishment and evaluation of a premenstrual syndrome rat model induced by progesterone-withdrawal and emotional-stimulation].

Premenstrual syndrome(PMS) lacks a highly consistent and feasible animal model that aligns with diagnostic and therapeutic standards in both traditional Chinese medicine(TCM) and western medicine, resulting in a lack of reliable experimental carriers for studying its pathogenesis and pharmacological effects. This study aims to systematically analyze the biological implications of PMS from the perspective of the "disease-syndrome-symptom" correlation and establish preparation and evaluation methods for an improved animal model of this disease. Firstly, clinical symptom gene sets related to the Qi stagnation syndromes due to liver depression and blood stasis in PMS in both modern medicine and TCM diagnostic standards were collected through GeneCards, DisGeNET, Mala-Cards, and the System of Foundational Diagnostic Association(SoFDA) database, as well as published literature. Based on the interaction information between genes, a "disease-syndrome-symptom" correlation network of PMS was established. Based on data mining results, an improved rat model of PMS was prepared by combining chronic restraint stress with the classical progesterone-withdrawal mo-del to simulate emotional depression caused by external environmental stimuli during the clinical onset process, inducing pathological damage from both physiological and emotional dimensions. The evaluation of the improved model before and after modification included open field experiment scores, organ indices, ovarian pathological changes, serum levels of estradiol(E_2), follicle-stimulating hormone/luteinizing hormone(FSH/LH), 5-hydroxytryptamine(5-HT), dopamine(DA), norepinephrine(NE), as well as coagulation parameters and hemorheology indexes. By calculating the degree, betweenness, and closeness centrality of nodes in the "disease-syndrome-symptom" correlation network, 163 core genes with topological importance were identified. Further biological function mining results indicated that core genes in PMS mainly participated in the regulation of the "nervous-endocrine-immune" system and pathways related to circulatory disorders. Mapping analysis of clinical phenotype symptom gene sets suggested significant correlations between core genes in PMS and depressive symptoms and pain symptoms caused by blood stasis. Compared with the simple progesterone withdrawal model, rats subjected to combined injections and restraint stress showed more significant abnormalities in open field experiment scores, ovarian tissue pathology, serum neurotransmitter levels of 5-HT and DA, as well as serum hormone levels of E_2 and FSH/LH. The modified modeling conditions exacerbated the pathological changes in blood rheology, coagulation function, and red blood cell morphology in model rats, confirming that the improved rat model could characterize the "nervous-endocrine-immune" system disorder and circulatory system disorders in the occurrence and progression of PMS, consistent with the clinical diagnostic and therapeutic standards of both TCM and western medicine. The establishment of the improved rat model of PMS can provide a reliable experimental carrier for elucidating the pathogenesis of PMS and discovering and evaluating therapeutic drugs. It also provides references for objectively reflecting the clinical characteristics of PMS in TCM and western medicine and precision treatment.

MicroRNA-205-5p: A potential therapeutic target for influenza A.

We are committed to finding host targets for influenza A therapeutics. The nucleoprotein (NP) plays an important role in influenza A virus replication and is an indispensable part of viral transcription and replication. Exploring endogenous substances that can modulate NP is critical for finding host targets. MicroRNAs (miRNAs, miR) are a novel class of powerful, endogenous gene expression regulators. Herein, we used miRanda to analyse the base complementarity between the NP gene and the 14 host miRNAs reported previously by us. MiRanda predicted that miR-431-5p, miR-744-3p and miR-205-5p could complement the NP gene. To understand the effect of these miRNAs on NP expression, we co-transfected 293 T cells with NP gene sequence containing above miRNAs binding site or full sequence of NP gene (transfected into pmirGlo or pcDNA3.1 vectors, respectively), and mimics of miR-205-5p, miR-431-5p and miR-744-3p. Dual luciferase reporter gene or Western blotting assays confirmed that miR-205-5p and miR-431-5p inhibit NP expression by binding with the miRNA binding site of NP gene. Further, we infected Mouse Lung Epithelial (MLE-12) cells overexpressing miR-205-5p and miR-431-5p with influenza A virus and performed Western blotting to examine NP expression. We found that NP expression was significantly reduced in MLE-12 cells overexpressing miR-205-5p during influenza A infection. The miR-205-5p overexpression-induced inhibition of influenza A replication could be attributed to the inhibition of NP expression. Further, we administered oseltamivir and Jinchai Antiviral Capsules (JC, an anti-influenza Chinese medicine) to influenza A virus-infected MLE-12 cells and mice. We found that miR-205-5p was significantly decreased increased in infected cells and lung tissues, and oseltamivir and JC could up-regulate miR-205-5p. In conclusion, we provide new evidence that miR-205-5p plays a role in regulating viral NP protein expression in combating influenza A and may be a potential target for influenza A therapy.

