Objective:To investigate the relationship between genotype and phenotype in children with CRB1 mutated Leber congenital amaurosis (LCA) and early onset retinal dystrophy (EOSRD). Methods:A retrospective clinical study. From January 2013 to December 2019, 10 children with CRB1 mutated LCA/EOSRD were enrolled in the study. The patients were identified as CRB1 mutation by the second generation targeted capture sequencing, Sanger sequencing and the family segregation analysis. All children underwent electroretinogram (ERG) and fundus examination. At the same time, 6 cases were examined by optical coherence tomography (OCT); 1 case was examined by fluorescein fundus angiography (FFA), 7 cases were examined by wide-angle laser scanning ophthalmoscope (UWF SLO). Results:There were 6 cases of LCA and 4 cases of EOSRD in 10 patients with CRB1 gene mutations. The average age of first visit was 3.61 years old. The light and dark wave of ERG was flat in 6 cases, and decreased in 4 cases. A total of 19 pathogenic mutations were detected. There were 1 homozygous mutation and 9 compound heterozygous mutations. There were 4, 2 and 1 cases of "copper-coin" like, "salt and pepper" like and "osteocyte" like pigment changes in retina, 1 case of "crystalline pigment" change and 2 cases of macular pigment scar. In 7 cases of UWF SLO examination, different degrees of para-arteriolar pigment epithelium retention (PPRPE) were found in the middle and peripheral fundus. In 6 cases examined by OCT, the outer layer of retina atrophied and the band of ellipsoid disappeared. Symmetrical cystoid macular edema, splitting cystoid macular degeneration and adhesion of epi-macular membrane to optic disc and macular area were found in 1 case, respectively, the retinal structure was rough and thickened, and the fovea became thinner in 3 cases. In FFA examination, 1 case showed uveitis-like changes with late optic disc fluorescein staining, macular fluorescence accumulation, strong fluorescence diffusing along the blood vessels in each quadrant, peripheral PPRPE of "frost-branch" like strong fluorescence. Conclusion:The relationship between genotype and phenotype of CRB1 mutation is complex, and PPRPE is a common characteristic change.

Objective To investigate the common types of fetal cardiac malformations and complicated malformations,and to assess the value of classifying on these types.Methods 3201 pregnant women were undergone with fetal echocardiography (FECG),239 fetuses of them were diagnosed to be suffered with congenital heart disease(CHD),and 8 cases were one of twins with abnormal heart confirmed by FECG.All new-births were examined by echocardiography within half year after their births.Results 155 complex CHD in 239 fetuses were diagnosed by FECG,in them the common malformations were in turn 59 cases with diagnosed univentricular heart,29 cases with double outlet right ventricle,19 with atrio ventricular septum defect,12 with tetralogy of Fallot or quinalogy of Fallot,11 with persistent truncus arteriosus,6 with right ventricular dysplasia syndrome,6 cases transposition of the great vessels.100 cases were induced labor,41 of them were comfirmed by pathology.16 fetuses were born,123 cases were being pregnanted or un-followed up.Conclusions Complex and multi-malformation were common in fetal cardiocascular abnormalities.Diagnosing rate of fetal CHD(FCHD) in our enroll fetuses was 7.47 ％,rate of complex CHD vs CHD was 64.85 ％.According different types of FCHD,able to be operated or not after birth,surgery methods,as well as prognosis evaluations,all FCHD cases were classified into three subtypes:curable type,curable palliative type and untreatable type.This newly viewpoint will help pregnant women and their family to make reasonable selection.

