Role of mitochondrial DNA in human retinal pigment epithelium cells apoptosis induced by blue light / 中华实验眼科杂志

Objective To research the role of mitochondrial DNA mediate the cultured human retinal pigment epithelium (hRPE) cell apoptosis induced by blue light and the relationship with time.Methods Established the blue light damage model of cultured hRPE cells in vitro with light emitting diode (LED) blue light density of (4.0-±0.5)mW/cm2 adjusted by FL-1D blue light illumination meter,and the illumination time was set as 0,0.5,1,2,4,6,12 and 24 hours,then the cells were grouped according to the illumination time.Immunofluorescence were used to identify the cells;the expressions of caspase-3,cleaved caspase-3,caspase-9,cleaved caspase-9,bax and bcl-2 were detected with Western blot.Quantitative PCR was used to detect the copy number of mitochondrial DNA and PCR was used to detect mitochondrial DNA 4977bp common deletion.Results Immunofluorescence results showed that the RPE65 protein was expressed in the cytoplasm.The expressions of bax were upregulated after illumination for 1 hour,cleaved caspase-3 were upregulated after illumination for 2 hours,caspase-3,caspase-9,cleaved caspase-9 were upregulated after illumination for 4 hours,while the expression of bcl-2 was downregulated after illuminated for 2 hours,with significant differences compared to the normal control group (all at P＜0.05).The copy number of mitochondrial DNA in 0.5,1,2,4,6,12 and 24 hours groups was downregulated,with significant differences compared to the normal control group (all at P＜0.05).The expressions of 4977bp common deletion in 0.5,1,2,4,6,12 and 24 hours groups were increased,with significant differences compared with the normal control group (all at P＜0.05).Conclusions Blue light can cause cell apoptosis,especially mitochondrial apoptosis,in hRPE probably motivated by mitochondrial DNA damage.

Effects of artesunate on cell proliferation and apoptosis of human Tenon's capsule fibroblasts / 眼科新进展

Objective To investigate the effects of artesunate (Art) on cell proliferation and apoptosis of human Tenon's capsule fibroblasts (HTFs),and discuss the countermeasures of bleb scarfing in glaucoma.Methods In vitro,HTFs were cultivated and applicated by different concentrations (50 μg · mL-1,100 μg · mL-1,150 μg ·mL-1,200 μg · mL-1) of Art for 48 hours.The effect of Art on cell proliferation was assessed by MTT method.The rate of apoptosis induced by Art was determined by flow cytometry.Western Blot was performed to detect the relative expression levels of Bax and Bcl-2 after Art was treated.Results After treated with Art for 48 hours,compared with blank control group,Art (50 μg · mL-1,100 μg · mL-1,150 μg · mL-1,200 μg · mL-1) group exhibited notable anti-proliferative effect on HTFs with concentration-dependence (all P ＜ 0.05).The results of flow cytometry showed that the apoptosis rates (8.80％ ±0.88％,11.60％ ±0.56％,16.30％ ±1.03％,23.40％ ±1.62％) of HTFs were significantly enhanced with the increase of Art concentration (all P ＜ 0.05).The relative expression levels of Bax were obviously high with the increase of Art concentration,while Bcl-2 levels were significantly low with the increase of Art concentration (all P ＜ 0.05).Conclusion Art can inhibit the proliferation and induce cell apoptosis of HTFs possibly by enhancing the expression of Bax and reducing the expression of Bcl2.Art may be a potential drug in preventing fibrous scar formation after glaucoma filtration surgery.

Protective effects of tissue factor targeting peptide on human retinal pigment epithelial cell damage induced by blue light / 中华实验眼科杂志