Identification and evaluation of the chemical similarity of Yindan xinnaotong samples by ultra high performance liquid chromatography with quadrupole time-of-flight mass spectrometry fingerprinting.

Yindan xinnaotong, a compound preparation used in traditional Chinese medicine, is composed of eight herbs: Ginkgo biloba leaf (yinxingye), Salvia miltiorrhizae (danshen), Herba gynostemmatis (jiaogulan), Erigerontis herba (dengzhanxixin), Allii sativi bulbus (dasuan), Notoginseng radixe rhizoma (sanqi), Crataegi fructus (shanzha), and Borneolum (tianranbingpian). Yindan xinnaotong is primarily used to treat cardiovascular and cerebrovascular diseases. However, to date, no scientific methods have been established to assess the quality of Yindan xinnaotong. Therefore, a combinatorial method was developed based on chemical constituent identification and fingerprint analysis to assess the consistency of Yindan xinnaotong quality. In this study, ultra high performance liquid chromatography coupled with time-of-flight mass spectrometry was used to identify the chemical components of Yindan xinnaotong soft capsules. Approximately 74 components were detected, of which 70, including flavonoids, ginkgolide, phenolic acid, diterpenoid tanshinones, and ginsenoside, were tentatively identified. A fingerprint analysis was also conducted to evaluate the uniformity of the quality of Yindan xinnaotong soft capsules. Ten batches of Yindan xinnaotong soft capsules were analyzed. All of the resulting chromatograms were imported into the "Similarity Evaluation System for Chromatographic Fingerprints of TCM" (Chinese Pharmacopoeia Commission, version 2004A). The similarity scores of common peaks from these samples ranged from 0.903-1.000, indicating that samples from different batches were highly correlated.

Prevention of atherosclerosis by Yindan Xinnaotong capsule combined with swimming in rats.

BACKGROUND: Yindan Xinnaotong capsule has been used for treating cardio-cerebrovascular diseases for several decades in China. Exercise training can protect against the development of atherosclerosis. The aim of the present study is to evaluate the joint effect of YXC and exercise on atherosclerosis in rats. METHODS: A combined method involving low shear stress and a high-fat diet was used to establish the atherosclerosis model in rats. Partial ligation of the left common carotid artery was performed, and then the rats were divided into 9 treatment groups according to a 3 × 3 factorial design with two factors and three levels for each factor, swimming of 0, 0.5, 1 h daily and YXC administration of 0, 1, 2 g/kg p.o. daily. Next the interventions of swimming and YXC were executed for 8 weeks. After that, blood samples were collected to determine blood viscosity, plasma viscosity, haematocrit (HCT), fibrinogen (FIB), blood lipid profile (including total cholesterol (TC), low-density lipoprotein-cholesterol (LDL-C), triglyceride (TG) and high-density lipoprotein-cholesterol (HDL-C)), nitric oxide (NO), 6-keto- prostaglandin (PG) F1α, endothelin (ET) and thromboxane (TX) B2. The common carotid arteries of the rats were harvested to examine pathological changes, wall thickness and circumference, and the expression of SM22αwas assayed via immune-histochemistry. RESULTS: The early pathological changes were observed. The joint effects of YXC and swimming showed significant changes in the examined parameters: (1) decreases in plasma viscosity, blood viscosity and FIB; (2) increases in NO and 6-keto-PGF1α; (3) decreases in ET and TXB2; and (4) decreases in LDL-C and TG. The combination of 2 g/kg YXC and 1 h of swimming led to synergistic decreases in LDL-C and TG. The interactive effect between YXC and swimming was obvious in decreasing wall thickness. Swimming alone was able to up-regulate the expression of SM22α. CONCLUSIONS: In conclusion, this study indicates that the combination of YXC and swimming may prevent atherosclerosis through a synergistic effect between YXC and swimming in improving blood circulation, hemorheological parameters, blood lipids levels and the vascular endothelium in rats. The vascular remodeling may be contributed to the prevention effects on AS by up-regulating SM22α.

Pharmacokinetic comparison of berberine in rat plasma after oral administration of berberine hydrochloride in normal and post inflammation irritable bowel syndrome rats.

