Dynamics of axonal ß-actin mRNA in live hippocampal neurons.

Localization of mRNA facilitates spatiotemporally controlled protein expression in neurons. In axons, mRNA transport followed by local protein synthesis plays a critical role in axonal growth and guidance. However, it is not yet clearly understood how mRNA is transported to axonal subcellular sites and what regulates axonal mRNA localization. Using a transgenic mouse model in which endogenous ß-actin mRNA is fluorescently labeled, we investigated ß-actin mRNA movement in axons of hippocampal neurons. We cultured neurons in microfluidic devices to separate axons from dendrites and performed single-particle tracking of axonal ß-actin mRNA. Compared with dendritic ß-actin mRNA, axonal ß-actin mRNA showed less directed motion and exhibited mostly subdiffusive motion, especially near filopodia and boutons in mature dissociated hippocampal neurons. We found that axonal ß-actin mRNA was likely to colocalize with actin patches (APs), regions that have a high density of filamentous actin (F-actin) and are known to have a role in branch initiation. Moreover, simultaneous imaging of F-actin and axonal ß-actin mRNA in live neurons revealed that moving ß-actin mRNA tended to be docked in the APs. Our findings reveal that axonal ß-actin mRNA localization is facilitated by actin networks and suggest that localized ß-actin mRNA plays a potential role in axon branch formation.

All-in-one microfluidic design to integrate vascularized tumor spheroid into high-throughput platform.

The development of a scalable and highly reproducible in vitro tumor microenvironment (TME) platform still sheds light on new insights into cancer metastasis mechanisms and anticancer therapeutic strategies. Here, we present an all-in-one injection molded plastic array three-dimensional culture platform (All-in-One-IMPACT) that integrates vascularized tumor spheroids for highly reproducible, high-throughput experimentation. This device allows the formation of self-assembled cell spheroids on a chip by applying the hanging drop method to the cell culture channel. Then, when the hydrogel containing endothelial cells and fibroblasts is injected, the spheroid inside the droplet can be patterned together in three dimensions along the culture channel. In just two steps above, we can build a vascularized TME within a defined area. This process does not require specialized user skill and minimizes error-inducing steps, enabling both reproducibility and high throughput of the experiment. We have successfully demonstrated the process, from spheroid formation to tumor vascularization, using patient-derived cancer cells (PDCs) as well as various cancer cell lines. Furthermore, we performed combination therapies with Taxol (paclitaxel) and Avastin (bevacizumab), which are used in standard care for metastatic cancer. The All-in-One IMPACT is a powerful tool for establishing various anticancer treatment strategies through the development of a complex TME for use in high-throughput experiments.

Prevention of urethral fibrosis induced by transforming growth factor beta 1 using selective Wnt/ß-catenin signaling inhibitors in a rat model.

OBJECTIVES: To determine the anti-fibrotic effects of Wnt/ß-catenin signaling inhibitors on urethral stricture. METHODS: Human fibroblasts were exposed to transforming growth factor beta 1 combined with various concentrations of Wnt/ß-catenin inhibitors (ICG-001, IWR-1, and PRI-724), and cell proliferation and migration were evaluated. Urethral fibrosis was induced in male Sprague-Dawley rats by urethral injection of transforming growth factor beta 1 and co-treatement with inhibitors. Urethral tissues were harvested 2 weeks after the injection. The messenger ribonucleic acid and protein expression was examined for fibrosis markers Axin-1, collagen type 1, alpha smooth muscle actin, and ß-catenin. Histological analysis of fibrosis and collagen deposition was also performed. RESULTS: Cell migration was ameliorated by ICG-001 and PRI-724. Protein and messenger ribonucleic acid expression of collagen type 1 and alpha smooth muscle actin in transforming growth factor beta 1-treated fibroblasts decreased in a concentration-dependent manner with the ICG-001 and PRI-724 treatments (P < 0.05). However, there were no significant changes with the IWR-1 treatment. Collagen type I and alpha smooth muscle actin messenger ribonucleic acid and protein expression were both significantly increased in the urethral tissues of rats with transforming growth factor beta 1-induced urethral fibrosis. Rats co-treated with ICG-001 or PRI-724 showed relatively mild fibrosis and significantly reduced collagen type I and alpha smooth muscle actin messenger ribonucleic acid and protein expression (P < 0.05). CONCLUSIONS: ICG-001 and PRI-724 significantly ameliorated urethral fibrosis induced by transforming growth factor beta 1 in rats. These results suggest that ICG-001 and PRI-724 can be developed as therapeutics for treating urethral stricture.

