I would like to thank my supervisor, Wenhua Huang, for providing support conditions and guidance for this experiment. This research was funded by the Discipline construction project of Guangdong Medical University (4SG22260G), Young Innovative Talents Project of Guangdong Higher Education Institutions (2021KQNCX023), National Natural Science Foundation of China (82205301), and Futian Healthcare Research Project (FTWS2022051).

1. Printing and preparation of a titanium alloy workpiece
<ol>
	<li>Prepare a workpiece made of porous titanium alloy using the SLM printing technique. Import STL format files into the metal printer, add Ti-6Al-4V powder, install the build substrate, set up the wiper blade, set the laser spot size to 70 &#181;m, and set the layer thickness to 30 &#181;m (Figure 1).</li>
	<li>Grade 23 Ti-6Al-4V powder with chemical composition as shown in Table 1 and a powder particle size of 15-53 &#181;m.</li>
	<li>Design the porous titanium alloy structure with simulated trabecular bone based on Tyson polygon anisotropy using parametric modeling, with an aperture size of 400-600 &#181;m, small beam diameter of 100-300 &#181;m, and porosity of 70%31 .</li>
	<li>Ensure the porous titanium alloy workpiece is in the shape of the medical lumbar cage32. For the porous structure and the lumbar cage, use Boolean operations to obtain the porous workpiece structure.</li>
</ol>
2. Heat treatment
<ol>
	<li>A high temperature gradient during SLM printing will cause residual stress in the workpiece. Use heat treatment to eliminate the residual stress inside the workpiece and maintain the toughness, plasticity, tensile strength, and other physical properties of the workpiece.</li>
	<li>Separate the porous titanium alloy workpiece from the printing substrate after printing using a medium speed wire-cutting machine. Install the titanium plate on the medium speed wire-cutting machine, in order to make the plate perpendicular to the ground, and ensure that the wire just contacts the support surface. Then, cut along the support and titanium plate to separate the porous titanium alloy workpiece from the printing substrate.</li>
	<li>Place the porous titanium alloy workpiece into the ultrasonic cleaning machine with deionized water for 15 min and the temperature controlled at 30 &#176;C. Keep the ultrasonic frequency at 40,000 Hz. The ultrasonic cleaning aims to remove the titanium alloy powder remaining in the porous structure.</li>
	<li>Repeat the aforementioned ultrasonic cleaning procedure four times to remove residual titanium alloy powder and deionized water from the porous structure. After that, aim high-pressure air at the porous structure for 20 s to blow away the residual powder and liquid. The pressure of the high-pressure air is 0.71 MPa, which is generated by an air compressor and air dryer.</li>
	<li>Put the titanium basket into the heat treatment furnace at room temperature. The titanium basket is equipped with titanium alloy workpieces separated from the substrate. Keep different workpieces from touching each other and close the furnace door.</li>
	<li>Open the gas valve, take out the air, and keep the vacuum degree at 3.9 x 10-3 Pa.</li>
	<li>Set the heat treatment process. First, heat the furnace to 800 &#176;C for 1.5 h, maintain the temperature for 2 h, and then cool the workpiece inside the furnace. This process ensures that the vacuum pressure remains unchanged.</li>
	<li>After the heat treatment, cool the furnace to room temperature and fill the furnace with air. After returning to atmospheric pressure, as seen on the panel, take out the porous titanium alloy workpiece.</li>
</ol>
3. Removing the support
<ol>
	<li>After heat treatment, the porous titanium alloy workpieces have no internal residual stress, so the workpiece surface will not crack and/or fracture when removing the support.</li>
	<li>Measure the support thickness using a vernier caliper, fix the workpiece on the low-speed wire-cutting electrical discharge machining (EDM) machine, and ensure that the copper wire just contacts the support surface.</li>
