Objective To induce embryonic stem cell (ESC) to differentiate into endothel ioid cells using a simple adhesive culture method, and to provide a new cells seed source for vascular tissue engineering or cell therapy. Methods SV129-derived ESC were seeded at 2 × 104/cm2 and maintained undifferentiated on ESC culture medium in the presence of 1 000 U/mL leukaemia inhibitory factor (LIF). Embryoid body (EB) formatted when ESC cultured in suspension in the lack of LIF. At 4 days, EB was transferred to 0.1% gelatin coated dish and cultured with medium supplementary of VEGFto be induced differentiation. The characteristics of differentiated cells were determined by immunohistochemistry staining, flow cytometry (FCM), 1, 1-dioctadecyl-3, 3, 3, 3-tetramethyl indocarbocyanine-labeled acetylated low density l ipoprotein (DiIAc- LDL) takeup test, and TEM detection. Results Differentiated cells were morphologically characterized as endothel ial cells. They could takeup DiI-Ac-LDL, be stained positive by Flk-1 and CD31. The CD31 positive cells reached above 90% when measured by FCM. Furthermore, Weibel-Palade bodies were detected and tight junctions were found when differentiated cells were examined by TEM. Conclusion Using a simple adhesive culture method and by suppl ied with VEGF alone, ESCs can be induced to differentiate into endothel ioid cells. The differentiation method is simple and economic, and can provide seed cells for vascular tissue engineering or cell-therapy.
ObjectiveTo explore the relationship between the oxygen partial pressure of mice hindlimb muscles with normal blood supply or ischemia and expression of HIF-1αprotein, and to provide a theoretical basis for the study of angiogenesis in vitro hypoxia. MethodsMice hind limb ischemia model were established, tissue oxygen tension of gastrocnemius muscle and bone marrow were measured by micro electrode at different time points of ischemia (24 hours, 1 week, 2 weeks, 3 weeks, and unoperated as control). Protein level of hypoxia inducible factor-1αand histological examination were performed on gastrocnemius muscle as well. ResultsThe oxygen tension baselines of gastrocnemius muscle and femoral bone marrow was (47.78±4.37) mm Hg and (21±3.40) mm Hg, respectively. Muscle oxygen tension decreased significantly at all time points after modeling (P < 0.05), and reached lowest level in 1 week of ischemia. The inflammatory reaction was most serious and HIF-1αprotein reached highest level at the same time point. With the extension of ischemic time, the tissue oxygen tension recovered while HIF-1αlevel was down-regulated, however, There was no statistical correlation(r=-0.86, P > 0.05). Oxygen tension in bone marrow didn't show any significant change at all time points. ConclusionsThe expression level of HIF-1αprotein in ischemic tissue can reflect the degree of ischemic limb. The concept that physiological oxygen level differs in different tissue is highlighted, and may provide basis for ex vivo hypoxic research.
ObjectiveTo summarize the feasibility and effectiveness of endovascular treatment for vertebral artery stenosis ectopic originating from the aortic arch. MethodThe clinical data of 5 patients with vertebral artery stenosis ectopic originating from aortic arch admitted to Xuanwu Hospital of Capital Medical University from January 2020 to May 2023 were retrospectively analyzed. ResultsAll 5 patients underwent magnetic resonance angiography and CT angiography before treatment. The vertebral arteries of all patients originated from the aortic arch and were severe stenosis, with a median diameter of 1.3 mm (range, 1.1–1.5 mm) by CT angiography. All 5 patients were successfully treated with endovascular intervention. The operation time was 37–45 min, with a median of 39 min. There were no complications of vertebral artery rupture, embolism or occlusion, cerebral infarction, plaque abscission, hematoma at puncture site, pseudoaneurysm, postoperative hemorrhage, or perioperative pulmonary infection. The patients were followed-up for 13–30 months, with a median follow-up time of 20 months. The blood vessels of 5 patients remained unobstructed and no clinically significant vascular restenosis occurred. ConclusionsBased on the experiences of 5 cases of vertebral artery stenosis ectopic originating from the aortic arch, it is safe and effective to choose endovascular treatment for severe symptomatic vertebral artery stenosis. Especially, it is necessary to evaluate the stenosis degree of vertebral artery using CT angiography before treatment.
Objective To establish chronic hindlimb ischemia model with suture-occluded method in rats， and then compare the effects of chronic hindlimb ischemia model with acute ischemia model. Methods Models of chronic hindlimb ischemia were established by using suture-occluded femoral artery method. The laser Doppler blood flow analysis and angiography were performed on day 7， 14， 28， 42， and 49 after operation， and then the rats were sacrificed after angiography， respectively， the quadriceps and gastrocnemius of contralateral and ipsilateral （surgical side） were gotten， which were tested by HE staining and α-actin immunohistochemistry staining， and then calculate arteriolar density. Results There were no lameness and limb necrosis after operation in chronic hindlimb ischemia models. Laser Doppler analysis found that chronic hindlimb ischemia models were still maintained in ischemia state on day 49 after operation compared with acute ischemic models. The resluts of HE staining showed no acute necrosis and muscle fibrosis in chronic hindlimb ischemia model group. Chronic hindlimb ischemia models after operation did not appear obvious lameness and limb necrosis. The arteriolar density of quadriceps femoris on day 7 after operation in chronic hindlimb ischemia models were less than that in acute hindlimb ischemia models （0.015 2 vs. 0.036 4）. Conclusions Compared with the commonly used acute ischemic models， the duration of arterial limb ischemia in chronic hindlimb ischemia rats， which were established by suture-occluded method， is longer and less likely to be affected by the compensatory mechanisms. So suture-occluded method can provide a new animal hindlimb ischemia model for further study of ischemia angiogenesis mechanism and treatment of severe lower extremity ischemia.
