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Objective To compare the protective effects of pharmacological batch RC28-E1 and pilot batch RC28-E2 on retinal vascular endothelial cells (RF/6A) under the stimulation of vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF).Methods RF/6A cells were divided into normal control group,VEGF + FGF group and RC28-E1 groups with different concentrations.The optimal concentration of RC28-E1 was determined by cell counting kit-8 (CCK-8) method.Cells were divided into normal control group,VEGF+FGF group,RC28-E1 group,RC28-E2 group,conbercept group and FGF trap group,and cultured with serum-free culture medium,serum-free culture medium containing VEGF+FGF,serum-free culture medium containing VEGF+FGF+RC28-E1,serum-free culture medium containing VEGF+FGF+RC28-E2,and serum-free culture medium containing VEGF+FGF+ conbercept,serum-free medium containing VEGF+FGF+FGF trap,respectively.Cell proliferation rate was measured by CCK-8 method,cell migration ability was detected by Transwell test,and tube formation ability was detected by Matrigel assay.Results The cell proliferation rate of 0.080 mg/ml RC28-E1 group was significantly lower than that of VEGF+FGF group (P＜0.05).The cell proliferation rate of RC28-E1 group,RC28-E2 group and FGF trap group were significantly lower than that of VEGF+FGF group (P＜0.05).The number of migrated cells in RC28-E1 group,RC28-E2 group,conbercept group and FGF trap group were significantly lower than that in VEGF+FGF group (P=0.000).The numbers of meshes formed by retinal vascular endothelial cells in RC28-E1 group,RC28-E2 group,conbercept group and FGF trap group were significantly lower than that in VEGF+FGF group (P =0.003,0.001,0.009,0.018).The number of tube formation in FGF trap group was significantly higher than those in RC28-E1 group,RC28-E2 group,conbercept group and normal control group (P =0.014,0.000,0.008,0.014).Conclusions Under the stimulation of VEGF + FGF,the inhibitory effect of RC28-E on the proliferation of retinal vascular endothelial cells is greater than that of conbercept,and its inhibitory effect on the tube formation is superior to that of FGF trap.There is no significant difference in the effects of different batches of recombinant decoy receptor innovative drugs on retinal vascular endothelial cells.
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Objective To compare the pharmacodynamics between different batches of recombinant decoy receptor innovative drug RC28-E1 and RC28-E2 in retinal angiogenesis and neovascularization,and analyze its mechanism.Methods Sixty postnatal Day 4 (P4) C57BL/6J mice were randomly divided into normal control group,vascular endothelial growth factor (VEGF)+fibroblast growth factor 2 (FGF2) group,VEGF+FGF2+RC28-E1 group,VEGF+FGF2+RC28-E2 group,VEGF+FGF2+conbercept group and VEGF+FGF2+FGF trap group by using a random number table,with 10 mice in each group.The mouse retinal explant culture system was established,and stimulated with the corresponding factors and drugs prepared in the starving culture media.The normal controls were treated with the starving media.Then the retinal explants were stained with Isolectin B4 and imaged.The number of filopodia per vascular length was quantified.In addition,ninety-six P7 C57BL/6J mice were randomly divided into normal control group,oxygen-induced retinopathy (OIR) model control group,OIR + RC28-E1 group,OIR + RC28-E2 group,OIR+conbercept group and OIR+FGF trap group by using a random number table,with 16 mice in each group.The normal controls were raised under normoxia for 10 days,and the rest of the groups were raised under hyperoxia for 5 days,then returned to normoxia for another 5 days.On P17,the retinas were isolated and stained with Isolectin B4.The stained retinas were mountedon the slides and photographed.The relative vessel obliteration and neovascularization in retina were analyzed with computer software.Then the protein levels of VEGF and FGF2 were examined by Western blot in the retinas of each group in the OIR experiment.Finally,in the RF/6A cells stimulated with VEGF and FGF2,the activities of the signaling pathways,including MEK-extracellular regulated protein kinases (Erk),protein kinase C (PKC) and protein kinase B (Akt) pathways,were examined by Western blot.All experimental procedures were evaluated and approved by the Institutional Animal Care and Use Committee of Tianjin Medical University (SYXK 2009-0001),and were in accordance with the ARVO Statement for the Use of Animals in Ophthalmic and Vision Research.Results The results of retinal explant cultures showed that the numbers of filopodia per vascular length in VEGF + FGF2 + RC28-E1,VEGF + FGF2 + RC28-E2,VEGF + FGF2 + conbcrcept,and VEGF+FGF2+FGF trap groups were all significantly less than that in the VEGF+FGF2 group (all at P ＜ 0.001).The filopodia number in retinal vascular front in RC28-E1 group was similar to that in the RC28-E2 group (P =0.15),whereas the filopodia numbers in both groups were significantly decreased as compared to those in VEGF+ FGF2+conbercept group and VEGF+FGF2+FGF trap group (all at P＜0.001).The results from the OIR mouse model showed that the relative vessel obliteration area in OIR model control group was dramatically higher than those in the drug intervention groups (all at P＜0.05).There was no statistical significance in the relative vessel obliteration area between OIR+RC28-E1 group and OIR+RC28-E2 group (P =0.17),while the obliteration areas in both RC28-E-intervened groups were significantly lower than those in the OIR+conbercept group and OIR+FGF trap group (all at P＜0.05).The relative neovascular pixels in the intervention groups were significantly lower than those in the OIR model control group (all at P＜0.001).The neovascular pixels in OIR+RC28-E1 group were significantly lower than those in VEGF+FGF2+conbercept group and VEGF + FGF2 + FGF trap group (both at P ＜ 0.05),but comparable to those in OIR+RC28-E2 group (P =0.39).Western blot result showed that,the protein expression of VEGF and FGF2 in the OIR mouse retinas were significantly upregulated compared to those in the normal ones (both at P＜0.001).The upregulation of both genes were normalized by both RC28-E1 and RC28-E2.In addition,the stimulation of VEGF and FGF2 induced an enhanced activity in MEK-Erk pathway in RF/6A cells,whereas RC28-E1 inhibited the overactivation.Conclusions RC28-E1 and RC28-E2 both can inhibit angiogenesis in the retinal explants isolated from neonatal mice;they also reduce vessel obliteration and mitigate neovascularization in the OIR mouse model.Therefore,the pharmacology batch and pilot test batch of RC28-E have similar efficacies and reliable stability,and are superior in the anti-angiogenic and anti-neovascular efficacy to the currently clinically available drugs conbercept and FGF trap.RC28-E1 may suppress pathological neovascularization through inhibiting the overactivation of MEK-Erk pathway in retinal vascular endothelial cells.
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Objective To compare the protective effects of pharmacological batch RC28.E1 and pilot batch RC28.E2 on retinal vascular endothelial cells ( RF/6A) under the stimulation of vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF). Methods RF/6A cells were divided into normal control group, VEGF + FGF group and RC28.E1 groups with different concentrations. The optimal concentration of RC28.E1 was determined by cell counting kit.8 (CCK.8) method. Cells were divided into normal control group,VEGF+FGF group, RC28.E1 group,RC28.E2 group,conbercept group and FGF trap group,and cultured with serum.free culture medium, serum.free culture medium containing VEGF+FGF,serum.free culture medium containing VEGF+FGF+RC28.E1, serum.free culture medium containing VEGF+FGF+RC28.E2,and serum.free culture medium containing VEGF+FGF+conbercept,serum.free medium containing VEGF+FGF+FGF trap,respectively. Cell proliferation rate was measured by CCK.8 method, cell migration ability was detected by Transwell test, and tube formation ability was detected by Matrigel assay. Results The cell proliferation rate of 0. 080 mg/ml RC28.E1 group was significantly lower than that of VEGF+FGF group (P<0. 05). The cell proliferation rate of RC28.E1 group,RC28.E2 group and FGF trap group were significantly lower than that of VEGF+FGF group (P<0. 05). The number of migrated cells in RC28.E1 group,RC28.E2 group,conbercept group and FGF trap group were significantly lower than that in VEGF+FGF group (P=0. 000). The numbers of meshes formed by retinal vascular endothelial cells in RC28.E1 group,RC28.E2 group, conbercept group and FGF trap group were significantly lower than that in VEGF+FGF group ( P=0. 003,0. 001, 0. 009,0. 018). The number of tube formation in FGF trap group was significantly higher than those in RC28.E1 group,RC28.E2 group, conbercept group and normal control group ( P = 0. 014, 0. 000, 0. 008, 0. 014 ). Conclusions Under the stimulation of VEGF+FGF,the inhibitory effect of RC28.E on the proliferation of retinal vascular endothelial cells is greater than that of conbercept,and its inhibitory effect on the tube formation is superior to that of FGF trap. There is no significant difference in the effects of different batches of recombinant decoy receptor innovative drugs on retinal vascular endothelial cells.
