This study examined the expression and distribution of angiopoietin-1/-2 (Ang-1/-2) in the endometrium of early pregnant mice. The expression of Ang-1/-2 was detected by immunohisto-chemical staining and in situ hybridization respectively. Computerized image analysis system was used to measure the average optical intensity of Ang-1/-2 in endometria at different time points after gestation. Mice were randomly divided into 5 groups: control group, D2 group (2 days after pregnancy), D4 group (4 days after pregnancy), D6 group (6 days after pregnancy) and D8 group (8 days after pregnancy), each containing 15 mice. The results showed that the expression of Ang-1 and Ang-2 was very different among 4 groups (P<0.01). Immunohistochemical staining revealed that Ang-1 was localized in the cytoplasma of stromal cells 2 days after pregnancy (day 2), and in luminal epithelial cells on day 4. The protein of Ang-2 was mainly expressed in the cytoplasma of glandular epithelia and stromal cells. With gestation time, the positive reactions of Ang-1/-2 were stronger in the endometria of the pregnant mice (P<0.01). In situ hybridization showed Ang-1 mRNA in stromal cells on day 2. Hybridization signal was localized in both stromal cells and vessel epithelial cells on day 4; Ang-2 mRNA was expressed in stromal cells and glandular epithelia on day 2; high mRNA levels appeared in stromal cells, glandular epithelia and vascular endothelia on day 4; an increasing in mRNA expression of Ang-1/-2 was observed on day 6 and day 8 (P<0.01). It is suggested that Ang-1/-2 may play an important role in the cross-talk between blastocyst and maternal endometrium during the process of embryo implantation.

This study examined the expression and distribution of angiopoietin-1/-2 (Ang-1/-2) in the endometrium of early pregnant mice. The expression of Ang-1/-2 was detected by immunohistochemical staining and in situ hybridization respectively. Computerized image analysis system was used to measure the average optical intensity of Ang-1/-2 in endometria at different time points after gestation. Mice were randomly divided into 5 groups: control group, D2 group (2 days after pregnancy), D4 group (4 days after pregnancy), D6 group (6 days after pregnancy) and D8 group (8 days after pregnancy), each containing 15 mice. The results showed that the expression of Ang-1 and Ang-2 was very different among 4 groups (P＜0.01). Immunohistochemical staining revealed that Ang-1 was localized in the cytoplasma of stromal cells 2 days after pregnancy (day 2), and in luminal epithelial cells on day 4. The protein of Ang-2 was mainly expressed in the cytoplasma of glandular epithelia and stromal cells. With gestation time, the positive reactions of Ang-1/-2 were stronger in the endometria of the pregnant mice (P＜0.01). In situ hybridization showed Ang-1 mRNA in stromal cells on day 2. Hybridization signal was localized in both stromal cells and vessel epithelial cells on day 4; Ang-2 mRNA was expressed in stromal cells and glandular epithelia on day 2; high mRNA levels appeared in stromal cells, glandular epithelia and vascular endothelia on day 4; an increasing in mRNA expression of Ang-1/-2 was observed on day 6 and day 8 (P＜0.01). It is suggested that Ang-1/-2 may play an important role in the cross-talk between blastocyst and maternal endometrium during the process of embryo implantation.

OBJECTIVETo study the effect of rhTNF-alpha on human sperm mitochondrial function and motility in vitro.

METHODSFifty-six semen samples collected by masturbation were analyzed according to WHO protocols. Semen samples from 40 healthy men were prepared using Percoll centrifugation. Sperm suspension was diluted to a concentration of 10 x 10(6)/ml in Ham's F10 medium. Sperm samples were incubated with rhTNF-alpha solution (final concentration 0.03 microg/L, 0.06 microg/L, 0.09 microg/L and 0.27 microg/L, respectively) for 0.5 h, 1 h, 2 h, 3 h and 4 h at 37 degrees C in 5% CO2, and comparative studies were made with a control group. Ten microl sperm samples were examined with CASA technique, 250 microl stained in the presence of 10 microg/ml Rh123 and PI, and mitochondrial function analyzed by flow cytometry.

