Modulation of gut microbiota by crude mulberry polysaccharide attenuates knee osteoarthritis progression in rats.

Mulberry (Morus alba L.), a kind of common fruits widely cultivated worldwide, has been proven various biological activities. However, its potential role in the progression of knee osteoarthritis (KOA) remains unclear. This study aims to investigate the potential protective effects of crude polysaccharide extracted from mulberry fruit, referred to as a complex blend of polysaccharides and other unidentified extracted impurities, on KOA progression. The KOA rats were established by injection of 1 mg sodium monoiodoacetate into knee, and administrated with crude mulberry polysaccharide (Mup) by gastric gavage for 4 weeks. Furthermore, intestinal bacteria clearance assay (IBCA) and fecal microbiota transplantation were conducted for the evaluation of the effect of gut microbiota (GM) on KOA. Our findings demonstrated that Mup, particularly at a dosage of 200 mg/kg, effectively improved abnormal gait patterns, reduced the level of inflammation, mitigated subchondral bone loss, restored compromised joint surfaces, alleviated cartilage destruction, and positively modulated the dysregulated profile of GM in KOA rats. Moreover, IBCA compromised the protective effects of Mup, while transplantation of fecal bacteria from Mup-treated rats facilitated KOA recovery. Collectively, our study suggested that Mup had the potential to ameliorate the progression of KOA, potentially through its modulation of GM profile.

Schisandrae chinensis fructus extract protects against hepatorenal toxicity and changes metabolic ions in cyclosporine A rats.

While cyclosporin A (CsA) is an effective immunosuppressive agent, its clinical application is limited by serious hepatorenal toxicity. However, Schisandrae chinensis fructus extract (SCE) has been previously shown to alleviate the hepatorenal damage caused by CsA. In this study, we aimed to evaluate the protective effects of SCE against hepatorenal toxicity induced by CsA. Our results revealed that SCE can prevent and treat CsA-induced liver and kidney injury. Furthermore, SCE inhibited the upward trend of dUDP and CDP-ethanolamine in the urine of CsA rats, pathways of which are involved in pyrimidine and glycerophospholipid metabolism. We finally confirmed that this protection of SCE was regulated by the activation of Nrf2 signaling pathway and the inhibition of apoptosis. In summary, our findings indicated that SCE may effectively prevent and treat hepatorenal toxicity caused by CsA. In addition, metabolomic techniques identified potential biomarkers for the occurrence of hepatorenal toxicity in CsA rats.

IL-17 in pancreatic disease: pathogenesis and pharmacotherapy.

Increasing evidence highlights the role of the interleukin (IL)-17 family in pancreatic diseases. IL-17A induces acinar cell injury directly, recruits neutrophils, and cooperates with other inflammatory factors to exacerbate pancreatic inflammation. It also triggers islet ß-cell apoptosis and nitric oxide-dependent cytotoxicity, thus aggravating islet inflammation. IL-17A seems to have different roles in pancreatic intraepithelial neoplasia (PanIN) and pancreatic cancer (PC). IL-17A participates in the progression of acinar-ductal metaplasia (ADM) and PanIN, but not related to the characteristics of PC stem cells and the overall survival of patients. Acting similar to IL-17A, IL-17B accelerates the invasion and metastasis of PC, and predicts prognosis of PC and the therapeutic effect of gemcitabine. Herein, we review the current understanding of the pathogenesis of IL-17 in pancreatitis, type 1 diabetes mellitus (T1DM), and PC, as well as potential pharmacotherapy targeting IL-17 and its receptors in pancreatic diseases. The findings summarized in this article are of considerable significance for understanding the essential role of IL-17 in pancreatic diseases.

The comparison of extraction methods of ganjiang decoction based on fingerprint, quantitative analysis and pharmacodynamics.

