Objective To investigate the effect of recombinant human erythropoietin (rhEPO) on the expression of glutamine synthetase (GS) of cultured rat retinal Müller cells in high glucose environment in vitro. Methods Müller cells were isolated from retinas of 10 Sprague-Dawley rats at postnatal day three to five by trypsin digestion, and were randomly divided into six groups, including normal control group, high glucose group, high glucose +5 U/ml rhEPO group, high glucose+10 U/ml rhEPO group, high glucose +20 U/ml rhEPO group, high glucose+40 U/ml rhEPO groups. After 48 hours, the apoptosis of retinal Müller cells were assayed by terminal transferase-mediated DNA end labelling assay, and the expression levels of GS protein were detected with semi-quantitative immunocytochemistry. Results Compared with the normal control group, the cell viability and GS protein were reduced while the cell death increased in Müller cells cultured in high glucose, the difference was statistically significant (t=27.4,P＜0.01). Compared with the high glucose group, rhEPO treatment reduced the apoptotic Müller cells (t=857.2, 2 374.6, 2 473.2, 2 537.7;P＜0.01), induced the expression of GS proteins (t=3.2, 18.0, 22.5, 26.4; P＜0.05). Conclusions rhEPO can protect Müller cells from apoptosis under high glucose condition. The mechanism may be related to its function to up-regulate the GS protein expression, promote glutamic acid cycle, and reduce the excitotoxicity effects of high concentration of glutamate.
目的 总结反复黏膜感染的IgA肾病的临床病理特点。 方法 采用单中心流行病学调查及回顾性研究，收集2006年1月－2009年12月253例经肾活检确诊为IgA肾病的住院患者的临床病理资料，对114例反复黏膜感染的IgA肾病患者（A组）及139例偶发或从未发生黏膜感染的IgA肾病患者（B组）的临床病理指标进行比较。 结果 A组患者年龄较B组小（t=2.913，P=0.004），临床表现无明显症状者比例较B组多，临床分型以反复发作肉眼血尿型为多，病理分级较B组轻（Z=?2.042，P=0.041），IgA+IgM+C3沉积率高（P＜0.001）；B组IgA+IgG+C3沉积率高（P＜0.001），纤维连接蛋白沉积率高（P＜0.001）。 结论 反复黏膜感染的IgA肾病组患者年龄小，反复发作肉眼血尿型多，临床表现及病理分级较轻；两组FN沉积及免疫球蛋白沉积的主要类型不同，提示两组发病机制可能有所不同。
目的 探讨姑息性右室-肺动脉连接术在重症紫绀型先天性心脏病治疗中的临床应用。 方法 回顾性分析郑州市第七人民医院心脏外科 2011 年 1 月至 2015 年 1 月期间所有行姑息性右室-肺动脉连接术治疗的重症紫绀型先天性心脏病患者 25 例的临床资料，其中男 17 例、女 8 例，年龄 31（5～108）个月，体重 3.5～37.2（12.82±6.73）kg。 结果 25 例姑息性右室-肺动脉连接术后早期死亡 2 例（术后 30 d 内），早期死亡率 8.0%（2/25）。患者术后动脉血氧饱和度与术前差异有统计学意义（62.43%±7.83%vs. 81.62%±6.25%，P<0.05）。术后随访 6 个月至 3 年（每 3 个月复查一次超声心动图），23 例患者 McGoon 比值（1.05±0.14vs. 1.61±0.18，P<0.05）和 Nakata 指数[（112.37±14.38）mm2/m2 vs. （165.74±22.62） mm2/m2，P<0.05]均明显上升，且差异有统计学意义。17 例患者行二期根治手术治疗。 结论 姑息性右室-肺动脉连接术能够有效促进重症紫绀型先天性心脏病患者的自身肺血管床发育，为行二期根治术创造条件。
ObjectiveTo study the effect and mechanism of lipopolysaccharide (LPS) on osteoclasts formation and its bone resorption function.MethodsBone marrow-derived macrophages (BMMs) were extracted from the marrow of femur and tibia of 4-week-old male C57BL/6 mice. Flow cytometry was used to detect BMMs. The effect of different concentrations of LPS (0, 100, 200, 500, 1 000, 2 000 ng/mL) on BMMs activity was examined by cell counting kit 8 (CCK-8) activity test. In order to investigate the effect of LPS on osteoclastogenesis, BMMs were divided into macrophage colony-stimulating factor (M-CSF) group, M-CSF+receptor activator of nuclear factor κB ligand (RANKL) group, M-CSF+RANKL+50 ng/mL LPS group, M-CSF+RANKL+100 ng/mL LPS group. After the completion of culture, tartrate resistant acid phosphatase (TRAP) staining was used to observe the formation of osteoclasts. In order to investigate the effect of LPS on the expression of Connexin43, BMMs were divided into the control group (M-CSF+RANKL) and the LPS group (M-CSF+RANKL+100 ng/mL LPS); and the control group (M-CSF+RANKL), 50 ng/mL LPS group (M-CSF+RANKL+50 ng/mL LPS), and 100 ng/mL LPS group (M-CSF+RANKL+100 ng/mL LPS). The expressions of Connexin43 mRNA and protein were detected by Western blot and real-time fluorescent quantitative PCR, respectively. In order to investigate the effect of LPS on osteoclast bone resorption, BMMs were divided into M-CSF group, M-CSF+RANKL group, M-CSF+RANKL+50 ng/mL LPS group, and M-CSF+RANKL+100 ng/mL LPS group. Bone absorption test was used to detect the ratio of bone resorption area.ResultsThe flow cytometry test confirmed that the cultured cells were BMMs, and CCK-8 activity test proved that the 100 ng/mL LPS could promote the proliferation of BMMs, showing significant differences when compared with the 0, 200, 500, 1 000, and 2 000 ng/mL LPS (P<0.05). TRAP staining showed no osteoclast formation in M-CSF group. Compared with M-CSF+RANKL group, the osteoclasts in M-CSF+RANKL+50 ng/mL LPS group and M-CSF+RANKL+100 ng/mL LPS group were larger with more nuclei, while the osteoclasts in M-CSF+RANKL+100 ng/mL LPS group were more obvious, and the differences in the ratio of osteoclast area between groups were statistically significant (P<0.05). Western blot result showed that the relative expression of Connexin43 protein in LPS group was significantly higher than that in control group (P<0.05). Real-time fluorescent quantitative PCR showed that the relative expression of Connexin43 mRNA in control group, 50 ng/mL LPS group, and 100 ng/mL LPS group increased gradually, and the differences between groups were statistically significant (P<0.05). Bone resorption test showed that osteoclast bone resorption did not form in M-CSF group, but the ratio of bone resorption area increased gradually in M-CSF+RANKL group, M-CSF+RANKL+50 ng/mL LPS group, and M-CSF+RANKL+100 ng/mL LPS group, and the differences between groups were statistically significant (P<0.05).ConclusionLPS at concentration of 100 ng/mL can promote the expression of Connexin43, resulting in increased osteoclastogenesis and enhanced osteoclastic bone resorption.
OBJECTIVE To determine the characteristics and regularity of fibronectin mRNA expression in diabetic ulcers, and to investigate the relationship between the changes of fibronectin mRNA expression and pathogenesis of diabetic ulcer. METHODS Biopsies were removed from the margins of diabetic foot ulcers, included surrounding skin as experimental group, and the biopsies from normal skin of the same patients as control group. The mRNA expression of fibronectin was measured by quantitative RT-PCR technique. RESULTS The mRNA expression of fibronectin could be detected in both normal skin and diabetic foot ulcers, but the level of expression in diabetic ulcers was lower than that of normal skin. CONCLUSION The level of mRNA expression of fibronectin in diabetic ulcers is decreased, which suggest that the down-regulation of transcription may be one of the mechanisms of chronic impaired ulcers.