	<li>Set the cutting depth equal to the support thickness. It&#39;s inevitable that removing the support by the wire-cutting EDM machine will form an ablation oxide layer. When removing the support, ensure that the workpiece is immersed in deionized water to minimize burns to the workpiece surface.</li>
	<li>A reasonable support design ensures accuracy when removing the support. If there are still some support residues, polish the workpiece with sandpaper.</li>
</ol>
4. Ultrasonic cleaning
<ol>
	<li>Since the workpiece is immersed in deionized water during support removal, perform ultrasonic cleaning before plasma polishing to remove other impurities.</li>
	<li>Put the porous titanium alloy workpiece into the ultrasonic cleaning machine with deionized water, set the water temperature to 30 &#176;C, and clean it for 5 min. After 5 min, take out the workpiece and blow out residual liquid with high-pressure air.</li>
</ol>
5. First characterization
<ol>
	<li>Scanning electron microscope (SEM): Image the surfaces with a SEM at 15 and 20 kV accelerating voltage, after ultrasonic cleaning and before plasma polishing.</li>
	<li>Take images at 30x, 100x, and 500x visual fields. Observe the general surface morphology, particle adhesion, and pore size of the porous titanium alloy workpiece, and qualitatively evaluate the plasma polishing effect.</li>
	<li>Confocal microscope: Image the surfaces using a confocal microscope.</li>
	<li>Place the workpiece on the storage platform horizontally. Measure the surface arithmetic average roughness (Ra) parameter. Use ZEN core v3.0 and ConfoMap ST 8.0 software.
	<ol>
		<li>Select 2.5x magnification, choose Wide for live mode, click Auto intensity, and then go to 5x magnification to observe the overall situation. Click Auto intensity and set the live mode to Comp. Select the area of interest, click Set first at the lowest point and Set last at the highest point, and then set the acquisition to Normal.</li>
		<li>After about 5 min, import the results into a new document in ConfoMap ST 8.0. The Ra is easy to obtain in the parameters table in ConfoMap ST.</li>
	</ol>
	</li>
	<li>Observe the overall condition of the workpiece with a fivefold mirror, then switch to a high-power mirror and focus the field of vision on a trabecula. Evaluate the plasma polishing effect quantitatively by describing the Ra of the porous titanium alloy workpiece before plasma polishing.</li>
</ol>
6.&#160;Plasma polishing
<ol>
	<li>For this, use an electrolytic cell to immerse the workpiece in an electrolyte connected as an anode20. Use 4% ammonium sulfate solution [(NH4)2SO4], pH between 5.7-6.1, as the electrolyte. Preheat the polishing electrolyte to 80 &#176;C before plasma polishing.</li>
	<li>Set the polishing current to 59 A, the voltage to 313 V, and the polishing electrolyte temperature to 101.6 &#176;C (Figure 2A). Conduct plasma polishing according to these parameters.</li>
	<li>Place the surface of the porous titanium alloy workpiece to be polished horizontally and fix it on the fixture, and then put the fixture into the plasma polishing machine (Figure 2B). Conduct plasma polishing for 90 s, and then take the fixture out of the plasma polishing machine.</li>
	<li>Since the porous titanium alloy workpiece is fixed on the fixture through the clamping point, the clamping point is not in contact with the polishing solution, and the corresponding electrochemical reaction does not occur at the clamping point. Therefore, change the position of the clamping point slightly after the fixture is taken out.</li>
	<li>Conduct plasma polishing again for 90 s and take the fixture out of the plasma polishing machine. Remove the porous titanium alloy workpiece from the fixture and then put it into the ultrasonic cleaning machine with deionized water.</li>
	<li>Set the water temperature at 30 &#176;C and clean the workpiece for 2 min. After 2 min, take out the workpiece and blow out the residual liquid with high-pressure air.</li>
</ol>
7. Second characterization
<ol>
	<li>After the completion of plasma polishing, image the surfaces using a SEM and a confocal microscope in the same way as in step 5. Assess the influence of plasma polishing on the surface roughness and surface quality of 3D printing porous titanium alloy by comparing the above two shooting results.</li>
</ol>