Anchor-IMPACT: A standardized microfluidic platform for high-throughput antiangiogenic drug screening.

In vitro models are becoming more advanced to truly present physiological systems while enabling high-throughput screening and analysis. Organ-on-a-chip devices provide remarkable results through the reconstruction of a three-dimensional (3D) cellular microenvironment although they need to be further developed in terms of multiple liquid patterning principle, material properties, and scalability. Here we present a 3D anchor-based microfluidic injection-molded plastic array culture platform (Anchor-IMPACT) that enables selective, space-intensive patterning of hydrogels using anchor-island for high-throughput angiogenesis evaluation model. Anchor-IMPACT consists of a central channel and an anchor-island, integrating the array into an abbreviated 96-well plate format with a standard microscope slide size. The anchor-island enables selective 3D cell patterning without channel-to-channel contact or any hydrogel injection port using an anchor structure unlike conventional culture compartment. The hydrogel was patterned into defined regions by spontaneous capillary flow under hydrophilic conditions. We configured multiple cell patterning structures to investigate the angiogenic potency of colorectal cancer cells in Anchor-IMPACT and the morphological properties of the angiogenesis induced by the paracrine effect were evaluated. In addition, the efficacy of anticancer drugs against angiogenic sprouts was verified by following dose-dependent responses. Our results indicate that Anchor-IMPACT offers not only a model of high-throughput experimentation but also an advanced 3D cell culture platform and can significantly improve current in vitro models while providing the basis for developing predictive preclinical models for biopharmaceutical applications.

3D brain angiogenesis model to reconstitute functional human blood-brain barrier in vitro.

The human central nervous system (CNS) vasculature expresses a distinctive barrier phenotype, the blood-brain barrier (BBB). As the BBB contributes to low efficiency in CNS pharmacotherapy by restricting drug transport, the development of an in vitro human BBB model has been in demand. Here, we present a microfluidic model of CNS angiogenesis having three-dimensional (3D) lumenized vasculature in concert with perivascular cells. We confirmed the necessity of the angiogenic tri-culture system (brain endothelium in direct interaction with pericytes and astrocytes) to attain essential phenotypes of BBB vasculature, such as minimized vessel diameter and maximized junction expression. In addition, lower vascular permeability is achieved in the tri-culture condition compared to the monoculture condition. Notably, we focussed on reconstituting the functional efflux transporter system, including p-glycoprotein (p-gp), which is highly responsible for restrictive drug transport. By conducting the calcein-AM efflux assay on our 3D perfusable vasculature after treatment of efflux transporter inhibitors, we confirmed the higher efflux property and prominent effect of inhibitors in the tri-culture model. Taken together, we designed a 3D human BBB model with functional barrier properties based on a developmentally inspired CNS angiogenesis protocol. We expect the model to contribute to a deeper understanding of pathological CNS angiogenesis and the development of effective CNS medications.

Optogenetic neuronal stimulation promotes axon outgrowth and myelination of motor neurons in a three-dimensional motor neuron-Schwann cell coculture model on a microfluidic biochip.

Axonal regeneration and remyelination of peripheral motor neurons (MNs) are critical for restoring neuromuscular motor function after injury or peripheral neuropathy. We examined whether optogenetically mediated light stimulation (OMLS) could enhance the axon outgrowth and myelination of MNs using three-dimensional motor neuron-Schwann cell (MN-SC) coculture on a microfluidic biochip. The biochip was designed to allow SCs to interact with the axons of MNs, while preventing direct contact between SCs and the cell bodies of MNs. Following coculture with SCs on the microfluidic biochip, MNs were transfected with a light-sensitive channelrhodopsin gene. Transfected MNs subjected to repeated light stimulation (20 Hz, 1 hr) produced significantly longer axons than nontransfected MNs. OMLS of MNs greatly increased the number of myelin basic protein (MBP)-expressing SCs, promoting the initiation of myelination of MNs. Ultrastructurally, OMLS of MNs markedly enhanced the thickness of the compact myelin sheath around the MN axons such that the average thickness was closer to that of the theoretical estimates in vivo. Thus, the MN-SC coculture model on a microfluidic biochip augmented by OMLS of MNs is a feasible platform for studying the relationship of neuronal activity with regrowth and remyelination.

Three-dimensional reconstructive kidney volume analyses according to the endophytic degree of tumors during open partial or radical nephrectomy.