Objective To explore the mechanism of mesenchymal stem cells （MSCs） transplantation for chronic hindlimb ischemia in Lewis rats by using cell tracer technique. Methods MSCs were isolated and cultured by using density gradient centrifugation and adherence method respectively， then labeled by 5-bromo-2-deoxyuridine （BrdU）. Eight chronic hindlimb ischemia models of Lewis rats were prepared by using suture-occluded method and then divided randomly to MSCs transplantation group and control group， each group enrolled 4 rats， accepting MSCs transplantation and saline respectively. Then on 7 days and 14 days after transplantation， clinical observation， determination of blood flow， and angiography were performed on rats of the 2 groups. At the same time points after previous tests， rats of the 2 groups were sacrificed to get quadriceps tissues and gastrocnemius tissues to perform HE staining and BrdU immunohis-tochemistry. Results The 8 rats were all survived on 14 days after transplantation， with no tumor happened and necroses in the transplanted area. On 14 days after transplantation， the blood flow ratio of operated side to un-operated side in the hindlimb （1.773 vs. 1.279） of rats in MSCs transplantation group and control group increased， and the angiography results showed that there were no much increase in ratio of collateral vessels number （0.908 vs. 0.835）. There were no significant change in the quadriceps tissues and gastrocnemius tissues by HE staining. The BrdU positive kernels located in the inter-stitial substance cells and vascular endothelia cells， and divided differently in different parts of hindlimb at different time points， that the ratio of positive cells in gastrocnemius tissue was higher than those of quadriceps tissue on 7 days after transplantation， but lower on 14 days. Conclusions MSCs transplantation can increases the blood perfusion of hindlimb in the early stage of chronic hindlimb ischemia model， and the possible mechanism may be the paracrine effect of MSCs but not the number increase of collateral vessels.
ObjectiveTo evaluate the dynamic changes of blood flow and blood pressure of acute hindlimb ischemia of rats by laser Doppler flowmetry (LDF) and laser Doppler perfusion imaging (LDPI). MethodsThe acute hindlimb ischemia model of rats was established by resection of rats femoral arteries of left hindlimb. The blood flow and blood pressure between operated and nonoperated hindlimbs were examined by LDF on 2, 7, 14, 28, and 49 d after operation. And the blood flow was evaluated by LDPI on 7 d after operation. ResultsAll rats survived after operation and no hindlimb necrosis occurred. The mean score was 2 on 14 d after operation and 1 on 49 d after operation. The ratio of blood flow between operated and nonoperated hindlimbs on 2 d after operation significantly increased from 1 to 1.31±0.439 (P=0.021). The ratio of blood flow on 7 d (0.82±0.538) and 14 d (0.93±0.294) after operation was significantly lower than that on 2 d after operation (P=0.032 and P=0.019), although the difference between the two former was not significant (P=0.502). Furthermore, the ratio of blood flow on 28 d after operation reached the bottom (0.41±1.970), which was obviously lower than that on 2, 7, and 14 d after operation (P=0.004, P=0.007, and P=0.006). The blood flow of operated hindlimbs recovered approximately the value before operation (0.98±0.093), which was significantly lower than that on 2 d (P=0.010), higher than that on 28 d (P=0.005), but not different from that on 7 d and 14 d after operation (P=0.126 and P=0.382). The ratio of blood pressure between operated and nonoperated hindlimbs on 2 d after operation significantly increased from 1 to 0.47±0.375 (P=0.031). The ratio of blood pressure decreased on 7 d after operation (0.44±0.118), which was not different from that on 2 d after operation (P=0.203). Furthermore, the ratio of blood pressure on 14 d after operation reached the bottom (0.35±0.115), which was obviously lower than that on 2 d and 7 d after operation (P=0.001 and P=0.036). On 28 d after operation, the ratio of blood pressure increased (0.54±0.146), which was significantly higher than that on 14 d after operation (P=0.008), while not different from that on 2 d (P=0.493) and 7 d after operation (P=0.551). The ratio of blood pressure recovered approximately the value before operation (0.97±0.094), which was significantly higher than that on 2, 7, 14, and 28 d (P=0.013, P=0.021, P=0.002, and P=0.031). ConclusionAcute hindlimb ischemia model of rats can be established by resection of rats femoral arteries of left hindlimb and the most serious stage of hindlimb ischemia is on 14-28 d after operation. LDF and LDPI are of importance for monitoring the dynamic changes of rats hindlimb ischemia after operation.
目的 总结锁骨下动脉-颈内动脉人工血管通路辅助颈内动脉支架置入术治疗复杂颈总动脉闭塞的 可行性和有效性。 方法 在同侧锁骨下动脉-颈内动脉人工血管搭桥建立腔内治疗的通路后，行颈内动脉支架置 入治疗复杂颈总动脉闭塞，总结其可行性。 结果 该例患者的手术时间为125 min，术中出血量为10 mL。行锁骨 下动脉-颈内动脉人工血管通路辅助颈内动脉支架置入术后，颈内动脉血流恢复通畅。术后没有过度灌注综合征 及其他并发症发生。术后6 d 患者康复出院，出院后随访7 个月，未再发生头晕及头痛。 结论 锁骨下动脉- 颈内 动脉人工血管通路辅助颈内动脉支架置入术适用于治疗复杂颈总动脉闭塞患者。