HPV16 E5 peptide vaccine in treatment of cervical cancer in vitro and in vivo / 华中科技大学学报（医学）（英德文版）

Human papillomavirus (HPV)-induced cervical cancer is the second most common cancer among women worldwide. Despite the encouraging development of the preventive vaccine for HPV, a vaccine for both prevention and therapy or pre-cancerous lesions remains in high priority. Thus far, most of the HPV therapeutic vaccines are focused on HPV E6 and E7 oncogene. However these vaccines could not completely eradicate the lesions. Recently, HPV E5, which is considered as an oncogene, is getting more and more attention. In this study, we predicted the epitopes of HPV16 E5 by bioinformatics as candidate peptide, then, evaluated the efficacy and chose an effective one to do the further test. To evaluate the effect of vaccine, rTC-1 (TC-1 cells infected by rAAV-HPV16E5) served as cell tumor model and rTC-1 loading mice as an ectopic tumor model. We prepared vaccine by muscle injection. The vaccine effects were determined by evaluating the function of tumor-specific T cells by cell proliferation assay and ELISPOT, calculating the tumor volume in mice and estimating the survival time of mice. Our in vitro and in vivo studies revealed that injection of E5 peptide+CpG resulted in strong cell-mediated immunity (CMI) and protected mice from tumor growth, meanwhile, prolonged the survival time after tumor cell loading. This study provides new insights into HPV16 E5 as a possible target on the therapeutic strategies about cervical cancer.

HPV16 E5 peptide vaccine in treatment of cervical cancer in vitro and in vivo / 华中科技大学学报（医学）（英德文版）

Human papillomavirus (HPV)-induced cervical cancer is the second most common cancer among women worldwide. Despite the encouraging development of the preventive vaccine for HPV, a vaccine for both prevention and therapy or pre-cancerous lesions remains in high priority. Thus far, most of the HPV therapeutic vaccines are focused on HPV E6 and E7 oncogene. However these vaccines could not completely eradicate the lesions. Recently, HPV E5, which is considered as an oncogene, is getting more and more attention. In this study, we predicted the epitopes of HPV16 E5 by bioinformatics as candidate peptide, then, evaluated the efficacy and chose an effective one to do the further test. To evaluate the effect of vaccine, rTC-1 (TC-1 cells infected by rAAV-HPV16E5) served as cell tumor model and rTC-1 loading mice as an ectopic tumor model. We prepared vaccine by muscle injection. The vaccine effects were determined by evaluating the function of tumor-specific T cells by cell proliferation assay and ELISPOT, calculating the tumor volume in mice and estimating the survival time of mice. Our in vitro and in vivo studies revealed that injection of E5 peptide+CpG resulted in strong cell-mediated immunity (CMI) and protected mice from tumor growth, meanwhile, prolonged the survival time after tumor cell loading. This study provides new insights into HPV16 E5 as a possible target on the therapeutic strategies about cervical cancer.

Nuclear status of day 2 preembryos affects embryo quality and implantation potential / 中华男科学杂志

<p><b>OBJECTIVE</b>To assess the effects of the nuclear status of day 2 preembryos on day 3 embryo quality and implantation potential and to weigh its clinical value in the human in-vitro fertilization-embryo transfer (IVF-ET) program.</p><p><b>METHODS</b>Embryos obtained from 409 fresh conventional IVF-ET/ICSI cycles from July to October 2006 were assessed retrospectively. Day 2 preembryos were classified according to the number of nuclei in each blastomere in 3 groups: grade A with only mononucleated blastomeres, grade B with one or more blastomeres containing no visible nucleus, and grade C with one or more multinucleated blastomeres. Comparisons were made of the rates of arrested embryos and excellent embryos on day 3 as well as of the pregnancy outcome and implantation potential of those in whom cohorts of similar nuclear scoring embryos were transferred.</p><p><b>RESULTS</b>There were fewer arrested embryos and more excellent embryos on day 3 in grade A than in grade B and C (P < 0.01), and so were there in grade B than in grade C (P < 0.01). Among the 234 cycles in which all the transfer embryos were derived from a similar day 2 nuclear scoring, 51 cycles originated from grade A embryos (group A) and 183 from grade B (group B), with similar clinical pregnancy rates between the two groups, while the implantation rate was higher in group A than in B (P < 0.05).</p><p><b>CONCLUSION</b>Day 2 nuclear scoring can be used to predict the devel- opment and implantation potential of embryos. A combined evaluation of day 2 nuclear scoring and day 3 embryo morphology helps identify the most viable embryos and reduce the number of embryos for transfer.</p>