OBJECTIVES: To investigate the renal function outcomes and contralateral kidney volume change measured by using a 3-dimensional reconstructive method after open partial nephrectomy (PN) or open radical nephrectomy (RN) according to the endophytic degree of tumors. MATERIALS AND METHODS: We included 214 PN and 220 RN patients. According to the endophytic degree of the tumors, we divided patients into 3 groups. Patients were assessed for renal function and kidney volume change both preoperatively and postoperatively at 6 months. Kidney volume was calculated by using personal computer-based software. Subgroup analyses was performed for tumor >4cm. RESULTS: Larger and complex tumors were more frequent in the RN group than PN group. Among patients with exophytic and mild endophytic tumors, the mean postoperative renal function was well preserved in PN group and the mean contralateral kidney volume significantly increased in the RN compared to the PN group (PN, 145.55 to 149.98mL; 3.0% versus RN, 143.93 to 169.64mL;17.9% p=0.006). However, in fully endophytic tumors, compensatory hypertrophy of the contralateral kidney was similar between PN and RN (PN, 138.16 to 159.64mL; 15.5% versus RN, 138.65 to 168.04mL; 21.2% p=0.416) and renal functional outcomes were similar between both groups. These results were also confirmed in tumors >4cm in size. CONCLUSIONS: In fully endophytic tumors, especially large tumors, the postoperative renal function and contralateral kidney volume were similar; therefore, we should consider RN preferentially as surgical option for these tumors.

Phasing and climate forcing potential of the Millennium Eruption of Mt. Baekdu.

The Millennium Eruption of Mt. Baekdu, one of the largest volcanic eruptions in the Common Era, initiated in late 946. It remains uncertain whether its two main compositional phases, rhyolite and trachyte, were expelled in a single eruption or in two. Investigations based on proximal and medial ash have not resolved this question, prompting us to turn to high-resolution ice-core evidence. Here, we report a suite of glaciochemical and tephra analyses of a Greenlandic ice core, identifying the transition from rhyolitic to trachytic tephra with corresponding spikes in insoluble particle fallout. By modeling annual snow accumulation, we estimate an interval of one to two months between these spikes, which approximates the hiatus between two eruptive phases. Additionally, negligible sulfur mass-independent fractionation, near-synchroneity between particle and sulfate deposition, and peak sulfur fallout in winter all indicate an ephemeral aerosol veil. These factors limited the climate forcing potential of the Millennium Eruption.

Angio-Net: deep learning-based label-free detection and morphometric analysis of in vitro angiogenesis.

Despite significant advancements in three-dimensional (3D) cell culture technology and the acquisition of extensive data, there is an ongoing need for more effective and dependable data analysis methods. These concerns arise from the continued reliance on manual quantification techniques. In this study, we introduce a microphysiological system (MPS) that seamlessly integrates 3D cell culture to acquire large-scale imaging data and employs deep learning-based virtual staining for quantitative angiogenesis analysis. We utilize a standardized microfluidic device to obtain comprehensive angiogenesis data. Introducing Angio-Net, a novel solution that replaces conventional immunocytochemistry, we convert brightfield images into label-free virtual fluorescence images through the fusion of SegNet and cGAN. Moreover, we develop a tool capable of extracting morphological blood vessel features and automating their measurement, facilitating precise quantitative analysis. This integrated system proves to be invaluable for evaluating drug efficacy, including the assessment of anticancer drugs on targets such as the tumor microenvironment. Additionally, its unique ability to enable live cell imaging without the need for cell fixation promises to broaden the horizons of pharmaceutical and biological research. Our study pioneers a powerful approach to high-throughput angiogenesis analysis, marking a significant advancement in MPS.

The Prevalence of Sexually Transmitted Infections and Sociosexual Behaviors in the South Korean Military Before and During the COVID-19 Pandemic.

