PLOS ONE: [sortOrder=DATE_NEWEST_FIRST, sort=Date, newest first, q=subject:"Manufacturing processes"]PLOShttps://journals.plos.org/plosone/webmaster@plos.orgaccelerating the publication of peer-reviewed sciencehttps://journals.plos.org/plosone/search/feed/atom?sortOrder=DATE_NEWEST_FIRST&unformattedQuery=subject:%22Manufacturing+processes%22&sort=Date,+newest+firstAll PLOS articles are Open Access.https://journals.plos.org/plosone/resource/img/favicon.icohttps://journals.plos.org/plosone/resource/img/favicon.ico2024-03-29T12:44:44ZLow-cost, versatile, and highly reproducible microfabrication pipeline to generate 3D-printed customised cell culture devices with complex designsCathleen HagemannMatthew C. D. BaileyEugenia CarraroKsenia S. StankevichValentina Maria LionelloNoreen KhokharPacharaporn SuklaiCarmen Moreno-GonzalezKelly O’TooleGeorge KonstantinouChristina L. DixSudeep JoshiEleonora GiagnorioMads S. BergholtChristopher D. SpicerAlbane ImbertFrancesco Saverio TedescoAndrea Serio10.1371/journal.pbio.30025032024-03-13T14:00:00Z2024-03-13T14:00:00Z<p>by Cathleen Hagemann, Matthew C. D. Bailey, Eugenia Carraro, Ksenia S. Stankevich, Valentina Maria Lionello, Noreen Khokhar, Pacharaporn Suklai, Carmen Moreno-Gonzalez, Kelly O’Toole, George Konstantinou, Christina L. Dix, Sudeep Joshi, Eleonora Giagnorio, Mads S. Bergholt, Christopher D. Spicer, Albane Imbert, Francesco Saverio Tedesco, Andrea Serio</p>
Cell culture devices, such as microwells and microfluidic chips, are designed to increase the complexity of cell-based models while retaining control over culture conditions and have become indispensable platforms for biological systems modelling. From microtopography, microwells, plating devices, and microfluidic systems to larger constructs such as live imaging chamber slides, a wide variety of culture devices with different geometries have become indispensable in biology laboratories. However, while their application in biological projects is increasing exponentially, due to a combination of the techniques, equipment and tools required for their manufacture, and the expertise necessary, biological and biomedical labs tend more often to rely on already made devices. Indeed, commercially developed devices are available for a variety of applications but are often costly and, importantly, lack the potential for customisation by each individual lab. The last point is quite crucial, as often experiments in wet labs are adapted to whichever design is already available rather than designing and fabricating custom systems that perfectly fit the biological question. This combination of factors still restricts widespread application of microfabricated custom devices in most biological wet labs. Capitalising on recent advances in bioengineering and microfabrication aimed at solving these issues, and taking advantage of low-cost, high-resolution desktop resin 3D printers combined with PDMS soft lithography, we have developed an optimised a low-cost and highly reproducible microfabrication pipeline. This is thought specifically for biomedical and biological wet labs with not prior experience in the field, which will enable them to generate a wide variety of customisable devices for cell culture and tissue engineering in an easy, fast reproducible way for a fraction of the cost of conventional microfabrication or commercial alternatives. This protocol is designed specifically to be a resource for biological labs with limited expertise in those techniques and enables the manufacture of complex devices across the μm to cm scale. We provide a ready-to-go pipeline for the efficient treatment of resin-based 3D-printed constructs for PDMS curing, using a combination of polymerisation steps, washes, and surface treatments. Together with the extensive characterisation of the fabrication pipeline, we show the utilisation of this system to a variety of applications and use cases relevant to biological experiments, ranging from micro topographies for cell alignments to complex multipart hydrogel culturing systems. This methodology can be easily adopted by any wet lab, irrespective of prior expertise or resource availability and will enable the wide adoption of tailored microfabricated devices across many fields of biology.Research on surface treatment technology for quickly improving the skid resistance of tunnel concrete pavementJun’an LeiFujing ZhaoYuanyuan WangXiaofeng Ren10.1371/journal.pone.02959382024-03-11T14:00:00Z2024-03-11T14:00:00Z<p>by Jun’an Lei, Fujing Zhao, Yuanyuan Wang, Xiaofeng Ren</p>