Cryopreservation by cryoloop damages human immature oocytes / 中华男科学杂志

<p><b>OBJECTIVE</b>To examine the influence of cryoloop on the spindle and chromosome configurations of human oocytes cryopreserved in the germinal vesicle (GV) and metaphase II (M II) stages, as well as on the survival rate and potential for in vitro maturation (IVM).</p><p><b>METHODS</b>GV oocytes were randomly assigned into a control group (matured in vitro into the M II stage), a GV cryopreserved group (cryopreserved in the GV stage and then matured in vitro), and an M II cryopreserved group (matured in vitro and cryopreserved in the M II stage). After cryopreservation and IVM, immunostaining of the tubulin and chromatin was performed followed by visualization using laser scanning confocal microscopy (LSCM).</p><p><b>RESULTS</b>A significantly higher survival rate was observed in the GV cryopreserved group than in the M II , but the maturation rate showed no significant difference between the GV cryopreserved group and the control (P > 0.05). Compared with the control group, there was a statistically significant decrease in the rates of normal meiotic spindles and chromosomes in the GV cryopreserved group (P < 0.05). A significantly lower rate of normal spindles was noted in the M II cryopreserved group than in the control, but no statistical difference was shown in the rate of normal meiotic chromosomes between the two groups (P > 0.05).</p><p><b>CONCLUSION</b>Cryopreservation by cryoloop has a damaging effect on the spindle and chromosome of human oocytes in the GV and M II stages.</p>

Expression of cyclooxygenase-2 in the testes and epididymides of adult male rats / 中华男科学杂志

<p><b>OBJECTIVE</b>To investigate the expression of cyclooxygenase-2 (cox-2) in the testes and epididymides of adult male rats and its significance.</p><p><b>METHODS</b>Immunohistochemical staining was used to detect the expression and localization of cox-2 in the testicle and epididymal tissues of 40 adult male SD rats.</p><p><b>RESULTS</b>Strong cox-2 immunoreactivity was detected in the epididymides and testes of the rats. In the caput epididymides, cox-2 expressed mainly in the epithelial nuclei and partly in the cytoplasm. Cox-2 was also found positive in the testis nuclei and cytoplasm.</p><p><b>CONCLUSION</b>Immunohistochemical staining is a fairly sensitive method for detecting cox-2 expression in the testes and epididymides of adult male rats.</p>

Expression of p57 and homeobox A10 during decidualization of endometrial stromal cell in vitro / 生理学报

In order to elucidate the function of homeobox A10 gene (HOXA10) and p57 during decidualization our present study was designed to observe the change of HOXA10 and p57 expression and subcellular localization of HOXA10 in the process of endometrial stromal cell (ESC) differentiation in vitro. Decidualization was induced by 0.5 mmol/L 8-Bromo-cAMP (8-Br-cAMP) together with 1x10(-6) mol/L medroxyprogesterone acetate (MPA). Expression of p57 and HOXA10 was detected by RT-PCR and Western blot after 1-day, 2-day, and 4-day treatment (D1, D2, D4). ESCs cultured in 2%FBS for 1 and 4 d were used as control (C1, C4). The location of HOXA10 was detected by indirect immunofluorescence and HOXA10-GFP transfection. The results are as follows: (1) The expression of HOXA10 decreased progressively during the course of decidualization, and showed significant difference compared to control group C4 after 2-day treatment (D2). (2) On the contrary, the expression of p57 increased progressively and also showed significant difference compared to the control group C4 after 2-day treatment (D2). (3) There was no significant change of HOXA10 and p57 expression after culturing ESCs in 2%FBS for 4 d (C1, C4). (4) HOXA10 located in the nucleus throughout the course. Cytoplasm and nucleus shuttle was not detected in the experiment. Our results suggest that the down-regulation of HOXA10 may contribute to the increase of p57 and that the up-regulation of p57 likely plays an important role in ESC differentiation. Progesterone receptor (PR) pathway may participate in promoting ESCs to exit cell cycle and enter differentiation.

Establishment of two human embryonic stem cell lines from cleavage arrested embryos / 中华妇产科杂志

Objective To determine whether cleavage developmentally retarded embryos have not cleaved during a 24 hour period could develop into blastocysts and produce hESC cell lines.Methods A total of 120 such embryos were cultured to blastocyst stage by sequential culture.Blastocysts formation rate and quality of blastocyst were detected under microscope.The relation between blastocyst formation rate and blastomere number,the fragment of blastomere and blastomere symmetry were analyzed by stepwise Logistical regression analysis.Inner cell masses(ICMs)were isolated by immunosurgery.Colonies derived from the ICMs were passed every 4-7 days and the derivatives were passaged and identified.Results A total of 22 blastocysts were obtained from 120 embryos.The blastulation rate was 18.7%.Early blatocyst, blastocyst,full blastocyst,expanded blastocyst,hatching blastoeyst and hatched blastocyst accounted for 5.9%,23.5%,35.3%,23.5%,5.9%,and 5.9% respectively.The grade of ICM and trophoblast was mostly scored C or B.Blastocyst formation rate was related to cell number and blastomere symmetry but not fragment.Immunosurgery resulted in the formation of 7 ICMs and 3 primary colonies,which produced 2 cell lines.The cell lines satisfied the criteria that characterize pluripotent hESC cells.Undifferentiated cells were positive for AKP,SSEA-4,TRA-1-60,and TRA-1-81.It could continue to proliferate in vitro and form embryoid bodies when cultured in suspension.It had capability to form teratoma in SCID mice.Both cell lines had normal karyotypes after 45 and 34 passages respectively.Conclusions Our results suggest that a subset of developmentally retarded embryos can form blastocysts and give rise to hESC cell lines.