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The authors have nothing to disclose.

Surface roughness is used to describe the amount of undulation and unevenness of micro geometric shapes on workpiece surfaces within a small spacing range. A number of previous studies have reported how to polish metal surfaces using different procedures, such as mechanical polishing, chemical polishing, electrochemical polishing, and more22,33,34,35. Although numerous studies have showed prosp...

Titanium and titanium alloy materials have been widely used as dental and orthopedic implant materials because of their good biocompatibility, corrosion resistance, and mechanical strength1,2,3. However, due to the high elastic modulus of the compact titanium alloy produced by traditional processing methods, these plates are not suitable for bone repair, since close proximity to the bone surface for long periods can result in stress shielding and bone embrittlement4,5 . Therefore, the porous microstructure of simulated bone trabeculae should be used in titanium alloy implants in order to reduce its elastic modulus to the level matching the bone6,7. Many scaffolds have been used in the field of orthopedics to improve cell viability, attachment, proliferation and homing, osteogenic differentiation, angiogenesis, host integration, and weight bearing4,8,9. Traditional fabrication methods of porous metal structures include the structural template method, defect formation method, compression or supercritical carbon dioxide method, electro-deposition technique10,11, etc. Although these production techniques are highly traditional, they occasionally squander raw materials and have substantial preparatory costs when compared to 3D printing12,13. 3D printing is a technology that uses metal or plastic powder and other adhesive materials to build solid 3D objects from computer aided design (CAD) models via the deposition of overlying layers14,15 . 3D printing shows great potential in directly customizing metallic cellular scaffolds for orthopedic implants and opens up new possibilities for manufacturing customizable complex designs with highly interconnected pores. Among them, selective laser melting (SLM) is one of the most representative 3D printing and manufacturing technologies for porous titanium implant structures16 .
The SLM process uses titanium alloy powder as the raw material, essentially powder melting and forming the structure. Therefore, a large number of semi-molten powders and ablative oxide layers often adhere to the surface of titanium alloy implants, which leads to high surface roughness17. Poor surface quality of porous titanium orthopedic implants leads to inflammation, decreased fatigue performance, and even new biological risks18 . Since the internal pores of porous structures cannot be polished by conventional mechanical polishing, an alternative method needs to be found. Plasma polishing is a new green polishing method for metal workpieces that can efficiently polish workpieces with complex shapes without pollution19 . It has great development potential in the field of titanium alloy implant post-processing.
As a kind of surface technology, plasma polishing technology is particularly suitable for metal workpieces with complex shapes that are not easy to be mechanically polished. The overall goal of this polishing option is to obtain a porous titanium alloy surface with low roughness. The technology can effectively remove particles and fine splash residues attached to the surface of porous titanium orthopedic implants fabricated by 3D printing and reduce surface roughness20. The principle of plasma polishing is a composite reaction process based on a combination of current-induced chemical and physical removal21; the entire circuit forms a transient short circuit, forming a vapor plasma-surrounding layer on the workpiece surface20. This process breaks through the gas layer to form a discharge channel, impacting the workpiece surface. The higher current impacts the convex part of the workpiece surface, leading to the faster removal of semi-molten powder and the burnt oxide layer. The concavity and convexity are constantly changing, and the rough surface becomes gradually smoothed, improving the surface roughness of the workpiece to achieve the purpose of polishing.
At the same time, this technology is a green processing technology, causing no pollution to the environment, and has great advantages compared with other polishing methods. Conventional mechanical polishing techniques mainly include mechanical polishing, chemical polishing, and electrochemical polishing22. Mechanical polishing is the most widely used conventional polishing process; it has the disadvantages of low polishing efficiency, higher demand for manual labor, and inability to polish parts with complex geometries. The potential for employee injury and the likelihood of exceeding tolerances due to human factors are frequent drawbacks of mechanical polishing23. In contrast to chemical polishing, which is based on utilizing a chemical solution to remove parts of a workpiece&#39;s material, electrochemical polishing utilizes an electric current and chemical solution to obtain the same result. Unfortunately, both these processes produce hazardous gases and liquids as by-products of use, the composition of which being dependent on the strength of the acid or alkaline chemical reagent being used. As a result, not only are the workers present deemed to be at risk due to exposure, but there is also the potential for severe damage to the environment24. Aliakseyeu et al.25 proposed utilizing plasma polishing for polishing titanium alloy workpieces with simple electrolyte composition. They found that, after polishing titanium sample surface scratches are removed and the surface gloss is significantly improved. Smyslova et al.26 deliberated upon the prospects of applying plasma polishing technology to treat the surfaces of medical implants.
Theoretically, plasma polishing technology can be utilized to polish the structure of any metal part. It has been widely applied for coating, in metal finishing industries, and in 3C electronics, among others22,27,28. However, the present study has some limitations. First of all, the manuscript only focuses on the surface quality and surface roughness of 3D printing porous titanium alloy before and after plasma polishing; the remaining changes are not involved. Secondly, we didn&#39;t measure and record the results after heat treatment. Jinyoung Kim et al.29 compared titanium surface modification strategies for osseointegration enhancement. Another study shows that the target-ion induced plasma sputtering (TIPS) technique can impart excellent biological functions to the surface of metallic bio-implants30. In order to further investigate the polishing efficacy and safety of porous titanium alloy for 3D printing, the next step will be to further study SLM part&#39;s other properties, such as fatigue performance and osteogenic differentiation. These issues need further refinement. This work differs from earlier plasma polishing studies in that it focuses on 3D printing porous titanium alloy rather than compact titanium alloy. As a result, different manufacturing processes should adopt different polishing parameters. The purpose of this manuscript is to introduce the plasma polishing scheme of 3D printing porous titanium alloy in detail, so as to reduce the surface roughness of workpieces.

