Objective To investigate the effects of esculin and digitalisglycosides eye drops combined with 532 laser on the treatment of diabetic retinopathy.Methods 65 cases(130 eyes)with diabetic retinopathy in our hospital from November 2012 to November 2015 were selected and divided into observation group(33 cases)and control group(32 cases).The control group was treated with 532 laser treatment,and the observation group was combined with esculin and digitalisglycosides eye drops.Two groups of treatment were one months.Therapeutic effect of two groups were compared,improvement of visual acuity,retinal hemorrhage,exudation,edema absorption time,before and after macular retinal thickness changes.Results After treatment,the total efficiency of the observation group(96.97％)was higher than the control group(75.00％),the difference was statistically significant(P<0.05),the visual acuity of the observation group was significantly higher than that of the control group,and decreased significantly less than the control group,the difference was statistically significant(P<0.05),the retinal hemorrhage,exudation and edema were observed in the observation group faster than the control group,the difference was statistically significant(P<0.05),the macular retinal thickness decreased in the two groups after treatment,the difference was statistically significant(P<0.05),the retinal thickness of the macular area was lower in the observation group than in the control group,the difference was statistically significant(P<0.05).Conclusion Esculin and digitalisglycosides eye drops combined with 532 laser in the treatment of diabetic retinopathy has significant effect,and can reduce the macular retinal thickness,which has important clinical significance.

With the transformation of marketization of economy and legislation of society in recent 20 years in China,the traditional mechanism in which medical therapy is decided by patients' relatives and doctors has not been able to offer sufficient protection for the legal rights of patients.So the rights of informed consent and decision making have been transferred passively to the patients' side.But in the field of assisted reproduction technique(ART),informed consent has to face some specific and unique problems.Firstly,being fully informed is sharply against the privacy of his(her) spouse as reproduction involved both the couple and their families.Secondly,it is a question that whether a wife can make her freewill choice when she accepts an ART program,because of the pressures coming from her family,the society,and her low social status well.And thirdly,The ART has the characteristics of both clinical therapy and scientific research as it is imperfect and its protocols still need to be improved.Ethicists should focus on thiese challenges.In the aspect of ethics,the regulations established by government and occupational association are the safeguard of the commonweal of society and general rights and benefits,and the ethics review and informed consent offered the final protection for the patients.

Objective To investigate the long-term safety and efficacy of the domestic biodegradable drug-eluting cor?onary stents (BuMA) in treating coronary heart disease (CHD). Methods Patients (n=440) who received BUMA stents were designated as observation group while patients (n=460) received Resolutestents were designated as control group. The base?line clinical characteristics, extend of pathological change shown by Coronary Arteriography (CAG),the procedure of percu?taneous coronary intervention were similar between these two groups. The patients were followed up for a mean of 24 ± 4 months;the primary endpoint was the occurrence of definite or probable stent thrombosis;secondary endpoint was major ad?verse cardiac events (MACEs) including complex end such as recurrent angina, acute non-fatal myocardial infarction, death, target vessel revascularization (TVR); other endpoints include all- cause mortality, cardiac death, myocardial infarc?tion (MI), target lesion revascularization (TLR), non-target vessel revascularization and stroke. Some patients were lost dur?ing follow up, which include 5 in observation group and 26 in control group. Results Patients in observation groups were in?serted with 615 stents while patients in control group were implanted with 614 stents, both groups with average of implanting 1.41 stent/case. There are no statistical significance differences in the primary endpoint [1.4%(6/435) vs 1.8%(8/434), χ2=0.087], secondary endpoint [12.3%(54/435) vs 10.8%(47/434),χ2=0.524] and other endpoints between the two groups. Con?clusion These data suggest that domestic biodegradable drug-eluting coronary stents (BuMA) are with good long-term safe?ty and efficacy.

Objective To investigate the application of venereal disease research laboratory(VDRL) test and toluidine red unheated serum test(TRUST) in syphilis laboratory diagnosis.Methods Serum,plasma and cerebrospinal fluid (CSF) in syphilis patients and healthy controls were measured by VDRL and TRUST.Results The VDRL detection results in serum and CSF sepcimens were generally higher than the TRUST detection results by 1-2 titers,while in plasma specimen,the VDRL detection results were generally lower than the TRUST detection results by 1-2 titers than TRUST when using plasma specimen.In addition,the VDRL detection in normal control serum and plasma specimens all appeared different degrees of false positive,but in the detection of normal control CSF,the results of TRUST and VDRL were consistent.Conclusion TRUST is more suitable for serum and plasma specimens,and CSF is suitable for the CSF specimen,but not suitable for serum and plasma detection.

