Background Retinal pigment epithelial (RPE) cell transplantation is a novel approach to the treatment of hereditary retinal diseases, however, human-derived RPE cell line occurs de-differentiation during in vitro cell culture.Studies showed that early abnormal activation of Akt/mammalian target of rapamycin (mTOR) signal pathway is the primary cause of RPE cell line de-differentiation, therefore, the inhibition of mTOR pathway will be helpful for the retard of de-differentiation of RPE cells.Objective This study aimed to investigate whether rapamycin can suppress the activation of mTOR pathway and promote differentiation of ARPE19 cells.Methods ARPE-19 cells were incubated in 12-well plate and divided into control group and rapamycin-treated group.DMSO or rapamycin with the final concentration of 400 nmol/L was added in the medium of the control group and the rapamycin-treated group, respectively.The cells of each group were collected 24 hours and 48 hours after cultured.The expression of zonula occludens-1 (ZO-1) in the cells was examined by immunofluorescence.The relative expression levels of RPE cell specific genes and proteins were assayed by using real-time quantitative PCR and Western blot, respectively.The detected results were compared between the two groups.Results ZO-1 was expressed in both group,but the fluorescence intensity was evidently enhanced in the rapamycin-treated group.The relative expression levels of RPE65, MERKT and LRAT mRNA in the cells increased by 25.97％ , 29.71％ and 13.00％ in the rapamycin treated group compared with the control group 24 hours after cultured (P=0.04,0.04,0.04) , and the expression levels of RPE65, LRAT, rLBP1, BEST1 , keratin18 and MERKT mRNA elevated by 174.00％ , 88.00％ , 56.18％ ,193.81％ ,10.83％ and 35.02％ in the rapamycin-treated group in comparison with the control group 48 hours after cultured (P =0.00,0.04,0.01,0.04,0.04,0.03).In addition, the expressions of p-mTOR, p-P70S6 and p-S6 protein were weaker in the rapamyein-treated group than those in the control group both 24 hours and 48 hours after cultured.Twenty four hours after cultured,the expression level of ZO-1 protein raised by 40％ in the rapamycin-treated group compared with the control group (P =0.01);while 48 hours after cultured,the expression levels of ZO-1 ,MERKT, catenin and LRAT proteins elevated by 36.00％ ,57.37％, 13.68％ and 41.07％ in the rapamycintreated group in comparison with the control group (P=0.01,0.00,0.04,0.04).Conclusions Rapamycin can suppress the activation of mTOR signaling pathway and up-regulate the expressions of RPE specific genes in ARPE19 cells.Inhibition of mTOR pathway might be an effective way for culturing RPE cells in vitro.