BACKGROUND: Ulcerative colitis (UC) is a chronic nonspecific inflammatory disease of the colon and rectum with unknown etiology, and its symptoms include bloody diarrhea, abdominal pain, and hematochezia. Traditional Chinese medicine compound has a good therapeutic, multi-target effect on UC. Ganjiang decoction (GD), which is a traditional classic prescription in China, contains Zingiberis Rhizoma, Angelicae Sinensis Radix, Coptidis Rhizoma, Phellodendri Chinensis Cortex, Sanguisorbae Radix, Granati Pericarpium, and Asini Corii Colla and could be used to treat symptoms of UC. This study aimed to conduct a preliminary study before GD colon-targeted preparation, to explore the relationship between extraction method and efficacy of GD. METHODS: High-performance liquid chromatography (HPLC) was used for the fingerprinting of five preparation methods of GD. HPLC and gas chromatography were used to quantitatively analyze the important chemical components of GD and compare their differences. Mice with UC induced by dextran sulphate sodium salt received the extracts from the five preparation methods of GD via gavage. Disease activity index (DAI) score, colonic length, relative weight of spleen, pathological analysis results, inflammatory factors, therapeutic effect of the five preparation methods of GD, and their relationship with extraction process were compared. RESULTS: Cluster analysis revealed that the content of the components extracted by traditional extraction methods was significantly different from the other four methods. The third and fifth preparation methods extracted Coptidis Rhizoma and Phellodendri Chinensis Cortex with 50% ethanol to obtain more alkaloids. In the fourth and fifth methods, more volatile oils were detected by adding Zingiberis Rhizoma and Angelicae Sinensis Radix fine powder. According to DAI score, colonic length, relative weight of spleen, pathological analysis results, and inflammatory factors, the third method showed a good therapeutic effect, while the fifth method had the best therapeutic effect. CONCLUSIONS: The results showed that the difference of the five extracts of GD in the efficacy of DSS-induced UC in mice was closely related to the extraction method. Our study improved the extraction process of GD and provided a foundation for the process of enteric-soluble preparations and a new idea for traditional Chinese medicine compound preparation.

Experimental evidence and network pharmacology identify the molecular targets of Tong Sheng tablets in cerebral ischemia reperfusion injury.

PURPOSE: Tong Sheng tablets (TSTs) have long been used for treating cerebral ischemic reperfusion injury (CIRI) in clinic, but the underlying mechanism remains unknown. Therefore, in this study, TSTs were evaluated systematically using chemical analysis, network pharmacology and classical pharmacology. METHODS: The first part was TSTs quality control including TSTs fingerprint establishment and chemicals identification. In the second part, network pharmacology analysis and bioinformatics were combined to construct a compound-target-disease network, which can screen out key targets or pathways, revealing complex molecule mechanism of TSTs. The last part was experiment verification. Classical pharmacology of TSTs was investigated in vivo to verify the results of network pharmacology. RESULTS: (1) Fingerprints of TSTs were established, and 11 characteristic peaks were identified using HPLC. (2) Network pharmacology and bioinformatics suggested that the protection of TSTs in treating CIRI might be related to regulation of oxidative stress, inflammation and apoptosis, and some key molecules such as Nrf2, IL-1ß, TNF, Bcl-2 and Cyt-C involved in the pathways. (3) TSTs significantly improved neurologic behavior scores, decreased the areas of ischemic necrosis and neuronal necrosis, and increased Nissl body counts. Besides, TSTs significantly decreased pro-inflammatory cytokine (IL-1ß, TNF-α) and pro-oxidative product levels (LPO, MDA) and increased anti-oxidative product levels (NO, SOD). TSTs downregulated the protein expressions of Nrf2 and HO-1. Meanwhile, TSTs reduced apoptotic cell counts, downregulated the protein expressions of Cyt-C and Bax, and upregulated the protein expression of Bcl-2. In terms of autophagy, TSTs enhanced LC-3B protein expression. CONCLUSION: The present results illustrated that TSTs effectively alleviated CIRI, and the underlying mechanism might be associated with multiple molecular pathways. Herein, we established a primary pattern for studying Chinese herbal compounds and provided basic guidance for future investigation.

A Multidimensional Characterization of E3 Ubiquitin Ligase and Substrate Interaction Network.

E3 ubiquitin ligases (E3s) play a critical role in molecular and cellular mechanisms. However, a large number of E3-substrate interactions (ESIs) remain unrevealed. Here, we integrated the increasing omics data with biological knowledge to characterize and identify ESIs. Multidimensional features were computed to obtain the association patterns of ESIs, and an ensemble prediction model was constructed to identify ESIs. Comparison with non-ESI cases revealed the specific association patterns of ESIs, which provided meaningful insights into ESI interpretation. Reliability of the prediction model was confirmed from various perspectives. Notably, our evaluations on leucine-rich repeat family of F box (FBXL) family were consistent with a proteomic study, and several substrates for SKP2 and an orphan E3 FBXL6 were experimentally verified. Moreover, a cancer hallmark ESI landscape was studied. Taken together, our study catches a glimpse at the omics-driven ESI association patterns and provides a valuable resource (http://www.esinet.dicp.ac.cn/home.php) to assist ubiquitination research.