Background Light-induced retinal damage results in the damage of retinl pigment epithelial (RPE) cells and therefore affects the pathogenesis and development of age-related macular degeneration (AMD).Studies showed that tissue factor (TF) is overexpressed in oxidative damaged RPE cells and the choroidal neovascularization (CNV) of AMD,speculating that the suppression of TF can prevent the damage of RPE cells and inhibit CNV.Objective This study was conducted to observe the protective effects of TF targeting peptide (TFTP),a new drug of autologous synthesis,on human RPE-cells induced by blue light.Methods Human RPE cells were isolated from donor eye and cultured.Cultured cells were divided into blank control group,model group and TFTP treated group.Light-induced RPE cell damage model was established by exposuring the cells in the blue light of (4.0±-0.5) mW/cm2 for 12 hours in the model group,and different concentrations (10,100,150,200,300 μmol/L) of TF-TP were added into the medium to pretreat the cells for 24 hours and then exposed the cells to the blue light for 12 hours in the TF-TP groups.The cell viability was determined by CCK-8 assay.The morphology and ultrastructure in the cells were observed under the inverted microscope and transmission electron microscope.The apoptosis of the cells was assayed by Hoechst staining.The expressions of TF and apoptosis-related protein bax,bcl-2 in the cells were determined by Western blot.Results CCK-8 assay showed that there was no significant difference in the cell viability among blank control group and different concentrations TF-TP groups (F=2.15,P =0.11).The cell survival rate of blank control group,model group and 150 μmol/L TF-TP group was (100.0±0.00) ％,(43.79±6.55) ％ and (63.45±3.57) ％,and the survial rate was increased in the 150 μmol/L TF-TP group compared with the model group (P =0.00),and 150 μmol/L was detemined as a optimal concentration of TF-TP.A lot of shrinkage,deformation,suspension cells were exhibited under the optical microscope,and decrease of microvilli structure,rupture of mitochondrial cristae and vacuolar degeneration of the cells were found in the model group,and the damage of the cells were evidently lightened in the 150 μ mol/L TF-TP group.The apoptosis rate of the cells were (0.98 ±0.19)％,(9.98 ±0.82) ％ and (5.73 ±0.88) ％ in the blank group,model group and 150 μmol/L TF-TP group,respectively,with a significant difference among the groups (F =206.18,P =0.00),and the apoptosis rate of the cells in the 150 μmol/L TF-TP group was significantly lower than that in the model group (P＜0.05).Compared with the blank control group,the relative expression of bax and TF was obviously increased and that of bcl-2 was decreased in the model group;while the expression of bax and TF was lower,and that of bcl-2 was higher in the 150 μmol/L TF-TP group compared with the model group (all at P ＜ 0.05).Conclusions Pretreation of TF-TP can lessen cell apoptosis and increase cell survival rate and therefore plays a protective role to blue light-induced human RPE cells possibly by inhibiting bax/bcl-2 apoptotic pathways mediated by TF.

Oxidative damage of human retinal pigment epithelium cells induced by blue light irradiation and mitochondria-participated mechanism / 中华实验眼科杂志

Background Researches showed that mitochondria and oxidative stress play a crucial role in retinal photochemical injury,but the relationship between the damage of human retinal pigment epithelium (RPE) cell-induced by blue light and light-irradiated time is less studied.Objective The aim of this study was to research the possible mechanism of RPE oxidative damage induced by blue light in vitro.Methods Human RPE cells were isolated from healthy donors and cultured.The cells were divided into the normal control group and the light exposure group.The cells of light exposure group were irradiated using the blue light of (4.0±0.5) mW/cm2 for 0.5,1,2,3,4,5,6,12 and 24 hours,respectively,and the cells of the normal control group were cultured in dark environment.Cellular viability was detected by MTT method,and the ultrastructure change of subcellular organelles in RPE cells was examined under the transmission electron microscope (TEM).The content of reactive oxygen species (ROS) was assayed by flow cytometry for the assessment of oxidative stress reaction.The relative expressions of nicotinamide adenine dinucleotide phosphate (NADPH) mRNA and cyclooxygenase 1 (COX1) mRNA in the cells were detected by real-time fluorescence quantitative PCR to evaluate the mitochondria function.Results The percentages of cellular viability were (100.00±20.00) ％,(95.73±0.89) ％,(94.67±2.56) ％,(84.23±0.16) ％,(78.57±3.09)％,(75.43±2.18)％,(66.13±1.42)％,(53.43±1.91)％ and (47.97±1.36)％ in the normal control group and light exposure for 1-hour,2-hour,3-hour,4-hour,5-hour,6-hour,12-hour and 24-hour groups,respectively,showing a significant difference among the groups (F =172.270,P =0.000),and the percentages of light exposure for the more than 3 hours groups were significantly lower than those of the normal control group (all at P＜ 0.05).The vacuoles-like degeneration,mitochondrial swelling,decreased microvilli were seen under the TEM.The contents of ROS in RPE cells were (14.75±2.49)％,(19.04± 1.02) ％,(22.81 ±3.20)％,(28.75±2.15)％,(33.06±0.96) ％,(40.64±2.11) ％,(48.25±2.50) ％ and (60.44±2.68) ％ in the normal control group and light exposure for 0.5-hour,1-hour,2-hour,3-hour,4-hour,5-hour,6-hour groups,and with significant increases in ROS contents in various light exposure groups compared with the normal control group (all at P＜0.05).The relative expression levels of NAPDH mRNA in the cells were gradually elevated 3 hours after light exposure with the increase of time in comparison with the normal control group (all at P＜0.05),and the relative expression levels of COX1 mRNA in the cells were higher in the light exposure for 2-hour,3-hour,4-hour and 5-hour group compared with the normal control group (all at P＜0.05),and after that the COX1 mRNA levels were gradually declined and were close to the normal level.Conclusions Blue light irradiation for more than 3 hours causes oxidative stress damage of mitochondria in RPE in vitro,and the damage was more obvious after irradiation for 5-6 hours.