In the present study, post inflammation irritable bowel syndrome (PI-IBS) rats were firstly established by intracolonic instillation of acetic acid with restraint stress. Then the pharmacokinetics of berberine in the rat plasma were compared after oral administration of berberine hydrochloride (25 mg/kg) to normal rats and PI-IBS rats. Quantification of berberine in the rat plasma was achieved by using a sensitive and rapid UPLC-MS/MS method. Plasma samples were collected at 15 different points in time and the pharmacokinetic parameters were analyzed by WinNonlin software. Compared with the normal group, area under the plasma concentration vs. time curve from zero to last sampling time (AUC0-t) and total body clearance (CL/F) in the model group significantly increased or decreased, (2039.49 ± 492.24 vs. 2763.43 ± 203.14; 4999.34 ± 1198.79 vs. 3270.57 ± 58.32) respectively. The results indicated that the pharmacokinetic process of berberine could be altered in PI-IBS pathological conditions.

Osteoking promotes bone formation and bone defect repair through ZBP1-STAT1-PKR-MLKL-mediated necroptosis.

BACKGROUND: Osteoking has been used for fracture therapy with a satisfying clinical efficacy. However, its therapeutic properties and the underlying mechanisms remain elusive. METHOD: A bone defect rat model was established to evaluate the pharmacological effects of Osteoking by the dynamic observation of X-ray, micro-CT and histopathologic examination. Transcriptome profiling was performed to identify bone defect-related genes and Osteoking effective targets. Then, a "disease-related gene-drug target" interaction network was constructed and a list of key network targets were screened, which were experimentally verified. RESULTS: Osteoking effectively promoted bone defect repair in rats by accelerating the repair of cortical bone and the growth of trabeculae. Histopathologically, the bone defect rats displayed lower histopathologic scores in cortical bone, cancellous bone and bone connection than normal controls. In contrast, Osteoking exerted a favorable effect with a dose-dependent manner. The abnormal serum levels of bone turnover markers, bone growth factors and bone metabolism-related biochemical indexes in bone defect rats were also reversed by Osteoking treatment. Following the transcriptome-based network investigation, we hypothesized that osteoking might attenuate the levels of ZBP1-STAT1-PKR-MLKL-mediated necroptosis involved into bone defect. Experimentally, the expression levels of ZBP1, STAT1, PKR and the hallmark inflammatory cytokines for the end of necroptosis were distinctly elevated in bone defect rats, but were all effectively reversed by Osteoking treatment, which were also suppressed the activities of RIPK1, RIPK3 and MLKL in bone tissue supernatants. CONCLUSIONS: Osteoking may promote bone formation and bone defect repair by regulating ZBP1-STAT1-PKR axis, leading to inhibit RIPK1/RIPK3/MLKL activation-mediated necroptosis.

Study on the potential mechanism of Qingxin Lianzi Yin Decoction on renoprotection in db/db mice via network pharmacology and metabolomics.

BACKGROUND: Diabetic nephropathy (DN) was one of the most popular and most significant microvascular complications of diabetes mellitus. Qingxin Lianzi Yin Decoction (QXLZY) was a traditional Chinese classical formula, suitable for chronic urinary system diseases. QXLZY had good clinical efficacy in early DN, but the underlying molecular mechanism remained unrevealed. PURPOSE: This study aimed to establish the content determination method of QXLZY index components and explore the mechanism of QXLZY on DN by network pharmacology and metabolomics studies. METHODS: Firstly, the content determination methods of QXLZY were established with calycosin-7-O-ß-d-glucoside, acteoside, baicalin and glycyrrhizic acid as index components. Secondly, pharmacological experiments of QXLZY were evaluated using db/db mice. UHPLC-LTQ-Orbitrap MS was used to carry out untargeted urine metabolomics, serum metabolomics, and kidney metabolomics studies. Thirdly, employing network pharmacology, key components and targets were analyzed. Finally, targeted metabolomics studies were performed on the endogenous constituents in biological samples for validation based on untargeted metabolomics results. RESULTS: A method for the simultaneous determination of multiple index components in QXLZY was established, which passed the comprehensive methodological verification. It was simple, feasible, and scientific. The QXLZY treatment alleviated kidney injury of db/db mice, included the degree of histopathological damage and the level of urinary microalbumin/creatinine ratio. Untargeted metabolomics studies had identified metabolic dysfunction in pathways associated with amino acid metabolism in db/db mice. Treatment with QXLZY could reverse metabolite abnormalities and influence the pathways related to energy metabolism and amino acid metabolism. It had been found that pathways with a high degree were involved in signal transduction, prominently on amino acids metabolism and lipid metabolism, analyzed by network pharmacology. Disorders of amino acid metabolism did occur in db/db mice. QXLZY could revert the levels of metabolites, such as quinolinic acid, arginine, and asparagine. CONCLUSION: This study was the first time to demonstrate that QXLZY alleviated diabetes-induced pathological changes in the kidneys of db/db mice by correcting disturbances in amino acid metabolism. This work could provide a new experimental basis and theoretical guidance for the rational application of QXLZY on DN, exploring the new pharmacological effect of traditional Chinese medicine, and promoting in-depth research and development.