INTRODUCTION: This study evaluated the epidemiological factors of sexually transmitted infections (STIs) among South Korean troops including the prevalence, therapeutic methods, and sexual risk behaviors. MATERIAL AND METHODS: The medical records of the STIs diagnosed troops at the Armed Forces Capital Hospital (AFCH) for 36 months (between January 2018 and December 2020) were retrospectively reviewed. The data collection for the study began after obtaining research approvals from the institutional ethics committee of AFCH. The patients were classified into two subgroups, pre-coronavirus disease 2019 (COVID-19) and COVID-19 groups. The clinical parameters of the patients including STI-related symptoms and underlying diseases were analyzed. The sociosexual conduct of the two study groups was evaluated and compared by using a survey questionnaire. RESULTS: Overall, 138 STI patients with mean age of 21.2 years were included (pre-COVID-19: 106 patients/COVID-19: 32 patients). 32.6% of the patients received college education before the military service. Regarding previous history of STIs, 24 patients (17.4%) had previous experience of STIs, which occurred before participation in the current study. Initial urine analysis results showed that 95 (68.8%) and 79 patients (57.2%) showed pyuria and bacteriuria, respectively. Neisseria gonorrhoeae (29.7%) was the most commonly identified pathogen. Each pathogen was treated with the therapies recommended by the current treatment guidelines, and no patient showed any recurrence of the disease during follow-up. Both pre-COVID-19 (91.5%) and COVID-19 (93.8%) groups showed high rates of binge drinking during off-duty. The pre-COVID-19 group had a greater number of patients (21.7%) having multiple sex partners (during the past 12 months) than the COVID-19 group (15.6%). The COVID-19 group had 18.8% of the troops involved in sexual activity even after the onset of STI-related clinical symptoms, whereas the rate was significantly higher than 2.8% of the pre-COVID-19 group (P = .001). The COVID-19 groups showed a significantly higher number of patients (four patients, 12.5%) experiencing suicidal ideation than the pre-COVID-19 group (two patients, 1.9%) (P = .010). Both groups showed <40% of condom use rates at the last sexual intercourse. CONCLUSION: The soldiers with STIs showed high rates of binge alcohol consumption, while the rates of engaging in safe sex by using condoms are markedly low. Although the COVID-19 group was under influence of social distancing and military base lockdown, the soldiers' sociosexual conduct was not significantly different in the pre-COVID-19 group. The importance of education on alcohol misuse and safe sexual relationships should be taken more seriously within the military.

A dual-center study of predictive factors for sperm retrieval through microdissection testicular sperm extraction and intracytoplasmic sperm injection outcomes in men with non-mosaic Klinefelter syndrome.

PURPOSE: This study evaluated the predictors of sperm retrieval (SR) in non-mosaic Klinefelter syndrome (KS) patients undergoing microsurgical testicular sperm extraction (mTESE). The cutoff values of the predictors of SR and overall pregnancy rates after intracytoplasmic sperm injection (ICSI) were analyzed for the positive SR (PSR) cases. MATERIALS AND METHODS: The study was a dual-center retrospective study. Overall 118 patients with KS underwent mTESE between January 2011 and July 2021. Clinicopathological factors including comorbidities, endocrine profiles, and testicular volumes were analyzed. RESULTS: A total of 58 patients showed PSR (49.2%) and 60 patients (50.8%) had negative SR (NSR). The mean overall age of the patients was 32.5 years. The NSR patients had a significantly greater prevalence of obesity, diabetes mellitus, and cerebrovascular disease. The PSR group had a significantly higher left testis mean volume (p=0.039). The differences between the two study groups regarding follicular-stimulating hormone, luteinizing hormone, and testosterone variations at 1 and 3 months after mTESE were insignificant. Preoperative mean neutrophil-to-lymphocyte ratio was significantly greater in the NSR group (p=0.011), but the platelet-to-lymphocyte ratio showed no significant difference between the two study groups. A live child birth was achieved in 53.4% of the PSR patients. Multivariate logistic analysis showed that total testicular volume >3.93 mL, left testis volume >1.79 mL, and neutrophil-to-lymphocyte ratio ≤1.82 were significantly associated with PSR. CONCLUSIONS: mTESE-ICSI is a feasible method for KS patients to have a child, and total testicular volume, left testis volume, and neutrophil-to-lymphocyte ratio might be predictors of successful SR.

Relationship among total kidney volume, renal function and age.

