Objective　To observe the effects of epithelial growth factor (EGF) and basic fibroblast growth factor (bFGF) on wound healing of the penetration keratoplasty (PKP) with autograft in rabbits. Methods　After the establishment of penetrating keratic autograft model on 24 rabbits, they were equally divided into EGF-, bFGF-, EGF+bFGF-treated and control groups. The wound healing of all the animals were observed with healing intensity, liquid scintillation counter, AgNORs staining, VG staining and transmission electron microscopy. Results　①EGF, bFGF, and EGF+bFGF increased the limiting pressure of the wounds and the 3H-TdR incorporation. ②The fibroblast cells and its secreting proliferative collagen in both the bFGF group and EGF+bFGF group were more well-arranged. Conclusion　①EGF, bFGF, EGF+bFGF can obviously elevate the intensity of wound healing after PKP and enhance the synthesis of DNA. ②The effect of this agent combination is just as the same as bFGF applied alone 14 d after the operation. ③bFGF can improve the quality of wound healing after PKP.

Objective　To observe the effect of epidemic growth factor (EGF) combining with basic fibroblast growth factor (bFGF) on wound healing of the penetrative keratic autoplasy in rabbits. 　 Methods　All 18 rabbits (36 eyes) were randomly divided into 9 groups and undergone penetrative keratic autoplasy, eyedrop.Diameters of the trephines were all 6 mm. Liquid scintillation counter was used to measure the incorporation rate of 3H-TdR. Pressure-detector was used to survey the intensity of keratic wound healing. AgNORs staining was used to count the fibroblast cells at keratic wound. VG staining, AgNORs staining and transmission electron microscopy were used to observe the morphological changes of keratic wound.　Results　(1) Both the wound intensity and the number of fibroblast cells of combination groups were higher than the 2 control groups in 8 days. (2) From the 14 th day to 21 st day, the wound intensity, the number of fibroblast cells and the incorporation rate of 3H-TdR in wound healing had no obvious difference from the positive control groups. 　Conclusions　 At the primary time, EGF combining with bFGF topic is more efficient to accelerate the wound healing of PKP. But the effect of this combination topic is just the same as bFGF topic alone 14 days later.

Objective　To observe the effects of epithelial growth factor (EGF) and basic fibroblast growth factor (bFGF) on wound healing of the penetration keratoplasty (PKP) with autograft in rabbits. Methods　After the establishment of penetrating keratic autograft model on 24 rabbits, they were equally divided into EGF-, bFGF-, EGF+bFGF-treated and control groups. The wound healing of all the animals were observed with healing intensity, liquid scintillation counter, AgNORs staining, VG staining and transmission electron microscopy. Results　①EGF, bFGF, and EGF+bFGF increased the limiting pressure of the wounds and the 3H-TdR incorporation. ②The fibroblast cells and its secreting proliferative collagen in both the bFGF group and EGF+bFGF group were more well-arranged. Conclusion　①EGF, bFGF, EGF+bFGF can obviously elevate the intensity of wound healing after PKP and enhance the synthesis of DNA. ②The effect of this agent combination is just as the same as bFGF applied alone 14 d after the operation. ③bFGF can improve the quality of wound healing after PKP.

Different human thyroid carcinoma cell lines were treated with all-trans retinoic acid(RA). RA could inhibit cell growth,improve iodide uptake and increase some thyroid specific genes and retinoid acid receptor(RAR)mRNA expressions in FTC-133 cells.However,RA had no effect in C643,HTH74 and XTC. UC1 cells.These findings indicate that different thyroid carcinoma cells display diverse responses to RA.

Objective To isolate and identify viruses from fecal samples of tree shrew with diarrhea.Methods Fecal sample supernatant of tree shrew with diarrhea was inoculated to three cell lines ( Vero, LLC-MK2 and KMB17 ) , and the cytopathic effects on the cells were observed.The infectious particles in the culture supernatant were further ana-lyzed by transmission electron microscopy ( TEM) , genomic RNA-PAGE, rotavirus detection kit, amplification of S1 com-plete segment and bioinformatics analysis.Results Constant cytopathic effects were induced in Vero, LLC-MK2 and KMB17 cell lines after three passages of culture.The results from TEM, RNA-PAGE and rotavirus analysis indicated that they belong to reoviruses.Analysis of the S1 segments revealed that the S1 sequence from KMB17 cell culture had the high-est homology with that of prototype isolate T1L (85%nucleotide homology and 90%amino acid homology), therefore this isolate was named as type I reovirus.The other two S1 sequences from LLC-MK2 and Vero cell culture were identical to have 85%nucleotide homology and 92%amino acid homology with the prototype isolate T3D, named as type III reovirus. Phylogenetic analysis indicated that the isolates in this study are evolutionally adapted to tree shrews.Conclusions It is the first report here that 2 genotypes of Tupaia orthoreovirus are isolated and identified from one fecal sample via three cell lines and viral S1-specific primers, which provides useful guidelines for the isolation and identification of other reoviruses from tree shrew or other hosts.