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	<tbody>
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			<td>Confocal microscope: Smartproof-5</td>
			<td>ZEISS</td>
			<td>4702000198</td>
			<td></td>
		</tr>
		<tr>
			<td>ConfoMap ST 8.0</td>
			<td>ZEISS</td>
			<td>4702000198</td>
			<td></td>
		</tr>
		<tr>
			<td>Electrical discharge machining (EDM) machine: MV1200S</td>
			<td>Mitsubishi Electric Automation (China) Ltd.</td>
			<td>92U3038</td>
			<td></td>
		</tr>
		<tr>
			<td>Heat treatment furnace: HSQ1-644</td>
			<td>Jiangsu Huasu Industrial Furnace Manufacturing CO., LTD.</td>
			<td>HSD20190812403</td>
			<td></td>
		</tr>
		<tr>
			<td>Metal 3D printer: Renishaw AM400</td>
			<td>Renishaw plc</td>
			<td>1HGW89</td>
			<td></td>
		</tr>
		<tr>
			<td>Middle speed wire-cut machine: HQ-400EZ</td>
			<td>Suzhou Hanqi CNC Equipment CO., LTD.</td>
			<td>W40ES20005</td>
			<td></td>
		</tr>
		<tr>
			<td>Permanent magnet frequency conversion screw air compressor M7-Y75AZ</td>
			<td>KUNJI MACHINERY(SHANGHAI) MANUFACTURING CO.,LTD.&#160;</td>
			<td>19055065</td>
			<td></td>
		</tr>
		<tr>
			<td>Refrigeration compressed air dryer SY-230FG</td>
			<td>Shanghai TaiLin Compressor Co., Ltd.</td>
			<td>S190826698</td>
			<td></td>
		</tr>
		<tr>
			<td>Scanning electron microscope (SEM): JSM-IT100</td>
			<td>JEOL (BEIJING) CO., LTD.</td>
			<td>MP1030004260426</td>
			<td></td>
		</tr>
		<tr>
			<td>Titanium alloy powder</td>
			<td>Renishaw plc</td>
			<td>H-5800-1086-01-A</td>
			<td></td>
		</tr>
		<tr>
			<td>Ultrasonic cleaning machine: AK-030S</td>
			<td>Shenzhen Yujie Cleaning Equipment Co., Ltd</td>
			<td>30820004</td>
			<td></td>
		</tr>
		<tr>
			<td>ZEN core v3.0</td>
			<td>ZEISS</td>
			<td>4702000198</td>
			<td></td>
		</tr>
	</tbody>
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plasma polishing as a new polishing option to reduce the surface roughness of porous titanium alloy for 3d printing

Porous titanium alloy implants with simulated trabecular bone fabricated by 3D printing technology have broad prospects. However, due to the fact that some powder adheres to the surface of the workpiece during the manufacturing process, the surface roughness in direct printing pieces is relatively high. At the same time, since the internal pores of the porous structure cannot be polished by conventional mechanical polishing, an alternative method needs to be found. As a surface technology, plasma polishing technology is especially suitable for parts with complex shapes that are difficult to polish mechanically. It can effectively remove particles and fine splash residues attached to the surface of 3D printed porous titanium alloy workpieces. Therefore, it can reduce surface roughness. Firstly, titanium alloy powder is used to print the porous structure of the simulated trabecular bone with a metal 3D printer. After printing, heat treatment, removal of the supporting structure, and ultrasonic cleaning is carried out. Then, plasma polishing is performed, consisting of adding a polishing electrolyte with the pH set to 5.7, preheating the machine to 101.6 &#176;C, fixing the workpiece on the polishing fixture, and setting the voltage (313 V), current (59 A), and polishing time (3 min). After polishing, the surface of the porous titanium alloy workpiece is analyzed by a confocal microscope, and the surface roughness is measured. Scanning electron microscopy is used to characterize the surface condition of porous titanium. The results show that the surface roughness of the whole porous titanium alloy workpiece changed from Ra (average roughness) = 126.9 &#181;m to Ra = 56.28 &#181;m, and the surface roughness of the trabecular structure changed from Ra = 42.61 &#181;m to Ra = 26.25 &#181;m. Meanwhile, semi-molten powders and ablative oxide layers are removed, and surface quality is improved.

Surface morphology 
Figure 3 shows the SEM result of the surface morphology of the porous titanium alloy workpiece before and after plasma polishing. We observed that at 30x and 100x magnification, the surface of the porous titanium alloy workpiece before plasma polishing seems to be rougher (Figure 3A,B). When magnified to 500x, we found that a large amount of semi-molten powders and ablative oxide layers could be observed...

Watch this Scientific Journal Video about Plasma Polishing as a New Polishing Option to Reduce the Surface Roughness of Porous Titanium Alloy for 3D Printing at JoVE.com

Plasma Polishing as a New Polishing Option to Reduce the Surface Roughness of Porous Titanium Alloy for 3D Printing

Plasma polishing is a promising surface processing technology, especially suitable for 3D printing of porous titanium alloy workpieces. It can remove semi-molten powders and ablative oxide layers, thereby effectively reducing surface roughness and improving surface quality.

Porous titanium alloy implants with simulated trabecular bone fabricated by 3D printing technology have broad prospects. However, due to the fact that ...