PURPOSE: We measured kidney volume using software and investigated the relationship between kidney volume and renal function. MATERIALS AND METHODS: Age, gender, height, body weight, body mass index, body surface area and serum creatinine were recorded for 539 normal individuals. A tissue segmentation tool program was used to measure kidney volume from computerized tomography images. The glomerular filtration rate was calculated using the Cockcroft-Gault equation and an abbreviated modification of diet in renal disease equation. We looked for correlations of renal parenchymal volume with age and anthropometric measurements. We also tested for a correlation between kidney volume and renal function using the glomerular filtration rate according to the Cockcroft-Gault and modification of diet in renal disease equations. RESULTS: Mean kidney volume in all participants was 261.3 ± 58.1 ml. Mean volume in men was approximately 14 ml greater than in women (266.1 vs 251.8 ml, p = 0.004). Kidney volume correlated significantly with height (r = 0.344, p <0.001), body weight (r = 0.343, p <0.001), body mass index (r = 0.177, p <0.001), body surface area (r = 0.371, p <0.001) and age (r = -0.418, p <0.001). Kidney volume also correlated with the glomerular filtration rate according to the Cockcroft-Gault and modification of diet in renal disease equations (p <0.001, r = 0.615 and p <0.001, r = 0.432, respectively). Kidney volume and the glomerular filtration rate decreased in parallel with increasing age. CONCLUSIONS: Kidney volume correlates well with renal function and anthropometric measurements. Knowledge of these relationships will be valuable in clinical urology and nephrology.

Prognostic significance of preoperative kidney volume for predicting renal function in renal cell carcinoma patients receiving a radical or partial nephrectomy.

UNLABELLED: Study Type - Prognosis (case series) Level of Evidence 4 What's known on the subject? and What does the study add? At present, many studies have been executed to identify predictors for chronic kidney disease or renal insufficiency after radical nephrectomy or partial nephrectomy. We examined whether preoperative kidney volume is a predictor for renal function after radical or partial nephrectomies in RCC patients. To our knowledge, this is the first study to report on the relationship between preoperative kidney volume and changes in renal function in RCC patients who underwent radical nephrectomy or partial nephrectomy performed by one surgeon. OBJECTIVE: To investigate whether preoperative kidney volume is a prognostic factor for predicting the postoperative glomerular filtration rate (GFR) in renal cell carcinoma (RCC) patients. PATIENTS AND METHODS: We included 133 patients who underwent radical (n= 83) or partial (n= 50) nephrectomy for RCC. Kidney parenchymal volume was measured using personal computer-based software and GFR was estimated before and after surgery at 6 and 12 months. We evaluated the change in kidney volume after radical and partial nephrectomy and used regression analysis to identify predictors of lower post-surgical GFR at 12 months. RESULTS: The mean volume of the normal side kidney for the radical nephrectomy group increased from 142.4 mL to 166.0 mL (17.2%) and 171.5 mL (21.2%) after surgery at 6 and 12 months, respectively. In the partial nephrectomy group, the volume of the normal side kidney increased from 127.2 mL to 138.8 mL (9.1%) and 140.6 mL (10.9%) after surgery at 6 and 12 months, respectively. The volume of the operated side kidney decreased from 128.5 mL to 102.3 mL (20.1%) and 101.8 (20.6%) after surgery at 6 and 12 months, respectively. In the radical nephrectomy group, older age (P < 0.001), preoperative volume of the normal kidney (P= 0.022) and preoperative GFR for the normal side kidney (P= 0.045) were significant predictors of lower post-surgical GFR at 12 months. In the partial nephrectomy group, older age (P= 0.001) and preoperative volume for both kidneys (P= 0.037) were significant predictors of lower post-surgical GFR at 12 months. CONCLUSION: Preoperative kidney volume is an independent predictor of GFR in RCC patients who underwent radical or partial nephrectomy.

: A case report of re-challenge of immune checkpoint inhibitors after immune-related neurological adverse events: Review of literature.

INTRODUCTION: The indications for immune checkpoint inhibitors (ICIs) are expanding for various cancers because of their durable responses and tolerable safety profiles. Immune-related adverse events (irAEs), including neurological adverse events (nAEs), are associated with ICIs therapy. However, there have been few studies on whether re-challenge with ICIs can be clinically acceptable after neurological AE has improved. PATIENT CONCERNS: A 69-year-old woman with recurrent ovarian cancer undergoing palliative chemotherapy was admitted to our hospital with sudden development of diplopia, dizziness, and gait instability. The patient was administered ICI therapy with anti-angiogenic agents for 9 weeks for 3 cycles. DIAGNOSIS: We performed neurological examination, brain imaging, nerve conduction studies, and serology tests. The patient was diagnosed with Guillain-Barré syndrome variant, an immune-mediated polyneuropathy characterized by a triad of ataxia, areflexia, and ophthalmoplegia. INTERVENTION: After prompt discontinuation of pembrolizumab, the patient was taken intravenous methylprednisolone (2 mg/kg) was administered for 5 days, and her symptoms were partially resolved. With the addition of immunoglobulin 0.4 g/kg for 5 days, her symptoms gradually improved. OUTCOMES: The patient's neurological symptoms improved after immunosuppressive therapy, without sequelae. The NCV showed normal nerve conduction. Unfortunately, because there was little evidence for pembrolizumab rechallenge, pembrolizumab therapy was permanently discontinued, and the tumors eventually progressed.