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Objective To analyze the risk factors of post-stroke disability in patients with acute cerebral infarction.Methods Total 1 137 consecutive patients with acute cerebral infarction admitted in Department of Neurology, General Hospital of Jinan Military Region, were prospectively recruited from August 2010 to August 2014.According to Oxfordshire Community Stroke Project(OCSP), 275 patients were classified as total anterior circulation infarction, 377 as partial anterior circulation infarction,305 as posterior circulation infarction and 180 as lacunar infarction.The baseline data including age, gender, National Institute of Health Stroke Scale ( NIHSS) score were recorded.Recovery was assessed by modified Rankin Scale ( mRS) 6 months after stroke by telephone interview ( mRS≤2:good prognosis, 2
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Objective To investigate the correlation between chronic kidney disease (CKD) and long-term outcomes in a large cohort of unselected patients with acute cerebral infarction.Methods Consecutive acute cerebral infarction patients hospitalized in Department of Neurology,General Hospital of Jinan Military Region were prospectively recruited from August 2010 to November 2013.The baseline data including age,sex,the National Institute of Health Stroke Scale (NIHSS) scores,type of Oxfordshire Community Stroke Project (OCSP:total anterior circulation infarct,partial anterior circulation infart,posterior circulation infarct and lacunar infarct),serum creatinine were recorded.Estimated glomerular filtration rate (eGFR) was calculated according to CKD epidemiology collaboration (CKD-EPI) equation.CKD was defined as eGFR ＜ 60 ml · min-1 · 1.73 m-2 body surface area.Patients were divided into eGFR≥60 ml · min-1 · 1.73 m-2 group and eGFR ＜ 60 ml · min-1 · 1.73 m-2 group.Recovery was assessed by modified Rankin Scale (mRS) 180 days after stroke by telephone interview (mRS≤2 reflected good prognosis,and mRS ＞ 2 reflected unfavorable prognosis).Multinominal Logistic regression analysis,Kaplan-Meier curve and log rank test were used.Results Eight hundred and fifty-two patients were enrolled,among them 93 patients were with CKD.Compared to patients without CKD,acute ischemic patients with CKD were older ((70.56 ± 11.86) years vs (63.11 ± 12.15) years,t =5.60,P =0.000),more likely with NIHSS ≥7 (59.14％ (55/93) vs 32.54％ (247/759),x2 =25.61,P =0.000),more likely with hypertension (89.25％ (83/93) vs 77.34％ (587/759),x2 =6.99,P =0.007),more likely with atrial fibrillation (29.03 ％ (27/93) vs 9.5 ％ (72/759),x2 =30.82,P =0.000),more likely with congestive heart failure (13.98％ (13/93) vs 3.03％ (23/759),x2 =24.54,P =0.000),more likely with tumour (6.50％ (6/93) vs 2.24％ (17/759),x2 =5.59,P =0.031).CKD was a independent prognostic factor for long-term poor outcome (OR =2.034,95％ CI 1.194-3.468) and long term mortality (OR =2.657,95％ CI 1.450-4.870).Kaplan-Meier estimate of patients without CKD for cumulative 180 days survival function for all-cause mortality was higher than those with CKD (79.57％ (74/93) vs 93.54％ (710/759),Log rank test:x2 =23.602,P =0.000).Conclusions Acute ischemic stroke patients with CKD are with more comorbidities.CKD is a independent prognostic factor for long-term poor outcomes and long term mortality in patients with acute cerebral infarction.
