[Pilot Study on Start-up and Stable Operation at Low Temperature Based on Denitrifying Phosphorus Removal].

In order to understand the denitrifying and phosphorus removal characteristics of denitrifying phosphate-accumulating organisms (DPAOs), a pilot scale (20 m3) denitrifying and phosphorus removal experiment was carried out using a modified University of Cape Town (UCT) process at low temperatures of (6-16)℃. The test results show that at such temperatures, the hydraulic retention time (HRT) is 20 h and the solids retention time (SRT) is 35 days, and the modified UCT process can start up successfully and run steadily. When running steadily, the system can maintain nitrogen and phosphorus removal rates of 60%±5% and 80%±5%, respectively. The effluent concentrations of chemical oxygen demand (COD), ammonium nitrogen (NH4+-N), total nitrogen (TN), and total phosphorus (TP) were 20, 5, 11, and 0.5 mg·L-1, respectively, which meet the first A emission standard of "Pollutant Discharge Standard for Urban Sewage Treatment Plant" (GB 18918-2002). In order to further investigate the characteristics of nitrogen and phosphorus removal in the system, the reflux ratio from the aerobic tank to the anoxic tank was increased to 150%. After the system was stabilized, it obtained higher nitrogen and phosphorus removal rates of 80%±10% and 90%±5%, respectively. Among them, denitrifying phosphorus removal in the anoxic tank accounted for 80%±4% of the total biological phosphorus removal. The average effluent concentrations of COD, NH4+-N, TN, and TP were 19.55, 0.1, 7.8, and 0.15 mg·L-1, respectively, which meet the Beijing Standard A discharge standard.

Expression and regulation of stanniocalcin 1 and 2 in rat uterus during embryo implantation and decidualization.

Stanniocalcin-1 (STC-1) is a recently discovered polypeptide hormone, while stanniocalcin-2 (STC-2) is a subsequently identified homologue of stanniocalcin-1. Although previous studies have shown that both STC-1 and -2 are involved in various physiological processes, such as ion transport, reproduction and development, their expression in the uterus and roles in implantation and early pregnancy are unclear. Here we have investigated the expression and regulation of both STC-1 and STC-2 in rat uterus during early pregnancy under various physiological conditions. We show that only basal levels of STC-1 and STC-2 mRNA were detected in the uterus from day one (D1) to day five (D5) of pregnancy. STC-2 immunostaining was gradually increased in the glandular epithelium from day two (D2), with a peak occurring on D5. High levels of both STC-1 and STC-2 mRNA were observed in the stoma cells at the implantation site on day six (D6) of pregnancy, whereas their immunostaining signals were also significant in the luminal epithelium. Basal levels of both STC-1 and STC-2 mRNA and STC-1 immunostaining were detected in the uterus with delayed implantation. After the delayed implantation was terminated by estrogen treatment, both STC-1 and STC-2 mRNA signals were significantly induced in the stroma underlying the luminal epithelium at the implantation site, and STC-2 immunostaining was also observed in the luminal epithelium surrounding the implanting blastocyst. Embryo transfer experiments further confirmed that STC-1 and STC-2 expression at the implantation sites was induced by the implanting blastocyst. Both STC-1 mRNA and immunostaining were seen in the decidualized cells from day seven (D7) to day nine (D9) of pregnancy. STC-2 mRNA was also found in the whole decidua from D7 to D9 of pregnancy; STC-2 protein, however, was strictly localized to the primary deciduas on D7 and D8, with a weak expression in the whole deciduas on D9. Consistent with the normal pregnancy process, strong STC-1 and STC-2 mRNA signals were detected in the decidualized cells under artificial decidualization, whereas only basal levels of STC-1 mRNA and immunostaining were observed in the control horn. These data suggest, for the first time, that STC-1 together with STC-2 may play important roles in the processes of implantation and decidualization in the rat.

An oligonucleotide microarray study on gene expression profile in mouse testis of experimental cryptorchidism.

To investigate the germ cell apoptosis under body temperature in testis, we analyzed the gene expression patterns on day 1, day 4, day 7, day 14, day 28 and normal control adult mouse testis after experimental cryptorchidism (EC) using Affymetrix MOE430A microarray. Our data showed that EC led to the oxidative stress and gene expression fluctuation in the first 28 days, both of which were highly coincident in timing. Cryptorchid testis showed more effective antioxidative capability in the first 4 days, and suddenly lowered the capability from day 5 on, then gradually restored the antioxidation from day 10 to day 14, and turned to worse on day 28 again. The extensive high gene expression on day 4 after EC and the up-rising of oxidative stress level on day 5 and the abrupt down-regulation of the gene on day 7 were closely related. From the chip data, we have found that the high level of reactive oxidative species (ROS) was not only related to the dysfunction or abnormality of the direct origin of ROS generation, but also related to the abnormality of the more upstream physiological events in energy metabolism, lipid metabolism. The selective regulation of metabolic substrate transporter in different cell population implied the existence of various regulation of the selective signal pathways among different cell populations by EC.

Cloning and characterization of a novel spermiogenesis-related gene, T6441, in rat testis.

