3D Printing of Artificial Blood Vessel - MDPI

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Jul 31, 2017 - ... rotation speed remain unchanged, the displacement relation under the different nozzle moving speed can be calculated as. S1 > S1 > S1.
micromachines Article

3D Printing of Artificial Blood Vessel: Study on Multi-Parameter Optimization Design for Vascular Molding Effect in Alginate and Gelatin Huanbao Liu 1 , Huixing Zhou 1,2, *, Haiming Lan 1 , Tianyu Liu 1 , Xiaolong Liu 1 and Hejie Yu 1 1

2

*

College of Engineering, China Agricultural University, Beijing 100083, China; [email protected] (H.L.); [email protected] (H.L.); [email protected] (T.L.); [email protected] (X.L.); [email protected] (H.Y.) School of Mechanical-Electronic Vehicular Engineering, Beijing University of Civil Engineering and Architecture, Beijing 100044, China Correspondence: [email protected]; Tel.: +86-010-6273-7742

Received: 30 June 2017; Accepted: 29 July 2017; Published: 31 July 2017

Abstract: 3D printing has emerged as one of the modern tissue engineering techniques that could potentially form scaffolds (with or without cells), which is useful in treating cardiovascular diseases. This technology has attracted extensive attention due to its possibility of curing disease in tissue engineering and organ regeneration. In this paper, we have developed a novel rotary forming device, prepared an alginate–gelatin solution for the fabrication of vessel-like structures, and further proposed a theoretical model to analyze the parameters of motion synchronization. Using this rotary forming device, we firstly establish a theoretical model to analyze the thickness under the different nozzle extrusion speeds, nozzle speeds, and servo motor speeds. Secondly, the experiments with alginate–gelatin solution are carried out to construct the vessel-like structures under all sorts of conditions. The experiment results show that the thickness cannot be adequately predicted by the theoretical model and the thickness can be controlled by changing the parameters. Finally, the optimized parameters of thickness have been adjusted to estimate the real thickness in 3D printing. Keywords: cardiovascular disease; 3D printing; alginate–gelatin; optimized parameters

1. Introduction Cardiovascular disease is a common disease that seriously affects the health of human beings, especially those over 50 years old. Many scientists [1–6] have adopted a lot of measures to treat cardiovascular disease, but more than 50% of survivors cannot completely take care of themselves. The number of deaths from cardiovascular diseases is up to 15 million people each year globally, which top killer of human beings. Therefore, it is significant to develop an effective method to fabricate vessel-like structures to treat cardiovascular disease. 3D printing is a novel technology to be applied to regenerative medicine that has attracted extensive attention due to its potential to cure disease through tissue engineering and organ regeneration [7–12]. In the past few years, many functional tissues and organs were manufactured by 3D printing systems such as inkjet-based printing [13–15], micro-extrusion printing [16], and laser-assisted printing [17–19], which have different forming parameters in different 3D printing system. One important challenge is the material viscosity characteristic which can determine the types of 3D printers. The inkjet-based printing has viscosity a limit ranging from 3.5 to 12 mPa·s and the viscosity limit of laser-assisted printing ranges from 1 to 300 mPa·s, but the viscosity limit of extrusion-based printing ranges from 30 to >6 × 107 mPa·s. The different working mechanisms and parameters of nozzles results in different viscosity limits.

