A pilot study of microRNAs expression profile in plasma of patients with hydatid disease: potential immunomodulation of hydatid disease.

Echinococcosis is a zoonotic disease, which seriously endangers human health. The immune game between parasite and host is not fully understood. Exosomes are thought to be one of the ways of information communication between parasite and host. In this study, we attempted to explore the communication between Echinococcus granulosus and its host through the medium of exosomes. We collected plasma from E. granulosus patients (CE-EXO) and healthy donors (HD-EXO) and extracted exosomes from the plasma. The expression profile of miRNA in plasma was determined by second generation sequencing. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) were used to annotate the function of target genes of differential miRNAs. Meanwhile, we co-cultured plasma exosomes from healthy donors and plasma exosomes from E. granulosus patients with Jurkat T cells with or without phytohaemagglutinin (PHA) stimulation. The expression of CD69 on Jurkat T cells was detected by flow cytometry. The results showed that the miRNA of exosomes between healthy donors and E. granulosus patients was significantly different. GO and KEGG were used to annotate the function of target genes of differential miRNAs. The results indicate that many important pathways are involved in inflammation, metabolism, and immune response after parasite infection, such as p53 signaling pathway, PI3K-Akt signaling pathway, and glycolysis/gluconeogenesis. Flow cytometry showed that CE-EXO reduced the expression of CD69 + on Jurkat T cells. Our present results suggest that these differentially expressed miRNAs may be important regulators of parasite-host interactions. Meanwhile, functional prediction of its target genes provides valuable information for understanding the mechanism of host-parasite interactions. These results provide clues for future studies on E. granulosus escape from host immune attack, which could help control E. granulosus infection.

LncRNA KCNQ1OT1 contributes to oxygen-glucose-deprivation/reoxygenation-induced injury via sponging miR-9 in cultured neurons to regulate MMP8.

Our study proposed to investigate the function of potassium voltage-gated channel sub-family Q member 1 opposite strand 1 (KCNQ1OT1) in cerebral ischemia-reperfusion (I/R) injury and the underlying mechanism. We constructed an oxygen-glucose-deprivation/reoxygenation (OGD/R) model using the primary cortical neurons to mimic the cerebral I/R injury in vitro. Small inference RNA (siRNA) was used to silencing KCNQ1OT1. Dual luciferase assay was conducted to verify the interaction between KCNQ1OT1 and miR-9 and interaction between miR-9 and MMP8. CCK8 assay and flow cytometry analysis were applied for determing the viability and apoptosis of neurons, accordingly. QPCR and Western blot were performed to determine the RNA and protein expression. Our outcomes revealed that the expression of KCNQ1OT1 in cultured neurons was notably enhanced after suffered to OGD/R. Knockdown of KCNQ1OT1 weakened OGD/R-induced injury in neurons. Moreover, depletion of KCNQ1OT1 lead to the up-regulation of miR-9 and down-regulation of MMP8. Dual luciferase target validation assays demonstrated that KCNQ1OT1 directly interact with miR-9 and MMP8 is a direct target of miR-9, suggesting that KCNQ1OT1/miR-9/MMP8 might constitute the competing endogenous RNA (ceRNA) mechanism. Knockdown of MMP8 or up-regulation of miR-9 also could weaken OGD/R-induced injury. Furthermore, cells co-transfected with si-KCNQ1OT1, miR-9 mimic and si-MMP8 could significantly abolish the injury on neurons caused by OGD/R. Taken together, our data manifested that KCNQ1OT1 possibly acts as a facilitator in cerebral I/R injury through modulating miR-9/MMP8 axis as a ceRNA.

Development and validation of an UPLC-Q/TOF-MS assay for the quantitation of neopanaxadiol in beagle dog plasma: Application to a pharmacokinetic study.

Neopanaxadiol (NPD), the main panaxadiol constituent of Panax ginseng C. A. Meyer (Araliaceae), has been regarded as the active component for the treatment of Alzheimer's disease. However, few references are available about pharmacokinetic evaluation for NPD. Accordingly, a rapid and sensitive method for quantitative analysis of NPD in beagle dog plasma based on ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry was developed and validated. Analytes were extracted from plasma by liquid-liquid extraction and chromatographic separation was achieved on an Agilent Zorbax Stable Bond C18 column. Detection was performed in the positive ion mode using multiple reaction monitoring of the transitions both at m/z 461.4 → 425.4 for NPD and internal standard of panaxadiol. All validation parameters, such as lower limit of quantitation, linearity, specificity, precision, accuracy, extraction recovery, matrix effect and stability, were within acceptable ranges and the method was appropriate for multitude sample determination. After oral intake, NPD was slowly absorbed and eliminated from circulatory blood system and corresponding plasma exposure was low. Application of this quantitative method will yield the first pharmacokinetic profile after oral administration of NPD to beagle dog. The information obtained here will be useful to understand the pharmacological effects of NPD.

