Objective Researches showed that matrix metalloproteinase-2 (MMP-2) has a critical role in the neovascularization of tumor,and vascular endothelial growth factor(VEGF)is a promoting factor of new blood vessel formation.There still is little literature about the effect of MMP-2 in retinal neovascularization up to now.This study tried to explore the expression and significance of MMP-2 and VEGF in retinal neovascularization.MethodsA retinal neovascularization model was established in 30 7-day-old cleaning C57BL/6J mice exposed to an environment of high concentration of oxygen for 5 days,and 30 matched mice were raised in normal air environment.Fifteen mice from hyperoxic group and control group were sacrificed in 10 days after treatment and the eyeballs were enucleated to make retinal stretched preparation.Adenosine diphosphate-ase (ADPase) stained retina flat-mounts was performed to assess the retinal vascular profiles,and H&E staining was applied to count the number of new vascular cell nuclei.The expression of MMP-2 and VEGF was detected using immunohistochemistry by calculating the intergrated value of positive cells.ResultsThe retinal stretched preparation presented more neovascularization in mice from the hyperoxic group compared with control group.The number of nuclei from the vascular endothelial cells in the new vessels breaking through the internal limiting membrane in the hyperoxic group was 33.51±2.55,indicating a significant increase in comparison with control group (7.27±0.20)(t=9.345,P＜0.05).There were stronger expression of MMP-2 protein and the VEGF protein in the ganglion cell layer,inner plexiform layer and inner nuclear layer,and neovascularization breaking through the internal limiting membrane in the hyperoxic group compared with control group (t=4.25,P＜0.05;t=6.38,P＜0.05).Expression of MMP-2 showed the positive correlation with the expression of VEGF(r=0.825,P＜0.05).ConclusionBoth MMP-2 and VEGF promote retinal neovascularization.The overexpression of MMP-2 and VEGF play a synergistic role during the formation of neovascularization.

AIM: To study the inhibitory effect of captopril on retinal neovascularization (RNV).METHODS: Sixty seven-day-old mice were randomly divided into treated group and control group with thirty mice in each group. These mice were exposed to 750 50mL/L oxygen for 5 days and then to room air.The treated group had been injected captopril (2.7mL/kg), while control group had been injected 9g/L sodium chloride (2.7mL/kg) by intravitreal for 5 days.The mice were sacrificed at the 17th day after birth and the eyes were enucleated. Adenosine diphosphate-ase(ADPase) stained retina flat-mounts was performed to assess the retinal vascular profiles, Hematoxylin Eosin (HE)staining method was applied to count the number of new vascular cell nuclei and the expression of matrix metalloproteinase-2(MMP-2)and pigment epithelium derived factor (PEDF)was detected by immunohistochemical method.RESULTS: Comparing with control group,regular distributions and good branch and reduced density of RNV were observed in the treated group. The number of nucleus of new vessels vascular endothelial cells breaking through the internal limiting membrane was less in the treated group than in the control group (P<0.05). Stain of retinal MMP-2 was weaker in the treated group than in the control group and stain of retinal PEDF was stronger in the treated group than in the control group.CONCLUSION: Intravitreal injection of captopril (2.7mL/kg) may block the RNV in the oxygen-induced mouse model and may provide an effective method for prevent-ing RNV.

Sperm DNA fragmentation index (DFI) refers to the percentage of DNA strand breaks in the total sperm. Many studies suggest that elevated DFI can lead to male infertility and early spontaneous abortion. High-DFI patients are more likely to fail in assisted reproduction and preliminary treatment or prevention methods have been developed for this population. This review focuses on the impact of DFI on clinical pregnancy outcomes and progress in the studies of its treatment.
Abortion, Spontaneous ; Chromatin ; DNA Fragmentation ; Female ; Humans ; Infertility, Male ; Male ; Pregnancy ; Pregnancy Outcome ; Sperm Injections, Intracytoplasmic ; Spermatozoa