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limit of laser-assisted printing ranges from 1 to 300 mPa·s, but the viscosity limit of extrusion-based 2 of 10 printing ranges from 30 to >6 × 107 mPa·s. The different working mechanisms and parameters of nozzles results in different viscosity limits. Many research research teams teams have have adopted adopted the the above above 3D 3D printing printing strategies strategies to to fabricate fabricate vessel-like vessel-like Many structures with with deposition deposition modeling modeling methods methods [20–22]. [20–22]. Cui, et al. al. [23] [23] have have used used thermal thermal inkjet inkjet structures Cui, X. X. et printing to fabricate the human microvasculature. Then Hoch, E. et al. [24] have shown that inkjetprinting to fabricate the human microvasculature. Then Hoch, E. et al. [24] have shown that inkjet-based based 3D bioprinting be expected of great vessel-like structures.Gaebel, Gaebel,R.R.etetal. al. [25] [25] 3D bioprinting can becan expected to betoofbegreat use use for for vessel-like structures. have adopted adopted laser laser printing printing to to distribute distribute the the endothelial endothelial cells cells to to fabricate fabricate blood blood vessels. vessels. Kucukgul, Kucukgul, have Can et al. [26] have adopted a deposition modeling method to construct vessel-like structures by Can et al. [26] have adopted a deposition modeling method to construct vessel-like structures by extrusion-based3D 3Dbioprinting. bioprinting.InIn those methods, vessel-like structures poor modeling extrusion-based those methods, the the vessel-like structures have have poor modeling effect effect and material accumulation phenomena. However, there is little research into adopting the and material accumulation phenomena. However, there is little research into adopting the rotary rotary forming method to fabricate vessel-like structures. Gao, Q. et al. [27] have adopted the rotary forming method to fabricate vessel-like structures. Gao, Q. et al. [27] have adopted the rotary printing printingtomethod to vessel-like fabricate vessel-like it can that this the method fabricate structures;structures; it can proven thatproven this method can method improvecan the improve cell survival cell survival rate. Meanwhile, Liu, H. et al. [28,29] have adopted the rotary printing method to obtain rate. Meanwhile, Liu, H. et al. [28,29] have adopted the rotary printing method to obtain vessel-like vessel-like and structures andthe analyzed the synchronization among nozzle extrusion, nozzle speed, and structures analyzed synchronization among nozzle extrusion, nozzle speed, and rotating rotating speed on an extrusion-based 3D bioprinter, whichimprove could improve the modeling speed based onbased an extrusion-based 3D bioprinter, which could the modeling effect ineffect this in this forming method. forming method. In this ourour 3D3D printing device for fabricating vessel-like structures, and further In this paper, paper,we wepresent present printing device for fabricating vessel-like structures, and optimize the thickness parameters of nozzle extrusion speed nozzle speed and rotating speed. The further optimize the thickness parameters of nozzle extrusion speed nozzle speed and rotating speed. δ and nozzle moving speed parameter the pressure have been been adjusted adjusted to to estimate estimate The nozzle moving speed parameter δ and the pressureparameter parameter∇ ∇ have the real real thickness thickness in in the the experiment. experiment. the Micromachines 2017, 8, 237

2. 2. System System and and Process Process Figure Figure 11 shows shows the the extrusion-based extrusion-based 3D 3D printing printing system, system, which which is is mainly mainly composed composed of of aa motion motion control system, multinozzle distribution system, rotary forming system, and temperature control control system, multinozzle distribution system, rotary forming system, and temperature control system. system. The The multinozzle multinozzle 3D 3D printing printing system system is is aa solid solid free-form free-form fabrication fabrication system system that that performs performs extrusion-based Two nozzles extrusion-based processes. processes. Two nozzles with with different different biomaterials biomaterials can can work work effectively effectively in in this this 3D 3D printing printing system system which which can can be be controlled controlled by by computer. computer. The The specification specification of of the the 3D 3D printing printing system system is is shown shown in in Table Table1. 1.

Figure 1. 1. The The extrusion-based extrusion-based 3D 3D printing printing system system with with rotary rotary printing printing device. device. Figure

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Table 1. The specification of 3D printing system.

Table 1. The specification of 3D printing system Parameters Value Parameters Value Dimensions (X × Y × Z) 150 × 150 × 150 mm3 Dimensions (X × Y × Z) 150 × 150 × 150 mm3 Position resolution ±5 µm Position resolution ±5 μm◦ C Temperature range 0–60 Temperature 0–60 mm/s °C Print speed range 0.1–50 Printrange speed 0.1–50 mm/s Pressure 0–1 MPa Pressure range 0–1 MPa