Objective To improve detection rate and diagnostic accuracy of single ventricle(SV) in obstetric fetal echocardiography,to investigate the common types and complicated malformations of SV in the fetus,and to summarize the differential announcements in diagnosing fetal SV.Methods In 345 fetal hearts which were diagnosed as congenital heart disease by fetal echocardiography in our hospital,73 cases diagnosed as SV,including 3 cases appeared as ones of twins,were included in this study.Systemic scanning and multiple-views fetal echocardiography were used to examine these enrolled fetuses.Results In all 73 SV eases,3 cases were diagnosed as simple SV,the others were diagnosed as SV accompanied with other abnormalities,among them 44 cases accompanied with single atrium,18 cases with single atrium and persistent truncus arteriosus,2 cases with pulmonary atresia,20 cases with pulmonary artery stenosis,4 cases with partial atrioventricular septal defect,3 eases with aorta dysplasia or aortic valve dysplasia.SV types were classified as 24.7％ in type A,13.7％ in type B,46.6％ in type C and 15.0％ in type D respectively.68.2％ of the cases were diagnosed with aortic D-transposition,and 45.2 ％ with common inlet,42.5 ％ with single inlet and 12.3 ％ with double inlet respectively.42 SV cases were executed termination of pregnancy which 11 cases were confirmed by pathology and the other 31 cases were out of following-up.Conclusions Most cases of fetal SV were accompanied other abnormalities and simple SV was rare.Type C in which ventricular structure was combined with left and right ventricle was the most common type.To avoid the false diagnosis,much attention must be paid to distinguish big papillary muscle and abnormal muscle bundle from interventricular septum during ultrasonic examination.

Objective To explore the clinical effect of acupuncture on acute renal injury of sepsis.Methods From January 2015 to May 2017,adult patients with sepsis who received ≥ 7 days of treatment were collected from the Central Hospital of Zhuzhou.The participants were randomly divided into the control group (n =35) and the treatment group (n =37),and the control group was given routine treatment according to the sepsis guidelines.On the basis of control group,the treatment group was given acupuncture treatment,acupoints Shenshu and Sanyinjiao,Taixi,Zusanli.The clinical effects of the two groups of patients were compared.Results The levels of cystatin C,blood urea nitrogen and creatinine in the treatment group were significantly lower than those in the control group (P ＜ 0.05).There was no significant difference in the number and proportion of patients with blood purification between the two groups (P ＞ 0.05),while the frequency and time of blood purification treatment in the treatment group were significantly lower than those in the control group (P ＜ 0.05).The levels of blood interleukin-6 and tumor necrosis factor-alpha in the treatment group were significantly lower than those in the control group (P ＜ 0.05).From the fourth day of treatment,the Acute Physiology and Chronic Health Evaluation Ⅱ (APACHE Ⅱ) score in the treatment group was significantly lower than that in the control group (P ＜ 0.05).There was no significant difference in mortality between the two groups at 28 days (P ＞ 0.05).Conclusions Acupuncture of Shenshu,Sanyinjiao,Taixi,Zusanli can improve sepsis patients with acute kidney injury in renal function.

BACKGROUNDThough there have been various methods for harvesting and preserving descemet membrane (DM) and intact endothelium, there is no literature about the morphological evaluation of endothelium after graft preparation for descemet membrane endothelial keratoplasty (DMEK). The aim of this study was to establish and improve a simple method for preparing, preserving, and morphologically evaluating the donor graft for DMEK.

METHODSTo obtain a donor graft, an air bubble was formed by injecting a 29 G needle with 1 ml sterile air into a small edge created outside the Schwalbe line. Another needle was inserted into the bubble through the stroma to aspirate the air or replace half the air with organ culture medium. Trypan blue was used to mark the location for small incision to improve the success rate. Frozen sections were stained with hematoxylin and eosin (HE). Based on the air bubble, DM grafts were divided into four groups: group A (normal control), graft without any operative technique; group B, graft with zero-pressure air bubble; group C, graft with full-pressure air bubble; group D, graft with half-pressure air bubble. The four groups of grafts were preserved for 24 hours to observe the effect of bubbles on cells. The gross and ultrastructure morphologies were evaluated using alizarin red and scanning electron microscopy (SEM), respectively.

RESULTSDonor grafts were harvested via the air bubble technique, facilitated by prior trypan blue staining. HE-stained sections revealed a pure graft without stroma. There were no significant changes under light microscope. In group A, SEM revealed a confluent layer of polygonal endothelium with distributed microvilli exhibiting characteristics of interdigitating junctions. In group B, intercellular borders became thinner. In group C, interdigitations were almost flat and microvilli were observed less frequently. In group D, other than less microvilli, there were minimal changes.

