Background Age-related macular degeneration (AMD) is a group of vision-threatening eye disorders.Previous researches showed that the activation of Akt/mammalian target of rapamycin (Akt/mTOR) pathway closely related to the mechanism of AMD.A new specific method to inhibit Akt/mTOR pathway will become a breakthrough for the treatment of AMD.Objective This study was to establish a mouse fibroblast cell line (NIH/3T3) which can stably inhibit the expression of mTOR gene and provide a cell model for the study on the function of Akt/mTOR pathway in AMD and observe the influence of mTOR gene knockdown on related proteins.Methods Three short hairpin RNAs (shRNAs) targeting mTOR gene were designed and synthesized based on murine mTOR mRNA sequence.Double-strand shRNA hairpins were separately cloned into PIGZ-green fluorescent protein (GFP) +Puro vectors to produce recombination plasmids.The packaged lentiviral plasmids and RNAi plasmids were co-transfected into NIH/3T3 cells,a mouse fibroblast line.After puromycin selection and culture expansion,0.5 mg/L puromycin was added the culture medium to establish stable cell clones.The expressions of mTOR mRNA and protein in NIH/3T3 cells were detected by real-time PCR and Western blot respectively,and the inhibitory efficiency of interference was analyzed.Results Transfected GFP-labeled NIH/3T3 cells by lentiviral presented the green fluoresccence with the efficiency of infection of 90％ in the third day.Real-time PCR showed a distinct band of mTOR mRNA in 184 bp.The knockdown rate for sh1,sh2 and sh3 were respectively 31.3％,31.8％ and 45.3％ in the lower multipolicity of infection (MOI) group ;while in the higher MOI group,the knockdown rate for sh1,sh2 and sh3 were 47.1％,56.5％ and 71.6％ respectively.Western blot assay exhibited weakened expression band of mTOR protein in NIH/3T3 cell line for sh1,sh2 and sh3 in both lower and higher MOI groups with the weakest expression for sh3.Conclusions A stable mouse fibroblast cell line is established by the inhibition of mTOR with RNA interference technique,which can provide a cell model for studying the function of Akt/mTOR pathway in AMD.

Sarcopenia and osteoporosis are two common diseases with adverse effects on the health of the elderly and share many characteristics.As target organs of the two diseases,muscles and bones are closely connected,not only in anatomy and physiology,but also in pathophysiological processes.Initially,mechanical stimulation was conducted to explain the relationship between muscle and bone at the macroscopic level.Currently,research is increasingly focused at the cellular and molecular levels.Both muscle and bone tissues can secrete a number of growth factors,cytokines,polypeptides,and so on.Myokines include fibroblast growth factor-2 (FGF-2),myostatin,irisin,insulinlike growth factor-1 (IGF-1),interleukins,musclin,among others.Osteokines comprise osteocalcin,fibroblast growth factor-23,IGF-1,vascular endothelial growth factor,etc.In this review,we summarize the roles of myokines and osteokines in muscle-bone interactions.

Child ; China ; epidemiology ; Female ; Human bocavirus ; Humans ; Infant ; Infant, Newborn ; Male ; Parvoviridae Infections ; epidemiology ; virology ; Respiratory Tract Infections ; epidemiology ; virology

Anionic surfactants in waters were determined using on-line double microporous membrane extraction-flow injection analysis method. This method was based on the extraction of the ion-association complex (λ_(max)=650 nm) which formed from methylene blue cation and anionic surfactants. The on-line double microporous membrane liquid-liquid extraction phase separators were adopted. Optimization of variables of the flow injection manifold, groove mechanical dimension of the phase separator, and pore size of the membrane was performed. The proposed method had a linear range of 25.0-1000.0 μg/L(r≥0.999). The detection limit was 4.28 μg/L. The relative standard deviation(n=7) of different concentration was 0.7%-6.0%. The recovery was between 96%-110% and the sample throughput was 18 h-1. The method was applied to the analysis of standard Reference materials with satisfactory results. For practical application, two water samples were analyzed with the established method.

Adolescent ; Diagnosis, Differential ; Hepatectomy ; methods ; Humans ; Liver Cirrhosis ; congenital ; diagnostic imaging ; pathology ; surgery ; Male ; Tomography, X-Ray Computed

Objective To explore different socio-demographic and clinical characteristics for early onset patients (EOP) with psoriasis and late onset patients (LOP) with psoriasis in Chinese population and to provide scientific evidences for establishing comprehensive prevention and treatment strategy for psoriasis.Methods Cross-sectional study was performed.A total of 942 diagnosed psoriasis patients who paid a visit to outpatient clinic in a tertiary hospital in China from 12th September 2013 to 19th May 2015 was consecutively enrolled and investigated.Exploratory analysis was conducted to detect the association between disease onset age and patients features.Results Among 942 enrolled psoriasis outpatients,the average age was (40.6 ± 13.6) years with a range from 5 to 80 years.The sex ratio was 2.1 in favor of male.The most observed type of psoriasis in the present study was psoriasis vulgaris (98.3％).The average psoriasis area and severity index (PASI) for patients were 9.47 ±7.63.Comorbidity was combined in 20.2％ cases.Patients with EOP had a significantly higher likelihood of family history of disease.Compared EOP to LOP,LOP had a significantly higher likelihood of comorbidities involvement (P ＜ 0.05) and significant higher BMI index (P ＜ 0.05).Conclusions The present study supports the hypothesis that there are clinical differences between EOP and LOP in Chinese population.Both dermatologists and patients should pay more attention to psoriasis-associated features,such as family history and comorbidity diseases involvements.