Study on the therapeutic mechanism of HJ granules in a rat model of urinary tract infection caused by Escherichia coli.

ETHNOPHARMACOLOGICAL RELEVANCE: Urinary tract infections (UTIs) are globally prevalent infectious diseases, predominantly caused by uropathogenic Escherichia coli (UPEC). The misuse of antibiotics has led to the emergence of several drug-resistant strains. Traditional Chinese Medicine (TCM) has its own advantages in the treatment of UTIs. HJ granules is a herbal formula used for the treatment of UTIs. However, its mechanism of action is not clear. AIM OF THE STUDY: The aim of this study was to investigate the therapeutic efficacy and mechanism of action of HJ granules in a rat model of UTI caused by Escherichia coli (E coli) CFT073. MATERIALS AND METHODS: SD rats were selected to establish a rat UTI model by injecting UPEC strain CFT073 into the bladder using the transurethral placement method. HJ granules were administered to rats after modelling and the efficacy of HJ granule was investigated by measuring urinary decanalogue, inflammatory factors in bladder tissue and pathological changes in the bladder after 3d of administration. Expression of sonic hedgehog (SHH), NOD-like receptor thermoprotein domain 3 (NLRP3), apoptosis-associated speck-like protein (ASC) and activation of cysteinyl aspartate specific proteinase-1 (caspase-1) were detected by western blotting and immunofluorescence staining in rat bladder tissue. NLRP3, ASC and caspase-1, a cysteine-containing aspartic protein, were expressed and activated. RESULTS: The results showed that infection of rats with UPEC resulted in increased pH and erythrocytes in bladder irrigation fluid; increased expression of IL-1ß, IL-6 and SHH and decreased expression of IL-10 in bladder tissue; and significant upregulation of the expression of both SHH and NLRP3 inflammasom and significant activation of NLRP3 inflammasom. HJ granules significantly increased the concentration of IL-10 in the bladder, inhibited the expression of SHH and NLRP3 inflammasom in bladder tissue, and suppressed the activation of NLRP3 inflammasom, thereby reducing inflammatory lesions in bladder tissue. CONCLUSION: HJ granules may improve bladder injury and treat UTIs by inhibiting the expression and activation of NLRP3 inflammasom.

Protective effect of compatible herbs in Jin-Gu-Lian formula against Alangium chinense-induced neurotoxicity via oxidative stress, neurotransmitter metabolisms, and pharmacokinetics.

Background: A. chinense frequently used in Miao medicine to treat rheumatic diseases. However, as a famous toxic herb, Alangium chinense and its representative components exhibit ineluctable neurotoxicity, thus creating significant challenges for clinical application. The combined application with compatible herbs in Jin-Gu-Lian formula attenuates such neurotoxicity according to the compatible principle of traditional Chinese medicines. Purpose: We aimed to investigate the detoxification of the compatible herbs in Jin-Gu-Lian formula on A. chinense-induced neurotoxicity and investigate its mechanism. Methods: Neurobehavioral and pathohistological analysis were used to determine the neurotoxicity in rats administered with A. chinense extract (AC), extract of compatible herbs in Jin-Gu-Lian formula (CH) and combination of AC with CH for 14 days. The mechanism underlying the reduction of toxicity by combination with CH was assessed by enzyme-linked immunosorbent assays, spectrophotometric assays, liquid chromatography tandem-mass spectrometry and real-time reverse transcription-quantitative polymerase chain reaction. Results: Compatible herbs attenuated the AC-induced neurotoxicity as evidenced by increased locomotor activity, enhanced grip strength, the decreased frequency of AC-induced morphological damage in neurons, as well as a reduction of neuron-specific enolase (NSE) and neurofilament light chain (NEFL) levels. The combination of AC and CH ameliorated AC-induced oxidative damage by modulating the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px), and total antioxidant capacity (T-AOC). AC treatment significantly reduced the levels of monoamine and acetylcholine neurotransmitters in the brains of rats, including acetylcholine (Ach), dopamine (DA), 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), norepinephrine (NE), and serotonin (5-HT). Combined AC and CH treatment regulated the abnormal concentrations and metabolisms of neurotransmitters. Pharmacokinetic studies showed that the co-administration of AC and CH significantly decreased plasma exposure levels of two main components of AC, as evidenced by the reduction of maximum plasma concentration (Cmax), area under the plasma concentration-time curve (AUC) compared to AC. In addition, the AC-induced downregulation in mRNA expression of cytochrome P450 enzymes was significantly reduced in response to combined AC and CH treatment. Conclusion: Compatible herbs in Jin-Gu-Lian formula alleviated the neurotoxicity induced by A. chinense by ameliorating oxidative damage, preventing abnormality of neurotransmitters and modulating pharmacokinetics.