Toxicity assessment of 7 anticancer compounds in zebrafish.

Toxicity is one of the major reasons for failure in drug development. Zebrafish, as an ideal vertebrate model, could also be used to evaluate drug toxicity. In this study, we aimed to show the predictability and highlight novel findings of toxicity in zebrafish model. Seven anticancer compounds, including triptolide (TP), gambogic acid (GA), mycophenolic acid (MPA), curcumin, auranofin, thalidomide, and taxol, were assessed in zebrafish for their toxicity. Three compounds (GA, TP, and taxol) showed highest acute lethality, with 50% lethal concentration ≈ 1 µmol/L. Missing tails, severe pericardial edema, and enlarged yolk sacs were observed in MPA-treated embryos. The development of pectoral fins was severely disturbed in thalidomide-, GA-, and TP-treated embryos. Bradycardia was observed in MPA- and thalidomide-treated groups. Our findings suggested that the zebrafish are a good model for toxicity assessment of anticancer compounds.

[Effect of total saponins from Sanguisorba officinalis on megakaryocyte progenitor cells proliferation, differentiation and relative receptor expression].

OBJECTIVE: To investigate the effects of total saponins from Sanguisorba officinalis (DYS) on hematopoietic cell proliferation, differentiation and the expression level of IL-3R and c-kit. METHOD: Baf3 and 32D cells were cultured with or without IL-3, then the cells were exposed to DYS in different concentrations of 5, 10, 20, 30 and 40 mg x L(-1) for 24, 48, 72 and 96 hours separately. After that, the cell proliferation and differentiation capacity were determinated by the methods of CCK8 and Giemsa staining separately. The effects of DYS on the expression level of IL-3 receptor in Baf3 cells and the expression level of c-kit in 32D cells were determinated using RT-PCR. RESULT: DYS promotes alone proliferation of Baf3 cells and 32D cells after 48 h. In contrast to control cells, 32D cells containing DYS without IL-3 form many large clusters. DYS also increases the proliferation when cultured with IL-3. High concentration of DYS induce alone the differentiation of 32D cells and increase alone the number of the polyploidy megakaryocyte. Moreover, DYS increases alone the expression level of IL-3R in Baf3 cells and the expression level of c-kit in 32D cells separately. CONCLUSION: Our data shows DYS can promote alone proliferation and differentiation of megakaryocyte progenitor cells. The proliferative and differentiative effect of DYS on megakaryocyte progenitor cells is correlated to the up-regulation of IL-3 receptor and c-kit expression.

Green fabrication of modified lignin/zeolite/chitosan-based composite membranes for preservation of perishable foods.

Chitosan, as a kind of naturally occurring green and degradable material for the preservation of perishable foods, was investigated in this study with the objective of enhancing its preservation performances. Herein, lignin was modified using the solvent fractionation method (modified lignin, ML, including ML1-ML3), while natural clinoptilolite zeolite was modified using the alkali modification method (modified clinoptilolite zeolite, MCZ, including MCZ1-MCZ5). After optimizing the conditions, it was discovered that incorporating both ML3 and MCZ3 into pure chitosan-based membranes might be conducive to fabricate chitosan-based composite membranes for the preservation of perishable foods. As-prepared composite membranes possessed better visible light transmittance, antioxidant activity, and carbon dioxide/oxygen selectivity, resulting in improved preservation effects on the model perishable foods such as bananas, cherry tomatoes, and cheeses. These findings might indicate promising applications for chitosan-based composite membranes with modified lignin and zeolite in the field of eco-friendly degradable materials for the preservation of perishable foods.

Circadian disruption during fetal development promotes pathological cardiac remodeling in male mice.