Objective To study the inhibitory effect of fluorouracil combined with DDP on the growth of human osteosarcoma cell line MG-63,and to explore its influence on the expressions of transient receptor potential vanilloid 5 (TRPV5 )and transient receptor potential vanilloid 6 (TRPV6 )proteins.Methods The MG-63 cells were cultured by the density of 5 × 104 mL-1 .Fluorouracil group,DDP group,fluorouracil+ DDP group and control group containing 10% FBS were set up.The inhibitory rates of growth of MG-63 cells at different time were detected by CCK-8 assay.The apoptosis of MG-63 cells after treated with different drugs was determined by Hoechst staining Kit.The immunocytochemical staining was used to treatent to detect the expressions of TRPV5 and TRPV6 before and after treatment.Results Fluorouracil and DDP both inhibited the growth of MG-63 cells in a time-and dose- dependent manner.There were a lot of black particles in the MG-63 cells and the cells were smaller,aging or death when they were exposed to fluorouracil or DDP.Compared with 24 h group,the inhibitory rates of proliferation of MG-63 cells after treated with the sigle drug of fluorouracil or DDP for 48 and 72 h were increased significantly (P <0.05).Compared with control group,the apoptotic rates of MG-63 cells in fluorouracil group and DDP group 24,48,and 72 h after treatment were increased (P < 0.01)in a time-dependent manner. The expression levels of TRPV5 and TRPV6 in MG-63 cells 72 h after treatment of fluorouracil and DDP were decreased significantly compared with before treatment (P < 0.05 ). Conclusion Fluorouracil, DDP and fluorouracil combined with DDP could significantly inhibit the proliferation of MG-63 cells,induce the apoptosis, and decrease the expression levels of TRPV5 and TRPV6.

OBJECTIVETo observe the anti-inflammatory effect of total saponins of Panax japonicus on LPS-induced RAW264. 7 macrophages.

METHODThe effect of total saponins of P. japonicus of different concentrations on RAW264. 7 cell viability was determined with the MTT method. The NO kit assay was adopted to detect the NO release of total saponins of P. japonicus to LPS-induced RAW264. 7 cells. The enzyme linked immunosorbent assay (ELISA) was used to detect the secretion of tumor necrosis factor-alpha (TNF-alpha) and interleukin 1-beta (IL-1beta). The reverse transeriptase-polymerase chain reaction (RT-PCR) was used to determine the expression of inducible nitric oxide synthase (iNOS) ,TNF-alpha,IL-1beta. The protein expression of nuclear transcription factor-kappaB p65 (NF-kappaB p65) was tested by Western blot.

RESULTThe safe medication range of total saponins of P. japonicus was less than 80 mg x L(-1). Compared with the LPS model group, total saponins of P. japonicus high, middle and low dose groups (0.1, 1, 10, 40 mg x L(-1)) could significantly reduce the secretion of NO, TNF-alpha, IL-1beta of LPS-induced RAW264. 7 cells, and inhibit the expressions of iNOS, TNF-alpha and IL-1beta mRNA and the protein expression of NF-kappaB p65.

CONCLUSIONThis study preliminarily proves the protective effect of total saponins of P. japonicus on LPS-induced RAW264.7 macrophages. Its action mechanism may be related to NF-kappaB signal pathway.