We report in the present study the cloning and characterization of a novel gene, named T6441, initially derived by the suppressive subtracted hybridization (SSH) cDNA library. The full-length T6441cDNA was 664 bp long, containing a complete open-reading frame for a protein of 149 amino acids (aa). The protein bears no homology to any reported genes. It is predicted that the molecular mass was about 16.7 kDa. Northern blot analysis showed that the T6441 gene had about 4 transcripts in adult rat testis and was temporally regulated in a stage-dependent manner in the testis. In situ hybridization showed that T6441 mRNA was specifically localized in spermatids, and its expression level varied in the cells at different stages of the testicular development, with the highest level at steps 7-14. RT-PCR results showed that the T6441 mRNA was transcribed in most of the tested tissues with its strongest signal in the testis. Recombinant T6441 protein was prepared, purified, and was used to raise rabbit. Western blot analysis using the antiserum revealed four possible testicular specific proteins with their molecular weights being about 22, 25, 50 and 55 kDa respectively. The T6441 protein was expressed mainly in the cytoplasm of spermatids with the maximal levels at steps 12-19. At step 19 spermatid, the T6441 was mainly localized in the residual bodies. The cytoplasm localization of T6441 protein was supported by transient over expression of GFP-fusion protein in Hela cells. Interestingly, the expression of T6441 caused death of transfected cells within 48 h. Our preliminary experimental results suggest that the T6441 gene may play a role in cytoplasm movement and removal during spermiogenesis.

IFN-gamma and TNF-alpha inhibit expression of TGF-beta-1, its receptors TBETAR-I and TBETAR-II in the corpus luteum of PMSG/hCG treated rhesus monkey.

The corpus luteum (CL) is a transient endocrine organ that secretes progesterone to support early pregnancy. Using in situ hybridization, immunohistochemistry and computer imaging analysis, we have investigated the expression of transforming growth factor beta 1(TGF-beta 1) , its receptors, type I (TbetaR-I) and type II (TbetaR-II) as well as steroidogenic acute regulatory protein (StAR) in the corpus luteum (CL) of the rhesus monkey at various stages of CL development. The CL was induced by injection of pregnant mare serum gonadotropin (PMSG)/human chorionic gonadotropin (hCG). The expression of TGF-beta 1, TbetaR-I and TbetaR-II as well as StAR was detected in the CL in a time-dependent manner, reaching the maximum levels on D10 (functional stage), and decreased on Day 18 (regression stage). Injection of interferon-gamma (IFN-gamma) or tumor necrosis factor-alpha (TNF-alpha) at the functional stage of CL development significantly decreased the expression of StAR, as well as TGF-beta 1, and its receptors TbetaR-I and TbetaR-II. Our results suggest that TGF-beta 1 and its receptors may play an important regulatory role in maintaining CL function, and that IFN-gamma or TNF-alpha is capable of inhibiting their expression in the CL.

Disrupted expression of intermediate filaments in the testis of rhesus monkey after experimental cryptorchidism.

Cytoskeletons in Sertoli cell play an important role in process of spermatogenesis. The expression and distribution of the intermediate filaments, vimentin, keratin and desmin, were studied in the Sertoli cells of the cryptorchid testis of rhesus monkey. Vimentin was localized in the perinuclear region of Sertoli cells of the normal testis. An intense increase in vimentin immunoreactivity was observed with appearance of disorganized staining in the Sertoli cells of the cryptorchid testes. Cytokeratin 18, a marker of immature Sertoli cells, re-expressed in the cells of the adult cryptorchid testes. Desmin was also observed in the Sertoli cells in addition to the peritubular myoid cells on 30 days after the cryptorchid operation. These data suggest that Sertoli cells in primate can be affected by the heat stress. The altered changes in intermediate filaments could be possible to induce the Sertoli cell functional changes that would partially contribute to the germ cell apoptosis leading to azoospermia or oligozoospermia.

VEGF, bFGF, and their receptors in the endometrium of rhesus monkey during menstrual cycle and early pregnancy.

A number of cytokines and growth factors are known to modulate proliferation and differentiation of human endometrium. In this study, the expression of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and VEGF receptors, fms-like tyrosine kinase (Flt1) and kinase insert domain-containing region (KDR), and bFGF receptor 1 (Flg) were examined in the endometrium of rhesus monkey on Day 5, 10, 16, 20, 25 of menstrual cycle and on Day 19 of early pregnancy. Western blot analysis showed the specificity of the anti-human antibodies with the monkey tissue. The expression of mRNA and protein of VEGF was correlated with that of its receptor KDR, which was detected in epithelial, vascular, and myometrial cells. The localization of bFGF and its receptor Flg was similar to that of VEGF, except that the Flg was absent in the endothelial cells. Strong expression of VEGF and bFGF in the glandular epithelial cells was observed in the proliferative phase, declined in the secretory phase during the cycle. Stronger staining of these factors was also observed in the decidual cells of the pregnant uterus, as compared with the stromal cells of cycling uterus. No expression of Flt1 was detected in the tissue examined in this study. These data suggest that VEGF, bFGF, and their receptors play important roles in epithelial and stromal development, angiogenesis, and blood vessel function in the endometrium during the menstrual cycle and early pregnancy of the rhesus monkey.

Regulatory effect of IFN-gamma on expression of TGF-beta 1, T beta R-II, and StAR in corpus luteum of pregnant rhesus monkey.

AIM: To examine expression of transforming growth factor-beta 1 (TGF-beta 1) and TGF beta receptor II (T beta R-II) and steroidogenic acute regulatory protein (StAR) in the corpus luteum (CL) of pregnant monkeys at various stages and to study possible effect of IFN-gamma on their production. METHODS: In situ hybridization and immunohistochemistry were applied to detect mRNA and protein. RESULTS: The expression of StAR, TGF-beta 1, and T beta R-II in the pregnant monkey CL was progressively decreased from d 15 to d 35 of gestation. IFN-gamma down-regulated the expression of TGF-beta 1, T beta R-II, as well as StAR. CONCLUSION: TGF-beta 1 may play an important role in the CL formation and functional maintaining; IFN-gamma down-regulates the expression of TGF-beta 1, T beta R-II, and StAR.