Objective To investigate the influence of total progressively motile sperm count(TPMSC) after treatment on clinical outcomes of intrauterine insemination(IUI) with the husband′s sperm in ovulation-promoting cycles.Methods The clinical data in 4179 cases undergoing IUI with the husband′s sperm in ovulation-promoting cycles were retrospectively analyzed.The correlation between clinical pregnancy rate and TPMSC was analyzed.Results Among all the clinical data,TPMSC was to 100×106 in occasional live sperm.TPMSC<0.15×106 was in 15 cases,1 case had pregnancy (live sperm was occasionally seen on IUI day after sperm processing).Ten cases of TPMSC >60×106 had no pregnancy.A total of 4 154 cases of TPMSC (0.15-60.00)×106 were analyzed.The female age,duration of infertility,number of follicles and endometrial thickness(EDM) had no statistical differences among various groups.The clinical pregnancy rate was 13.5%(576/4 154),the group with the highest clinical pregnancy rate was (5.00-<10.00)×106.But there was no statistically significant difference in clinical pregnancy rate among groups(P=0.133).Conclusion Performing IUI in PMSC (0.15-60.00)×106 after processing can get preferable pregnancy rates.

BACKGROUND: Key point for subculture of human embryonic stem cells (hESCs) is to inhibit spontaneous differentiation and make sure totipotency of cells. Although mouse embryonic fibroblasts (MEF) or human foreskin fibroblasts (HFF) used as the feeder layer can maintain undifferentiated state of embryonic stem cells, cell clone is still imperfect and parallel arranged. OBJECTIVE: To establish mixed feeder layer of mouse embryonic fibroblasts plus human foreskin fibroblasts and to observe the hESCs growth.DESIGN: Multi-sample observation and comparison.SETTING: Medical Center of Reproduction, the Affiliated Hospital of Hainan Medical College. MATERIALS: This study was performed at the Medical Center of Reproduction, the Affiliated Hospital of Hainan Medical College from April 2006 to July 2007. Foreskin was derived from the children who underwent circumcision and came from Urinary Surgery, the Affiliated Hospital of Haihan Medical College. The children's family members provided the informed consent, and the experiment received confirmed consent from the local ethic committee. hESCs line HN-1 was separated from human blastula. Eleven 12.5-14.5-day-old fetal mice of clean grade were selected in this study. The experimental animals were disposed according to ethical criteria. METHODS: Heads, four extremities, and viscera were removed from fetal mice under anesthesia. Subsequently, cell suspension was prepared using routine trypsinase digestion and inoculated. When cells were cultured in confluent monolayer, some primary cells were frozen, processed with mitocin-C for 2.0-3.0 hours, and inoculated based on the density of 1×108 L-1 in gelatin-coated dish, I.e., MEF feeder layer. The HFF separation and culture and the preparation of HFF feeder layer were the same as above-mentioned processing. In addition, the two fibroblasts were respectively counted and mixed together according to the ratios of 1∶0, 3∶1, 1∶1, 1∶3, and 0:1. And then, the mixture was inoculated based on the density of 1×108 L-1 in gelatin-coated dish, I.e., mixed feeder layer. The growth of subcultured hESCs in vitro was observed in three different feeder layers, and hESCs in the mixed feeder layer underwent alkaline phosphatase test, OCT-4 expression immunohistochemical measurement, and OCT-4 and telomerase mRNA expression RT-PCR detection. Finally, differentiation in vitro of hESCs was observed after removing the feeder layer.MAIN OUTCOME MEASURES: ① Growth of hESCs in the three different feeder layers; ② Growth of hESCs in the mixed feeder layer based on different mixed ratios; ③ undifferentiated state of hESCs in the mixed feeder layer; ④ differentiation in vitro.RESULTS: ① hESCs clone in the MEF and HFF feeder layers was thin and flat and imperfect, but hESCs clone in the mixed feeder layer was perfect and thick and solid. Apparently, the clone form in the mixed feeder layer was superior to MEF and HFF feeder layers. ② When MEF and HFF was mixed together according to the ratio of 1∶1, hESCs grew in apparent accumulation; clone border was clear; eminentia was apparent and perfect. However, there were no changes according to the ratio of 1∶3. The ratio of 1∶1 was superior to the ratios of 1∶0, 3∶1, and 0∶1. ③ Alkaline phosphatase staining and OCT-4 antigen expression were strongly positive. Specific straps of OCT-4 and telomerase mRNA expression were observed at 200-300 bp and 300-400 bp, respectively. ④ Embryoid bodies were formed. hESCs could differentiate into multi-morphological cells after attachment.CONCLUSION: ① The mixed feeder layer may well support in vitro subculture of hESCs to acquire excellent clone form compared to MEF or HFF feeder layer. ② The mixture of MEF and HFF has excellent effect according to the ratio of 1∶1.

