组织工程软骨实验生物反应器的设计

目的:设计一套计算机控制的组织工程软骨实验用灌流型生物反应器。方法:通过计算机控制时间及流量,利用蠕动泵、密闭的硅胶管、玻璃管、储液瓶等串联设计一套无菌的灌流型软骨组织工程用生物反应器。结果:生物反应器由控制系统和培养系统组成,运行良好,能置于培养箱中对软骨细胞材料复合物进行动态培养。结论:反应器设计合理。整个生物反应器系统运行可靠。     

Early failure of the tissue engineered porcine heart valve SYNERGRAFT™ in pediatric patients

Objectives: The first tissue engineered decellularized porcine heart valve, Synergraft™ (Cryolife Inc., USA) was introduced in Europe as an alternative to conventional biological valves. This is the first report of the rapid failure of these new grafts in a small series. Materials and methods: In 2001, 2 model 500 and 2 model 700 Synergraft™ valves were implanted in four male children (age 2.5–11 years) in the right ventricular outflow tract as a root. Two patients had a Ross operation and two had a homograft replacement. Results: The cryopreserved Synergraft™ valves appeared macroscopically unremarkable at implantation. Recovery from surgery was uneventful and good valve function was demonstrated postoperatively. Three children died, two suddenly with severely degenerated Synergraft™ valves 6 weeks and 1 year after implantation. The third child died on the 7th day due to Synergraft™ rupture. Subsequently the fourth graft was explanted prophylactically 2 days after implantation. Macroscopically all four grafts showed severe inflammation starting on the outside (day 2 explant) leading to structural failure (day 7 explant) and severe degeneration of the leaflets and wall (6 weeks and 1 year explant). Histology demonstrated severe foreign body type reaction dominated by neutrophil granulocytes and macrophages in the early explants and a lymphocytic reaction at 1 year. In addition significant calcific deposits were demonstrated at all stages. Surprisingly pre-implant samples of the Synergraft™ revealed incomplete decellularization and calcific deposits. No cell repopulation of the porcine matrix occurred. Conclusion: The xenogenic collagen matrix of the Synergraft™ valve elicits a strong inflammatory response in humans which is non-specific early on and is followed by a lymphocyte response. Structural failure or rapid degeneration of the graft occurred within 1 year. Calcific deposits before implantation and incomplete decellularization may indicate manufacturing problems. The porcine Synergraft™ treated heart valves should not be implanted at this stage and has been stopped.     

Tricuspidisation of the aortic valve with creation of a crown-like annulus is able to restore a normal valve function in bicuspid aortic valves

Objective: To evaluate the early results of a new method to repair malfunctioning bicuspid aortic valves by creating a tricuspid valve with a crown-like (i.e. anatomic) annulus. Material and methods: Twelve patients (ages from 10 to 27 years) with chronic regurgitation (and flow-dependent stenosis) of a bicuspid aortic valve underwent repair with the principle of creating a tricuspid valve and a crown-like annulus. The fused leaflets were trimmed and reinserted underneath the existing aortic annulus to create one new native cusp. The third leaflet was fashioned out of a xenopericard patch and was inserted underneath the existing annulus as well to restore the crown-like anatomy of a normal aortic annulus. A tricuspid aortic valve with a morphologically normal annulus was thus created, which resulted in improved coaptation of the leaflets. The repair was immediately assessed by transesophageal echocardiography (TEE) with the heart loaded at 50%. In two patients, a second run helped fine-tune the repair. Median cross-clamping time was 82 min. Follow-up ranged from 3 to 46 months (median 13 months). Results: No significant complication occurred. The function of the aortic valve was excellent with trivial or mild regurgitation in 11 patients and moderate regurgitation in 1 patient. There was no stenosis across the valve. The repair remained stable over time. Remodelling of the left ventricle occurred as expected. Conclusions: Aortic valve repair is feasible in some dysfunctioning bicuspid aortic valves. Tricuspidisation of the valve can result in excellent systolic and diastolic functions. The creation of a crown-like annulus results in improved coaptation of the cusps and could lead to more reliable outcome. Although long-term results are needed, this anatomic correction seems to be a good alternative to valvular replacement in certain sub-groups of patients.     