Objective To explore the efficacy of GM6001,tissue inhibitor expression and significance of matrix metalloproteinase?9(MMP?9)in mice model of oxygen?induced retinal neovascularization(RNV)and evaluate the inhibition effect of MMP?9 inhibitor(GM6001)on RNV. Meth?ods Mice were placed in oxygen boxes to establish oxygen?induced RNV animal models. The GM6001 treated or hyperxia control groups received an intravitreal injection of 1μL GM6001(100μmol/L)or PBS at day 11 after birth. The normal control and hyperxia group were not treated. HE staining was used to detect RNV in retinal whole mounts,the mRNA level and protein expression of MMP?9 were measured by RT?PCR,Western blot and immunohistochemistry,respectively. Results RNV in the GM6001 treated group was decreased significantly compared with the hyperxia group and hyperxia control group. Compared with the normal control group,higher protein and mRNA expression of MMP?9 were observed in the hy?perxia group and hyperxia control group. The expression of MMP?9 protein and mRNA were decreased in the GM6001 treated group compared with the hyperxia control group(P<0.05). Conclusion The abnormal expression of MMP?9 was closely correlated with RNV. The development of RNV can be markedly inhibited by MMP?9 inhibitor(GM6001),which,we believe,will provide new molecular targets and therapeutic strategy for retinopathy of prematurity treatment.

Objective To explore the inhibition effect of Cysteine-rich 61 (CCN1; Cyr61) specific siRNA expression vector on RNV in a mouse model of oxygen-induced retinopathy (OIR).Methods One hundred and twenty healthy C57BL/6J mice were chosen and randomly divided into the experimental group and control group,with 60 mice in each group.The experimental group was intravitreously injected with CCN1siRNA recombinant plasmids.The control group was injected with vector plasmids.Adenosine diphosphate-ase stained retina flat-mounts was performed to assess the retinal vascular profiles,retinal section with HE staining was applied to count the number of new vascular cell nuclei and the protein and mRNA expression of CCN1 and vascular endothelial growth factor (VEGF) were detected by immunohistochemistry,Western blot and Real-time RT-PCR.Results Compared with control group,regular distributions,good branches and reduced density of retinal neovascularization were observed in the experimental group.The number of nucleus of vascular endothelial cells breaking through the inner limiting membrane was obviously less in the experimental group than that in the control group (t=8.756,P＜ 0.05).The expression of CCN1 and VEGF were obviously decreased in the experimental group compared with the control group (all P＜0.05).Conclusion The development of RNV of ROP can be markedly inhibited by RNA interference targeting CCN1,and CCNlsiRNA may provide an effective method for preventing vascular proliferative retinopathy.

Objective To evaluate the effects of dexmedetomidine combined with subanesthetic dose of ketamine on the emergence agitation in the patients undergoing thoracotomy.Methods Eighty ASA physical status Ⅱ or Ⅲ patients,aged 55-75 yr,weighing 50-75 kg,scheduled for elective esophageal cancer resection,were randomly divided into 4 groups (n =20 each) using a random number table:normal saline group (NS group),dexmedetomidine group (group D),subanesthetic dose of ketamine group (group K),and dexmedetomidine combined with ketamine group (group DK).In DK and K groups,ketamine 0.5 mg/kg was injected intravenously (within 1 min) at 10 min before the end of the operation.In DK and D groups,dexmedetomidine 0.5 μg/kg was infused intravenously over 10 min starting from 10 min before the end of operation.In group NS,the equal volume of normal saline was infused intravenously over 10 min starting from l0 min before the end of operation.The emergence time,extubation time,duration of ICU stay,occurrence and degree of agitation,and development of cardiovascular events and hypoxemia within 24 h after operation were recorded.Ramsay sedation scores were recorded before induction of anesthesia (T1),immediately after completion of administration at the end of surgery (T2),and at 0,5,10 and 30 min after extubation (T3-6).Results There was no significant difference in the emergence time,extubation time,and duration of ICU stay between the four groups.Compared with group NS,Ramsay sedation scores were significantly increased at T3-6,the incidence and degree of agitation were decreased,and the incidence of cardiovascular events and hyoxemia was decreased in D,K and DK groups.Compared with D or K group,Ramsay sedation scores were significantly increased at T3-6,the incidence and degree of agitation were decreased,and the incidence of cardiovascular events and hyoxemia was decreased in group DK.Conclusion Dexmedetomidine combined with subanesthetic dose of ketamine can prevent the emergence agitation in the patients undergoing thoracotomy,which provides better efficacy than either alone.