Identification and characterization of differentially expressed genes in hepatopancreas of oriental river prawn Macrobrachium nipponense under nitrite stress.

The oriental river prawn Macrobrachium nipponense is a highly adaptable, tolerant, and fecund freshwater prawn that inhabits a wide range of aquatic environments. The hepatopancreas of crustaceans is not only a site for secretion of digestive enzymes, and also plays important roles in several metabolic processes, such as lipid and carbohydrate metabolism. It is the main organ for the detoxification and immunity. In this study, high-throughput sequencing techniques were used to detect the effect of nitrite stress (10 mg/L nitrite-N for 48 h) on gene expression in the hepatopancreas of M. nipponense. A total of 13,769 million reads were harvested, and 94,534 transcripts were de novo assembled using Trinity software and produced 56,054 non-redundant transcripts. A total of 825 differentially expressed genes were obtained comparing 48 h nitrite stress with control group. In the analysis of GO and KEGG database, significant differences were found in 49 pathways. Immune-related pathways under nitrite stress included arginine and proline metabolism, glutamate metabolism, Jak-Stat signaling pathway, endocytosis, wnt signaling pathway, RIG-I-like receptor signaling pathway, TGF-beta signaling pathway, GnRH signaling pathway and phagosome. Apoptosis-related pathway was also significantly altered, such as lysosome and apoptosis. Remarkably, nitrite stress altered the expression patterns of key apoptosis genes (tetraspanins-like protein, LAMP, CD63, caspase 3C and Caspase 1) and immune genes (Serine proteinase-like protein, C-type lectin, daf-36, SOCS-2, alpha-2-macroglobulin), confirmed that nitrite-stress induce immune response and eventually even apoptosis. This study provided a new insight into the role of hepatopancreas in crustaceans, and further investigation will continue.

Metabolomics profiling of metformin-mediated metabolic reprogramming bypassing AMPKα.

BACKGROUND: Metformin is a first-line drug for treating type 2 diabetes and has gained considerable interest as a potential anticancer agent. Increasing evidence suggests that metformin antagonizes diabetes and tumors through disrupting metabolic homeostasis and altering energy state. However, whether AMP activated protein kinase (AMPK) contributes to such effects of metformin remains controversial. METHODS: We performed integrative metabolomics analyses to systematically examine the effects of metformin on metabolic pathways in Prkaa1 wild type (WT) and knock-out (KO) mouse embryonic fibroblast (MEF) cells as well as human cells based on gas chromatography-mass spectrometry and capillary electrophoresis-mass spectrometry (CE-MS). RESULTS: Metformin treatment induced metabolic reprogramming and reduced the energy state of both Prkaa1 WT and KO MEF cells, as evidenced by suppressed tricarboxylic acid (TCA) cycle, elevated lactate production as well as decreased NAD+/NADH ratio. Additionally, metabolic flux analysis also showed that metformin Ampkα-independently increased metabolic flux from glucose to lactate and decreased metabolic flux from acetyl-CoA to TCA cycle as well as from pyruvate to malate. Moreover, metformin Ampkα-dependently upregulated P-Acc but Ampkα-independently inhibited the levels of P-mTor, P-S6, Lc3, Atgl and P-Erk in MEF cells. Similarly, we demonstrated that a commonly used AMPK agonist 5-Aminoimidazole-4-carboxamide ribonucleotide (AICAR) and fetal bovine serum (FBS) starvation, as a common model for energy stress, both led to Ampkα-independent metabolism alterations in MEF cells. Furthermore, these effects of metformin were also confirmed in human hepatocellular carcinoma (HCC) cells as well as in MCF10A shControl and shPRKAA1 cells. Importantly, we found that metformin could obviously inhibit colony conformation of HCC cells in an Ampkα-independent manner. CONCLUSIONS: Our data highlight a comprehensive view of metabolic reprogramming mediated by metformin as well as AICAR. These observations suggest that metformin could affect cellular metabolism largely bypassing Ampkα, and may provide a new insight for its clinical usage.

Effects of berberine on tumor growth and intestinal permeability in HCT116 tumor-bearing mice using polyamines as targets.