组织工程心脏瓣叶的体外培养

目的 探讨瓣叶支架种植细胞后,不同体外培养时间瓣叶的生成情况. 方法杂种猪9只,取主动脉,分离内皮细胞、成纤维细胞和平滑肌细胞,培养扩增.将成纤维细胞、平滑肌细胞和内皮细胞按顺序种植在预湿处理的支架材料上,以首次种植成纤维细胞、平滑肌细胞为起点,取7天、14天和28天样本使用戊二醛固定,用扫描电子显微镜(SEM)检测,对培养28天的样本进行组织学检查. 结果 SEM检测显示:随着培养时间的延长,细胞数量增加,细胞间连接出现,并有基质生成;至28天,瓣膜支架上可见大量细胞黏附,细胞融合成片,表面可见基质形成.支架的细胞覆盖率也随时间的延长明显增加.组织学检测见大量细胞黏附材料,细胞已长入材料中部. 结论体外培养的瓣叶存在生长活性,使用组织工程方法有可能生成组织工程心脏瓣膜.     

Surgical treatment of aortic valve endocarditis: a 26-year experience

Objective

We have retrospectively analyzed the results of the operations made for aortic valve endocarditis in a single center in 26 years.

Methods

From June 1985 to January 2011, 174 patients were operated for aortic valve endocarditis. One hundred and thirty-eight (79.3%) patients were male and the mean age was 39.3±14.4 (9-77) years. Twenty-seven (15.5%) patients had prosthetic valve endocarditis. The mean duration of follow-up was 7.3±4.2 years (0.1-18.2) adding up to a total of 1030.8 patient/years.

Results

Two hundred and eighty-two procedures were performed. The most frequently performed procedure was aortic valve replacement with mechanical prosthesis (81.6%). In-hospital mortality occurred in 27 (15.5%) cases. Postoperatively, 25 (14.4%) patients had low cardiac output and 17 (9.8%) heart block. The actuarial survival rates for 10 and 15 years were 74.6±3.7% and 61.1±10.3%, respectively. In-hospital mortality was found to be associated with female gender, emergency operation, postoperative renal failure and low cardiac output. The long term mortality was significantly associated with mitral valve involvement. Male gender was found to be a significant risk factor for recurrence in the follow-up.

Conclusion

Surgery for aortic valve endocarditis has significant mortality. Emergency operation, female gender, postoperative renal failure and low cardiac output are significant risk factors. Risk for recurrence and need for reoperation is low.     

Aortic valve calcification is mediated by a differential response of aortic valve interstitial cells to inflammation

Background

Although calcific aortic stenosis is common, calcification of the other three heart valves is not. The aortic valve interstitial cell (VIC) has been implicated in the pathogenesis of aortic stenosis. Proinflammatory stimulation of aortic VICs induces an osteogenic and inflammatory phenotypic change. We hypothesized that the VICs of the other heart valves do not undergo these changes. Using isolated human VICs from normal aortic, mitral, pulmonary, and tricuspid valves, our purpose was to compare the osteogenic response to proinflammatory stimulation via toll-like receptor 4 (TLR-4).

Materials and methods

Aortic, pulmonic, mitral, and tricuspid (n = 4 for each valve type) VICs were isolated from hearts valves explanted from patients undergoing cardiac transplantation. Cells were cultured and grown to confluence in passage 2–6 before treatment with Lipopolysaccharide (LPS) (100–200 ng/mL) for 24 or 48 h. Cells were characterized by immunofluorescent staining. TLR-4 expression was analyzed (immunoblotting, flow cytometry). Bone morphogenetic protein 2 and intercellular adhesion molecule 1 production were determined (immunoblotting). Monocyte chemoattractant protein 1 levels were determined by enzyme-linked immunosorbent assay. Statistics were by Mann–Whitney U test.

Results

TLR-4 stimulation induced bone morphogenetic protein 2 production only in aortic VICs (P < 0.05). Intra-cellular adhesion molecule 1 production and monocyte chemoattractant protein 1 secretion increased in a similar fashion among TLR-4-stimulated VICs from all four valves.

Conclusions

Proinflammatory stimulation induces an osteogenic phenotype in aortic VICs but not mitral, pulmonic, or tricuspid VICs. We conclude that this differential osteogenic response of aortic VICs contributes to the pathogenesis of calcific aortic stenosis.     