In this 3D printing system, 3D physical models (STL) were divided into different regions, In this 3D printing system, 3D physical models (STL) were divided into different regions, which which were sliced to generate the G-code in the computer, and then each region was filled with were sliced to generate the G-code in the computer, and then each region was filled with different different materials. The STL files can be obtained by CAD/CAM software or 3D scanners. materials. The STL files can be obtained by CAD/CAM software or 3D scanners. It is difficult to extrude biomaterials with a solid state or high viscosity. In order to reconstruct the It is difficult to extrude biomaterials with a solid state or high viscosity. In order to reconstruct digital model, the materials should be transformed into a liquid state or to a proper viscosity to ensure the digital model, the materials should be transformed into a liquid state or to a proper viscosity to extrusion of biomaterials in the process of 3D printing. To that end, the biomaterials firstly have the ensure extrusion of biomaterials in the process of 3D printing. To that end, the biomaterials firstly characteristic of thermosensitivity, which can be liquefied in suitable temperatures (35–40 ◦ C). Secondly, have the characteristic of thermosensitivity, which can be liquefied in suitable temperatures (35–40 the temperature system cancontrol controlsystem the material temperature to achieve the conversion between °C). Secondly, control the temperature can control the material temperature to achieve the solid and liquid states. In addition, there is no heating damage for bioactivity in this temperature conversion between solid and liquid states. In addition, there is no heating damage for bioactivity in control system. control system. this temperature 3. Theory 3. Theory Figure 2 shows the schematic diagram of rotary printing device under the different nozzle speed. Figure 2 shows the schematic diagram of rotary printing device under the different nozzle speed. The biomaterials can be squeezed by pressure control system, the rotating rod is controlled by the The biomaterials can be squeezed by pressure control system, the rotating rod is controlled by the motor, and the nozzle speed is controlled by linear motor. In the Figure 2a, V 1 , V 2 , and V 3 denote motor, and the nozzle speed is controlled by linear motor. In the Figure 2a, V1, V2, and V3 denote the the nozzle speed, the extrusion speed, and the motor speed respectively. When the nozzle speed is nozzle speed, the extrusion speed, and the motor speed respectively. When the nozzle speed is greater than V 1 , the biomaterials will demonstrate the dispersion phenomenon, as shown in Figure 2b. greater than V1, the biomaterials will demonstrate the dispersion phenomenon, as shown in Figure When the nozzle speed is less than V 1 , the biomaterials will demonstrate accumulation, as shown 2b. When the nozzle speed is less than V1, the biomaterials will demonstrate accumulation, as shown in Figure 2c. in Figure 2c.

Figure 2. The schematic diagram of rotary printing device under the different nozzle speed: (a) ideal Figure 2. The schematic diagram of rotary printing device under the different nozzle speed: (a) ideal state; state; (b) nozzle moving speed greater than V1; (c) nozzle moving speed less than V1. (b) nozzle moving speed greater than V 1 ; (c) nozzle moving speed less than V 1 .

In order to realize the reconstruction of vessel-like structures and avoid the phenomenon of In order to realize the of vessel-like structures and avoid the phenomenon of material accumulation andreconstruction material dispersion, the theoretical model can be obtained by this rotary material material the theoretical model can be obtained by this rotary printingaccumulation device, whichand is given thatdispersion, [28] printing device, which is given that [28] → → → (1) V2 →V3 =→V1 +→ V3 = V1 + V2 (1) Because equal time is consumed in the building process of vessel-like structures, the relationship among nozzle speed, extrusion speed, and motor speed is calculated as

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Because equal time is consumed in the building S1 Sprocess S of vessel-like structures, the relationship = 2 = 3 (2) among nozzle speed, extrusion speed, and motor speed is V1 V2 V3calculated as

where S1 , S 2 , and

S3

S2 S 1 refer to linear Smotor rotating rod perimeter, and material = displacement, = 3 (2) V1 V2 V3