The relationship between mitochondrial DNA damage and retinal pigment epithelium cells / 天津医药

Mitochondrial DNA (mtDNA) is a genetic effect DNA molecule of double closed loop, and is crucial for cells and their functions. Mitochondria take an active part in physiological activities of retinal pigment epithelium (RPE) cells. The oxidative stress is usually occurred in RPE for its active metabolism, which can lead to mitochondria and mtDNA dam?age. Once mitochondria and mtDNA lesions have not been repaired timely, the lesions can be accumulated, which can cause dysfunctions and damaged-structures of RPE and mitochondria, and can motivate the progression of cell apoptosis. In the end it can result in some ocular related diseases such as aged-related macular degeneration (AMD). This study reviewed the functional relationship between mtDNA and RPE, and repair and detection methods of mtDNA damage.

Anti-apoptosis effect of lipoic acid-niacin diad on acrolein-induced apoptosis in ARPE-19 cells / 实用医学杂志

Objective To investigate the regulations of Bax , Bcl-2 in the protection of lipoic acid-niacin diad in acrolein-induced apoptosis in ARPE-19 cells. Methods The ARPE-19 cells were cultured in medium containing 10% fetal bovine serum , at 37 ℃ with 5% CO2. The ARPE-19 was transferred to 6-well plate after reaching to 70% confluence. After starvation for 24 h , the cells in 6-well plates were divided into three groups , including the blank control group , the acrolein treatment group with 50 μmol/L acrolein for 24 h , and the protection group with 100 μmol/L lipoic acid-niacin diad for 24 h and with the acrolein for another 24 h. The apoptotic cells were detected by flow cytometry assay , and expressions of Bcl-2 , Bax protein were detected by Western Blot assay. Results The percentages of normal healthy cells were 94.8%, 60.98%, and 91.34% in the blank control group , 50 μmol/L acrolein group and 100 μmol/L diad contained of lipoic acid and niacin group , respectively. The ratios of Bax/Bcl-2 protein expression were 0.293 9, 1.389 2, and 0.555 8 in the blank control group, 50 μmol/L acrolein group and 100 μmol/L diad contained of lipoic acid and niacin group, respectively. Conclusion The protective effect of lipoic acid-niacin diad on acrolein-induced apoptosis in ARPE-19 cell through promoting Bcl-2 expression and inhibiting Bax expression.

Transforming growth factor-β1 level in tears and corneal haze formation following flap-on or flap-off epi-LASIK / 南方医科大学学报

<p><b>OBJECTIVE</b>To observe the influence of flap-on or flap-off Epipolis laser in situ keratomileusis (epi-LASIK) on the release of transforming growth factor-β1 (TGF-β1) in tear fluid and corneal haze formation.</p><p><b>METHODS</b>Thirty patients (60 eyes) with myopia underwent epi-LASIK surgery with epithelial flap repositioning (flap-on) in the right eyes and epithelial flap removal (flap-off) in the left eyes. The level of TGF-β1 in tears was measured preoperatively and on days 1, 3, and 7 postoperatively. Corneal haze was graded at 1, 3 and 6 months after surgery.</p><p><b>RESULTS</b>The mean preoperative spherical equivalent refraction was -4.98∓2.28 D (-2.50 to -7.25 D) in flap-on group and -5.20∓4.02 D (-1.75 to -7.00 D) in flap-off group, showing no significant difference between the two groups (P=0.80). TGF-β1 levels in the tear fluid were similar in the two groups preoperatively (P=0.11) and at 1, 3, and 7 days postoperatively (P=0.55, 0.45, 0.19, respectively). TGF-β1 levels in tears gradually decreased after the first postoperative day in both groups, but were still higher than the preoperative value till the 7th postoperative day. Corneal haze scores in the two groups were similar at 1 month (P=0.98), 3 months (P=0.52), and 6 months (P=0.72) after the operation.</p><p><b>CONCLUSION</b>Flap-on and flap-off epi-LASIK surgeries do not differ significantly in postoperative TGF-β1 levels in the tear fluid or in the postoperative haze scores. TGF-β1 may play a role in corneal wound healing.</p>