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1.6K Views.  Affiliated Hospital of Guangdong Medical University. Plasma polishing is a promising surface processing technology, especially suitable for 3D printing of porous titanium alloy workpieces. It can remove semi-molten powders and ablative oxide layers, thereby effectively reducing surface roughness and improving surface quality.

Video: Plasma Polishing as a New Polishing Option to Reduce the Surface Roughness of Porous Titanium Alloy for 3D Printing

Human Neuroendocrine Tumor Cell Lines as a Three-Dimensional Model for the Study of Human Neuroendocrine Tumor Therapy

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human malignancies.1 Other than surgery for the minority of patients who present with localized disease, there is little or no survival benefit of systemic therapy. Therefore, there is a great need to better understand the biology of NETs, and in particular define new therapeutic targets for patients with nonresectable or metastatic neuroendocrine tumors. 3D cell culture is becoming a popular method for drug screening due to its relevance in modeling the in vivo tumor tissue organization and microenvironment.2,3 The 3D multicellular spheroids could provide valuable information in a more timely and less expensive manner than directly proceeding from 2D cell culture experiments to animal (murine) models.
To facilitate the discovery of new therapeutics for NET patients, we have developed an in vitro 3D multicellular spheroids model using the human NET cell lines. The NET cells are plated in a non-adhesive agarose-coated 24-well plate and incubated under physiological conditions (5% CO2, 37 &deg;C) with a very slow agitation for 16-24 hr after plating. The cells form multicellular spheroids starting on the 3rd or 4th day. The spheroids become more spherical by the 6th day, at which point the drug treatments are initiated. The efficacy of the drug treatments on the NET spheroids is monitored based on the morphology, shape and size of the spheroids with a phase-contrast light microscope. The size of the spheroids is estimated automatically using a custom-developed MATLAB program based on an active contour algorithm. Further, we demonstrate a simple method to process the HistoGel embedding on these 3D spheroids, allowing the use of standard histological and immunohistochemical techniques. 
This is the first report on generating 3D spheroids using NET cell lines to examine the effect of therapeutic drugs. We have also performed histology on these 3D spheroids, and displayed an example of a single drug's effect on growth and proliferation of the NET spheroids. Our results support that the NET spheroids are valuable for further studies of NET biology and drug development.

We present a simple agarose overlay platform to grow 3D multicellular spheroids using neuroendocrine cancer cell lines. This method provides a very convenient way to examine the effect of therapeutic drugs on the neuroendocrine tumor cells. It could also help us establish human neuroendocrine tumor spheroids for cancer therapy.

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human ...

Osteopathic Manipulative Treatment as a Useful Adjunctive Tool for Pneumonia

Pneumonia, the inflammatory state of lung tissue primarily due to microbial infection, claimed 52,306 lives in the United States in 20071 and resulted in the hospitalization of 1.1 million patients2. With an average length of in-patient hospital stay of five days2, pneumonia and influenza comprise significant financial burden costing the United States $40.2 billion in 20053. Under the current Infectious Disease Society of America/American Thoracic Society guidelines, standard-of-care recommendations include the rapid administration of an appropriate antibiotic regiment, fluid replacement, and ventilation (if necessary). Non-standard therapies include the use of corticosteroids and statins; however, these therapies lack conclusive supporting evidence4. (Figure 1)

Osteopathic Manipulative Treatment (OMT) is a cost-effective adjunctive treatment of pneumonia that has been shown to reduce patients&#8217; length of hospital stay, duration of intravenous antibiotics, and incidence of respiratory failure or death when compared to subjects who received conventional care alone5. The use of manual manipulation techniques for pneumonia was first recorded as early as the Spanish influenza pandemic of 1918, when patients treated with standard medical care had an estimated mortality rate of 33%, compared to a 10% mortality rate in patients treated by osteopathic physicians6. When applied to the management of pneumonia, manual manipulation techniques bolster lymphatic flow, respiratory function, and immunological defense by targeting anatomical structures involved in the these systems7,8, 9, 10.

The objective of this review video-article is three-fold: a) summarize the findings of randomized controlled studies on the efficacy of OMT in adult patients with diagnosed pneumonia, b) demonstrate established protocols utilized by osteopathic physicians treating pneumonia, c) elucidate the physiological mechanisms behind manual manipulation of the respiratory and lymphatic systems. Specifically, we will discuss and demonstrate four routine techniques that address autonomics, lymph drainage, and rib cage mobility: 1) Rib Raising, 2) Thoracic Pump, 3) Doming of the Thoracic Diaphragm, and 4) Muscle Energy for Rib 1.5,11

Pneumonia is one of the top ten leading causes of death in the U.S. Osteopathic manipulative techniques (OMT) may be utilized as an adjunctive therapy in the treatment of pneumonia to enhance biomechanical and immune function; and ultimately, to reduce hospital stay, duration of antibiotics, incidence of respiratory failure, and mortality. In this video-article, we will review randomized controlled studies on the use of OMT in pneumonia patients, as well as demonstrate specific hands-on techniques that are routinely practiced by osteopathic physicians when treating pulmonary infections.