Objective To develop a PCR-mverse dot blot hybridization(RDB)assay to rapidly detect pathogenic mycoplasmas in genitourinary tract.Methods Universal primers were designed and applied to amplify the 16S rRNA gone of ureaplasma parvum(Up),ureaplasma urealyticum(Uu),Mycoplasma genitalium(Mg),Mycoplasma hominis(Mh)by using nestcd PCR.Specific nucleotide probes of Up,Uu,Mg and Mh Were constructed and immobilized onto nylon membranes.PCR products were denatured and hybridized、with specific oligonucleofide probes on nylon membrane.The sensitivity and specificity of the PCR-RDB assay were evaluated based.on the hybddizafion results.Also,PCR-RDB Was utilized to detect pathogenic mycoplasmas from 60 clinical samples.Results The four probes selectively hybridized with the PCR product of corresponding mycoplasmas,and no cross hybridization was observed.The detection limit of PCR-RDB Was one colony forming unit(CFU)of mycoplasma.Out of the 60 clinical samples、19were positive for mycoplasm,Mixed infections were found in three samples,including two coinfected with Up and Uu and one with Uu and Mg.Conclusion PCR-RDB is a rapid,specific and sensitive approach to the identification of pathogenic mycoplasmas in urogenital tract.

Objective To assess the vitro susceptibility to 6 antimicrobial agents and genotypes of clinical isolates of Chlamydia trachomatis (Ct) from Guangzhou region. Methods Ct was isolated from clinical specimens by using McCoy cell culture and subjected to propagation. The minimum inhibitory concentration (MIC) and minimal bactericidal concentration (MBC) of 6 antimicrobial agents (clarithromycin, roxithromycin, azithromycin, doxycycline, tetracycline, ofloxacin) against Ct isolates were determined in McCoy cell culture. Nested PCR was performed to amplify the outer membrane protein 1 (omp1) VS1-2 gene followed by sequencing. Results Seventy-six Ct strains were isolated from 346 urogenital specimens, and 40 strains met the require ments for susceptibility testing after serial propagation. The MIC50/MIC90 of clarithromycin, azithromycin, roxi thromycin, doxycycline, tetracycline and ofloxacin were as follows: 0.008/0.032, 0.080/0.160, 0.125/0.500, 0.032/0.064, 0.250/0.500 and 0.500/1.000 mg/L. Seven genotypes were observed. The most prevalent geno types in decreasing order were E (14, 35%), J (10, 25%)and F (6, 15%). The MIC50 was consistent for azithromycin among the 7 genotypes, but varied by 1 - 4 folds for doxycycline, ofloxacin and roxithromycin. Conclusions Clarithromycin, doxycycline and azithromycin exhibit an excellent activity against Ct, and the activity of azithromycin is consistent among the 7 genotypes of Ct.

Objective To develop a nested PCR for the detection of early syphilis and genotyping of Treponema pallidum (TP), and to investigate the distribution of genotypes of TP in Guangzhou. Methods Specimens were consecutively collected from genital ulcers of patients with suspected chancre during 2002-2004, and were detected by dark-field microscopy and nested PCR. The acidic repeat protein (arp) gene and the T. pallidum repeat (tpr) gene family were amplified with the positive specimens above. The number of repeats presented in the arp gene and the restriction fragment length polymorphism by Mse I in the tpr gene were analyzed by electrophoresis. The strains were genotyped according to Pillay's criteria. Results Out of 62 patients with suspected chancre, 33 cases (53.2%) were positive by dark-field microscopy and 54 cases (87.1%) by nested PCR. Of 47 TP-positive specimens genotyped by arp gene, 36 (76.6%) were type 14, while of 49 cases genotyped by tpr gene 39 (79.6%) were type d. By combining genotypes of arp and tpr genes, 7 genotypes were found, including 14d (31, 66.0%), 13d (5, 10.6%), 14b (4, 8.5%), 12b (3, 6.4%), 12d (2, 4.3%), 15d(l, 2.2%) and 14i (1, 2.2%). Conclusions Nested PCR shows a high sensitivity in early detection of TP. Genotype 14d seems the predominant type of TP in Guangzhou.

AIM: To investigate the effect of antisense RNA on osteopontin (OPN) expression in renal tubular epithelial cells. METHODS: Cell clone expressing stably OPN antisense RNA was formed by transfering retroviral vector expressing OPN antisense RNA into renal tubular epithelial cells, NRK52E cells, using liposome, with cell clones transfected by empty vector and vector expressing OPN sense RNA as controls. Ribonuclease protection assay(RPA), Western Blot, ELISA and assay of OPN activity were performed to detect expression of OPN mRNA and protein in above clones cultured with or without epidermal growth factor(EGF). RESULTS: The antisense RNA was only expressed by antisense clone. Antisense clone, sense clone and empty clone all expressed OPN mRNA. EGF enhanced expression of OPN mRNA, but not OPN antisense RNA or OPN sense RNA in above clones. OPN protein was not expressed in antisense clone cultured with or without EGF and empty clone cultured without EGF, but was expressed in sense clone cultured with or without EGF and empty clone cultured with EGF. CONCLUSION: Antisense RNA can inhibit OPN protein expression by means of preventing OPN mRNA translation, but not inhibit OPN mRNA transcription in renal tubular epithelial cells.