extrusion displacement. be derived bymotor Equations (1) and (2) rotating rod perimeter, and material where The S1 , Sthickness to linear displacement, 2 , and Scan 3 refer extrusion displacement. 2V1(2) ) 2 − L2 − r  H = α (V(1) The thickness can be derived by Equations 3 L and (3)  H =qα (V3C 2V2 2 ) 2 − L2 −r  H = α (V3 L/2V1 ) − L2 − r  q (3) where H is the thickness, L is H the = αdisplacement (V3 C/2V2 )2of − nozzle, L2 − r α is an invariant constant, and r is the rotating rod radius. The extrusion forming principle under of thenozzle, different moving speedsand is shown Figure 3. where H is the thickness, L is the displacement α isnozzle an invariant constant, r is theinrotating The thickness can be changed under the different control parameters. When the diameter of rotating rod radius. rodThe and motor rotation remain displacement relation under in theFigure different extrusion forming speed principle underunchanged, the differentthe nozzle moving speeds is shown 3. nozzle moving can beunder calculated as The thickness can speed be changed the different control parameters. When the diameter of rotating rod and motor rotation speed remain unchanged, the displacement relation under the different nozzle S1" > S1 ' > S1  moving speed can be calculated as  (4) S1 00 S>2 "S=1 0S> S1S 2'= 2  0 00  (4) S2 S="S>2 S=' > S2S 3 3 3  S3 00 > S3 0 > S3  The goal is to predict displacement changes which have different forming effects under different The goal is to predict displacement changes which have different forming effects under different nozzle moving speeds. In order to obtain the real thickness, the new thickness coefficient can be nozzle moving speeds. In order to obtain the real thickness, the new thickness coefficient can be adjusted to measure the thickness range. adjusted to measure the thickness range.

Figure 3. The relationship between thethe displacement and thickness under thethe different nozzle moving Figure 3. The relationship between displacement and thickness under different nozzle moving speed due to surface wettability. speed due to surface wettability.

4. 4. Experiment Procedure Experiment Procedure 4.1. Materials 4.1. Materials In order to optimize the above mentioned theoretical model, we designed the experiment to In order to optimize the above mentioned theoretical model, we designed the experiment to fabricate the vessel-like structures. The rotating rods with different diameters of 3, 4.26, and 6.9 mm were fabricate the vessel-like structures. The rotating rods with different diameters of 3, 4.26, and 6.9 mm used in this experiment. We selected alginate and gelatin as experiment materials. In the experiment, were used in this experiment. We selected alginate and gelatin as experiment materials. In the alginate and gelatin were premixed, extruded, and post crosslinked after depositing on the rotating rod experiment, alginate and gelatin were premixed, extruded, and post crosslinked after depositing on in the 3D printing system. The alginate and gelatin solution was cross-linked with calcium chloride the rotating rod in the 3D printing system. The alginate and gelatin solution was cross-linked with (5% w/w). The parameters of experiment materials are shown in Table 2. All of the above materials calcium chloride (5% w/w). The parameters of experiment materials are shown in Table 2. All of the were purchased from Xilong Scientific Co., Ltd. (Guangdong, China). above materials were purchased from Xilong Scientific Co., Ltd. (Guangdong, China).

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Table 2. The parameters of experiment materials. Micromachines 2017, 8, 237

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Concentration of Alginate Concentration of Gelatin Diameter (% w/w) (% w/w)of experiment materials (mm) Table 2. The parameters 3%