组织工程心脏瓣膜内皮细胞的整合素β1表达

目的体外构建组织工程心脏瓣膜（TEHV）,初步探讨内皮细胞黏附生长的分子机制。方法猪主动脉瓣膜经胰酶-EDTA、表面活性剂、核酸酶处理,去除猪主动脉瓣叶的细胞成分,测定瓣叶脱细胞后的生物力学特性;将扩增的人脐静脉血管内皮细胞（HUVECs）种植在瓣叶上,体外静态构建TEHV,观察内皮细胞的生长状态。消化瓣膜内皮细胞,半定量RT-PCR检测内皮细胞整合素＆mRNA的表达,Western-Blot检测内皮细胞膜上整合素＆蛋白的表达。结果猪主动脉瓣膜中的细胞成分能完全去除,脱细胞瓣叶的生物力学特性同新鲜瓣叶相比无明显变化;种植的HUVECs在瓣叶表面生长状态良好,长成一层连续的细胞层。瓣膜内皮细胞可检测到整合素岛mRNA和蛋白的表达。结论脱细胞猪主动脉瓣膜作为支架,HUVECs做种子细胞可以成功构建TEHV,瓣膜内皮细胞可以表达整合素岛。     

Computed tomography detects tissue formation in a stented engineered heart valve

Tissue-engineered heart valves (TEHV) are being explored as an alternative to conventional heart valve prostheses. Using the classic tissue engineering paradigm, a stented tri-leaflet valve is fabricated. Subsequently, the construct is implanted into the pulmonary position in a sheep. Follow-up by means of computed tomography, magnetic resonance imaging, and echocardiography was used to assess tissue formation. After 4 weeks, the scaffold of the TEHV has degraded and new tissue is formed. However, small areas without tissue formation were present at macroscopic inspection. This phenomenon was only visible on computed tomographic images. Therefore, computed tomography appears a promising technique for in vivo follow-up of tissue formation in tissue-engineered heart valves.     

反应器内构建组织工程血管平滑肌层的实验研究

目的 探讨生物反应器内构建组织工程血管的可行性.方法 于6个月龄犬颈总动脉血管获取平滑肌细胞,经体外培养扩增后接种于聚羟基乙酸(PGA)上,形成细胞-材料复合物,并将其置于生物反应器内培养.实验组模拟成年哺乳动物循环系统的参数予以动态力学刺激培养(搏动频率:75次/分;扩张量＜5%);对照组为静态培养,其余与实验组相同,分别于3与6周后取材检测.结果 生物反应器内培养的血管大体观察具有良好的弹性,管腔圆,色泽光亮;HE染色显示平滑肌纤维成分排列较规则,有层次感;弹力纤维染色显示弹力纤维成分较多而密;免疫组织化学检测显示为棕黄色层状排列的平滑肌纤维.对照组弹性欠佳,管腔塌陷,色泽暗淡;平滑肌纤维与弹力纤维成分较少且排列紊乱,层次感差.结论 应用生物反应器可构建具有良好结构的血管样结构的平滑肌层组织.     

Post hoc patient satisfaction with the choice of valve prosthesis for aortic valve replacement: results of a single-centre survey

Open in a separate window OBJECTIVESOur goal was to examine post hoc patient satisfaction and the decision-making process of choosing a prosthesis for aortic valve replacement (AVR).METHODSWe surveyed 113 patients who were operated on for AVR at 60–70 years of age, including 74 patients with a mechanical valve (MECH) and 39 with a bioprosthesis (BIO). The study focused on quality of life and the decision pathway in relation to prosthesis choice and valve-related complications. Decisional conflict was defined as the post hoc uncertainty perceived by patients regarding their choice of prosthesis.RESULTSThe survey was performed at a median of 5.2 (3.2–8.1) years after the AVR. Patients with a biological valve were older (BIO: 68.4 years [66.2–69.4] vs MECH: 63.9 [61.9–66.7]; P < 0.001). Global post hoc satisfaction with prosthesis choice was high in both groups (MECH: 95.9%; BIO: 100%), and 85.1% (MECH) and 92.3% (BIO) of them would repeat their choice. Conflict about their decision was equal (MECH: 30.3%; BIO: 32.6%) for different reasons: MECH patients experienced more anticoagulation-related inconvenience (25.9% vs 0%), fear of bleeding (31.1% vs 0%) and prosthesis noise (26.2% vs 0%), whereas more BIO patients feared prosthesis failure (39.7% vs 17.4%) or reoperation (43.5% vs 18.1%). Active involvement in the decision (odds ratio 0.37, 95% confidence interval 0.16–0.85; P = 0.029) and adequate information about the prosthesis (odds ratio 0.34, 95% confidence interval 0.14–0.86; P = 0.020) decreased the risk of conflict about the decision.CONCLUSIONSAlthough 30% of the responders showed a decisional conflict related to prosthesis-specific interferences, global patient satisfaction with the prosthesis choice for AVR is excellent. Increasing the patient’s involvement in the prosthesis choice through shared accountability and improved information is recommended to decrease the choice-related uncertainty.     