Disruption of circadian rhythms during fetal development may predispose mice to developing heart disease later in life. Here, we report that male, but not female, mice that had experienced chronic circadian disturbance (CCD) in utero were more susceptible to pathological cardiac remodeling compared with mice that had developed under normal intrauterine conditions. CCD-treated males showed ventricular chamber dilatation, enhanced myocardial fibrosis, decreased contractility, higher rates of induced tachyarrhythmia, and elevated expression of biomarkers for heart failure and myocardial remodeling. In utero CCD exposure also triggered sex-dependent changes in cardiac gene expression, including upregulation of the secretoglobin gene, Scgb1a1, in males. Importantly, cardiac overexpression of Scgb1a1 was sufficient to induce myocardial hypertrophy in otherwise naive male mice. Our findings reveal that in utero CCD exposure predisposes male mice to pathological remodeling of the heart later in life, likely as a consequence of SCGB1A1 upregulation.

Lignin-based covalent organic polymers with improved crystallinity for non-targeted analysis of chemical hazards in food samples.

Lignin, the most abundant source of renewable aromatic compounds derived from natural lignocellulosic biomass, has great potential for various applications as green materials due to its abundant active groups. However, it is still challenging to quickly construct green polymers with a certain crystallinity by utilizing lignin as a building block. Herein, new green lignin-based covalent organic polymers (LIGOPD-COPs) were one-pot fabricated with water as the reaction solvent and natural lignin as the raw material. Furthermore, by using paraformaldehyde as a protector and modulator, the LIGOPD-COPs prepared under optimized conditions displayed better crystallinity than reported lignin-based polymers, demonstrating the feasibility of preparing lignin-based polymers with improved crystallinity. The improved crystallinity confers LIGOPD-COPs with enhanced application performance, which was demonstrated by their excellent performances in sample treatment of non-targeted food safety analysis. Under optimized conditions, phytochromes, the main interfering matrices, were almost completely removed from different phytochromes-rich vegetables by LIGOPD-COPs, accompanied by "full recovery" of 90 chemical hazards. Green, low-cost, and reusable properties, together with improved crystallinity, will accelerate the industrialization and marketization of lignin-based COPs, and promote their applications in many fields.

[Simple nucleus pulposus removal for the treatment of prolapsed and displaced lumbar disc herniation].

OBJECTIVE: To explore the clinical effect of the simple nucleus pulposus removal and small incision interlaminar window in the treatment of prolapsed and displaced lumbar disc herniation. METHODS: From February 2016 to February 2018, 35 patients with single-segment prolapse and displaced lumbar disc herniation were treated by the simple nucleus pulposus removal and small incision interlaminar window under general anesthesia. Among them, there were 21 males and 14 females;aged (42±17) years;27 cases of L4,5 segment, 6 cases of L5S1 segment, 2 cases of L3,4 segment;20 cases on the left side, 13 cases on the right side. Modified Macnab standard was used to evaluate postoperative symptoms and functional recovery. RESULTS: All the operations were successful and the operation time was 30 to 60 min with an average of 40 min, the intraoperative blood loss was 10 to 30 ml with an average of 20 ml. All the patients were followed up for 1 to 3 years with an average of 1.2 years. Thirty-five patients with low back pain and lower limb symptoms were significantly relieved or disappeared. According to modified Macnab standard, 29 cases obtained excellent results, 5 good, and 1 fair. CONCLUSION: Applying the concept of minimally invasive operation, small incision interlaminar window and simple nucleus pulposus removal for the treatment of prolapsed and displaced lumbar disc herniation has the advantages of short operation time, definite curative effect, and less trauma. And it is a safe and effective surgical method under the premise of strict control of the indications.

[KeLuoXin medicated serum extract inhibits high glucose-induced proliferation and phosphorylation of ERK1/2 in rat mesangial cell].

OBJECTIVE: To investigate the effects and mechanism of KeLuoXin (KLX) capsule on rat mesangial cell proliferation induced by high glucose. METHODS: Rat mesangial cells (HBZY-1) were cultured in vitro and stimulated with high glucose(30 mmol/L glucose) for different time after the treatment with or without KLX medicated serum extract, the proliferation was assessed by cell counting Kit-8 (CCK-8) and the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK1/2) was detected by Western blot. RESULTS: KLX could significantly inhibit HBZY-1 cell proliferation induced by high glucose (P < 0.05). In addition, KLX could reduce the increase of the phosphorylation level of ERK1/2 (P < 0.01), which was induced by high glucose in HBZY-1 cells. CONCLUSION: KLX can inhibit rat mesangial cell proliferation, and the mechanisms may relate to the interruption of ERK1/2-Mitogen-activated protein kinase (MAPK) pathway.