Concentration of Alginate 3% (% w/w) 3% 3% 3% 3%

4.2. Experimental Procedure

4%

Concentration of Gelatin 6% (% w/w) 8% 4% 6% 8%

3

Diameter 4.26 (mm) 6.9 3 4.26 6.9

Calcium Chloride (% w/w) Calcium Chloride 5% (% w/w) 5%

The novel 3D Procedure printing system has been designed for the preliminary experiment. In this 4.2. Experimental 3D printing system, temperature control system can control the temperature The novel 3Dthe printing system has been designed for the preliminary experiment.ofInbiomaterials this 3D ◦ C to 45 ◦ C. Then, the motion control system can control the nozzle motion according in a range from 2 printing system, the temperature control system can control the temperature of biomaterials in a to G-code biomaterials be extruded bycontrol pressure controlmotion systemaccording ranged from range document. from 2 °C toNext, 45 °C.the Then, the motion can control system can the nozzle document. the biomaterials can be extruded by pressure system ranged from 0.1 toto1G-code MPa. Finally, theNext, biomaterials are extruded and distributed bycontrol the above-mentioned motion 0.1 to 1 MPa. Finally, the biomaterials are extruded and distributed by the above-mentioned motion control system and pressure control system. Based on this, vessel-like structures can be constructed in control system and pressure control system. Based on this, vessel-like structures can be constructed this 3D printing system. in this 3D printing system. In order to verify the theoretical model, we used a rotating rod with different diameters to analyze In order to verify the theoretical model, we used a rotating rod with different diameters to the motion synchronization and the related parameters of molding effect. In this experiment, the rotating analyze the motion synchronization and the related parameters of molding effect. In this experiment, rod was set to 4.25 r/s and the moving speed of nozzle ranged from 0.8 to 8.3 mm/s. The pressure was the rotating rod was set to 4.25 r/s and the moving speed of nozzle ranged from 0.8 to 8.3 mm/s. The set topressure 0.168–0.421 couldMPa, achieve biomaterial extrusion with different extrusion speeds. was MPa, set towhich 0.168–0.421 which could achieve biomaterial extrusion with different extrusion speeds.

5. Results and Discussion 5. Results and Discussion

5.1. Material Viscosity

5.1. Material Viscosity is Viscosity an important factor which plays a major role in fabricating the vessel-like structures. Viscosity an importantbetween factor which playsand a major role inconcentration. fabricating the In vessel-like structures. Figure 4 shows theisrelationship viscosity material Figure 4a, the viscosity Figure 4 shows the relationship between viscosity and material concentration. In Figure 4a, the of alginate solution increases with the increase of alginate concentration. In Figure 4b, we can see that viscosity of alginate solution increases with the increase of alginate concentration. In Figure 4b,viscosity we the 3% alginate with different concentrations of gelatin (4%, 6% and 8% w/w) have different can see thatThe the 3% alginate withincreased different concentrations of gelatin (4, 6, and 8%of w/w) haveconcentration. different characteristics. viscosity value gradually along with the increase gelatin viscosity characteristics. The viscosity value increased gradually along with the increase of gelatin Therefore, the alginate solution with gelatin can improve the material characteristics such as viscosity, concentration. Therefore, the alginate solution with gelatin can improve the material characteristics mechanical properties, and printable. such as viscosity, mechanical properties, and printable.

Figure 4. Dependence of material concentration on viscosity: (a) the relationship between viscosity

Figure 4. Dependence of material concentration on viscosity: (a) the relationship between viscosity and concentration of alginate; (b) the relationship between viscosity and concentration of gelatin and concentration of alginate; (b) the relationship between viscosity and concentration of gelatin under under the concentration of alginate (3% w/w). the concentration of alginate (3% w/w).

In this experiment, the extrusion-based nozzle has been developed to fabricate the vessel-like structures. Comparedthe with the deposition modeling, this method does not the scaffold to In this experiment, extrusion-based nozzle has been developed to build fabricate the vessel-like structures. Compared with the deposition modeling, this method does not build the scaffold to sustain

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the vessel-like structures, which can reduce time consumption in the build process of vessel-like sustain vessel-like structures, which can reduce consumption in the build process vesselstructure. Inthe order to improve the forming effect, thetime adaptive parameters should be setofto avoid the sustain the vessel-like structures, which can reduce time in the build process of avoid vessellike structure. In order to improve the forming effect, theconsumption adaptive parameters should be set to problems of material dispersion and material accumulation, as shown in Figure 5.be Those likethe structure. Inoforder to improve theand forming effect, the adaptive parameters should set toproblems avoid problems material dispersion material accumulation, as shown in Figure 5. Those problems will reduce the forming accuracy and mechanical properties. the will problems material dispersion andmechanical material accumulation, reduceofthe forming accuracy and properties. as shown in Figure 5. Those problems will reduce the forming accuracy and mechanical properties.

Figure 5. The problems of material dispersion and material accumulation.

Figure 5. The problems of material dispersion and material accumulation.