In order to solve the problem that the skid resistance of concrete pavement in tunnel deteriorates rapidly, which is easy to cause traffic accidents, the anti-skid rapid elevation technology of surface treatment is proposed. Wear tests were used to investigate the effects of concrete surface roughness, properties of modified emulsified asphalt binder and anti-skid fine aggregate type on long-term skid resistance of treated surfaces. The results show that the four coarsening methods of fine milling, milling, grooving and brooming can improve the skid resistance of concrete, and the skid resistance durability of fine milling and milling is better. The adhesive property of modified emulsified asphalt is the best when the content of water-based epoxy resin is 20%. In different aggregates, the anti-skid effect is better when silicon carbide is used as anti-skid aggregate and the particle size is 0.6mm:0.3mm = 2:3. The method of fine milling of concrete surface + spraying epoxy emulsified asphalt + spreading silicon carbide can effectively improve the anti-skid performance of the original concrete pavement, and the feasibility of the scheme is verified by the test road. The research results have a good reference value for improving the skid resistance of tunnel concrete pavement.Persistence comparison of two Shiga-toxin producing <i>Escherichia coli</i> (STEC) serovars during long-term storage and thermal inactivation in various wheat floursIan S. HinesTom JurkiwEmily NguyenMartine FergusonSultana SolaimanElizabeth ReedMaria HoffmannJie Zheng10.1371/journal.pone.02999222024-03-08T14:00:00Z2024-03-08T14:00:00Z<p>by Ian S. Hines, Tom Jurkiw, Emily Nguyen, Martine Ferguson, Sultana Solaiman, Elizabeth Reed, Maria Hoffmann, Jie Zheng</p>
Foodborne outbreaks associated with Shiga toxin-producing <i>Escherichia coli</i> (STEC) contaminated wheat flour have been an increasing food safety concern in recent decades. However, there is little literature aimed at investigating the impact of different flour types on the persistence of STEC during storage and thermal inactivation. Therefore, two serovars of STEC, O121 and O157, were selected to inoculate each of five different types of common wheat flours: whole wheat, bleached, unbleached, bread, and self-rising. Inoculated flours were examined for the stability of STEC during storage for up to 42 days at room temperature (RT) and a<sub>w</sub> ~0.56. Additionally, the thermal resistance of O121 and O157 under isothermal conditions at 60, 70, 80, and 90°C was analyzed for the inoculated flours. STEC storage persistence at RT was generally not affected by flour type, however, decreases of 1.2 and 2.4 log CFU/day within whole wheat flour for O121 and O157, respectively, were significantly lower than other flours. Though few differences were identified in relation to flour type, O121 exhibited significantly better survival rates than O157 during both equilibrium and storage periods. Compared to an approximate 6 log reduction in the population of O157, O121 population levels were reduced by a significantly lower amount (~3 log) during the entire storage period at RT. At each isothermal temperature, the impact of flour type on the thermal resistance capabilities of O121 or O157 was not a significant factor and resulted in similar survival curves regardless of serovar. Instead of exhibiting linear survival curves, both O121 and O157 displayed nonlinear curves with some shoulder/tail effect. Similar for both O121 and O157, the predicted decimal reduction time (D-value) decreased from approximately 25 min to around 8 min as the isothermal temperature increased from 60°C to 90°C. Results reported here can contribute to risk assessment models concerning contamination of STEC in wheat flour and add to our understanding of the impacts of flour type and STEC serovar on desiccation stability during storage and isothermal inactivation during thermal treatment.Low-power and area-efficient memristor based non-volatile D latch and flip-flop: Design and analysisHaroon Rasheed S.Rajeev Pankaj Nelapati10.1371/journal.pone.03000732024-03-07T14:00:00Z2024-03-07T14:00:00Z<p>by Haroon Rasheed S., Rajeev Pankaj Nelapati</p>