Allergic sensitization in patients with rhinitis and bronchial asthma in Ningxia region of China.

BACKGROUND: Allergic rhinitis and bronchial asthma are common allergic diseases. The pattern of dominant allergens depends on the degree of urbanization and the geographic region. The present study characterized the allergens of patients with allergic rhinitis and bronchial asthma in Ningxia region of China. METHODS: A total of 309 patients were enrolled in this study. Western blotting assays were performed of the serum samples to evaluate allergen-specific IgE antibody for inhaled and ingested allergens. Statistical analysis was performed to compare the positive rate among different subgroups. RESULTS: Among the 309 patients, 221 of them had positive test results. There were 157 positive cases for ingested allergens and 174 positive cases for inhaled allergens. No significant differences in positive rates were found between the ingested and inhaled allergens. Among the inhaled allergens, Artemisia was the most frequent allergen, followed by fungi and dog hair. Cashew was the most common ingested allergen, followed by crab, mango, and beef. Further analysis showed no significant differences in positive rate between males and females. However, significant differences in positive rate of inhaled and ingested allergens were found between children (1-13 years old) and adults (above 18 years old) (P < .05), while no significant differences were found between the children and teenagers (14-18 years old). For the comparison between teenagers and adults, significant difference in positive rate was found only in the ingested allergen. CONCLUSION: This study provided the characteristics of allergens in Ningxia population, providing clinical and epidemiological data for prevention and treatment of the diseases in the region.

Lycium barbarum Polysaccharide Extracted from Lycium barbarum Leaves Ameliorates Asthma in Mice by Reducing Inflammation and Modulating Gut Microbiota.

This study was designed to explore the impact of Lycium barbarum polysaccharide (LBP) on inflammation and gut microbiota in mice with allergic asthma. Mice were divided into four groups: control group, OVA (ovalbumin) group, Con+LBP group, OVA+LBP group. After 28 days of LBP intervention, mice were euthanized and associated indications were investigated. Histopathological examination demonstrated that LBP reduced lung injury. The results of our current study provide evidence that supplementation with LBP in asthmatic mice decreases TNF, IL-4, IL-6, MCP-1, and IL-17A in plasma and bronchoalveolar lavage fluid (BALF). Sequencing and analysis of gut microbiota indicated that compared with the OVA group, Lactobacillus and Bifidobacterium were increased, but Firmicutes, Actinobacteria, Alistipes, and Clostridiales were decreased in the OVA+LBP group. We also found that gut microbiota were related to inflammation-related factors. Therefore, we speculate that LBP may improve allergic asthma by altering gut microbiota and inhibiting inflammation in mice.

Examining and monitoring paretic muscle changes during stroke rehabilitation using surface electromyography: A pilot study.

Complex neuromuscular changes have been reported to occur in paretic muscles following stroke, but whether and how they can recover under rehabilitation therapy remain unclear. A tracking analysis protocol needs to be designed involving multiple sessions of surface electromyography (sEMG) examinations during the rehabilitation procedure. Following such a protocol, this pilot study is aimed to monitor paretic muscle changes using three sEMG indicators namely clustering index (CI), root mean square (RMS) and medium frequency (MDF). Initially, a single sEMG examination was performed on the abductor pollicis brevis (APB) muscle on both sides of 23 subjects with stroke and one side of 18 healthy control subjects. With these data to establish CI diagnostic criterion, the paretic muscles of all subjects with stroke showed a very board CI distribution pattern from abnormally low values through normality to abnormally high values. Afterwards, 9 out of 23 subjects with stroke had their paretic muscles examined at least twice before and after the treatment. Almost all paretic muscles had an increase of the RMS, a change of the MDF approaching to the value of the contralateral muscle, and a change of the CI returning to its normal range after common rehabilitation treatments. Finally, 4 of the 9 subjects with stroke participated into repeated examinations of their paretic muscles. The combined use of three indicators helped to reveal specific neuromuscular processes contributing to recovery of paretic muscles, due to their complementary diagnostic powers. Furthermore, neuromuscular processes were found to vary across subjects in type, order and timing during rehabilitation. In conclusion, given the 4 cases following the tracking analysis protocol, this pilot study preliminarily demonstrates usability of three sEMG indicators as tools for examining and monitoring stroke rehabilitation procedure in terms of improvements of paretic muscle changes. All the revealed complex neuromuscular processes imply the necessity of applying sEMG examinations in monitoring rehabilitation procedure, with the potential of offering important guidelines for designing better and individualized protocols toward improved stroke rehabilitation.