RESULTSSignificant differences were found between the experimental groups (final concentration 0.06 microg/L, 0.09 microg/L and 0.27 microg/L) and the control group in viability, straight line velocity, curvilinear velocity, average path velocity, progressive motility of human sperm and the number of spermatozoa with normal mitochondrial function (P < 0.01) except the final concentration 0.03 microg/L group (P > 0.05). Motility of human sperm lowered with the increase of rhTNF-alpha concentration and incubation time, and r values were 0.675, 0.691, 0.762, 0.693, 0.724 and 0.571, 0.594, 0.752, 0.791, 0.816, respectively (P < 0.01). The number of spermatozoa with normal mitochondrial function decreased with the increased rhTNF-alpha concentration and incubation time, and r values were 0.615, 0.643, 0.752, 0.691, 0.754 and 0.532, 0.567, 0.782, 0.692, 0.854, respectively (P < 0.01).

CONCLUSIONrhTNF-alpha can reduce human sperm motility function in vitro, possibly by interfering with human sperm mitochondrial function.

Adult ; Dose-Response Relationship, Drug ; Humans ; Male ; Mitochondria ; drug effects ; physiology ; Recombinant Proteins ; pharmacology ; Sperm Motility ; drug effects ; Tumor Necrosis Factor-alpha ; pharmacology

Objective:To investigate the effect of rare ginsenosides (RGS) on reproductive injury induced by cyclophosphamide (CP) in female rats.Methods:Twenty-four female rats were divided into four groups [normal control (NC), RGS, CP, and CP+RGS group] with 6 rats in each group. CP group (the model group) and CP+RGS group (the treatment group) were intraperitoneally injected with CP 30 mg/kg for 5 days for modeling, and CP+RGS group was given RGS intragastric intervention. General growth status of rats in each group was observed, the organ index was calculated, and the pathological changes of ovary, uterus, liver and kidney were observed by hematoxylin-eosin staining. Serum levels of estradiol, follicle stimulating hormone (FSH), luteinizing hormone (LH), pro-inflammatory factors interleukin (IL) 6, IL-1β, tumor necrosis factor-α were detected. The urine samples were collected after RGS treatment for metabonomics analysis. Metabolomic profiling based on ultra performance liquid chromatography (UPLC) coupled with mass spectrometry (MS) was used to analyze and determine the urine metabolites of rats in each group.Results:Compared with NC group, the ovary index of CP group [(0.054±0.015) %] was significantly decreased ( P<0.05), the uterus index [(0.293±0.036) %] and estradiol level [(62.9±6.4) pmol/L] were significantly decreased (all P<0.01), serum levels of FSH, LH, IL-6 and IL-1β [(20.4±1.0) U/L, (29.0±3.0) U/L, (185.4±28.6) ng/L, (72.9±2.0) ng/L, respectively] were significantly increased (all P<0.01). Compared with CP group, the ovary index in CP+RGS group [(0.075±0.010) %] was significantly increased ( P<0.05), serum estradiol level [(122.1±16.2) pmol/L] was significantly increased ( P<0.01), serum FSH, IL-1β and IL-6 levels [(16.7±1.0) U/L, (111.8±17.4) ng/L, (60.1±2.2) ng/L, respectively] were significantly decreased (all P<0.01). Metabonomics analysis results showed that, a total of 352 metabolites were detected in urine, of which 12 were found to be potential markers associated with reproductive injury according to the screening standard. After treatment with RGS, differential metabolites were improved in the direction of NC group. Pathway enrichment suggests that the therapeutic effect of RGS was related to multiple metabolic pathways, including purine metabolism and taurine and hypotaurine metabolism. Conclusion:RGS might reduce inflammation and thus ameliorate the damage caused by CP to the reproductive system of female rats by affecting purine metabolism and other pathways.