In recent years, non-volatile memory elements have become highly appealing for memory applications to implement a new class of storage memory that could replace flash memories in sequential logic applications, with features such as compactness, low power, fast processing speed, high endurance, and retention. The memristor is one such non-volatile element that fits the fundamental blocks of sequential logic circuits, the latch and flip-flop; hence, in this article, a non-volatile latch architecture using memristor ratioed logic (MRL) inverter and CMOS components is focused, with an additional memristor as a memory element. A Verilog-A model was used to create the memristor element. The simulation findings validated the compact, low-voltage, and reliable design of the latch design. We evolved in technology enough to create a master-slave flip-flop and arrange it to function as a counter and a shift register. Power, number of elements, cell size, energy, programming time, and robustness are compared to comparable non-volatile topologies. The proposed non-volatile latch proves non-volatility and can store data with a 24% reduction in power consumption and a near 10% reduction in area.Mining co-location patterns of manufacturing firms using Q statistic and additive color mixingYi SongGuanglei LiYihan WangYiheng WangChang Ren10.1371/journal.pone.02990462024-03-06T14:00:00Z2024-03-06T14:00:00Z<p>by Yi Song, Guanglei Li, Yihan Wang, Yiheng Wang, Chang Ren</p>
The agglomeration effect significantly influences firms’ site selection. Manufacturing firms often exhibit intricate spatial co-location patterns that are indicative of agglomerations due to their reliance on material input and product output across various subdivisions of manufacture. In this study, we present an analytical approach employing the Q statistic and additive color mixing visualization to assess co-location patterns of manufacturing firms. We identified frequent pairs and triplets of manufacturing divisions, mapping them to reveal distinct categories: labor-intensive clusters, upstream/downstream industrial chains, and technology-spillover clusters. These agglomeration categories concentrate in different regions of the city. Policy implications are proposed to promote the upgrade of labor-intensive divisions, enhance the operational efficiency of upstream/downstream industrial chains, and reinforce the spillover effects of technology-intensive divisions.Immunoinformatics-guided recombinant polypeptide-based enzyme-linked immunosorbent assay for seromonitoring of laboratory animals for minute virus of mice and Kilham rat virusCharanpreet KaurKandala Pavan AsrithS. G. RamachandraNagendra R. Hegde10.1371/journal.pone.02987422024-02-27T14:00:00Z2024-02-27T14:00:00Z<p>by Charanpreet Kaur, Kandala Pavan Asrith, S. G. Ramachandra, Nagendra R. Hegde</p>
Subclinical infection of laboratory animals with one or more of several pathogens affects the results of experiments on animals. Monitoring the health of laboratory animals encompasses routine surveillance for pathogens, including several viruses. This study aimed to explore the development of an alternative assay to the existing ones for detecting infection of mice and rats with the parvoviruses minute virus of mice (MVM) and Kilham rat virus (KRV), respectively. Full-length VP2 and NS1 proteins of these parvoviruses, besides fragments containing multiple predicted epitopes stitched together, were studied for serological detection. The optimal dilution of full-length proteins and antigenic regions containing predicted epitopes for coating, test sera, and conjugate was determined using a checkerboard titration at each step. The assays were evaluated vis-à-vis commercially available ELISA kits. The results showed that an engineered fusion of fragments containing multiple predicted MVM VP2 and NS1 epitopes was better than either of the full-length proteins for detecting antibodies in 90% of the tested sera samples. For KRV ELISA, full-length VP2 was better compared to other individual recombinant protein fragments or combinations thereof for the detection of antibodies in sera. This report is the first description of an ELISA for KRV and an improved assay for MVM. Importantly, our assays could be exploited with small volumes of sera. The results also demonstrate the utility of immunoinformatics-driven polypeptide engineering in the development of diagnostic assays and the potential to develop better tests for monitoring the health status of laboratory animals.Impact of air-polishing using erythritol on surface roughness and substance loss in dental hard tissue: An <i>ex vivo</i> studyAnne B. KruseStephan FortmeierKirstin VachElmar HellwigPetra Ratka-KrügerNadine Schlueter10.1371/journal.pone.02866722024-02-26T14:00:00Z2024-02-26T14:00:00Z<p>by Anne B. Kruse, Stephan Fortmeier, Kirstin Vach, Elmar Hellwig, Petra Ratka-Krüger, Nadine Schlueter</p>