Pneumonia, the inflammatory state of lung tissue primarily due to microbial infection, claimed 52,306 lives in the United States in 20071 and resulted in ...

Construction of Vapor Chambers Used to Expose Mice to Alcohol During the Equivalent of all Three Trimesters of Human Development

Exposure to alcohol during development can result in a constellation of morphological and behavioral abnormalities that are collectively known as Fetal Alcohol Spectrum Disorders (FASDs). At the most severe end of the spectrum is Fetal Alcohol Syndrome (FAS), characterized by growth retardation, craniofacial dysmorphology, and neurobehavioral deficits. Studies with animal models, including rodents, have elucidated many molecular and cellular mechanisms involved in the pathophysiology of FASDs. Ethanol administration to pregnant rodents has been used to model human exposure during the first and second trimesters of pregnancy. Third trimester ethanol consumption in humans has been modeled using neonatal rodents. However, few rodent studies have characterized the effect of ethanol exposure during the equivalent to all three trimesters of human pregnancy, a pattern of exposure that is common in pregnant women. Here, we show how to build vapor chambers from readily obtainable materials that can each accommodate up to six standard mouse cages. We describe a vapor chamber paradigm that can be used to model exposure to ethanol, with minimal handling, during all three trimesters. Our studies demonstrate that pregnant dams developed significant metabolic tolerance to ethanol. However, neonatal mice did not develop metabolic tolerance and the number of fetuses, fetus weight, placenta weight, number of pups/litter, number of dead pups/litter, and pup weight were not significantly affected by ethanol exposure. An important advantage of this paradigm is its applicability to studies with genetically-modified mice. Additionally, this paradigm minimizes handling of animals, a major confound in fetal alcohol research.

We demonstrate the construction of alcohol vapor chambers using readily available materials that simultaneously house 6 mouse cages. We further describe their use in a mouse model of fetal alcohol exposure equivalent to all 3&#160;trimesters of human pregnancy. This paradigm exposes animals during gestation and postnatal days 1-12.

Exposure to alcohol during development can result in a constellation of morphological and behavioral abnormalities that are collectively known as Fetal ...

Multicolor 3D Printing of Complex Intracranial Tumors in Neurosurgery

Three-dimensional (3D) printing technologies offer the possibility of visualizing patient-specific pathologies in a physical model of correct dimensions. The model can be used for planning and simulating critical steps of a surgical approach. Therefore, it is important that anatomical structures such as blood vessels inside a tumor can be printed to be colored not only on their surface, but throughout their whole volume. During simulation this allows for the removal of certain parts (e.g., with a high-speed drill) and revealing internally located structures of a different color. Thus, diagnostic information from various imaging modalities (e.g., CT, MRI) can be combined in a single compact and tangible object.
However, preparation and printing of such a fully colored anatomical model remains a difficult task. Therefore, a step-by-step guide is provided, demonstrating the fusion of different cross-sectional imaging data sets, segmentation of anatomical structures, and creation of a virtual model. In a second step the virtual model is printed with volumetrically colored anatomical structures using a plaster-based color 3D binder jetting technique. This method allows highly accurate reproduction of patient-specific anatomy as shown in a series of 3D-printed petrous apex chondrosarcomas. Furthermore, the models created can be cut and drilled, revealing internal structures that allow for simulation of surgical procedures.

The protocol describes the fabrication of fully colored three-dimensional prints of patient-specific, anatomical skull models to be used for surgical simulation. The crucial steps of combining different imaging modalities, image segmentation, three-dimensional model extraction, and production of the prints are explained.

Three-dimensional (3D) printing technologies offer the possibility of visualizing patient-specific pathologies in a physical model of correct dimensions. ...

Combining Augmented Reality and 3D Printing to Display Patient Models on a Smartphone

Augmented reality (AR) has great potential in education, training, and surgical guidance in the medical field. Its combination with three-dimensional (3D) printing (3DP) opens new possibilities in clinical applications. Although these technologies have grown exponentially in recent years, their adoption by physicians is still limited, since they require extensive knowledge of engineering and software development. Therefore, the purpose of this protocol is to describe a step-by-step methodology enabling inexperienced users to create a smartphone app, which combines AR and 3DP for the visualization of anatomical 3D models of patients with a 3D-printed reference marker. The protocol describes how to create 3D virtual models of a patient&rsquo;s anatomy derived from 3D medical images. It then explains how to perform positioning of the 3D models with respect to marker references. Also provided are instructions for how to 3D print the required tools and models. Finally, steps to deploy the app are provided. The protocol is based on free and multi-platform software and can be applied to any medical imaging modality or patient. An alternative approach is described to provide automatic registration between a 3D-printed model created from a patient&rsquo;s anatomy and the projected holograms. As an example, a clinical case of a patient suffering from distal leg sarcoma is provided to illustrate the methodology. It is expected that this protocol will accelerate the adoption of AR and 3DP technologies by medical professionals.