CONCLUSIONSThe donor graft preparation method appears to be effective and convenient. Properly decreasing the air pressure could protect and preserve the endothelium.

Objectives To investigate the homology and drug resistance gene of Group B Streptococcus ( GBS) Resistance induced by Clindamycin and provide basic data for clinical prevention and treatment of GBS Resistance infection induced by Clindamycin .Methods 921 strains of GBS were isolated at Obstetrics&Gynecology Hospital of Fudan University from January , 2014 to December , 2015.VITEK2-compact automatic bacterial susceptibility instrument was used to test their sensitivity to 7 antibacterial drugs.63 positive strains were chosen through D-inhibition zone trial which were drug resistant to Erythromycin and susceptible or intermediary to Clindamycin .The strain′s sequence type was identified by the method of multilocus sequence typing ( MLST typing ) .The drug resistance genes mefA & ermB to Erythromycin were detected by using PCR method .The analysis was carried out to reveal the relevance to drug resistance , multilocus sequence typing and drug resistance gene .Results Among 921 strains of GBS , the drug resistance rate was respectively 53.4％ ( 492/921 strains ) to Erythromycin , 50.2％ ( 462/921 strains) to Clindamycin, 34.7％ ( 320/921 strains ) to Levofloxacin and 7.5％ ( 69/921 strains ) to Nitrofurantoin.The drug resistance rate of Levofloxacin for 63 GBS strains was 27.0％(17/63 strains) and no drug resistant strain was found to Penicillin , Vancomycin & Nitrofurantoin.12 different ST types were involved in total, including a new ST type:ST1072.The most common ones were ST12 (30.1％) (20/63 strains) &ST19 (25.4％) (16/63 strains).The drug resistance rate of Levofloxacin with ST 19 (75.0％) (12/16 strains) was much higher than that of other ST types .The relevance ratio of mefA and ermB among 63 GBS strains was respectively 27.0％(17/63 strains) and 41.3％(26/63 strains).Conclusions The genetic diversity existed in Group B Streptococcus resistance induced by Clindamycin detected in this study . There was significant difference on drug resistance and relevant drug resistant genes among different ST types.

Objective To investigate the efficacy and safety of early goal-directed sedation with dexme-detomidine in patients with septic shock.Methods This study is selected from August 2015 to December 2016 in our department 70 patients receiving treatment of septic shock,and through random sample table method divided into the control group and the study group two,there were 32 cases in control group,the control group selected ob-ject according to the conventional sedation treatment,namely the application of midazolam extraction treatment, the study group of 38 patients with septic shock in patients with application of dexmedetomidine sedation,analysis and application of different sedation method analysis of two groups of patients with septic shock. The efficacy and safety of observation and etc. Results Two groups of patients with different treatment methods,The onset time of the study group of patients with septic shock after sedation,h recovery time and time of mechanical ventilation was obviously better than the control group,the effect of difference(P < 0.05),has significant research value. Conclusions For patients with septic shock early goal orientation selection and application of sedation of dexme-detomidine can play a better effect,the more safe and effective,sedation score is high,it is worth of application in clinical treatment.