This <i>ex vivo</i> study aimed to investigate surface roughness and substance loss after treatment with different professional cleaning methods and to determine whether subsequent polishing with a rubber cup and polishing paste is necessary. Samples (flat and natural surfaces) of human enamel and dentin were prepared (baseline) and treated with either a curette, air-polishing with erythritol, a rubber cup and polishing paste, or a combination thereof (treatment). Subsequently, all samples were immersed in an ultrasonic bath (ultrasonic) to remove residues from the treatment procedures. The surface roughness values sRa and sRz as well as tissue loss were measured profilometrically. Linear regression models were used to compare group differences (roughness and loss) considering the corresponding baseline value. The significance level was set at <i>p<0</i>.<i>05</i>. sRa increased significantly after treatment with curettes or air-polishing with erythritol in both enamel (<i>p<0</i>.<i>001</i>) and dentin (<i>p<0</i>.<i>001</i>) of flat samples. The same effect was observed for sRz in dentin (<i>p<0</i>.<i>001</i>) but not for enamel compared to negative control. Polishing with a rubber cup and paste alone had no significant effect on roughness values. When combined with other treatments, the effect of curette or air-polishing with erythritol dominated the effect. In enamel, none of the tested methods led to measurable tissue loss. In dentin, air-polishing with erythritol caused ≤50% tissue loss compared to the curette. Conclusively, for enamel, treatment effects on roughness were measurable but of limited clinical relevance. For dentin, air-polishing resulted in a smaller but insignificant roughness increase and less tissue loss compared to the curette. Polishing with a rubber cup and paste did not affect surface roughness. Regarding the clinical application, the use of air-polishing seems to be a less invasive procedure than using a curette; polishing with rubber cup and paste offers no advantage in terms of reducing roughness as a final procedure.Sublethal thermal stress promotes migration and invasion of thyroid cancer cellsChi-Yu KuoChung-Hsin TsaiJun Kui WuShih-Ping Cheng10.1371/journal.pone.02989032024-02-23T14:00:00Z2024-02-23T14:00:00Z<p>by Chi-Yu Kuo, Chung-Hsin Tsai, Jun Kui Wu, Shih-Ping Cheng</p>
Objective <p>Radiofrequency ablation is a viable option in the treatment of benign thyroid nodules. Some reports suggest that thermal ablation may also be safe for the management of low-risk thyroid cancer. In this study, we applied transient heat treatment to thyroid cancer cells to mimic clinical scenarios in which insufficient ablation leads to incomplete eradication of thyroid cancer.</p> Methods <p>Differentiated thyroid cancer cell lines B-CPAP, TPC-1, and FTC-133 were subjected to heat treatment at different temperatures for 10 min. Effects on cell growth, clonogenicity, wound healing assay, and Transwell invasion were determined.</p> Results <p>Heat treatment at 45°C or higher reduced cell growth, whereas viability of thyroid cancer cells was not changed after heat treatment at 37, 40, or 42°C. Heat treatment at 40°C increased the number of colony formations by 16% to 39%. Additionally, transient heat treatment at 40°C resulted in a 1.75-fold to 2.56-fold higher migratory activity than treatment at 37°C. Invasive capacity was increased after heat treatment, ranging from 115% to 126%. Expression of several epithelial-mesenchymal transition markers, including ZEB1, N-cadherin, and MMP2, was upregulated following heat treatment at 40°C.</p> Conclusion <p>We for the first time demonstrate that sublethal thermal stress may increase clonogenicity, migration, and invasion of thyroid cancer cells.</p>Sustained antibacterial activity of orthodontic elastomeric ligature ties coated with a novel kombucha-derived bacterial nanocellulose: An <i>in-vitro</i> studyFateme EskandariSusan BorzouAlireza RazavianNeda BabanouriKhadije Yousefi10.1371/journal.pone.02929662024-02-08T14:00:00Z2024-02-08T14:00:00Z<p>by Fateme Eskandari, Susan Borzou, Alireza Razavian, Neda Babanouri, Khadije Yousefi</p>