Outcomes of Liu’s aortic root repair and valve preservation in patients with type A dissection and aortic regurgitation

 Open in a separate windowOBJECTIVESTo evaluate short- and medium-term outcomes following Liu’s aortic root repair and valve preservation in patients with acute type A aortic dissection complicated by moderate-to-severe aortic regurgitation (AR). METHODSFrom October 2011 to July 2018, a total of 324 consecutive patients underwent emergency surgery for acute type A aortic dissection. There were 122 patients (38%) with moderate-to-severe AR, of whom 82 (67%) underwent Liu’s aortic root repair and valve preservation. Aortic computed tomography angiography and echocardiography were performed at discharge, 6 and 12 months postoperatively, and annually thereafter. We focused on assessing the survival and aortic root and valve durability in the 82 patients.RESULTSThe 30-day, 1-year, 3-year and 6-year survival estimates were 94%, 90%, 85% and 81%, respectively. At a median follow-up of 36.5 (interquartile range 24.9–50.9) months, all patients were free from reoperation. No residual false lumens in the aortic root, recurrent aortic root dissections or aortic root pseudoaneurysms were observed during the follow-up period. Only 1 patient (1%) presented with moderate AR at 6 months, which remained asymptomatic with no significant changes over a 3-year period. The remaining patients showed satisfactory valve function with an AR grade of mild (27%) or trace or none (72%). In the competing risk analysis, the incidence of recurrence of AR was 2% at 8 years.CONCLUSIONSLiu’s aortic root repair and valve preservation is a safe and effective operative strategy that achieves favourable short- and medium-term outcomes for acute type A aortic dissection with moderate-to-severe AR.     

尿道种子细胞与生物可降解材料的生物相容性研究

目的 观察生物可降解材料聚β-羟基丁酸酯(PHB)与种子细胞即兔膀胱移行上皮细胞及平滑肌细胞间的生物相容性,探讨其作为种子细胞载体构建组织工程化尿道的可行性.方法 经过传代培养,分别在PHB与上皮细胞共同培养2、7 d,与平滑肌细胞共同培养1、5 d,每组各取6个样本,用噻唑蓝(MTT)比色法检测细胞的增殖情况,并利用扫描电镜检测细胞在材料上的空间生长情况,评估移行上皮细胞和平滑肌细胞与聚β-羟基丁酸酯的生物相容性.结果 平滑肌细胞能够在该材料上正常生长,各组间A值差异无统计学意义(P>0.05);而移行上皮细胞在该材料上只有少量生长,且实验组与对照组间A值差异有统计学意义(P<0.05),而A、B两实验组间A值差异无统计学意义(P>0.05).结论 聚β-羟基丁酸酯与平滑肌细胞具有良好的生物相容性,但其对移行上皮细胞的生长具有一定的抑制作用,生物相容性不甚理想.     