Background Retinal pigment epithelium (RPE) cell transplantation is the primary means of human trial for the treatment of retinal degeneration.Induced pluripotent stem cells (iPSCs) will become an important source for cell transplantation.In addition,iPS-RPE cells may provide a personalized treatment platform for the patient's own cells treatment.Objective This study was to evaluate the feasibility of human fibroblasts differentiate toward iPSCs from retinitis pigmentosa (RP) patients and toward iPSC-RPE cells from non-RP individual by retroviral transfection of Oct4,Sox2,c-Myc and KLF4 genes.Methods Human thigh skin tissues were obtained from a RP patient with hotspot mutation of SNRNP200 p.S1087L and individual without SNRNP200 p.S1087L mutation,respectively,with the size 2 c m×3 cm.Human dermal fibroblasts were isolated and cultured by trypsin digestion and explant method.The fibroblasts were transfected by a series of retrovirus and cultured by human embyonic cellconditioned medium to generate and induce iPSCs,and then the iPSCs were identified by morphology,alkaline phosphatase (AP) staining and immunofluorescence assay of specific markers of pluripotent stem cells.iPSCs suspension were injected into SCID mouse to observe the tumorigenesis.The iPSCs from non-RP subject were induced to differentiate toward iPS-RPE cells by embryonic body (EB) inducing method,and iPS-RPE cells were identified by detecting the expression of RPE65,zonula occludens protein 1 (ZO-1) and lecithin retinol acyltransferase (LRAT).Results Cultured human dermal fibroblasts showed fusiform or polygon shape and intercellular close arrangement,and Vimentin was positively expressed in the cells.Small cell colonies were harvested 5-7 days after infected by retroviruses,and the morphology changed from spindle into round mass.The hESC-like iPSCs clonies appeared 20 days after cultivation,and the positive expressions of hESC-specific surface antigens including SSEA3,SSEA4,TRA-1-60,TRA-1-81 and Nanog were found in the cells 25-30 days after cultivation,and the positive staining of AP was obtained 12 weeks after cultivation.A teratoma was formed at the injection site of iPSCs suspension in SCID mouse.Immunofluorescence technique showed that RPE cell-specific proteins including RPE65,ZO-1 and LRAT proteins were positively expressed in iPS-RPE cells at 30 days after differentiation.Conclusions Mutation SNRNP200 p.S1087L of RP patient-specific iPSCs can be induced from human dermal fibroblast by retrovirus infection method.The function and morphology of the iPSCs were similar to hESCs.Human iPSCs cell line generated from the dermal fibroblasts of non-RP individuals can differentiate into iPS-RPE cells.

Objective To investigate the popularization of cardiopulmonary resuscitation(CPR)knowledge and science popularization needs among urban and rural residents in Tonghai County,Yuxi City,Yunnan Province,so as to explore the establishment of an efficient and appropriate science popularization model.Methods A total of 300 residents aged 15-60 years old were selected from Tonghai County,Yuxi City,Yunnan Province using stratified and simple random sampling methods.A self-designed questionnaire was used to conduct an anonymous questionnaire survey.Results Only 20.3%of Tonghai County residents master CPR skills,and 26.2%of Tonghai County residents have never heard of CPR.There is a statistically significant difference in the awareness rate of CPR between rural residents and non-rural residents(P<0.01).There are differences in residents'age and CPR awareness(P<0.01),the age and CPR are inversely proportional.The residents have a higher willingness to perform chest compressions and mouth-to-mouth resuscitation on strangers,66.2%and 68.6%respectively.63.79%of residents have never attended relevant training.But 92.76%of the people said they were willing to participate in the relevant training when they learned the training news.Conclusion Residents in Tonghai County generally lack knowledge of CPR first aid,but the demand for first aid knowledge of residential CPR is high and the attitude towards rescue is positive.It is recommended that relevant departments increase CPR science popularization and training efforts,and popularize CPR into villages.

Objective:To summarize the clinical manifestations, imaging characteristics, and diagnoses basis of adrenomyeloneuropathy (AMN).Methods:The clinical data of 3 patients with AMN, admitted to our hospital from November 2016 to April 2019, were retrospectively collected. The clinical manifestations, imaging features, and diagnostic process of these patients were analyzed.Results:Three young male patients had onset with gradual aggravation of unilateral or bilateral lower limb insufficiency. MR imaging showed symmetrical abnormal signals in brainstem in 2 patients, and atrophy of thoracic spinal cord in 1 patient. By target region capture sequencing, mutations in the ABCD1 gene were found in all 3 patients; 2 underwent pedigree validation; the remaining one patient and his mother had failed Sanger sequencing validation due to pseudogene interference, and elevated plasma level of very long chain fatty acid (VLCFA) was noted in this patient. Conclusions:AMN usually initiates in the adulthood with spastic paraplegia as onset. Symmetrical lesions in brainstem or atrophy of spinal cord can be manifested on MR imaging; some patients may be accompanied by adrenocortical insufficiency. The definite diagnosis mainly depends on genetic screening and determination of VLCFA level in the blood.