Applications of Different Insulin Therapies on type 2 diabetic patients in Fasting State / 中国基层医药

Objective To investigate the status of diabetes in fasting state on different therapies.Methods 244 type 2 diabetic patients were divided into four groups according to different insulins:60 for insulin pump group(A),64 cases for insulin glargine group(B),59 cases for microinjection pump group(C),61 cases for isophane insulin suspension group.Then the time when blood glucose was up to standard,the usage of insulin and the rate of hypoglycemia were abserved.Results A group was(2.89 ± 1.32)d on the time when blood glucose was up to standard,B group was(3.14 ±1.25)d,C group was(4.91 ±2.81)d and D group was(5.62 ±2.52)d.There were significant differences between A and other groups( P ＜ 0.05 ) on time.There were significant differences between B and C group,B and D group,but there was no difference between C and D group.A group was(30.61 ±2.21) IU/don the usage of insulin,B group was ( 31.12 ± 3.38 ) IU/d,C group was ( 42.25 ± 4.01 ) IU/d and D group was (44.31 ±3.22)IU/d.There were significant differences between A and C group,A and D group,B and C group,B and D group(P ＜0.05)on usage of insulin,but there were no differences between A and B group,C and D group.A group was 3.3％ on the rate of hypoglycemia,B group was 4.7％,C group was 15.2％ and D group was 16.4％.There were significant differences between A and C group,A and D group,B and C group,B and D group ( all P ＜ 0.05 ) on usage of insulin,but there were no differences between A and B group,C and D group.Conclusion Compared to other insulin therapies,insulin pump had obviously superiority,which could be generalized to diabetics in fasting state.

Protective effects of alpha-lipoic acid against beta-cell damage in streptozotocin induced diabetes In rats / 中华胰腺病杂志

Objective To investigate the protective effects of alpha-lipoie acid (ALA) against beta-cell damage in streptozotocin (STZ)-induced diabetes in rats. Methods Thirty SD rats were randomly divided into three groups: normal control (NC) group, STZ group and ALA + STZ group, with 10 rats in each group. mg/kg, intraperitoneal injection), till the end of the study (4 weeks later). Blood glucose were measured every 3 days after STZ injection. Malondialdehyde (MDA) and reduced glutathione (GSH) levels were measured in pancreatic homogenates. Pancreatic beta-cells were examined by immunohistocbemical methods, Results STZ induced a significant increase of the level of blood glucose. Body weight of rats in ALA + STZ group was (341±26)g, which significantly lower than (368±3)g in NC group, and high than (301±2)g in STZ group with stas(P < 0. 05). Meanwhile the MDA levels in STZ group and NC group were(1.22 ± 0. 14) and(0.57 ± 0.04)nmoL/mg prot, respectively, and there was significant difference between the two groups (P < 0.05) ; the GSH levels in STZ group and NC group were(16.54 ± 1.10) and(25.46 ± 0.62) mg/g prot (P < 0.05), respectively; degeneration of islet cells and decreased blood glucose were observed in STZ + ALA-pretreated rats; MDA level in pancreatic homogenates was(0.72 ± 0. 23)nmoL/mg prot, which was significantly lower than that in STZ group (P < 0.05) ; the GSH level was (35.33 ± 2.66) mg/g prot, which was significantly higher than that in STZ group (P < 0.05) ; increased staining of insulin and preservation of islet ceils functions were more obvious in the STZ + ALA-pretreated rats. Conclusions ALA exerted its protective effect through reducing the oxidative stress and preserving pancreatic beta-cell integrity.

The analysis on clinical data of 22 patients with preliminarily diagnosed islet cell adenoma / 中国医师进修杂志

Objective To study and evaluate islet cell adenoma of preliminary diagnosis.Method Retrospective analysis Wag done on 22 patients clinical data of preliminarily diagnosed islet cell adenoma.They were divided into two groups according to final diagnosis,group of islet cell adenoma(n=16)and group of reacfional hypoglycemia n=6).Results There were significant differences between the two groups in term of fames,cold sweat,cataphora,Whipple triad,hungry test,the ratio offasting blood insulin and fasting blood glucose exceeding 0.3 (I/G＞0.3) (P＜0.05).No statistical differences were found on blood glucose level in hypoglycemia onset,psychiatric symptom.Conclusion The clinic data is helpful for preliminary diagnosis,and Whipple triad,hungry test,I/G＞0.3 above all.