Dependence of the extrusion pressure on molding effect: this experiment, we have adopted a Figure 5. The problems of material dispersion andIn material accumulation. Dependence of the extrusion pressure on initial molding effect: In this have solution with 3% alginate and 8% gelatin for research. When theexperiment, rotating speedwe was 4.25adopted r/s Dependence of thespeed extrusion pressure on molding effect:structures In When this experiment, wespeed have adopted and nozzle moving was 1.62 mm/s, vessel-like exhibited different molding a solution with 3% alginate and 8% gelatin forthe initial research. the rotating was 4.25ar/s effectswith under pressures, as shown invessel-like Figure 6. structures The different external diameters orr/s solution 3% different alginate gelatin for the initial research. When the rotating speed was 4.25 and nozzle moving speed and was8% 1.62 mm/s, exhibited different molding thicknesses can be achieved under the different extrusion pressures. Figure 6a shows the relationship and nozzle moving speed was 1.62 mm/s, the vessel-like structures exhibited different molding effects under different pressures, as shown in Figure 6. The different external diameters or thicknesses between thickness andpressures, extrusion pressure. We in canFigure see that6.theThe thickness is onexternal the increase with theor effects under different shown different diameters can be achieved under the different as extrusion pressures. Figure 6a shows the relationship between increase of pressure. The molding effects are shown in Figure 6 b. When the pressure exceeds the 0.31 thicknesses can be achieved under the different extrusion pressures. Figure 6a shows the relationship thickness and extrusion pressure. We can see that the thickness is on the increase with the increase Mpa, thickness the molding appears as material accumulation, which leads istoon a poor formingwith effect. andeffect extrusion pressure. We can see that the thickness the increase ofbetween pressure. Thewhen molding effects isare shown in Figure 6b.vessel-like When the pressure exceeds the 0.31the Mpa, Conversely, the pressure below the 0.16 MPa, the structures will not be fabricated increase of pressure. The molding effects are shown in Figure 6 b. When the pressure exceeds the 0.31 the molding effect appears as material accumulation, which leads to a poor forming effect. Conversely, by this 3D printing system.

Mpa, the molding effect appears as material accumulation, which leads to a poor forming effect. when the pressure is below the 0.16 MPa, the vessel-like structures will not be fabricated by this 3D Conversely, when the pressure is below the 0.16 MPa, the vessel-like structures will not be fabricated printing system. by this 3D printing system.

Figure 6. The molding relationship and effect: (a) dependence of the extrusion pressure on thickness under different diameters; (b) the relationship between molding effect and diameter.

Figure 7 shows the relationship between the printable area and extrusion pressure under the Figure6.6.The Themolding molding relationship relationship and effect: (a) ofof the extrusion pressure on on thickness Figure (a)dependence dependence the extrusion pressure different diameters. We can see thatand theeffect: relatively stable thickness region can be derived tothickness analyze under different diameters; (b) the relationship between molding effect and diameter. under different diameters; (b) the relationship between molding effect and the forming state of vessel-like structures. Based on this, the parameter of diameter. the theoretical model can

Figure 7 shows the relationship between the printable area and extrusion pressure under the Figure 7 shows the relationship between the printable area and extrusion pressure under the different diameters. We can see that the relatively stable thickness region can be derived to analyze different diameters. We can see that the relatively stable thickness region can be derived to analyze the the forming state of vessel-like structures. Based on this, the parameter of the theoretical model can

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forming state of vessel-like structures. Based on this, the parameter of the theoretical model can be readjusted to estimate the thickness of vessel-like structures. Based onon these analysis be readjusted to estimate the thickness of vessel-like structures. Based these analysisresults, results,we wecan draw conclusion that the model parameter ∇ is∇ derived can a draw a conclusion thatoptimized the optimized model parameter is derived Micromachines 2017, 8, 237

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H = ∇β

H = ∇β (5) (5) be readjusted to estimate the thickness of vessel-like structures. Based on these analysis results, we qa conclusion that the optimized model parameter ∇ is derived can draw

∇ optimized where is the optimized parameter ranged 0.45 to 2.45. α3 C/2V (V3C2 )22V − ris, the where β isβα is(V −2 L) 2 − −Lr, ∇ parameter ranged from from 0.45 to 2.45. 2

where

β

is

2

α (V3C 2V2 ) 2 − L2 − r ,

H = ∇β



(5)

is the optimized parameter ranged from 0.45 to 2.45.