Modules in robust but low-efficiency phyllosphere fungal networks drive saponin accumulation in leaves of different Panax species.

BACKGROUND: The phyllosphere mycobiome plays a crucial role in plant fitness and ecosystem functions. The complex microbial ecological networks (MEN) formed by these fungi remain poorly understood, particularly with regard to their organization strategy and their contributions to plant secondary metabolites such as saponin. RESULTS: In this study, we constructed six MENs from leaf epiphytic and endophytic mycobiomes of three Panax species distributed in the northeast and southwest ends of mainland China. Hub nodes were absent in these MENs, which were significantly more complex, robust, and less efficient compared to random graphs (P < 0.05), indicating a hub-independent high-robustness strategy to maintain structural homeostasis. The important roles of specific MEN modules in shaping leaf saponin profiles of each Panax species were proved by multiple machine learning algorithms. Positive regulation modules (PRMs) of total saponin content were further identified, which exhibited more deterministic ecological assembly and comprised of highly connected nodes as well as higher proportion of plant-associated fungal guilds compared to other network members, indicating their tight links with host plant. The significant and direct effects (P < 0.05) of PRMs on total saponin accumulation were validated by well-fitted structural equation models (χ2 < 0.3, P > 0.5). Taxonomic analysis revealed that Pleosporales and Chaetothyriales were significantly overrepresented by positive regulation taxa (PRT) of total saponin content (FDR < 0.05). Across PRT identified in three Panax species, Epicoccum and Coniothyrium were conservatively present, while species-specific taxa such as Agaricales were also found, indicating the conservatism and specificity of plant-fungi interactions associated with leaf saponin accumulation in Panax genus. CONCLUSIONS: These findings provide a foundation for understanding mechanisms maintaining the steady state of phyllosphere mycobiome in healthy plant, and offer clues for engineering phyllosphere mycobiome to improve the accumulation of bioactive secondary metabolites on the basis of network modules.

[Development of gastric precancerous lesion animal model].

OBJECTIVE: To establish a model of gastric precancerous lesion by using Aristolochic manshuriensis which contains aristolochic acids. METHOD: The SD rats were randomly divided into four groups: control and three different doses of ethanol extractive of A. manshuriensis (EEA) (corresponding to aristolochic acid I 2.5, 5.0, 10.0 mg x kg(-1)), respectively. EEA was intragastrically given to rats every other day. At the end of the 10th, 15th, 20th week, part of the rats in each group was sacrificed and the stomachs were weighed. The gastric tumor was assessed by the weight and the relative stomach weight to the body weight. The stomachs were fixed in 4% neutral formalin, and the paraffin imbedding tissues were sliced and HE stained. Histomorphology was observed under the light microscope to determine gastric hyperplasia, mucosa precancerosis (atypical hyperplasia) and gastric cancer formation. RESULT: The rats treated with different doses of EEA for 10 weeks induced mucosa papillary, epithelioma hyperplasia. Histological observation showed mucosa precancerosis lesions characterized as atypical hyperplasia at the dose levels corresponding to aristolochic acid I 5.0 and 10.0 mg x kg(-1) treated for 10 weeks. The incidence rate of gastric precancerosis in those two groups was 100% at the 15th week. Malignant tumors were observed in most of the animals in 10.0 mg x kg(-1) group. The animals in 5.0 mg x kg(-1) group were well tolerant compared to 10.0 mg x kg(-1) group during the course of experiment, so the dose of aristolochic acid I 5.0 mg x kg(-1) and 10-15 weeks treatment were considered to be optimum to establish the model of gastric precancerosis. CONCLUSION: A rat model of gastric precancerosis can be induced within a short duration by giving an oral administration of the ethanol extract of A. manshuriensis which contains aristolochic acids.