Animals ; Anti-Inflammatory Agents ; pharmacology ; Drugs, Chinese Herbal ; pharmacology ; Humans ; Inflammation ; drug therapy ; genetics ; immunology ; Interleukin-1beta ; genetics ; immunology ; Lipopolysaccharides ; adverse effects ; Macrophages ; drug effects ; immunology ; Mice ; NF-kappa B ; genetics ; immunology ; Nitric Oxide ; immunology ; Nitric Oxide Synthase Type II ; genetics ; immunology ; Panax ; chemistry ; Protective Agents ; pharmacology ; Saponins ; pharmacology

BACKGROUND: There are currently many cryopreservation methods for the aminotic membrane, which have varying effects on the ultrastructure and biological activity of amniotic membrane, but on no one is effective.
OBJECTIVE: To compare the effects of different cryopreservation methods on the ultrastructure and viability of aminotic membrane and to seek the ideal cryopreservation method.
METHODS: Aminotic membrane separated from the fresh placenta was preserved respectively with deep-frozen cryopreservation and vitrification, and everyway was run for 3 and 6 months. Fresh aminotic membrane was used as control. The ultrastructure of aminotic membrane was observed by transmission electron microscopy, and the viability of aminotic membrane was assessed by microcomputer analysis system for biological oxygen consumption, and immunohistochemical staining combined with image analysis system was used for lactate dehydrogenase activity.
RESULTS AND CONCLUSION:After 3 and 6 months of crypreservation, the damage to the ultrastructure of aminotic membrane by vitreous cryopreservation was slighter than that of amniotic membrane cryopreserved at-80℃. Compared with the fresh aminotic membrane, the gray value of lactate dehydrogenase and partial pressure of oxygen were significantly decreased in the cryopreserved aminotic membrane by deep-frozen cryopreservation at 3 and 6 months (P < 0.05) and by vitreous cryopreservation at 6 months (P < 0.05), but there was no statisticaly significant difference in the change rate of oxygen partial pressure and the gray value of lactate dehydrogenase between the fresh aminotic membrane and the cryopreserved aminotic membrane by vitreous cryopreservation at 3 months. The present study led to the conclusion that vitreous cryopreservation protocol alows to not only maintain the integrity of AM, but also to preserve the viability of the cels. So the vitreous cryopreservation is superior to the deep-frozen cryopreservation for cryopreservation of aminotic membrane.

Objective To explore the relationship between plasma homocysteine levels and diabetic retinopathy (DR).MethodIn 40 DR patients and 63 non-DR patients,the indexes including plasma homocysteine concentrations,duration of diabetes,ages,blood pressure,preprandial blood glucose(PBG),plasma GHbA,e,plasma lipids,serum folie acid and semm vitamin B12 were analyzed.ResultsPlasma homocysteine level was significantly higher in DR patients (12.9μmol/L) than that in non-DR patients (7.8μ.mol/L) (P＜0.01),and the relationship remained significant correlation after adjusted for duration of diabetes,GHbA1c,ages,serum folic acid and serum vitamin B12(OR = 1.23,P＜0.05).ConclusionHyperhomocysteinimia is a risk factor and an independent predieator of DR.

Objective: To study the relationship between serum levels of insulin-like growth factor-1 (IGF-1), cystation-C (Cys-C), interleukin-6 (IL-6) and atherosclerosis in middle and old age patient with newly diagnosed type 2 diabetes mellitus (T2DM). Methods: A total of 206 patients with newly diagnosed T2DM at the age of (67.3±10.4) years were enrolled. Based on color Doppler ultrasound examination, the patients were divided into 2 groups: Control group, the patient without carotid plaque or increased intima thickening, n=105 and Experiment group, patient with carotid plaque or increased intima thickening, n=101. The general information, fasting blood glucose, 2h postprandial blood glucose, glycosylated hemoglobin, LDL-C, HDL-C, TG, TC and IGF-1, Cys-C, IL-6, hs-CRP were recorded and compared between 2 groups, BMI was calculated in all patients. Results: Compared with Control group, Experiment group had increased carotid intima-media thickness (CIMT), elevated serum levels of Cys-C, IL-6, hs-CRP and reduced IGF-1, allP<0.05. Multiple Logistic regression analysis showed that CIMT was negatively related to IGF-1 (r=-0.493,P<0.01), positively related to Cys-C, IL-6 and hs-CRP (r=0.464,r=0.219 andr=0.618, allP<0.01). Conclusion: Serum levels of Cys-C and IL-6 might be the independent risk factors for atherosclerosis occurrence in meddle and old patients with T2DM; combined detection of IGF-1, Cys-C and IL-6 could help clinical diagnosis in relevant patients.