Figure Figure7.7.The Therelationship relationshipbetween betweenthe theprintable printablearea areaand anddiameter diameterunder underthe thedifferent differentextrusion extrusionpressure. pressure.

Dependence of the moving speed of nozzle molding The controllability of materials Figure 7. The relationship between the printable areaon and diametereffect: under the different extrusion pressure. Dependence of the moving speed of nozzle on molding effect: The controllability of materials plays a significant role in the construction of vessel-like structures. In the case of high pressure, the plays a significant role in moving the construction of vessel-like In the case ofofhigh pressure, Dependence of the speed of onprinting moldingstructures. effect: The controllability materials materials are difficult to be controlled in nozzle this 3D system. Meanwhile, the materials are the materials are difficult tothebeconstruction controlledofinvessel-like this 3D printing system. Meanwhile, the materials plays atosignificant role in structures. the case of pressure, the difficult be extruded under low pressure circumstances. To In overcome thehigh above-mentioned aredrawbacks, difficult bepressure extruded low pressure circumstances. To Meanwhile, overcome the above-mentioned materialstothe are difficult to under bethe controlled in this were 3D printing system. the materials are and rotating speed set to 0.224 MPa and 4.25 r/s, respectively. The difficult to be extruded under low pressure circumstances. To overcome the above-mentioned drawbacks, the pressure and the rotating speed were set to 0.224 MPa and 4.25 r/s,speed. respectively. vessel-like structures have the different thicknesses under the different nozzle moving The drawbacks, the pressurehave and the speed were set tounder 0.224 MPa 4.25 r/s, respectively. Thespeed. The vessel-like structures therotating different thicknesses the and different moving relationship between thickness and moving speed of nozzle is shown in Figure nozzle 8. The conclusion is vessel-like structures have the different thicknesses under the different nozzle moving speed. The The relationship between andwith moving speed of of nozzle is shown FigureWhen 8. Thethe conclusion that the thickness is onthickness the decline the increase nozzle movinginspeed. nozzle is relationship between thickness and moving speed of nozzle is shown in Figure 8. The conclusion is that the thickness on the with the increase of structure nozzle moving speed. material When the nozzle moving moving speed isisless thandecline 0.75 mm/s, vessel-like will present accumulation. that the thickness is on the decline with the increase of nozzle moving speed. When the nozzle speed is less than 0.75 vessel-like structurethe will present material accumulation. Conversely, when themm/s, nozzlethe moving speed exceeds certain values, vessel-like structuresConversely, will not moving speed is less than 0.75 mm/s, the vessel-like structure will present material accumulation. when the nozzle moving speed exceeds the certain values, vessel-like structures will not form. form. Conversely, when the nozzle moving speed exceeds the certain values, vessel-like structures will not form.

Figure 8. The molding relationship and effect: (a) dependence of the nozzle moving speed on Figure 8. molding The molding relationship and effect: (a) dependence ofnozzle the nozzle moving speed on Figure 8. The relationship and effect: (a)relationship dependence of the moving speed on thickness thickness under the different diameters; (b) the between molding effect and diameter. thickness under the different diameters; (b) the relationship between molding effect and diameter. under the different diameters; (b) the relationship between molding effect and diameter.

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Parameter optimization optimization design: design: Figure Figure 99 shows shows the the relationship relationship between between the the printable printable area area and and Parameter the different diameters under different nozzle moving speeds. Similarly to the Figure 7, the vessel-like the different diameters under different nozzle moving speeds. Similarly to the Figure 7, the vesselstructures have have a relatively stable thickness region. region. Based onBased this, the of the theoretical like structures a relatively stable thickness on parameters this, the parameters of the model can be readjusted estimate to theestimate thickness vessel-like theoretical model can be to readjusted theofthickness of structures. vessel-like structures.