Danlu tongdu tablets: Preclinical safety evaluation and mechanism of hepatotoxicity.

Danlu tongdu tablets (DLTD) is a listed Chinese patent medicine collected in the Pharmacopoeia of the People's Republic of China (version 2020). This prescription has been applied in clinics in China for lumbar spinal stenosis and lumbosacral disc herniations. The wide application of Danlu tongdu in therapy has raised some clinical adverse reactions, such as significant elevation of alanine transaminase (ALT) and aspartate transaminase (AST) in individual patients after use. The present study aimed to investigate the safety of Danlu tongdu and analyze its adverse effects on the liver. The maximum feasible dose (MFD) was used to carry out the acute toxicity tests. Mortality, adverse effects, body weight and food consumption were recorded for up to 14 days post treatment. In the 6-month chronic toxicity test, sprague-dawley rats were randomly divided into four groups according body weight, the experimental groups were administrated to rats at the concentrations of 1.67, 3.34 and 6.67 g/kg/day, whereas the control group was received the ultrapure water (vehicle) only, 10 ml/kg, once a day. The animal's body weight, food consumption was monitored weekly. In addition, their hematological and biochemical parameters, body and organ weights and histopathology, were all measured at specific observation time points. Additionally, we further explored the adverse effects mechanism of Danlu tongdu on the liver through transcriptome analysis. No deaths or substance-relative toxicity were observed in the acute toxicity study or the 6-month chronic toxicity study with doses of 1.67 g/kg and 3.34 g/kg, respectively. We found that mild hypertrophy and hyperplasia of hepatic interlobular bile ducts were detected in some rats with doses of 6.67 g/kg after repeated oral administration of Danlu tongdu for 13 and 26 weeks, but the above changes in liver were reversible. The results of transcriptome sequencing showed that Danlu tongdu had a significant effect on cytochrome P450 enzymes in rat liver, especially cytochrome P450 1 (CYP1) subtype. Therefore, the toxic target organ of Danlu tongdu is the liver and the mechanism of mild liver injury is closely related to the up-regulation of cytochrome P450 1A1 (CYP1A1) and cytochrome P450 1A2 (CYP1A2) expression.

[Research progress of especial toxicity and of pyrrolizidine alkaloids].

Pyrrolizidine alkaloids (PAs) are widely distributed in many plants including medicinal herbs. The hepatotoxicity of PAs has been known academically for a long time, however, their reproductive toxicity, mutagenesis and carcinogenicity have been less researched. This article is an overview of the clinical and experimental reports of the reproductive toxicity, mutagenesis and carcinogenicity of PAs, the effective factors and generating mechanism of the toxicity.

[Toxicity of monocrotaline on in vitro cultured mouse embryos].

OBJECTIVE: To investigate the fetotoxicity of monocrotaline. METHOD: Mouse whole embryo culture (WEC) was applied. Post-implantation (8.5 d) mouse embryos were isolated from their mothers and put into the medium of immediately centrifuged serum (ICS) prepared from rats. Different concentrations of monocrotaline (100, 50, 25, 12.5 mg x L(-1)) were added into the WEC. Development (yolk sac diameter, crown-rump length, head length, somite number) and organic morphodifferentiation (yolk sac circulation, allantois, embryonic flexion, heart, brain, optic-otic-olfactory organ, branchial arch, maxillary, mandible, bud) of embryos were observed at 48 h after treatment. RESULT: Obvious fetotoxicity could be observed in various monocrotaline treatment groups in a dose-dependent manner. Development of embryos was delayed significantly at dose 12.5-100 mg x L(-1). Malformations were shown in all organic morphodifferentiation indice, especially in opti-otic organ, mandible and bud. CONCLUSION: Monocrotaline had obvious fetotoxicity in vitro WEC, indicating that exposure of pregnant mice to monocrotaline may have potential risk on fetus.

[Antiatherogenic effect of Radix Salviae Miltiorrhizae and Fructus Crataegi on experimental atherosclerosis in rats].

