Quantification And Evaluation Of Mechanical Stimulation In Tissue Engineered Blood Vessel Culture

Background:In recent years,due to the Westernization of life style and the acceleration of the aging process of social population,the incidence of cardiovascular disease has increased gradually,which has seriously threatened human health and social and economic stability.Although interventional surgery is becoming more and more popular,surgical vascular transplantation is still the main way to treat coronary heart disease.Currently,there are autologous saphenous vein or internal mammary artery for coronary artery bypass grafting.Autologous blood vessels are damaged themselves,while the source of autologous blood vessels is very limited.Vascular graft with synthetic material in small artery(diameter less than 6 mm)has a low patency rate,leading to high failure rate of arterioles transplantation.So far,the industry recognized that tissue-engineered blood vessel is the most promising way to solve this problem.The ideal tissue engineering blood vessel(TEBV)construction method can form abundant collagen fibers and elastic fibers to enhance mechanical properties.In addition to sufficient mechanical strength,the mechanical properties of tissue engineered blood vessels should be in line with the physiological needs of the human body,such as anisotropy like primary blood vessels;stress and strain meet the nonlinear relationship;enough sutures retain strength.In vitro construction of tissue engineered blood vessels,mechanical microenvironment has an important influence on the mechanical properties of tissue engineered blood vessels,such as stimulating seed cells to secrete extracellular matrix,collagen fibers and elastic fibers.Therefore,it is necessary to quantify the stress culture environment of tissue engineered blood vessels.There are two main methods used at present.The first one is based on high speed camera(CCD).It is easily affected by vibration,unclear edge,high requirement and low resolution in the process of shooting.It is not able to accurately detect the direct transmission medium silicon tube under pulsating pressure.Jing Kuozhang.The other is based on the simple linear Hooke's law,which can not accurately reflect the stress-strain relationship between the nonlinear elastic body and the viscoelastic body by calculating the strain through the pressure of the cavity.Based on the above background,the design of this study mainly includes two aspects: 1.in view of the difficulty of accurate quantification of the stress environment in TEBV,a mechanical monitoring platform based on the laser diameter measuring system and pressure sensor is set up to obtain the deformation value of the mechanical transfer medium and the lame method of elastic mechanics.The size of the stress is calculated accurately;2.this new mechanical monitoring method is applied to the actual TEBV construction process,and the practicability and feasibility of this method is preliminarily evaluated through the construction of the results.1.Build mechanical monitoring platform based on laser diameter measuring system and pressure sensorMethods: The data acquisition system is built by high-precision pressure sensor and laser diameter measuring system,and its accuracy is as high as 2 microns.The system uses analog voltage output and data acquisition card to collect voltage signals.The laser acquisition position is the middle point of the silicon rubber tube.The high-precision pressure sensor accurately capture the pressure of the fluid on the silica tube wall under the pulsating flow field,and the location is the end of the silicone tube.The inner diameter of the silicon rubber tube is calculated through the external diameter data and the incompressible characteristic of the silicon rubber tube itself,and the strain is accurately calculated from the strain definition.The two driving pressures of the power drive system are measured at 120 mmHg and 180 mm Hg,respectively.Results: Through the construction of the mechanical monitoring platform,the external diameter data and pressure data in the tube can be quickly and quickly obtained.The interference caused by the silicone tube vibration is very small,and the repeatability of the numerical value is good.After obtaining the deformation value by the mechanical monitoring platform,the surface stress distribution of the mechanical transfer medium(silicone tube)can be accurately quantified according to the lame method.Under the pressure pulsation of the120 mmHg peak of the pulsating pump,the silicon rubber tube with a hardness of 55 A,a diameter of 5 mm and a wall thickness of 0.3 mm can reach 2.53% strain and 115.67 KPa stress.Under 180 mm Hg peak pressure pulsation,the same silicone tube can reach 4.90%strain and 165.38 KPa stress.2.Preliminary application of new mechanical detection system in tissue engineeringblood vessel constructionMethods: Two kinds of silicon rubber tubes with the same outer diameter and material hardness but the tube wall thickness of 0.3 mm and 0.4 mm were selected.The pulsating flow of the peak pressure of 120 mmHg and 180 mmHg was formed by pulsating pump respectively.The deformation of the two kinds of silicone tubes with different wall thickness were detected on the mechanical monitoring platform,and the respective stresses were calculated and compared with Hu Keding.The diameter of the silica gel calculated from the law and the measured diameter of the diameter measured by laser can be used to understand their errors.In the same bioreactor,the mechanical environment of two different stress intensities was formed in the same bioreactor with a pulse flow of 180 mmHg peak pressure.Then the tissue engineering blood vessel culture method was established before the research group,and the total culture time was 4 weeks.After culture,the tissue engineered blood vessels were removed and stained.It was observed mainly by HE staining and Masson tricolor staining.Results: Under 180 mmHg peak pressure pulsation,0.4 mm wall thick silicone tube can reach 2.90% strain and 116.54 KPa stress,while 0.3 mm wall thick silicone tube can reach4.9% strain and 165.38 KPa stress.Under 120 mm Hg peak pressure pulsation,0.4 mm wall thick silicone tube can reach 1.93% strain and 80.65 KPa stress,while 0.3 mm wall thick silicone tube can reach 2.53% strain and 115.67 KPa stress.The measurement of the outer diameter of the silicon rubber tube by laser diameter measuring system is compared with the linear Hooke law.Under 120 mmHg pulse pressure,the maximum error of Hooke's law of 0.3mm wall thick silicone tube can reach 34.15%,and the maximum error of 0.4 mm wall thick silicone tube can reach 93.1%.Under 180 mm Hg pulsating pressure,the maximum error of Hooke's law of 0.3 mm wall thick silicone tube can reach 47.06%,and the maximum error of0.4 mm wall thick silicone tube can reach 50%.The tissue engineering vessels of 4 weeks were stained with HE and Masson,and the silicone tube with 0.3 mm wall thickness was more compact than the tissue engineered blood vessel under the stress environment provided by 0.4mm wall thick silicone tube,the cell proliferation was more obvious,the cell migration was uniform,and the extracellular matrix was evenly distributed in the inner and outer layers of the vessel.And the degradation of PGAis more complete.Conclusions:1.The mechanical monitoring platform built by this study can quickly,stably and accurately obtain the key mechanical parameters.The stress of the mechanical transfer medium(silicone tube)can be accurately quantified in the tissue engineering blood vessel culture by the lame method of elastic mechanics,but the calculation of the Hooke law can make a great deviation.2.By accurately quantifying the stress changes of mechanical transfer medium(silicone tube)in the construction of tissue engineering vessels,it can accurately guide the cultivation of tissue engineering vessels in the future,and find out the ideal mechanical microenvironment parameters.3.Tiny changes in the mechanical microenvironment greatly affect the function of vascular smooth muscle cells to secrete extracellular matrix.