Objective To investigate the relationship between the ultrasonography diagnosis in prenatal and the postnatal placental vascular casts of the twins reversed arterial perfusion(TRAP) sequence. Methods Six cases of TRAP sequence underwent the two-dimensional and color Doppler ultrasonography, and performed placental vascular casts after termination of pregnance. Results According to the anatomical features and ultrasonography, there were 3 cases of acardius acephalus( IV) ,2 cases of acardius anceps( II ) , and 1 case of acardius acormus( I ). In all 6 cases,there were arterio-arterial and veno-venous anastomoses at the level of the chorionic plate between the normal co-twin and the acardiac twin,these anastmosis vessels were superficial and large calibre, and the distances of the placental insertions of the umbilical cords were different,4 cases <0. 5 cm, 2 cases were 4-5 cm. In 4 cases, the vessels entered and went out of the acardiac twin were just one large umbilical artery and one large umbilical vein. In 2 cases, the umbilical artery and umbilical vein had many vascular branches in the placenta. Out of 6 cases of the pump twin, 4 cases died,and 2 cases survived. Conclusions Ultrasonography was consistent with the placental vascular casts,the contrast observing gives us new information for the further research of the special type of the vascular anastmosis and prognosis of the TRAP sequence.

Animals ; Carcinoma, Squamous Cell ; genetics ; Colorectal Neoplasms ; genetics ; metabolism ; Enzyme Activation ; Esophageal Neoplasms ; genetics ; Genome-Wide Association Study ; Head and Neck Neoplasms ; genetics ; Humans ; Neoplasms ; chemically induced ; enzymology ; genetics ; Phosphoinositide Phospholipase C ; chemistry ; genetics ; metabolism ; physiology ; Signal Transduction ; Skin Neoplasms ; chemically induced ; enzymology ; Stomach Neoplasms ; genetics ; Urinary Bladder Neoplasms ; metabolism ; pathology ; ras Proteins ; metabolism

Objective To investigate the clinical characteristics and mutations of guanosine triphosphate eyclohydrolase (GCH) Ⅰ gene in patients with dopa-responsive dystonia (DRD). Methods Five families with 18 affected family members and 17 patients with sporadic DRD were examined. Patients were allocated into 3 groups according to onset time, either in childhood, or in adolescence or adult. Interview, physical examination, psychologic testings and CT/MR scan were performed. Mutation screening was performed on 26 patients and 1 normal family nember. Thirty-five healthy control subjects were matched for age and sex. Statistical analysis were conducted with the use of SPSS 13.0 computer software. Results(1)Most of patients started with dystonia. The main clinical manifestation was dystonia too. There was no difference among 3 groups.(2) There were significant differences in diurnal fluctuation among 3 groups(15/15,6/6,7/14, χ2=13.125,P=0.001). Diurnal fluctuation negatively correlated with age (r=-0.720, P<0.01).(3)The differences in postural tremor were also found among 3 groups (7/15,5/6,1/14, χ2=8.073, P=0.018). Postural tremor positively correlated with age (r=0.399, P=0.018).(4)There were differences in exaggeration of tendon among three groups(11/15,1/6,4/14, χ2=8.309, P=0.016). Exaggeration of tendon reflexes negatively correlated with age (r=-0.429, P=0.010).(5)The scores of Hamilton Depression Scale and Hamilton Anxiety Scale in patients were higher than those in controls.(6)DNA sequencing revealed a heterozygous A224G missense mutation(Tyr75Cys)located within exon 1 in one autosomal dominant inheritance family. Conclusions The manifestations of DRD varies. The clinical course is closely correlated with age. A missense mutation(A224G)in coding region of the GCH 1 gene probablyleads to the occurrence of DRD.

Objecfive To study the effects of some cytokines such as TNF-α,IL-6 and IFN-γ as well as lipopolysaccharide on CD68 expression in HaCaT cells.Methods Human HaCaT keratinocytes were randomly divided into natural proliferation group (without stimulation),IFN-γ-stimulated group,TNF-α-stimulated group,LPS-stimulated group and IL-6 stimulated group.The work concentration of TNF-α,IL-6,IFN-γ and LPS was 50 mg/L.HaCaT cells were collected after 24-hour treatment with the cytokines followed by the examination of CD68 expression with flow cytometry,immunohistochemistry and reverse transcription(RT)-PCR,respectively.Results Compared with untreated HaCaT cells,the count of CD68-positive cells was elevated in cells stimulated by TNF-α(t=3.60,P<0.01),IL-6(t=3.93,P<0.01),IFN-γ(t=2.38,P<0.05)and LPS(t=2.52,P<0.05),and the effect of TNF-α and IL-6 was stronger than that of IFN-γ and LPS.Among the four cytokines,only IL-6 enhanced the mean fluorescence intensity of CD68-positive cells (t=8.34,P<0.01).After 24-hour treatment with TNF-α,IFN-γ and IL-6,CD68 expression was observed in the cytoplasm and on the membrane of HaCaT cells and was stronger in cells treated with TNF-α and IL-6 than in those with the other cytokines.A significant increase was observed in the CD68 mRNA expression after 24-hour treatment with TNF-α (t=4.34,P<0.01),IL-6 (t=7.52,P<0.01)and IFN-γ (t=2.81,P<0.05);TNF-α and IL-6showed a stronger promotive effect than IFN-γ.Conclusion IL-6,TNF-α,IFN-γ and LPS can upregulate the CD68 expression in HaCaT cells.