[Abstract ] Objective High-mobility group box-1 (HMGB1) is abundantly released in the epileptogenic brain tissue , but few reports are seen about the effect of HMGB 1 on the expression of P-glycoprotein ( P-gp) in the vascular endothelial cells of the epi-leptogenic tissue .This study is to explore whether HMGB 1 can regulate P-gp expression in the brain microvascular endothelial cells of the mouse in vitro . Methods Immortalized brain microvascular endothelial bEnd .3 cells of the mouse were cultured in vitro and al-located to different concentration groups ( treated with culture medium containing 10 , 100 , 500 , and 1000 ng/mL HMGB1 for 8 hours), treatment duration groups (treated with culture medium containing 100 ng/mL HMGB1 for 4, 8, 16, 24, and 32 hours), and a control group ( treated with culture medium without HMGB 1 ) .The mRNA expression of P-gp-encoding gene-multidrug resistance gene 1a (mdr1a) was detected by real-time qPCR, and its protein expression determined by Western blot and immunocytochemistry . Results The results of qPCR manifested that the expressions of mdr 1a mRNA were 1.646 ±0.176, 1.777 ±0.135, 1.617 ±0.043, and 1.398 ±0.182 in the 10, 100, 500, and 1000 ng/mL HMGB1 groups, respectively, significantly higher than 1.030 ±0.284 in the control group (P<0.05), and so were those in the 4, 8, 16, 24 h, and 32 h groups (2.655 ±0.112, 2.168 ±0.212, 1.823 ± 0.232, 1.418 ±0.376, and 1.445 ±0.123) than in the control (1.010 ±0.164) (P <0.05).Western blot showed a significant increase in the P-gp protein expression in all the concentration groups (P<0.05) as well as in the 8 h and 16 h treatment duration groups as compared with the control group (P<0.05).Immunocytochemis-try also revealed a higher P-gp expression in the HMGB1-treated than in the control cells (P<0.01). Conclusion HMGB1 can upregu-late the expressions of mdr1a mRNA and P-gp protein in the brain microvascular endothelial cells of the mouse , which may associated with drug resistance of central nervous system diseases , especially that of epilepsy .

Objective: To investigate the relationship between Bilirubin blood lipid comprehensive index and ifbrinogen (FIB) to severity of coronary lesions in patients with coronary artery disease (CAD).
Methods: A total of 324 patients with angiography (CAG) conifrmed diagnosis were divided into 2 sets of groups.①By CAG examination, the patients were divided into 2 groups: CAD group,n=262 and Non-CAD group,n=62.②By Gensini scoring system, the patients were divided into 4 quartile groups: 1st quartile group,n=58, 2nd quartile group,n=110, 3rd group, n=80 and 4th quartile group,n=76. The blood levels of total bilirubin (TBIL), total cholesterol (TC), LDL-C, HDL-C, TG and ifbrinogen were measured and bilirubin blood lipid comprehensive index, TC/(HDL-C+TBIL) and LDL-C/(HDL-C+TBIL) were calculated respectively.
Results:①By CAG examination, compared with Non-CAD group, CAD group had increased TC, LDL-C, ratios of TC/(HDL-C+TBIL), LDL-C/(HDL-C+TBIL) and FIB,P<0.05; decreased TBIL and indirect bilirubin (IBIL),P<0.05.②By Gensini scoring system, the level of TBIL, IBIL, ratios of LDL-C/(HDL-C+TBIL), TC/(HDL-C+TBIL), HDL-C and FIB were different among 4 groups,P<0.05.③ Logistic regression analysis indicated that with excluded interactional factors, the bilirubin blood lipid comprehensive index was the independent risk factor for CAD[TC/(HDL-C+TBIL): OR: 1.668, 95% CI:1.065~2.784,P=0.028; LDL-C/(HDL-C+TBIL ): OR: 1.786, 95% CI: 1.021~2.879,P=0.030].④Correlation analysis presented that TC/(HDL-C+TBIL), LDL-C/(HDL-C+TBIL) and FIB were positively related to Gensini scoring system (r=0.423,P<0.01), (r=0.417,P<0.01) and (r= 0.293,P <0.01) respectively.
Conclusion: Bilirubin blood lipid comprehensive index and ifbrinogen were positively related to severity of coronary lesions in CAD patients.