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Objective To investigate the correlation between anemia and outcome in a large cohort of unselected patients with acute cerebral infarction.Methods Consecutive acute cerebral infarction patients who were hospitalized were prospectively recruited from August 2010 to November 2013.Eight hundred and fifty-eight patients were enrolled,and the baseline data including age,sex,National Institute of Health Stroke Scale(NIHSS) scores,type of Oxfordshire Community Stroke Project(OCSP:total anterior circulation infarct,partial anterior circulation infarct,posterior circulation infarct and lacunar infarct),serum creatinine,initial hemoglobin level,initial hematocrit level,etc,were recorded.Hemoglobin level and hematocrit level during hospitalization were also recorded.Domestic criteria were used to define if the patient had anemia on admission.Recovery was assessed by modified Rankin Scale (mRS) 180 days after stroke by telephone interview (mRS scores ≤ 2 reflected good prognosis,and mRS scores ＞ 2 reflected unfavorable prognosis).The influence on outcome by anemia on admission,initial hemoglobin level,nadir hemoglobin level,nadir hematocrit level was analyzed by multinomial Logistic regression analysis.Results Odds ratio of initial hemoglobin level for poor outcome was 1.013 (95％ CI 1.001-1.024,P =0.027) with each decrease in hemoglobin of 1 g/dl.Initial anemia(OR =2.417,95％ CI 1.202-4.859,P =0.013) was a independent prognostic factor for mortality;odds ratio of nadir hemoglobin level for mortality was 1.016(95％ CI 1.002-1.030,P =0.026) with each decrease in hemoglobin of 1 g/dl;odds ratio of nadir hematocrit level for mortality was 1.047(95％ CI 1.003-1.093,P =0.037) with decrease in hematocrit of one percentage point.Conclusions Initial hemoglobin level was a independent prognostic factor for poor outcome in patients with acute cerebral infarction.Anemia on admission,nadir hemoglobin level,nadir hematocrit level were independent prognostic factors for mortality in patients with acute cerebral infarction.
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Objective To investigate the correlation between prestroke comorbidity and long-term outcomes in patients with acute cerebral infarction.Methods Consecutive acute ischemic stroke patients who were hospitalized were prospectively recruited from August 2010 to November 2012.Six hundred and forty-four patients were enrolled,the baseline data including Charlson Comorbidity Index (CCI),National Institute of Health Stroke Scale (NIHSS) score,type of Oxfordshire Community Stroke Project (OCSP:total anterior circulation infarct,partial anterior circulation infarct,posterior circulation infarct and lacunar infarct) were recorded.And recovery was assessed by modified Rankin Scale (mRS) 90 days after stroke by telephone interview (mRS score ≤ 2 reflected good prognosis,and mRS score ＞ 2 reflected unfavorable prognosis).Because CCI included specific comorbidity,we considered CCI,CCI without specific comorbidity and specific comorbidity as variable respectively.After screening the risk factors affecting prognosis using univariate analysis,the relationship between comorbidity and prognosis was estimated using multinomial logistic regression model.Results CCI was an independent predictor of good prognosis and unfavorable prognosis (OR =3.446,95％ CI 1.662-7.417; P =0.001).Congestive heart failure and diabetes were each independent predictor of good prognosis and unfavorable prognosis also (diabetes:OR =2.584,95％ CI 1.709-3.906,P =0.000; congestive heart failure:OR =6.229,95％ CI 1.705-22.755,P =0.006).Conclusions After acute ischemic stroke,the patients with the higher CCI score,diabetes and congestive heart failure are more likely to achieve unfavorable outcome.CCI,diabetes and congestive heart failure can each be used as a sensitive index to evaluate the 90 d prognosis of patients.Trial registration Clinical Research Center of China (CHiCTR-OCH-14004228)
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Objective To analyse the clinical features and prognosis of multiple sclerosis (MS). Methods Clinical data of 117 patients with MS were analyzed retrospectively. The clinical and imaging characters in types of optico-spinal(OSMS) and conventional MS(CMS) were compared.Results Among the 117 MS patients, 42 cases(35.9%) were OSMS. 75 cases(64.1%) were CMS. About the clinical manifertation, the incedence of extremital weakness(88.1%), anesthesia(85.7%),abnormal sensation(57.1%), vision(76.2%), urination disorder (73.8%) in OSMS patients were significantly higher than those in CMS patients(70.7%,56.0%,20.0%,45.3% and 26.7%)(allP