组织工程心脏瓣膜的初步构建

目的探讨用体外培养的犬骨髓基质干细胞（MSC）和由聚4-羟基丁酸酯（P4HB）制作的心脏瓣膜支架构建组织工程心脏瓣膜（TEHv）的可行性。方法取年轻健康犬髂前上嵴的骨髓，分离培养出骨髓基质干细胞，通过苏木素-伊红（HE）染色、Mallory染色、Masson染色及免疫组织化学鉴定细胞，取第3代细胞种植在P4HB构建的心脏瓣膜支架上，培养2周后，用扫描电镜观察。结果MSCs经20～25d的培养后，梭形细胞已达到90％以上融合；经HE染色、Mallory染色、Masson染色及免疫组织化学证实培养出来的细胞为MSCs。扫描电镜下，未种植细胞的瓣膜呈蜂窝状，孔径在89～150btm之间，适宜MSCs的爬行；种植细胞后，瓣膜表面有大量MSCs黏附，且爬行长入蜂窝状孔径内。结论MSCs和以P4HB为材料的瓣膜支架构建TEHV具有可行性。     

Direct transaortic transcather valve-in-valve implantation into a mechanical aortic valve prosthesis during left ventricular assist device implantation: description of a surgical technique

The presence of a mechanical aortic valve prosthesis is considered a relative contraindication for left ventricular assist device implantation (LVAD) due to the occurrence of thromboembolic events. Five patients were operated on for LVAD implantation with status post mechanical aortic valve implantation (n = 3 with status post Bentall procedure). After removal of the leaflets, a transcatheter balloon-expandable valve was placed within the mechanical ring in all patients. Three patients were discharged from hospital with a maximum follow-up of 3.3 years. Transaortic transcatheter valve implantation into a mechanical aortic valve during LVAD implantation is a feasible option. It reduces operative times and might also prevent thromboembolic events.     

生物素化去细胞猪主动脉瓣构建组织工程心脏瓣膜研究

目的 制备生物素化去细胞猪主动脉瓣,通过生物素-亲和素系统提高去细胞瓣膜的细胞黏附效率.方法 将10 mmol/L的生物素和5 g/L亲和素分别依次与去细胞猪主动脉瓣反应,制备表面含亲和素的生物素化去细胞猪主动脉瓣.将1.0×105生物素化和未生物素化的骨髓基质干细胞,分别种植于上述改性前后的去细胞瓣膜,应用Hoechst 33258试验确定细胞的黏附效率,扫描电镜观察细胞在瓣膜上的生长.结果 生物素和亲和素能链接到去细胞猪主动脉瓣,生物素化骨髓基质于细胞在表面含亲和素的生物素化去细胞猪主动脉瓣上的黏附效率(55.73±4.53)%显著高于各对照组分别为(31.20±3.94)%、(30.75±3.15)%和(28.51±4.52)%,且扫描电镜示细胞在去细胞猪主动脉瓣膜表面形成一细胞层,生长良好.结论 去细胞猪主动脉瓣可生物素化,并能以亲和素为中介,与细胞表面生物素的结合,增加细胞的黏附效率,且不影响细胞的生长.     

New Pulsatile Hydrostatic Pressure Bioreactor for Vascular Tissue‐engineered Constructs

Mechanical conditioning represents a potential means to enhance the biochemical and biomechanical properties of tissue‐engineered cell constructs. Bioreactors that can simulate physiologic conditions can play an important role in the preparation of tissue‐engineered constructs. Although various forms of bioreactor systems are currently available, these have certain limitations, particularly when these are used for the creation of vascular constructs. The aim of the present report is to describe and validate a novel pressure bioreactor system for the creation of vascular tissue. Here, we present and discuss the design concepts, criteria, as well as the development of a novel pressure bioreactor. The system is compact and easily housed in an incubator to maintain sterility of the construct. Moreover, the proposed bioreactor, in addition to mimicking in vivo pressure conditions, is flexible, allowing different types of constructs to be exposed to various physiologic pressure conditions. The core bioreactor elements can be easily sterilized and have good ergonomic assembly characteristics. This system is a fundamental tool, which may enable us to make further advances in bioreactor technology and tissue engineering. The novel system allows for the application of pressure that may facilitate the growth and development of constructs needed to produce a tissue‐engineered vascular graft.     