Presented here is a method to design an augmented reality smartphone application for the visualization of anatomical three-dimensional models of patients using a 3D-printed reference marker.

Augmented reality (AR) has great potential in education, training, and surgical guidance in the medical field. Its combination with three-dimensional (3D) ...

3D Planning and Printing of Patient Specific Implants for Reconstruction of Bony Defects

We are in the midst of the 3D era in most aspects of life, and especially in medicine. The surgical discipline is one of the major players in the medical field using the constantly developing 3D planning and printing capabilities. Computer-assisted design (CAD) and computer assisted manufacturing (CAM) are used to describe the 3D planning and manufacturing of the product. The planning and manufacturing of 3D surgical guides and reconstruction implants is performed almost exclusively by engineers. As technology advances and software interfaces become more user-friendly, it raises a question regarding the possibility of transferring the planning and manufacturing to the clinician. The reasons for such a shift are clear: the surgeon has the idea of what he wants to design, and he also knows what is feasible and could be used in the operating room. It allows him to be prepared for any scenario/unexpected results during the operation and allows the surgeon to be creative and express his new ideas using the CAD software. The purpose of this method is to provide clinicians with the ability to create their own surgical guides and reconstruction implants. In this manuscript, a detailed protocol will provide a simple method for segmentation using segmentation software and implant planning using a 3D design software. Following the segmentation and stl file production using segmentation software, the clinician could create a simple patient specific reconstruction plate or a more complex plate with a cradle for bone graft positioning. Surgical guides can be created for accurate resection, hole preparation for proper reconstruction plate positioning or for bone graft harvesting and re-contouring. A case of lower jaw reconstruction following plate fracture and nonunion healing of a trauma sustained injury is detailed.

This protocol describes the use of 3D planning and printing for reconstruction of bony defects. We use segmentation tools to create 3D models followed by 3D design software to create patient specific implants for reconstruction purposes concomitant to ablative surgery or as a second stage.

We are in the midst of the 3D era in most aspects of life, and especially in medicine. The surgical discipline is one of the major players in the medical ...

Creation of a High-Fidelity, Low-Cost, Intraosseous Line Placement Task Trainer via 3D Printing

The description of procedural task trainers includes their use as a training tool to hone technical skills through repetition and rehearsal of procedures in a safe environment before ultimately performing the procedure on a patient. Many procedural task trainers available to date suffer from several drawbacks, including unrealistic anatomy and the tendency to develop user-created &#39;landmarks&#39; after the trainer tissue undergoes repeated manipulations, potentially leading to inappropriate psychomotor skill development. To ameliorate these drawbacks, a process was created to produce a high-fidelity procedural task trainer, created from anatomy obtained from computed tomography (CT) scans, that utilize ubiquitous three-dimensional (3D) printing technology and off-the-shelf commodity supplies.
This method includes creating a 3D printed tissue mold capturing the tissue structure surrounding the skeletal element of interest to encase the bony skeletal structure suspended within the tissue, which is also 3D printed. A tissue medium mixture, which approximates tissue in both high-fidelity geometry and tissue density, is then poured into a mold and allowed to set. After a task trainer has been used to practice a procedure, such as intraosseous line placement, the tissue media, molds, and bones are reclaimable and may be reused to create a fresh task trainer, free of puncture sites and manipulation defects, for use in subsequent training sessions.

We describe a procedure to process computed tomography (CT) scans into high-fidelity, reclaimable, and low-cost procedural task trainers. The CT scan identification processes, export, segmentation, modeling, and 3D printing are all described, along with the issues and lessons learned in the process.

The description of procedural task trainers includes their use as a training tool to hone technical skills through repetition and rehearsal of procedures ...