Incipient carious lesions, the most common complication in orthodontic patients with fixed appliances, call for the development of novel preventive dental materials that do not rely on patient adherence. The present study aimed to assess the ability of elastomeric ligatures coated with bacterial nanocellulose (BNC) to deliver sustained antibacterial activity, during the standard 28-day interval between orthodontic appointments, without compromising their mechanical properties. Kombucha membrane was used to produce cellulose as a secondary product from the fermentation of tea broth with symbiotic bacteria and yeast culture. Characterization of BNC-coated elastomeric ligatures was performed using Fourier Transform Infrared Spectroscopy and Energy Dispersive Spectroscopy analysis. The samples were pre-treated by immersion first in isopropyl alcohol, then in 8 mL nanocellulose solution for 7 days. Tensile strain and strength of the BNC-coated and conventional ligatures were evaluated using a tensile testing machine. Direct contact and agar diffusion tests were performed to assess the antibacterial activity of nanocellulose. In addition, the release profile of BNC was evaluated. Data analysis was performed by one-way analysis of variance (ANOVA) followed by post-hoc Tukey’s test and Wilcoxon signed-rank test. P values less than 0.05 was regarded as significant. There was no statistically significant difference in tensile strain and strength between the BNC-coated and conventional ligatures. The coated ligatures provided sustained antibacterial activity during the required 28 days. The use of BNC-coated elastomeric ligatures in patients with fixed orthodontic appliances might be a promising solution to plaque formation and subsequent enamel decalcification.Multi-domain automated patterning of DNA-functionalized hydrogelsMoshe RubanovJoshua ColeHeon-Joon LeeLeandro G. Soto CordovaZachary ChenElia GonzalezRebecca Schulman10.1371/journal.pone.02959232024-02-02T14:00:00Z2024-02-02T14:00:00Z<p>by Moshe Rubanov, Joshua Cole, Heon-Joon Lee, Leandro G. Soto Cordova, Zachary Chen, Elia Gonzalez, Rebecca Schulman</p>
DNA-functionalized hydrogels are capable of sensing oligonucleotides, proteins, and small molecules, and specific DNA sequences sensed in the hydrogels’ environment can induce changes in these hydrogels’ shape and fluorescence. Fabricating DNA-functionalized hydrogel architectures with multiple domains could make it possible to sense multiple molecules and undergo more complicated macroscopic changes, such as changing fluorescence or changing the shapes of regions of the hydrogel architecture. However, automatically fabricating multi-domain DNA-functionalized hydrogel architectures, capable of enabling the construction of hydrogel architectures with tens to hundreds of different domains, presents a significant challenge. We describe a platform for fabricating multi-domain DNA-functionalized hydrogels automatically at the micron scale, where reaction and diffusion processes can be coupled to program material behavior. Using this platform, the hydrogels’ material properties, such as shape and fluorescence, can be programmed, and the fabricated hydrogels can sense their environment. DNA-functionalized hydrogel architectures with domain sizes as small as 10 microns and with up to 4 different types of domains can be automatically fabricated using ink volumes as low as 50 μL. We also demonstrate that hydrogels fabricated using this platform exhibit responses similar to those of DNA-functionalized hydrogels fabricated using other methods by demonstrating that DNA sequences can hybridize within them and that they can undergo DNA sequence-induced shape change.From birth to lying on- or under a supplemental heat source: How long does it take for half the litter to lie down and how long do they stay?Cassandra R. StambukAnna K. JohnsonKarli J. LaneKenneth J. Stalder10.1371/journal.pone.02943362023-12-21T14:00:00Z2023-12-21T14:00:00Z<p>by Cassandra R. Stambuk, Anna K. Johnson, Karli J. Lane, Kenneth J. Stalder</p>
Piglets are susceptible to hypothermia because they lack hair and energy reserves, have a large surface area to body weight ratio, and have poor body thermostability. Different heat sources are used on farm, but it is not known how long it takes half a litter to locate it and lie down. The objectives of this study were to determine 1) how long it took for ≥ 50% of piglets to locate and lie on- or under the supplemental heat source for ≥ 5 min after the birth of the last-born pig and, 2) how long this cohort of piglets laid on- or under the heat source. A total of 12 sows were enrolled in the study (parity 1 = 4; 3 = 2; 4 = 2; 5 = 2; and 7 = 2). The stall containing one sow and her litter was the experimental unit. Two treatments were compared: 1) Baby Pig Heat Mat—Single 48 (MAT) and 2) Poly Heat Lamp (LAMP). Temperature was 32°C for both heat source treatments. Sow and litter video recordings