Objective This study intends to analyze the radioactivity level of slag and its building materials, estimate the effective dose of the residents caused by the comprehensive utilization of slag in building materials, and provide basis for rational utilization of slag. Methods Data of slag production and its comprehensive utilization in building materials, radioactive level investigation and detection of slag and building materials in different periods and regions were collected from 2005 to 2016. Using the room model of concrete structure and the model of calculating equilibrium equivalent radon concentration with the specific activity of ²²⁶Ra in indoor building materials, the external and internal radiation doses of residents caused by comprehensive utilization of slag for building materials were estimated respectively, and then the annual additional effective dose and collective dose of residents were estimated. Results The contents of ²²⁶Ra and ²³²Th in the slag were relatively high, while the contents of ⁴⁰K were relatively low. The radionuclide content in slag cement decreased significantly. The annual additional effective doses of slag used in concrete and cement mixture relative to ordinary cement were 0.40 mSv·a⁻¹ and 0.20 mSv·a⁻¹ respectively. The annual additional collective dose and 50-year collective dose caused by slag used in building materials were 3.87 × 10³～1.84 × 10⁴ man·Sv and 1.94 × 10⁵～9.20 × 10⁵ man·Sv respectively. Conclusion The comprehensive utilization of slag in building materials increases the additional effective dose of residents, so the use of some slag in building materials needs to be paid more attention to. In addition, the influence of comprehensive utilization of slag in building materials, such as the way, the amount of slag and the amount of building materials, on the annual additional effective dose of residents caused should also be paid attention to.

Objective:To understand the distribution and epidemiological characteristics of chronic lymphocytic leukemia (CLL) in Hunan Province.Methods:According to the audit methods and evaluation criteria specified by the National Cancer Registration Center, the registration data of CLL reported by 24 tumor registries was included. Through the research method of retrospective analysis, the selected registry data was calculated and analyzed according to the year, administrative division, urban and rural areas, gender and age.Results:A total of 104 newly diagnosed CLL patients were diagnosed in Hunan Province from 2014 to 2015, with an average annual morbidity of 0.39/100, 000. The morbidity in 2014 and 2015 was 0.39/100, 000 and 0.39/100, 000, respectively. The annual average morbidity in Zhuzhou was 0.8/100, 000, which was the highest among municipalities. The annual average morbidity in Kaifu District of Changsha was 1.65/100, 000, which was the highest among district-level administrative divisions. The morbidity of urban was higher than that of rural (Urban vs Rural, P=0.006). The male to female morbidity was 1.7∶1. The cases were mainly concentrated in the 61-70-year-old population, accounting for 33.65% of all cases (35/104). There were 64 patients died of CLL in Hunan Province from 2014 to 2015, and the average annual mortality was 0.24/100, 000. The mortality in 2014 and 2015 was 0.22/100, 000 and 0.26/100, 000, respectively. The average annual mortality in Hengyang was 0.53/100, 000, which was the highest among municipalities. The average annual mortality in Furong District of Changsha was 0.74/100, 000, which was the highest among district-level administrative divisions. The mortality of urban was higher than that of rural but with no significant difference ( P=0.006). The male to female mortality rate was 1.4∶1. The deaths were mainly concentrated in the 71-80-year-old population, accounting for 29.69% of all deaths (19/64). Conclusions:The morbidity of CLL in Hunan Province is much lower than that of European and American populations, and it mainly occurs in the elderly people. It is more common in men. The morbidity of urban is higher than that of rural and morbidity in Zhuzhou is the highest. The death of CLL patients was mainly in middle-aged and elderly population, with more males. The mortality of urban is slightly higher than that of rural and the mortality in Hengyang is the highest.

Objective: To establish a method for in vitro expansion of regulatory T cells (Tregs) for clinical study and immunotherapy. Methods: CD4⁺ T cells were isolated from BALB/c spleens by magnetic activated cell sorting (MACS), then cultured in 24-well plates coated with anti-CD3/CD28 microbeads in the presence of 100 U/ml interleukin-2 (IL-2) and 5 ng/ml recombinant human transforming growth factor-β (rhTGF-β). The purity of the expanded Tregs were tested by flow cytometry. In vitro inhibition of CD4⁺ CD25^-T cells response by expanded Tregs was measured by mixed lymphocyte reaction test. The number and the viability of the expanded Tregs were detemined by trypan blue staining. Results: Tregs were expanded up to 20 folds after 5 days in culture, and the activity was (93±4)%. The purity of expanded Tregs were (79.1±1.5)%, and maintained suppressive ability. Conclusions We can obtain large numbers of Tregs with highly purity and suppressive ability, thereby providing a solution to the availability of sufficient Tregs in clinical study and immunotherapy.