OBJECTIVE: To investigate the antiatherogenic effect and possible mechanisms of the extracts of Radix Salviae Miltiorrhizae (RSM) or Fructus Crataegi (FC), as well as their interaction. METHOD: Wistar rats were randomly divided into 2 groups: normal group and model group. The atherosclerotic model rats were injected VD3 and ovalbumin, while fed with high cholesterol diet. After the model was determined successfully, all model rats were divided into normal group, model group, Xuezhikang group, RSM group, FC group, mixture of RSM and FC group. Each group was given the corresponding drugs for 4 weeks. After 12 weeks, blood serum were analyzed for total cholesterol (TC), triglyceride (TG), low density lipoprotein cholesterol (LDL-C) and high density lipoprotein cholesterol (HDL-C), superoxide dismutase ( SOD), malondialdehyde (MDA) and nitric oxide (NO). And the blood plasma also analyzed for levels of endothelin (ET), 6-keto prostaglandin F1alpha (6-keto-PGF1alpha), thromboxane B2 (TXB2), C-reactive protein (CRP), interleukin 6 (IL-6), interleukin 8 (IL-8), tumor necrosis factor alpha (TNF-alpha) and so on. At last, the pathological observation of aorta was carried out. RESULT: Compared with those in model group, the TC, TG, LDL-C, ET, TXB2 and MDA levels and TXB2/PGF1alpha ratio were reduced, while the HDL-C, the serum SOD, No and 6-keto-PGF1alpha level were raised in the intervention groups. Although the levels of CRP, IL-6 and IL-8 were lower than model group, there was no obvious effect on the releasing of TNF-alpha. CONCLUSION: RSM and FC could inhibit the atherogenesis formation and development, which might be due to regulating the lipid metabolism, enhancing the antioxidation, and reducing the release of inflammatory factors.

[Development of animal model for anaphylactoid test of rodent].

OBJECTIVE: To establish a simple and feasible method of anaphylactoid test on awaked small animals for screening and assessing anaphylactoid reaction of traditional Chinese medicine (TCM) injection with different concentration of tween 80. METHOD: Test substances containing 0.4% Evans blue were intravenously injected into mice at volume of 20 mL x kg(-1) or guinea pigs at a volume of 30 mL x kg(-1). The behaviors were observed and the vascular permeability of ears evaluated by the extent of ear blue staining and absorbance of Evans blue extraction of ears were tested at 30 min after injection. RESULT: Tween 80 solution, Yuxingcao injection with tween 80, and Shuanghuanglian powder injection obviously increased vascular permeability of ears characterized as ear blue staining and increased absorbance of the Evans blue extract from ears extracted by acetone saline both in mice and in guinea pigs in a concentration-dependent (in the case of tween 80) or a dose-dependent (Shuanghuanglian) manner. CONCLUSION: Ear vascular permeability test in mice and guinea pigs can be used as animal models to screen and test anaphylactoid reaction induced by injections.

[Evaluation of vitro hepatotoxicity of monocrotaline by precision-cut liver slice technique].

OBJECTIVE: To modify the empirical method of precision-cut liver slice technique, and study the hepatotoxicity of monocrotaline by this technique. METHOD: Liver slices were prepared by the domestic shaking slicer. The technique of precision-cut liver slice was established by detecting MTT reduction used as the slice viability under different culture medium, thickness of slices, pH and culture temperature. After monocrotaline and liver slices co-culture for 6, 24 h, the slice viability, enzyme activity of GPT, GOT, LDH, GGT and protein concentration were detected by MTT reduction, enzyme kinetics method and BCA protein assay method, respectively. RESULT: When the thickness of slices was 200 microm and pH of medium was 6.8, culture temperature was 37 degrees C, BPM culture medium, the viability of slices could maintain on a steady level. LDH leakage was significantly increased and protein content was obviously decreased after monocrotaline co-culture for 24 h with final concentration 0.02, 0.1 and 0.5 g x L(-1). No statistically significant difference between control group and monocrotaline 3 dose groups was observed in the slice viability and the content of GPT, GOT, LDH, GGT and protein after monocrotaline co-culture for 6 h. CONCLUSION: The slice viability could retain 24 h in modified BPM medium surroundings; monocrotaline displayed liver toxicity in some degree after co-culture for 24 hours in 0.02, 0.1 and 0.5 g x L(-1) concentration.

[Material and mechanisms for evaluation of Shuanghuanglian injection induced pseudoanaphylactoid reactions].