occurred continually over a 24-h period on the day of farrowing. Two measures were determined 1) how long it took for ≥ 50% of piglets to locate and lie on- or under the supplemental heat source for ≥ 5 min after the birth of the last-born pig (h:min), and 2) how long this cohort of piglets laid on- or under the heat source (min:sec). Lying was defined as either sternal or lateral recumbency with ≥ 75% of the piglet’s body touching the heat mat or inside the lamp heat circle. Production records were used to verify farrowing date, total number of piglets born, and born alive. No cross fostering occurred during this study. All data will be presented descriptively. On average, sows assigned to the LAMP treatment took ~2 ½-h to farrow, and for sows assigned to the MAT ~3 ½-h, respectively. Piglets took between ~5-h (LAMP) and ~9-h (MAT) for ≥ 50% of piglets to locate and lie on- or under the supplemental heat source for ≥ 5 min after the birth of the last piglet. Cohort of piglets laid on- or under the heat source as follows, LAMP piglets spent ~29 mins lying and for MAT piglets ~42 mins, respectively. Average pre-weaning mortality was 11% (LAMP) and 18% (MAT). The MAT heat source used less energy than the LAMP (16 vs. 63 kWh) over the study duration. To the authors’ knowledge, this is the first published study using a continuous sampling method to precisely examine a new measure (time needed for ≥ 50% of piglets to locate and lie on- or under the supplemental heat source for ≥ 5 min after the birth of the last-born piglet) and to determine how long this cohort of piglets laid on- or under the heat source. Our findings show an immense range in locating and lying under- or on the heat source. Therefore, we suggest that caretakers should assist all piglets to locate the heat source after farrowing is complete to improve piglet livability.Design of vacuum annealing furnace temperature control system based on GA-Fuzzy-PID algorithmJintao MengHaitao GaoMixue RuanHai GuoXiaojie ZhouDi Zhang10.1371/journal.pone.02938232023-11-29T14:00:00Z2023-11-29T14:00:00Z<p>by Jintao Meng, Haitao Gao, Mixue Ruan, Hai Guo, Xiaojie Zhou, Di Zhang</p>
As is well known, the metal annealing process has the characteristics of heat concentration and rapid heating. Traditional vacuum annealing furnaces use PID control method, which has problems such as high temperature fluctuation, large overshoot, and long response time during the heating and heating process. Based on this situation, some domestic scholars have adopted fuzzy PID control algorithm in the temperature control of vacuum annealing furnaces. Due to the fact that fuzzy rules are formulated through a large amount of on-site temperature data and experience summary, there is a certain degree of subjectivity, which cannot ensure that each rule is optimal. In response to this drawback, the author combined the technical parameters of vacuum annealing furnace equipment, The fuzzy PID temperature control of the vacuum annealing furnace is optimized using genetic algorithm. Through simulation and comparative analysis, it is concluded that the design of the fuzzy PID vacuum annealing furnace temperature control system based on GA optimization is superior to fuzzy PID and traditional PID control in terms of temperature accuracy, rise time, and overshoot control. Finally, it was verified through offline experiments that the fuzzy PID temperature control system based on GA optimization meets the annealing temperature requirements of metal workpieces and can be applied to the temperature control system of vacuum annealing furnaces.What drives researcher preferences for chemical compounds? Evidence from conjoint analysisBongsuk SungKang-Min ParkChun Gun ParkYong-Hee KimJaeyong LeeTae-Eun Jin10.1371/journal.pone.02945762023-11-27T14:00:00Z2023-11-27T14:00:00Z<p>by Bongsuk Sung, Kang-Min Park, Chun Gun Park, Yong-Hee Kim, Jaeyong Lee, Tae-Eun Jin</p>