The prognosis of colorectal cancer (CRC) is seriously affected by high intestinal mucosal permeability accompanied by increasing tumor load. Berberine, a natural plant-derived product, can protect the intestinal mucosal barrier and suppress tumor growth, but its effects on the intestinal mucosal barrier dysfunction of CRC have not yet been evaluated. Herein, we assessed the effects of berberine on the intestinal mucosal permeability of HCT116 tumor-bearing mice and the underlying mechanism. Berberine (6.25, 12.5, 25 mg/kg) was administered to tumor-bearing mice for 3 weeks by intraperitoneal injection, and saline was given to controls and models. Compared with the control group, tumor-bearing mice had increased intestinal mucosal permeability in the third week. Meanwhile, the body weight decreased by 4%-7%, the concentration of D-lactic acid in plasma increased, and the expressions of ZO1 and Occludin were down-regulated. The intestinal mucosa was impaired. Compared with the model group, berberine inhibited tumor growth in a dose-dependent manner (6.25, 12.5, 25 mg/kg), reduced the permeability of intestinal mucosa, and alleviated intestinal mucosal damage. HPLC showed that berberine decreased the content of polyamines in tumor tissue, whereas increased that in intestinal mucosa tissue. Western blot showed that berberine inhibited the expressions of ODC, C-MYC and HIF-1α, but up-regulated those of OAZ1 and SSAT. In short, berberine may exert antitumor effects by suppressing tumor growth and elevating the intestinal mucosal permeability.

USP10 suppresses tumor progression by inhibiting mTOR activation in hepatocellular carcinoma.

Dysregulation of deubiquitination pathway is associated with poor prognosis in cancers such as hepatocellular carcinoma (HCC). The mammalian target of rapamycin, mTOR, has become an attractive cancer therapeutic target in HCC. However, whether and how aberrant expression of deubiquitination pathway regulates mTOR pathway has remained elusive. Here we report that ubiquitin-specific protease 10 (USP10) functions as a tumor suppressor which inhibits mTOR pathway by stabilizing PTEN and AMPKα in HCC cells. Mechanistically, USP10 interacts and stabilizes PTEN and AMPKα by inhibiting their polyubiquitylation. This stabilization in turn inhibits AKT phosphorylation and mTOR Complex1 (mTORC1) activation. In human liver cancer, USP10 expression is downregulated in HCC tumor tissues across three independent HCC cohorts, and lower-expression of USP10 will generate poor prognosis outcome. Collectively, our results uncover an undescribed mechanism where USP10, as a tumor suppressor, negatively regulates mTORC1 activation and AKT phosphorylation by stabilizing AMPKα and PTEN in HCC cells. This study sheds light on the theoretical basis of mTOR signaling pathway-oriented targeting treatment in clinic.

Identification of SPOP related metabolic pathways in prostate cancer.

Speckle-type POZ protein (SPOP), as a cullin-based E3 ubiquitin ligase, has been identified as one of the most frequently mutated genes in prostate cancer (PCa). However, whether SPOP mutations contribute to metabolic reprogramming in PCa remains unknown. Here, integrated studies of transcriptomics and metabolomics as well as lipidomics were performed in matched PCa tumor (PCT) and adjacent non-tumor (ANT) tissues, followed by correlation analysis of SPOP mutations with altered metabolic pathways in SPOP-mutated PCa patients. Interestingly, transcriptomics profiling showed that all SPOP mutations (with 16.7% frequency, 11/66) occurred at the conserved residues in the substrate binding domain of meprin and TRAF homology (MATH). The results of integrated analysis indicated that three metabolic pathways, including tricarboxylic acid (TCA) cycle, fatty acid metabolism and glycerophospholipid metabolism, exhibited obvious upregulation in SPOP-mutated PCT tissues. Furthermore, both correlation analyses based on integrated data and cBioportal revealed that FH, ELOVL2 and ACADL genes might be involved in SPOP-mutation-related upregulation of these metabolic pathways. Taken together, our study provided new insights in understanding the relationship between metabolic pathways and SPOP mutations in PCa.

Attenuation of cyclosporine A induced nephrotoxicity by schisandrin B through suppression of oxidative stress, apoptosis and autophagy.