Objective:To investigate the clinical characteristics of Guillain-Barré syndrome (GBS) combined with hyponatremia in Southern China and its risk factors for prognosis.Methods:The retrospective cohort study involved patients who met the diagnostic criteria of GBS from 18 upper first-class hospitals of 6 provinces/cities in southern China (south of Huaihe River) from January 1, 2013 to September 30, 2016. The clinical data of these patients were collected. According to serum sodium levels, they were divided into hyponatremia group (serum sodium concentration<135 mmol/L) and normal serum sodium group (serum sodium concentrations≥135 mmol/L). Based on Medical Research Coucil sum scores at nadir, these patients were divided into mild GBS group (>40), moderate GBS group (30-40), and severe GBS group (<30). Furthermore, according to the Hughes GBS disability scale (H-GBS-DS) scores at discharge, these GBS patients with hyponatremia were divided into favorable prognosis group (H-GBS-DS<3) and poor prognosis group (H-GBS-DS≥3). The incidence of hyponatremia in patients from the mild GBS group, moderate GBS group, and severe GBS group were compared. Multivariate Logistic regression analysis was performed to determine the clinical risk factors for hyponatremia in GBS patients. The clinical data of hyponatremia patients from favorable prognosis group and poor prognosis group were compared; multivariate Logistic regression analysis was used to determine the risk factors for poor prognosis in GBS patients with hyponatremia.Results:(1) Among the 570 patients, 354 had mild GBS, 94 had moderate GBS, and 122 had severe GBS; 134 GBS patients were combined with hyponatremia, 436 GBS patients had normal serum sodium. The hyponatremia incidence in mild, moderate and severe GBS groups increased successively, ( P<0.05). Multivariate Logistic regression analysis showed that facial paralysis ( OR=1.979, 95%CI: 1.172-3.342, P=0.011), respiratory muscle paralysis ( OR=3.218, 95%CI: 1.611-6.428, P=0.001), secondary pulmonary infection ( OR=4.822, 95%CI: 2.835-8.201, P=0.000), severe GBS ( OR=2.611, 95%CI: 1.444-4.721, P=0.001) and length of hospital stay ( OR=1.029, 95%CI: 1.009-1.050, P=0.004) were risk factors for hyponatremia in GBS patients. (2) Among 134 GBS patients with hyponatremia, 80 had poor prognosis and 54 had favorable prognosis. As compared with the favorable group, the poor prognosis group had significantly lower proportion of patients with extraocular muscle paralysis, statistically higher proportions of patients with respiratory muscle paralysis and secondary pulmonary infection, significantly different severities of GBS, signficantly higher proportion of patients accepted intravenous immunoglobulin (IVIG) and hormone treatments, statistically longer length of hospital stay ( P<0.05). Respiratory muscle paralysis ( OR=25.590, 95%CI: 9.433-69.423, P=0.000), moderate GBS ( OR=17.030, 95%CI: 8.441-34.361, P=0.000), and severe GBS ( OR=51.042, 95%CI: 24.596-105.926, P=0.000) were independent risk factors for poor short-term prognosis of GBS patients with hyponatremia. Conclusions:Severe GBS patients with facial paralysis, respiratory muscle palsy, secondary pulmonary infection, and long hospital stay trend to have hyponatremia. Hyponatremia patients with respiratory muscle paralysis and moderate/severe GBS have poor short-term prognosis.

Hepatic fibrosis is a pathological process in which the liver is subjected to various acute and chronic injuries for a long time, resulting in activation of hepatic stellate cells, the imbalance between the production and degradation of extracellular matrix, and the deposition of extracellular matrix in the liver, and it is jointly controlled by multiple cellular signal transduction pathways and a series of cellular information molecular networks. If there is no effective treatment, with the progression of the disease, liver fibrous nodules will form, destroy normal liver structure and function, and finally develop into liver cirrhosis, the decline of liver function, and even liver cancer. This article summarizes the research advances in the signaling pathways, receptors, and non-coding RNAs involved in liver fibrosis and the corresponding anti-hepatic fibrosis drugs/molecules.