We investigated the attributes and attribute levels that affect researcher preferences for chemical compounds. We conducted a conjoint analysis on survey data of Korean researchers using chemical compounds from the Korean Chemical Bank (KCB). The analysis estimated the part-worth utility for each attribute’s level, calculated relative importance of attributes, and classified user segmentation with different patterns. The results show that the structure database offers the highest part-worth utility to researchers, followed by high new functionality, price, screening service, and drug action data provided only by the KCB. Notably, researchers view the offer of a structured database and high new functionality as more important than other attributes in decision-making about research and development of chemical compounds. Furthermore, the results of segmentation analysis demonstrated that researchers have distinct usage patterns of chemical compounds: researchers consider structure database and high new functionality in cluster 1; and high new functionality and price in cluster 2, to be the most appealing. We discussed some policy and strategic implications based on the findings of this study and proposed some limitations.HSP70 via HIF-1 α SUMOylation inhibits ferroptosis inducing lung cancer recurrence after insufficient radiofrequency ablationBin PengXiean LingTonghai HuangJun Wan10.1371/journal.pone.02942632023-11-10T14:00:00Z2023-11-10T14:00:00Z<p>by Bin Peng, Xiean Ling, Tonghai Huang, Jun Wan</p>
Radiofrequency ablation (RFA) is an effective and feasible therapy for lung cancer, but accelerated progression of residual non-small cell lung cancer (NSCLC) after incomplete radiofrequency ablation (RFA) has frequently been reported. A previous study reported that HSP70 and HIF-1α were highly expressed in areas with incomplete RFA. Therefore, we sought to elucidate the regulatory effect of the HIF-1α/HSP70 pathway on lung cancer recurrence after incomplete radiofrequency ablation. In this study, we found that knockdown of HSP70 can reduce sumo 1, sumo 2/3 (marker of SUMOylation) of HIF-1α and inhibit A549 cell proliferation and migration under heat stress conditions (used to simulate incomplete RFA in vitro). We observed that knockdown of HSP70 altered the expression of ferroptosis-related proteins and genes (SLC7A11 and ACSL3), and the RNA-seq results showed that knockdown of HSP70 activated the ferroptosis pathway, further confirming that HSP70 regulates ferroptosis. In summary, HSP70, via HIF-1α SUMOylation, inhibited ferroptosis, inducing lung cancer recurrence after radiofrequency ablation. The study reveals a new direction for further research on therapeutic targets to suppress lung cancer recurrence and provides a theoretical foundation for further clinical studies.<i>In vitro</i> formation and extended culture of highly metabolically active and contractile tissuesIsabella A. BagdasarianThamidul Islam TonmoyB. Hyle ParkJoshua T. Morgan10.1371/journal.pone.02936092023-11-01T14:00:00Z2023-11-01T14:00:00Z<p>by Isabella A. Bagdasarian, Thamidul Islam Tonmoy, B. Hyle Park, Joshua T. Morgan</p>
3D cell culture models have gained popularity in recent years as an alternative to animal and 2D cell culture models for pharmaceutical testing and disease modeling. Polydimethylsiloxane (PDMS) is a cost-effective and accessible molding material for 3D cultures; however, routine PDMS molding may not be appropriate for extended culture of contractile and metabolically active tissues. Failures can include loss of culture adhesion to the PDMS mold and limited culture surfaces for nutrient and waste diffusion. In this study, we evaluated PDMS molding materials and surface treatments for highly contractile and metabolically active 3D cell cultures. PDMS functionalized with polydopamine allowed for extended culture duration (14.8 ± 3.97 days) when compared to polyethylamine/glutaraldehyde functionalization (6.94 ± 2.74 days); Additionally, porous PDMS extended culture duration (16.7 ± 3.51 days) compared to smooth PDMS (6.33 ± 2.05 days) after treatment with TGF-β2 to increase culture contraction. Porous PDMS additionally allowed for large (13 mm tall × 8 mm diameter) constructs to be fed by diffusion through the mold, resulting in increased cell density (0.0210 ± 0.0049 mean nuclear fraction) compared to controls (0.0045 ± 0.0016 mean nuclear fraction). As a practical demonstration of the flexibility of porous PDMS, we engineered a vascular bioartificial muscle model (VBAM) and demonstrated extended culture of VBAMs anchored with porous PDMS posts. Using this model, we assessed the effect of feeding frequency on VBAM cellularity. Feeding 3×/week significantly increased nuclear fraction at multiple tissue depths relative to 2×/day. VBAM maturation was similarly improved in 3×/week feeding as measured by nuclear alignment (23.49° ± 3.644) and nuclear aspect ratio (2.274 ± 0.0643) relative to 2x/day (35.93° ± 2.942) and (1.371 ± 0.1127), respectively. The described techniques are designed to be simple and easy to implement with minimal training or expense, improving access to dense and/or metabolically active 3D cell culture models.