Quantitative ultrasound measurement and evaluation of bone quality in normal Hubei Yichang subjects / 中国医师进修杂志

Objective To measure the parameters of ealeaneal quantitative ultrasound(QUS)in nomal Hubei Yichang subjects.which were compared between different age groups after standardization by body mass index(BMI).Methods Calcaneal parameters were measured by UBIS5000 in 2912 normal Hubei Yichang subjects,from 20 to 82 years old women 1450,men 1462).Results The peak values of broadband ultrasound attenuation(BUA)and stiffness(STI)of the calcaneal were both in 45-49 year group for women.The peak value of BUA was in 50-54 year group for men.Standardized by BMI,the peak value of BUA、STI and sound of speed(SOS)were both in 20-24 year groups for women and men.Conclusion BUA、SOS and STI of the calcaneal may change with age.After the influence of body mass is eliminated,the change still exists obviously,and the parameters may gradually reduce with increasing age.

Clinical study of continuous subcutaneous Insulin infusion combined with mefformln hydrochloride in the in-teusive treating of type 2 diabetes mellitus / 中国基层医药

Objective To evaluate the efleet of continuous subcutaneous insulin infusion (CSII)combined with mefformin hydroehloride( Met H)on type 2 diabetic patients. Methods 240 patients with type 2 diabetes melli-tus (T2DM) were divided into 3 groups randomly: CSII combined with Met H ( A group: n=80) ; simple CSII ( B group:n=80) ; multiple deliver insulin (MDI) ( C group: n=80). The levels of blood glucose, HbA1 e, therapeutic time, a-mount of insulin, Fins ,C-P, HOMA-IR and IAI were compared among them before and after 10d of treatment. Results GroupA, B eornpared with group C, the levels of HbA1 e significantly deeroased in other two groups [ ( 8.0±3.2)%vs. (6.9±1.4)% and(7.0±1.2)% ] (aU P<0.05),so did the therapeutic time[(6.0±1.9)d vs. (3.0±1.4) and(3.5±1.5) d] and insulin dose[ (55.0±17.0) U/d vs. (38.0±15.0) U/d and(45.0±16) U/d] (all P<0.05). The insulin dose was much less in group A than in group B. Versus group B and C,the HOMA-IR of group A (0.8±0.3 vs. 1.5±0.4 and 1.6±0.4)decreased(P<0.05) ,and IAI(-2.0±4.3 vs. -3.6±4. 2 and -4.1±4.3) increased (all P<0.05), without the function impairments of heart, liver or kidney. Conclusion CSII in combi-nation with mefformin hydreehloride can significantly enhance the treatment effect of the patients with type 2 diabetes mellitns; reduce blood glucose and HbA1 e, improve insulin resistenee,lessen insulin need.

Effect of insulin pump in treatment of type two diabetic patients with pulmonary infections / 中国医师进修杂志

Objective To study the effect of continuous subcutaneous insulin injection(CSII) in treatment of type two diabetic patients with pulmonary infections. Methods From January 2001 to December 2005,83 type two diabetic patients with pulmonary infections were randomized to group CSII (42 cases) and group MSII(41 cases). The period of normalization of the blood glucose and cure of the pulmonary infections, cost and morbidity of low blood glucose were observed. Results Blood glucose of all patients in two groups reduced significantly. Significant difference was observed in the period of cure of the pulmonary infections, usage amount of insulin and morbidity of low blood glucose. In group CSII HbA1c, CRP and CHOL reduced significantly. Fasting CRP and high density lipoprotein-C (HDL-C) were remarkably increased. Conclusions Consecutive subcutaneous insulin pump injection is able to control the blood glucose better, to correct metabolic disorder, reduce the cost and hospital day and decrease the morbidity of low blood glucose. CSII is a better of method.

Clinical application of insulin pump in treatment of diabetic ketoaciduria / 中国医师进修杂志

Objective To evaluate the difference of efficacy between continuous subcutaneous insulin infusion(CSII)and continuous intravenous insulin infusion(CVII)with human insulin in patients with diabetic ketoaciduria or ketoacidosis(DKA). Methods 120 patients with DKA were randomized into two groups,one was CSII group by a portable insulin pump(60 cases),the other was CVII group(60 cases).Results Both of CSII and CVII were effective in controlling blood glucose levels.CSII therapy provided better glycemic control(P