Combining 3D-Printing and Electrospinning to Manufacture Biomimetic Heart Valve Leaflets

Electrospinning has become a widely used technique in cardiovascular tissue engineering as it offers the possibility to create (micro-)fibrous scaffolds with adjustable properties. The aim of this study was to create multilayered scaffolds mimicking the architectural fiber characteristics of human heart valve leaflets using conductive 3D-printed collectors.
Models of aortic valve cusps were created using commercial computer-aided design (CAD) software. Conductive polylactic acid was used to fabricate 3D-printed leaflet templates. These cusp negatives were integrated into a specifically designed, rotating electrospinning mandrel. Three layers of polyurethane were spun onto the collector, mimicking the fiber orientation of human heart valves. Surface and fiber structure was assessed with a scanning electron microscope (SEM). The application of fluorescent dye additionally permitted the microscopic visualization of the multilayered fiber structure. Tensile testing was performed to assess the biomechanical properties of the scaffolds.
3D-printing of essential parts for the electrospinning rig was possible in a short time for a low budget. The aortic valve cusps created following this protocol were three-layered, with a fiber diameter of 4.1 &#177; 1.6 &#181;m. SEM imaging revealed an even distribution of fibers. Fluorescence microscopy revealed individual layers with differently aligned fibers, with each layer precisely reaching the desired fiber configuration. The produced scaffolds showed high tensile strength, especially along the direction of alignment. The printing files for the different collectors are available as Supplemental File 1, Supplemental File 2, Supplemental File 3, Supplemental File 4, and Supplemental File 5.
With a highly specialized setup and workflow protocol, it is possible to mimic tissues with complex fiber structures over multiple layers. Spinning directly on 3D-printed collectors creates considerable flexibility in manufacturing 3D shapes at low production costs.

The presented method offers an innovative way for engineering biomimetic fiber structures in three-dimensional (3D) scaffolds (e.g., heart valve leaflets). 3D-printed, conductive geometries were used to determine shape and dimensions. Fiber orientation and characteristics were individually adjustable for each layer. Multiple samples could be manufactured in one setup.

Electrospinning has become a widely used technique in cardiovascular tissue engineering as it offers the possibility to create (micro-)fibrous scaffolds ...

Platelet-Rich Plasma Lysate for Treatment of Eye Surface Diseases

Various ocular surface diseases are treated with blood-derived eye drops. Their use has been introduced in clinical practice because of their metabolite and growth factor content, which promotes eye surface regeneration. Blood-based eye drops can be prepared from different sources (i.e., whole blood or platelet apheresis donation), as well as with different protocols (e.g., different dilutions and freeze/thaw cycles). This variability hampers the standardization of clinical protocols and, consequently, the evaluation of their clinical efficacy. Detailing and sharing the methodological procedures may contribute to defining common guidelines. Over the last years, allogenic products have been diffusing as an alternative to the autologous treatments since they guarantee higher efficacy standards; among them, the platelet-rich plasma lysate (PRP-L) eye drops are prepared with simple manufacturing procedures. In the transfusion medicine unit at AUSL-IRCCS di Reggio Emilia, Italy, PRP-L is obtained from platelet-apheresis donation. This product is initially diluted to 0.3 x 109 platelets/mL (starting from an average concentration of 1 x 109 platelets/mL) in 0.9% NaCl. Diluted platelets are frozen/thawed and, subsequently, centrifuged to eliminate debris. The final volume is split into 1.45 mL aliquots and stored at &#8722;80 &#176;C. Before being dispensed to patients, eye drops are tested for sterility. Patients may store platelet lysates at &#8722;15 &#176;C for up to 1 month. The growth factor composition is also assessed from randomly selected aliquots, and the mean values are reported here.

Platelet lysates represent an emerging tool for the treatment of ocular surface diseases. Here, we propose a method for the preparation, dispensation, storage, and characterization of platelet lysate collected from platelet donors.

Various ocular surface diseases are treated with blood-derived eye drops. Their use has been introduced in clinical practice because of their metabolite ...

A Personalized 3D-Printed Model for Preoperative Evaluation in Thyroid Surgery

The anatomic structure of the surgical area of thyroid cancer is complex. It is very important to comprehensively and carefully evaluate the tumor location and its relation with the capsule, trachea, esophagus, nerves, and blood vessels before operation. This paper introduces an innovative 3D-printed model establishment method based on computerized tomography (CT) DICOM images. We established a personalized 3D-printed model of the cervical thyroid surgery field for each patient who needed thyroid surgery to help clinicians evaluate the key points and difficulties of the surgery and select the operation methods of key parts as a basis. The results showed that this model is conducive to preoperative discussion and the formulation of operation strategies. In particular, as a result of the clear display of the recurrent laryngeal nerve and parathyroid gland locations in the thyroid operation field, injury to them can be avoided during surgery, the difficulty of thyroid surgery reduced, and the incidence of postoperative hypoparathyroidism and complications related to recurrent laryngeal nerve injury reduced too. Moreover, this 3D-printed model is intuitive and aids communication for the signing of informed consent by patients before surgery.

Here, a new method of establishing a personalized 3D-printed model for preoperative evaluation of thyroid surgery is proposed. It is conducive to preoperative discussion, reducing the difficulty of thyroid surgery.