17ß-Estradiol Inhibits PCSK9-Mediated LDLR Degradation Through GPER/PLC Activation in HepG2 Cells.

Plasma levels of PCSK9 are significantly higher in postmenopausal women. Pharmacologically increased estrogen levels have been shown to lower PCSK9 and LDL-C levels in animals and humans. The action of estrogen suggests that it has the ability to prevent PCSK9-mediated LDLR degradation in liver cells. However, little is known about how estrogen alters PCSK9-mediated LDLR degradation. Here, we report that 17ß-estradiol (ßE2) reduces PCSK9-mediated LDLR degradation by a mechanism that involves activation of the G protein-coupled estrogen receptor (GPER). In cultured HepG2 cells, ßE2 prevented the internalization of PCSK9, which subsequently lead to PCSK9-mediated LDLR degradation. The altered LDLR levels also resulted in an increase in LDL uptake that was not observed in the absence of PCSK9. In addition, we showed that clathrin was rapidly increased in the presence of PCSK9, and this increase was blocked by ßE2 incubation, suggesting rapid recruitment of clathrin in HepG2 cells. PLCγ activation and intracellular Ca2+ release were both increased due to the rapid effect of estrogen. By using a GPER antagonist G15, we demonstrated that the GPER mediates the action of estrogen. Together, the data from this in vitro study demonstrate that estrogen can regulate LDLR levels mainly through GPER activation, which prevents PCSK9-dependent LDLR degradation in HepG2 cells.

[Mechanism of rosiglitasone to improve glucose-uptake of 3T3-L1 adipocyte with insulin resistance induced by dexamethasone and insulin].

OBJECTIVE: To study the mechanism of rosiglitasone to improve glucose-uptake of 3T3-L1 adipocyte with insulin resistance induced by dexamethasone and insulin. METHODS: Insulin resistance was induced to 3T3-L1 adipocyte after chronic treatment of dexamethasone and insulin. The insulin resisted 3T3-L1 adipocyte was treated with 10(-5) mol/L of rosiglitasone for 48 h. The mRNA, protein of glucose transporter GLUT4 and CAP gene were then examined. RESULTS: (1) Rosiglitasone increased the expression of GLUT4 mRNA and protein inhibited by dexamethasone and insulin although it had not reached a normal level. (2) Rosiglitasone increased the mRNA of CAP, which remained unchanged during insulin resistance. CONCLUSION: Rosiglitasone, an insulin sensitizer, might up-regulate GLUT4 and CAP along with the peroxisome proliferators activated receptor gamma (PPAR-gamma), which not only increases the GLUT4, but also activates the CAP-depended signaling pathway; improves the translocating of GLUT4 to the cell membrane; and increases the ability of glucose-uptake of cells.

[Synthesis of phenyloxyisobutyric acid derivatives and their antidiabetic activity in vitro].

AIM: To design and synthesize new phenyloxyisobutyric acid analogues as antidiabetic compounds. METHODS: Eight new target compounds were synthesized by combination of lipophilic moieties and acidic moiety with nucleophilic replacement or Mitsunobu condensation. The eight compounds were confirmed by 1H NMR, 13C NMR, IR and MS. RESULTS: In vitro insulin-sensitizing activity (3T3-L1 adipocyte) demonstrated, that the cultured glucose concentration of up-clear solution detected with GOD-POD assay were 5.942, 6.339, 6.226 and 6.512 mmol x L(-1), respectively, when rosiglitazone, pioglitazone, compounds A and B were added to the insulin-resistant system. CONCLUSION: In vitro insulin-sensitizing activity of target compound A is in between that of rosiglitazone and pioglitazone, and activity of target compound B is slightly less than that of pioglitazone.

[Perioperative intensive statin therapy improves outcomes in patients with ischemic stroke undergoing middle cerebral artery stent implantation].