Objective To study the effect of down-regulation of Claudin-3 mediated by adeno-associated virus (AAV) of shRNA on the cultured retinal ganglion cells (RGCs) in vitro. Methods RGCs isolated from mouse eyes were divided into normal control group, AAV-shScramble group, and AAV-shClaudin-3 group. The RGCs in AAV-shScramble group and AAV-shClaudin3 group were treated with AAV-shScramble and AAV-shClaudin-3 respectively 24 hours after cell seeding. Dynamic live cell fluorescence microscopy was used to observe the transfection efficiency 96 hours after transfection. Immunofluorescent staining of β-tubulin was used to measure the length of RGCs′ axon. 4′, 6-diamidino-2-phenylindole staining was used to observe the nuclei of apoptotic cells. The mRNA level of Claudin-3 and VEGF was measured by real-time polymerase chain reaction. The protein levels of Claudin-3, vascular endothelial growth factor (VEGF), Bcl-2 and Caspase-3 was determined by Western blot. Results The positive transfection rate was more than 50% in both AAV-shScramble group and AAV-shClaudin-3 group. The length of RGCs' axon in AAV-shClaudin-3 group was shorter than that in normal control group and AAV-shScramble group (F=22 363.274,P＜0.05). Down-regulation of Claudin-3 accelerated RGCs' apoptosis with nuclei shrinkage, tapering, and nucleolus formation of apoptotic bodies. The mRNA levels of Claudin-3 and VEGF in AAV-shClaudin-3 group were lower than those in normal control group and AAV-shScramble group (F=257.408, 160.533;P＜0.05). The protein levels of Claudin-3, VEGF and Bcl-2 in AAV-shClaudin-3 group were lower than those in normal control group and AAV-shScramble group (F=129.671, 420.552, 62.669;P＜0.05), while the protein level of Caspase-3 in AAV-shClaudin-3 group was higher than that in normal control group and AAV-shScramble group (F=231.348,P＜0.05). Conclusion Down-regulation of Claudin-3 increases the expression of Caspase-3, reduces the expression of VEGF and Bcl-2, accelerates RGCs' apoptosis and inhibit the RGCs' axon growth.
Objective To evaluate the clinical outcomes of total cavopulmonary connection (TCPC) and bidirectional Glenn shunt for treating complex congenital heart diseases with single functional ventricles. Methods From January 2002 to May 2004, twelve children, who had complex congenital heart diseases with single functional ventricles, underwent TCPC and bidirectional Glenn shunt. Among them, male was 3 and female was 9. Ages were from 4 to 13 years and body weights were from 14 to 34 kilograms. The diseases included mitral atresia 1 case, tricuspid atresia 3 cases, right ectopic heart with transposition of great arteries 3 cases, D-transposition of great arteries 3 cases, and single ventricle 2 cases. Results Eleven children survived and one child died in acute renal failure 19 hours after operation. The hospital mortality was 8.3%. Four children had chyle-thorax postoperatively, and eight children had uneventful recovery. In the follow-up period, one child died 12 months postoperatively for pulmomary arteriovenous fistula, and there were no complications like severe arrhythmia, thrombosis and cerebral problems. Conclusions TCPC and bidirectional Glenn shunt are safe and effective techniques for treating complex congenital heart diseases with single functional ventricles, and the clinical outcomes are satisfactory. The key points for the successful operation are big enough cava-pulmonary anastomosis as well as aggressive perioperative management.
Objective To investigate the effect of hypoxia on the exp ression and function of integrin receptor αvβ3 of bovine retinal vascular endotheliocytes. Methods Bovine retinal vascular endotheliocy tes in the culture dishes coated by vitronectin was put into the normal and hypoxemic condition, respectively. Enzyme linked immunosorbent assay and cell adhesion analysis were used to detect the expression and function of integrin receptor αvβ3 in bovine retinal vascular endotheliocytes, respectively. Results Under the condition of hypoxia, the expression of αvβ3 increased gradually, and reached the peak at the 48th hour. The expression of αvβ3 at the 60th and 72nd hour in hypoxia group was higher than that in the normal group. Bovine retinal vascular endotheliocytes absorbed more Vn of extra-cellular matrixes (ECM) after cultured under hypoxemic condition for 24 hours.Conclusion Hypoxia may up-regulate the expression of αvβ3, which promote the adsorbability of endotheliocytes.(Chin J Ocul Fundus Dis，2004，20：360-363)
OBJECTIVE To investigate the effect of allogeneic decalcified bone graft in the treatment of nonunion in children. METHODS From April 1990 to September 1997, 17 children with nonunion were adopted in this study. Among them, there were 10 boys and 7 girls, the age ranged from 2 to 13 years. The allogeneic decalcified bone graft taken from fresh corpse by aseptic manipulation were used to repair 3 cases of congenital nonunion and 14 cases of acquired nonunion. RESULTS All cases were followed up 2 to 9 years after operation, 9 cases were cured successfully and 7 cases were cured after twice operations. One case of congenital pseudoarthrosis of the tibia was operated twice and there were callus grown half years after the second operation, but reoccurred after one year. Although there were 1.5 cm to 3 cm shortening deformity of extremity including another 2 cases of congenital pseudoarthrosis of the tibia and 5 cases of nonunion caused by chronic osteomyelitis, the function of joint was normal. CONCLUSION Treatment of nonunion in children by allogeneic decalcified bone graft is a valuable technique in clinical practice. It is characterised by high capacity of osteoinduction, low antigenicity, rapid bony union, plentiful source of bone graft and convenient utilization.