Cyclosporine A (CsA) is a potent immunosuppressive agent whose clinical usage is limited by nephrotoxicity. Schisandrin B (SchB), isolated from the fruit of Schisandra chinensis, is a natural compound with multiple pharmacological activities that has been shown to attenuate organ injury caused by CsA. Hence, the primary objective of the current study was to evaluate whether SchB has a cytoprotective effect on CsA-induced nephrotoxicity in human proximal tubular epithelial cell line (HK-2). This study demonstrated that pre-incubation of HK-2 cells with 2.5-10.0µM SchB ameliorated CsA induced cytotoxicity caused by oxidative stress as evidenced by reduced levels of intracellular reactive oxygen species (ROS) and LDH release along with increased levels of mitochondrial membrane potential (ΔΨm) and glutathione (GSH). Also, it was demonstrated that nuclear factor erythroid 2-related factor 2 (Nrf2) activation was involved in modulating cellular oxidative stress, where SchB promoted Nrf2 translocation into the nucleus and downstream target gene expression of heme oxygenase-1 (HO-1), NAD(P)H:quinone oxidoreductase 1 (NQO1) and Glutamate-cysteine ligase modifier subunit (GCLM). Additionally, SchB was found to enhance cell survival via reducing apoptosis rate as well as recover the CsA induced blockade of autophagic flux. Collectively, these findings demonstrated that SchB mediated alleviation of CsA induced nephrotoxicity by preventing the accumulation of ROS by way of suppressing oxidative stress, apoptosis and autophagy.

Apigenin Inhibits Human SW620 Cell Growth by Targeting Polyamine Catabolism.

Apigenin is a nonmutagenic flavonoid that has antitumor properties. Polyamines are ubiquitous cellular polycations, which play an important role in the proliferation and differentiation of cancer cells. Highly regulated pathways control the biosynthesis and degradation of polyamines. Ornithine decarboxylase (ODC) is the rate-limiting enzyme in the metabolism, and spermidine/spermine-N1-Acetyl transferase (SSAT) is the rate-limiting enzyme in the catabolism of polyamines. In the current study, the effect of increasing concentrations of apigenin on polyamine levels, ODC and SSAT protein expression, mRNA expression, cell proliferation and apoptosis, and the production of reactive oxygen species (ROS) was investigated in SW620 colon cancer cells. The results showed that apigenin significantly reduced cell proliferation, decreased the levels of spermidine and spermine, and increased previously downregulated putrescine contents. Apigenin also enhanced SSAT protein and mRNA levels and the production of reactive oxygen species in SW620 cells, though it had no significant effect on the levels of ODC protein or mRNA. Apigenin appears to decrease the proliferation rate of human SW620 cells by facilitating SSAT expression to induce polyamine catabolism and increasing ROS levels to induce cell apoptosis.

Pharmacokinetic and nephroprotective benefits of using Schisandra chinensis extracts in a cyclosporine A-based immune-suppressive regime.

Cyclosporine A (CsA) is a powerful immunosuppressive drug. However, nephrotoxicity resulting from its long-term usage has hampered its prolonged therapeutic usage. Schisandra chinensis extracts (SCE) have previously been used in traditional Chinese medicine and more recently coadministered with Western medicine for the treatment of CsA-induced side effects in the People's Republic of China. This study aimed to investigate the possible effects of SCE on the pharmacokinetics of CsA in rats and elucidate the potential mechanisms by which it hinders the development of CsA-induced nephrotoxicity. A liquid chromatography/tandem mass spectrometry method was developed and validated for determining the effect of SCE on the pharmacokinetics of CsA. Male Sprague Dawley rats, which were administered with CsA (25 mg/kg/d) alone or in combination with SCE (54 mg/kg/d and 108 mg/kg/d) for 28 days, were used to evaluate the nephroprotective effects of SCE. Our study showed that SCE increased the mean blood concentration of CsA. Furthermore, we found that the concomitant administration of SCE alongside CsA prevented the disruption of catalase activity and reduction in creatinine, urea, renal malondialdehyde, and glutathione peroxidase levels that would have otherwise occurred in the absence of SCE administration. SCE treatment markedly suppressed the expression of 4-hydroxynonenal, Bcl-2-associated X protein, cleaved caspase 3, and autophagy-related protein LC3 A/B. On the other hand, the expression of heme oxygenase-1, nuclear factor erythroid 2-related factor 2 (Nrf2), and P-glycoprotein was enhanced by the very same addition of SCE. SCE was also able to increase the systemic exposure of CsA in rats. The renoprotective effects of SCE were thought to be mediated by its antiapoptotic and antioxidant abilities, which caused the attenuation of CsA-induced autophagic cell death. All in all, these findings suggest the prospective use of SCE as an effective adjunct in a CsA-based immunosuppressive regimen.

[Effect of dendrobium mixture on hypoglycemic and the apoptosis of islet in rats with type 2 diabetic mellitus].