OBJECTIVE: To investigate the substance basis and the mechanism of pseudoanaphylactoid reactions (PR) induced by Shuanghuanglian injection (SHLI). METHOD: (1)The study of PR and the substance basis of PR of SHLI: ICR mice were divided into different test groups, the mice were intravenously injected with solutions of different concentration of SHLI, baicalin, forsythin, caffeotannic acid, positive control Compound 48/80 and normal sodium. All test substances were mixed with 0.4% Evans blue. The reaction and vascular permeability of the ears were observed and measured 30 min after SHLI injection. (2) The study of mechanisms: Mice were pretreated with an oral administration of Astemizol, intraperitoneal injection of cyclophosphamide 75 mg x kg(-1) or Compound 48/80 4 mg x kg(-1), then mice were intravenously injected with SHLI. At last, vascular permeability of the ears in pretreated groups was compared with SHLI treatment alone group. RESULT: SHLI of 300 mg x kg(-1) and 600 mg x kg(-1) caused obvious vascular hyperpermeability, but baicalin, forsythin and caffeotannic didn't cause vascular hyperpermeability in the ears. The Astemizol can decrease the degree of SHLI-induced vascular hyperpermeability of the ears in the mice. After intraperitoneal injected with cyclophosphamide, there was a slight decrease in the degree of SHLI-induced vascular hyperpermeability, but there was no marked changes in the degree of the SHLI-induced vascular hyperpermeability after the mice were pretreated with Compound 48/80. CONCLUSION: SHLI in clinic equivalent dose can cause vascular hyperpermeability. Baicalin, forsythin and caffeotannic may not result in the PR of SHLI. The mechanism of the PR maybe relate to that SHLI stimulates histamine release, the activation of leucocyte maybe take part in the SHLI-induced PR, too. Antihistamine drug can prevent the genesis of PR which induced by SHLI.

[Content detection of bacterial endotoxin in two kinds of injection by gelatin technique].

OBJECTIVE: To detect content of bacterial endotoxin in Yuxingcao and Qingkailing injections by specific and nonspecific tachypleus amebocyte lysate technique for in order to investigate the feasibility of specific tachypleus amebocyte lysate technique for detecting bacterial endotoxin in traditional Chinese drug injections. METHOD: Different batches of Yuxingcao and Qingkailing injections were detected by specific and nonspecific tachypleus amebocyte lysate kits. RESULT: Yuxingcao injection could be detected by specific and nonspecific tachypleus amebocyte lysate technique, Whereas Qingkailing injection could be detected only by specific tachypleus amebocyte lysate. CONCLUSION: Using specific tachypleus amebocyte lysate as a substitute for nonspecific tachypleus amebocyte lysate is an effective method for detecting content of bacterial endotoxin in Qingkailing injection.

[Material and mechanisms induced pseudo allergic reactions of Yuxingcao injection].

OBJECTIVE: To investigate the characteristics, sensitizin and the mechanism of pseudo allergic reaction induced by Yuxingcao injection. METHOD: Beagle dogs were randomly assigned to control group, 0.5% tween 80 group, Yuxingcao injection without tween 80 group, Yuxingcao injection included 0.5% tween 80 group. The animals in control group were intravenously injected with saline. The other group were intravenously injected with the corresponding test substances. Observe pseudo anaphylaxis of Beagle dogs within 30 min after administration. Blood pressure and respiration rate of Beagle dogs were measured before and after injection drugs 10 min and 30 min respectively. The pseudo allergic reactions were scored at same time points, and the sera of animals were collected to determine the HIS, CH50 and C5b-9 concentration using ELISA. RESULT: The scores of allergic reaction in 0.5% tween 80 group and Yuxingcao injection included 0.5% tween 80 group was evidently higher than that in control group in 2-5 min after administration. Animals of above two groups showed the symptoms of red swelling on ear part, pruritus, throwing the head, nausea, lapping the tongue, dysphoria and bradykinesia. Some of them had behaved with repose, urination, defecation, cyanosis, the frequency of breathes accelerating and blood pressure decreasing. The rate of pseudo allergic reactions was 100%. Serum CH50 concentration of 0.5% tween 80 group decreased 10 min after injection, while C5b-9 concentration increased. No obvious differences were observed 30 min after injection. There was no significant difference in HIS concentration between control group and treatment groups. CONCLUSION: The pseudo allergic reactions appeared after intravenous 0.5% tween 80 and Yuxingcao injection when mixed with tween 80. Furthermore, Yuxingcao injection without tween 80 did not induce pseudo allergic reactions. It was suggested that the pseudo allergic reactions of Yu Xing Cao Injection was related to the cosolvent tween 80. The pseudo allergic reactions of tween 80 may relate to the activation of complement.