骨髓间充质干细胞和内皮祖细胞作为组织工程瓣种子细胞的对比

目的 诱导分化骨髓间充质干细胞(BMSC)及内皮祖细胞(EPC),进行BMSC及EPC作为组织工程瓣膜(TEHV)种子细胞的对比.方法 分离扩增BMSC及EPC,分别定向诱导分化为内皮细胞,比较两种细胞在形态学、增殖能力、黏附力以及细胞冻存和复苏率方面的特点与区别.结果 光镜下,两种细胞均为贴壁生长细胞,BMSC的形态为梭形或多角型,EPC的形态为圆形及不规则形状,第2代诱导的BMSCs、EPCs的倍增时间分别为34 h和35 h,诱导后的BMSC冻存和复苏率以及黏附力与EPC之间差异无统计学意义;电镜示两种细胞均能够种植在去细胞猪主动脉瓣上.免疫组织化学结果显示两种细胞种植后的瓣膜上均有间质细胞生长.结论 BMSC和EPC诱导后可分化为内皮细胞,两者之间差异无统计学意义,都是合适的TEHV种子细胞.     

主动脉成形术治疗主动脉瓣病变伴升主动脉扩张的疗效分析

目的总结主动脉成形术治疗主动脉瓣病变伴升主动脉扩张患者的临床经验及疗效。方法1998年2月至2004年5月共对23例主动脉瓣病变伴升主动脉扩张的患者行主动脉瓣膜置换和纵行切除部分升主动脉壁的主动脉成形术,术后随访4～78个月,平均(36±25)个月,分别于术前、术后出院前及术后随访中,通过心脏超声检查测量主动脉直径。结果主动脉直径术前为(4.8±0.5)cm,术后出院前为(3.6±0.4)cm,两者比较差异有统计学意义(P<0.01)。术后随访中,主动脉直径为(3.7±0.4)cm,与术前比较差异有统计学意义(P<0.01),与术后出院前比较差异无统计学意义(P>0.05)。结论应用切除部分升主动脉壁的主动脉成形术治疗主动脉瓣病变伴升主动脉扩张或升主动脉瘤,中期疗效良好。     

A difficult case of retrieval of an aortic valve and balloon during a transcatheter aortic valve implantation

INTRODUCTIONTranscatheter aortic valve implantation (TAVI) represents an emerging therapy for valve replacement in patients not suitable for traditional open repair. As awareness of the procedure grows, case numbers are increasing worldwide. Though this procedure represents a less invasive approach to aortic valve replacement, it is not without complications.PRESENTATION OF CASEThis case presentation describes a serious, previously unreported, complication incurred in an 83-year-old male in whom TAVI was attempted. During deployment of the valve at the aortic annulus, both the valve and accompanying balloon embolised into the thoracic aorta and this was further complicated by migration of the balloon into the abdominal aorta and an aortic dissection. The false lumen of the dissection at the level of the infrarenal aorta was tacked to the aortic adventitial wall using interrupted sutures through a laparotomy. A completion angiogram demonstrated that a flow limited dissection did extend up to both common iliac arteries. This was managed with balloon-expandable covered stents deployed in both common iliac arteries with satisfactory outcome.DISCUSSIONThis case occurred as a combination of multiple factors that include lack of burst pacing and poor timing of the balloon inflation. The aortic balloon and the valve had to be removed urgently to avoid ventricular embolization of these structures that can result in a fatal situation.CONCLUSIONThis case presentation describes the management of these complications using a combined open and endovascular approach in a well-equipped hybrid operating theatre, resulting in the patient survival.     

去细胞组织工程同种心脏瓣膜的生物力学研究

目的　观察去细胞组织工程同种心脏瓣膜的生物力学特性。方法　取液氮保存的人同种主动脉带瓣管道 ,采用低渗液 去污剂 (1%去氧胆酸 ) 核酸酶法去除了同种心脏瓣膜组织中所有的细胞成分 ,组织基因组DNA含量下降 91% ,保留了完整的细胞外基质。测定去细胞前后组织厚度、组织含水量、热皱缩温度、应力应变曲线、破坏强度及组织伸长比。结果　与去细胞前相比 ,只有管壁组织的含水量去细胞前与去细胞后比较明显增加 (P <0 .0 1) ,瓣叶组织含水量、组织厚度、热皱缩温度、应力应变曲线、破坏强度及组织伸长比差异无显著性 (P >0 .0 5 )。结论　去细胞组织工程同种心脏瓣膜的生物力学特性稳定 ,符合机体的要求 ,且为受体细胞化组织工程心脏瓣膜的研制提供了可靠的天然的纤维支架材料。