HH = = δµ δµ

(6) (6)

q δ isoptimized where µµis is the optimized parameter ranged from to 0.447 to 2.44. V3 L 12)V2 1−) 2L−2 − L2 r,−δris, the where α α(V3(L/2V parameter ranged from 0.447 2.44.

Figure Therelationship relationshipbetween betweenthe theprintable printablearea areaand anddiameter diameterunder under the the nozzle nozzle moving Figure 9.9.The movingspeed. speed.

Based Based on on the the optimized optimized analysis analysis results results under under the the different different pressure pressure and and nozzle nozzle moving moving speed, speed, the optimized parameters of the theoretical model can be derived from the rotating forming device. the optimized parameters of the theoretical model can be derived from the rotating forming device. δ ≈ δ∇ ≈ The parameter thethe parameter of nozzle moving speed, namely . It ∇ is. The parameter of ofpressure pressureisissimilar similartoto parameter of nozzle moving speed, namely proven that the rods with diameters have a similar under constant It is proven thatrotating the rotating rodsdifferent with different diameters have arange similar range under viscosity. constant AlthoughAlthough the thickness characteristics of vessel-like structure are are similar between viscosity. the thickness characteristics of vessel-like structure similar betweenthe the different different pressures and a pressures and nozzle nozzle moving moving speeds, speeds, we wewill willintend intendtotocontrol controlthe thenozzle nozzlemoving movingspeed speedunder under constant pressure due to the time delay in the process of material extrusion. a constant pressure due to the time delay in the process of material extrusion. 6. Conclusions Conclusions 6. In conclusion, conclusion, we we have have designed designed aa novel novel 3D 3D printing printing system system using using aa rotary rotary forming device to to In forming device fabricate vessel-like vessel-like structures structures for for theoretical theoretical analysis, analysis, and and its its feasibility feasibility has has been beenshown shownin inthis thisstudy. study. fabricate Compared with with the the deposition modeling method, method, those those vessel-like vessel-like structures structures produced produced by by rotary rotary Compared deposition modeling forming method accuracy, butbut also have faster molding speed. Using this forming methodnot notonly onlyhave havehigher higherforming forming accuracy, also have faster molding speed. Using rotary forming device, wewe have proposed a theoretical synchronization. this rotary forming device, have proposed a theoreticalmodel modelto toanalyze analyze motion motion synchronization. The experiments with withalginate–gelatin alginate–gelatinsolution solution carried to verify the theoretical results. The experiments areare carried outout to verify the theoretical results. The The experiment results show that the thickness cannot be adequately predicted by the theoretical experiment results show that the thickness cannot be adequately predicted by the theoretical model model the thickness be controlled by changing the correlation parameters. We have optimized and theand thickness can becan controlled by changing the correlation parameters. We have optimized the the parameters under different pressures nozzle moving speeds in the process of 3D printing, parameters under different pressures andand nozzle moving speeds in the process of 3D printing, in in which parameter of pressure is similar toparameter the parameter of nozzle moving namely δ andδ which thethe parameter of pressure is similar to the of nozzle moving speed,speed, namely and ∇ . This optimized theoretical model can be used to predict the thickness of the alginate–gelatin ∇ . This optimized theoretical model can be used to predict the thickness of the alginate–gelatin material. Meanwhile, Meanwhile, it it has has been been proven proven that that extrusion-based extrusion-based 3D 3D printing printing with with alginate alginate and and gelatin material. gelatin could afford afford an to treat disease due due to to the in could an opportunity opportunity to treat cardiovascular cardiovascular disease the possibility possibility of of angiogenesis angiogenesis in tissue engineering and organ regeneration. tissue engineering and organ regeneration.

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Acknowledgments: This work is supported by the national high technology research and development program of China (863 program) under Grant 2015AA020305-01. Author Contributions: Huanbao Liu conceived the problem and designed the solution; Huixing Zhou conceived and designed the experiments; Haiming Lan and Tianyu Liu prepared the biological materials; Xiaolong Liu revised and improved the manuscript for the final submission; Huanbao Liu and Hejie Yu analyzed the data. Conflicts of Interest: The authors declare no conflict of interest.

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