OBJECTIVE: To study the effect of Dendrobium mixture on hypoglycemic and the apoptosis of islet in rats with type 2 diabetic mellitus. METHODS: Type 2 diabetes mellitus models were induced by high sugar and fat diet and low dose intraperitoneal injection of streptozotocin (STZ) in rats, and treated with Dendrobium mixture (5, 10, 20 g/kg) by intragastric administration. Observed islet cell morphology with histopathological techniques and tested the apoptosis of islet cells by MTT and Annexin V/PI method. CONCLUSION: Dendrobium mixture could reduce the levels of blood glucose, triglyceride and glucosylated serum protein effectively and significantly improve the modeling structure and function of rat pancreatic tissue. The apoptotic islet cells was significantly reduced (P < 0.01) in treatment group compared with the model group. RESULTS: Dendrobium mixture have a hypoglycemic effect on rat models of type 2 diabetes. It can protect and restore the structure and function of pancreatic tissue.

Molecular cloning, characterization and expression of cathepsin D from grass carp (Ctenopharyngodon idella).

Cathepsin D is a lysosomal aspartic proteinase which participates in various degradation functions within the cell. In this current study, we cloned and characterized the complete cDNA of grass carp cathepsin D through 5'- and 3'-RACE. The cathepsin D contained a 56 bp 5' terminal untranslated region (5'-UTR), a 1197 bp open reading frame encoding 398 amino acids, and a 394 bp 3'-UTR. Grass carp cathepsin D shared high similarity with those from other species, and showed the highest amino acid identity of 91% to Danio rerio. Unlike many other organisms, the grass carp cathepsin D contains only one N-glycosylation site closest to the N-terminal. Real-time quantitative RT-PCR demonstrated that Cathepsin D expressed in all twelve tissues (bladder, brain, liver, heart, gill, muscle, fin, eye, intestines, spleen, gonad and head kidney). The relative expression levels of Cathepsin D in gonad and liver were 26.58 and 24.95 times as much as those in fin, respectively. The expression level of Cathepsin D in muscle approximately 16-fold higher, in intestines and spleen were 12-fold higher. The cathepsin D expression showed an upward trend during embryonic development. After challenged with Aeromonas hydrophil, the expression of grass carp cathepsin D gene showed significant changes in the four test tissues (liver, head kidney, spleen and intestines). The fact that the bacterial infection can obviously improve the cathepsin D expression in immune-related organs, may suggest that cathepsin D plays an important role in the innate immune response of grass carp.

[Relieving pre-exam anxiety syndrome with wrist-ankle acupuncture: a randomized controlled trial].

BACKGROUND: Pre-exam anxiety syndrome is a common condition occurring in pre-exam students and directly affects their examination performance and physical state. Wrist-ankle acupuncture has significant therapeutic effects in treating mental disorders and may also relieve the symptoms of pre-exam anxiety syndrome. OBJECTIVE: To assess the therapeutic effect of wrist-ankle acupuncture on pre-exam anxiety syndrome. DESIGN, SETTING, PARTICIPANTS AND INTERVENTIONS: A total of 60 students who met the inclusion criteria of pre-exam anxiety syndrome were enrolled from a university in Shanghai and they were randomly divided into treatment group and control group. There were 30 cases in each group, and no case failed to follow-up. In the treatment group, wrist-ankle acupuncture was adopted to point upper 1 bilaterally (impression between flexor carpi ulnaris tendon and ulnar margin), and there was no requirement for Deqi (arrival of qi). In the control group, sham acupuncture was adopted. The treatment was applied 3 times totally in both groups one week before the exam, once every other day, each time with the needles retained for 30 min. MAIN OUTCOME MEASURES: The therapeutic effects were compared between two groups. Before and after 3 treatments, Sarason Test Anxiety Scale (TAS) and Expectation and Treatment Credibility Scale (ETCS) were measured and evaluated. RESULTS: The therapeutic effect experienced by the treatment group was better than that of the control group (P<0.05). There were no statistically significant differences in TAS and ETCS before treatment between the two groups. The scores of TAS after treatment in two groups were higher than those before treatment (P<0.05, P<0.01). There were statistical differences in TAS absolute difference and TAS relative difference between the two groups and the treatment group had better results (P<0.05, P<0.01). After treatment, patients in the treatment group had higher scores in ETCS than those in the control group (P<0.05, P<0.01). No adverse reaction was reported. CONCLUSION: Wrist-ankle acupuncture can relieve the symptoms of pre-exam anxiety syndrome significantly, and this therapy is highly safe.