AlM: To explore the expression and significance of cysteine- rich 61 ( CCN1/Cyr61 ) in oxygen - induced retinal neovascularization ( RNV) of mice and study the inhibition effect of CCN1 specific siRNA on RNV.
METHODS:Two hundred healthy C57BL/6J mice were chosen and randomly divided into control group, hyperxia group, hyperxia control group and CCN1 treated group, with 50 mice in each group. The hyperxia control group was treated with vector plasmids by intravitreal injection. The CCN1 treated group received CCN1 siRNA recombinant plasmids by intravitreal injection. Adenosine diphosphate-ase ( ADPase) stained retina flat-mounts was performed to assess the retinal vascular profiles, HE staining was applied to count the number of vascular endothelial cell nuclei breaking through the internal limiting membrane, protein and mRNA level expression of CCN1 were measured by immunohistochemistry, Western blot and RT-PCR.
RESULTS: There were large nonperfusion area and a large number of vascular endothelial cell nuclei breaking through the internal limiting membrane ( 25. 25 ± 1. 26;23. 12 ± 1. 16 ) in the hyperxia group and the hyperxia control group. Regions of nonperfusion and vascular endothelial cell nuclei (8. 47±1. 15) were decreased in the CCN1 treated group compared to the hyperxia group and the hyperxia control group. Compared with the control group, there were high protein and mRNA expression of CCN1 in the hyperxia group and the hyperxia control group. The expression of CCN1 protein and mRNA were decreased in the CCN1 treated group compared with the hyperxia group and hyperxia control group (all P<0. 05).
CONCLUSlON: The abnormal expression of CCN1 has close relation with RNV. The development of RNV can be markedly inhibited by RNA interference targeting CCN1, which, we believe, will provide new molecular targets and a rationale for clinical developing new strategy for ROP therapy.

Background Cysteine-rich 61 (Cyr61)/CCN1 has been reported to stimulate retinal neovascularization (RNV) in retinopathy of prematurity (ROP).However, whether CCN1 small interfering RNA (CCN1 siRNA) can inhibit or cure ROP has not been extensively investigated.Objective This study was to investigate the regulation effect of CCN1 specific siRNA expression vector on retinal endothelial cells.Methods Rhesus choroid-retinal vascular endothelial cells (RF/6A) were cultured under the normoxic (normoxia control group) and hypoxic condition (1％ O2,5％ CO2 with 94％ N2) in vitro, and then lipofectamineTM 2000 (LF2000) vector plasmid with or without CCN1 siRNA was transiently transfected in the hypoxic-cultured cells as the CCN1 siRNA transfected group and hypoxic control group, respectively.Reverse transcription PCR was employed to detect the expression of CCN1 siRNA plasmid 24 hours after transfection.The vatality of the cells was assayed by cell counting kit-8 (CCK-8) 0,24,48,72 and 96 hours after cultured.Twenty-four hours after cultured,the apoptosis of the cells was evaluated by flow cytometry, and the expressions of CCN1 and vascular endothelial growth factor (VEGF) proteins were detected by immunofluorescence technique and Western blot assay.Results The expression band of CCN1 siRNA was detected in the cells 24 hours after transfection of CCN1 siRNA.CCK-8 assay showed that RF/6A cells were significantly increased over time, and the proliferating value (absorbancy) of the cells was significantly reduced in the CCN1 siRNA transfected group compared with in the normoxia control group and hypoxic control group (Fgroup =198.45, P＜0.05;Ftime =39.26, P＜ 0.05).The apoptosis rates of the cells were (68.9± 1.1) ％ , (18.9±1.3)％ and (39.6± 1.8)％ in the CCN1 siRNA transfected group, normoxia control group and hypoxic control group,and the apoptosis rates of the CCN1 siRNA transfected group were evidently higher than those of the normoxia control group and hypoxic control group (t =2.93 ,t=2.56 ,both at P＜0.05).CCN1 and VEGF proteins were weakly expressed in the normoxia control group and strongly expressed in the hypoxic control group,however,their expression intensity was evidently weakened in the CCN1 siRNA transfected group.The related expression levels of CCN1 and VEGF proteins in the CCN1 siRNA transfected group were significantly lower than those in the hypoxic control group (both at P＜0.05).Conclusions RNA interference targeting CCN1 can inhibit proliferation and promote apoptosis of RF/6A cells.CCN1 siRNA can arrest RNV probably by downregulating the expression levels of CCN1 and VEGF in the cells.