OBJECTIVE: To investigate whether intensive statin therapy during the perioperative period improves outcomes in patients undergoing middle cerebral artery (MCA) stent implantation for ischemic stroke. METHODS: Forty patients with ischemic stroke undergoing delayed stent implantation in our department from January, 2010 to November, 2014 were randomized to intensive statin group (atorvastatin, 80 mg/day, 3 days before till 3 days after intervention; n=20) and standard therapy group (atorvastatin, 20 mg/day, n=20). All the patients received long-term atorvastatin treatment thereafter (20 mg/day). Serum levels of C-reactive protein (CRP), vascular cell adhesion molecule-1 (VCAM-1), and soluble extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) were measured at 24 h before and 24 h after the intervention. The primary end point was procedure-related intra-stent thrombosis, 1-month incidence of major adverse cerebrovascular events (stroke, transient ischemic attack, in-stent restenosis, death or unplanned revascularization). RESULTS: The basic clinical data were similar between the two groups before the intervention (P>0.05). In the intensive therapy group, the levels of CRP, VCAM-1, and sCD147 were significantly lower at 24 h after the intervention than the levels before intervention (P<0.05) and the postoperative levels in the standard therapy group (P<0.05). The levels of CRP, VCAM-1, and sCD147 were all increased after the intervention in the standard therapy group (P>0.05). The incidence of primary end point was lower in intensive therapy group than in standard therapy group (P<0.05). CONCLUSION: In patients undergoing MCA intravascular stent implantation for ischemic stroke, perioperative intensive statin therapy improves the patients' outcomes, reduces the levels of CRP, VCAM-1 and sCD147 molecules, and lowers the incidences of cerebrovascular events.

Identification of TMEM230 mutations in familial Parkinson's disease.

Parkinson's disease is the second most common neurodegenerative disorder without effective treatment. It is generally sporadic with unknown etiology. However, genetic studies of rare familial forms have led to the identification of mutations in several genes, which are linked to typical Parkinson's disease or parkinsonian disorders. The pathogenesis of Parkinson's disease remains largely elusive. Here we report a locus for autosomal dominant, clinically typical and Lewy body-confirmed Parkinson's disease on the short arm of chromosome 20 (20pter-p12) and identify TMEM230 as the disease-causing gene. We show that TMEM230 encodes a transmembrane protein of secretory/recycling vesicles, including synaptic vesicles in neurons. Disease-linked TMEM230 mutants impair synaptic vesicle trafficking. Our data provide genetic evidence that a mutant transmembrane protein of synaptic vesicles in neurons is etiologically linked to Parkinson's disease, with implications for understanding the pathogenic mechanism of Parkinson's disease and for developing rational therapies.

[Design, synthesis and insulin-sensitizing activity of some peroxisome proliferator-activated gamma agonists].

AIM: To find new peroxisome proliferator-activated y agonists with high activity and low toxicity. METHODS: Based on JTT-501 and JTT-20993, new isoxazolidine-3,5-dione and noncyclic 1,3-dicarbonyl compounds were designed and synthesized. Their insulin-sensitizing activities were evaluated. RESULTS: Eight new compounds were obtained. The structures of synthesized compounds were characterized by NMR, MS and IR. Four compounds (1A-4A) showed insulin-sensitizing activities. CONCLUSION: Compounds (1A and 3A) showed excellent insulin-sensitizing activities and should be worth further investigation.

[Effects of estrogen on acrosome reaction and intracellular calcium in human spermatozoa and the possible mechanism concerned].

OBJECTIVE: To study the effects of 17beta-estradiol (E2) on the activation of human spermatozoa and the possible mechanism. METHODS: Spermatozoa of fertile men were stimulated by E2 and the membrane impermeable conjugated 17beta-estradiol (E2-BSA) respectively. Then the acrosme reaction (AR) rate was determined by triple-stain technique and the intracellular calcium concentration ([Ca2+]i) in spermatozoa was measured by flow cytometry. The changes of [Ca2+]i were measured again following the addition of E2 and E2-BSA in the calcium-free medium. RESULTS: E2 significantly increased the AR rate and the [Ca2+]i in capacitated human spermatozoa, while it had no effects on the uncapacitated spermatozoa. E2-BSA also promoted AR and increased [Ca2+]i in the capacitated human spermatozoa. Additionally, the change of [Ca2+]i was absent in case that human spermatozoa were stimulated by E2 or E2-BSA in the calcium-free medium. CONCLUSION: Estrogen can activate human spermatozoa and such activation is probably mediated by the integrating of estrogen with the specific sites on human spermatozoa and thus causing the influx of extracellular calcium.