Perfusable micro-vascularized 3D tissue array for high-throughput vascular phenotypic screening.

Microfluidic organ-on-a-chip technologies have enabled construction of biomimetic physiologically and pathologically relevant models. This paper describes an injection molded microfluidic platform that utilizes a novel sequential edge-guided patterning method based on spontaneous capillary flow to realize three-dimensional co-culture models and form an array of micro-vascularized tissues (28 per 1 × 2-inch slide format). The MicroVascular Injection-Molded Plastic Array 3D Culture (MV-IMPACT) platform is fabricated by injection molding, resulting in devices that are reliable and easy to use. By patterning hydrogels containing human umbilical endothelial cells and fibroblasts in close proximity and allowing them to form vasculogenic networks, an array of perfusable vascularized micro-tissues can be formed in a highly efficient manner. The high-throughput generation of angiogenic sprouts was quantified and their uniformity was characterized. Due to its compact design (half the size of a 96-well microtiter plate), it requires small amount of reagents and cells per device. In addition, the device design is compatible with a high content imaging machine such as Yokogawa CQ-1. Furthermore, we demonstrated the potential of our platform for high-throughput phenotypic screening by testing the effect of DAPT, a chemical known to affect angiogenesis. The MV-IMPACT represent a significant improvement over our previous PDMS-based devices in terms of molding 3D co-culture conditions at much higher throughput with added reliability and robustness in obtaining vascular micro-tissues and will provide a platform for developing applications in drug screening and development.

Combination of Abemaciclib following Eribulin Overcomes Palbociclib-Resistant Breast Cancer by Inhibiting the G2/M Cell Cycle Phase.

Breast cancer remains a leading cancer burden among women worldwide. Acquired resistance of cyclin-dependent kinase (CDK) 4/6 inhibitors occurs in almost all hormone receptor (HR)-positive subtype cases, comprising 70% of breast cancers, although CDK4/6 inhibitors combined with endocrine therapy are highly effective. CDK4/6 inhibitors are not expected to cooperate with cytotoxic chemotherapy based on the basic cytotoxic chemotherapy mode of action that inhibits rapidly proliferating cells. The palbociclib-resistant preclinical model developed in the current study investigated whether the combination of abemaciclib, CDK4/6 inhibitor with eribulin, an antimitotic chemotherapy could be a strategy to overcome palbociclib-resistant HR-positive breast cancer. The current study demonstrated that sequential abemaciclib treatment following eribulin synergistically suppressed CDK4/6 inhibitor-resistant cells by inhibiting the G2/M cell cycle phase more effectively. The current study showed the significant association of the pole-like kinase 1 (PLK1) level and palbociclib resistance. Moreover, the cumulative PLK1 inhibition in the G2/M phase by each eribulin or abemaciclib proved to be a mechanism of the synergistic effect. The synergistic antitumor effect was also supported by in vivo study. The sequential combination of abemaciclib following eribulin merits further clinical trials to overcome resistance to CDK4/6 inhibitors in HR-positive breast cancer.

U-IMPACT: a universal 3D microfluidic cell culture platform.

The development of organs-on-a-chip has resulted in advances in the reconstruction of 3D cellular microenvironments. However, there remain limitations regarding applicability and manufacturability. Here, we present an injection-molded plastic array 3D universal culture platform (U-IMPACT) for various biological applications in a single platform, such as cocultures of various cell types, and spheroids (e.g., tumor spheroids, neurospheres) and tissues (e.g., microvessels). The U-IMPACT consists of three channels and a spheroid zone with a 96-well plate form factor. Specifically, organoids or spheroids (~500 µm) can be located in designated areas, while cell suspensions or cell-laden hydrogels can be selectively placed in three channels. For stable multichannel patterning, we developed a new patterning method based on capillary action, utilizing capillary channels and the native contact angle of the materials without any modification. We derived the optimal material hydrophilicity (contact angle of the body, 45-90°; substrate, <30°) for robust patterning through experiments and theoretical calculations. We demonstrated that the U-IMPACT can implement 3D tumor microenvironments for angiogenesis, vascularization, and tumor cell migration. Furthermore, we cultured neurospheres from induced neural stem cells. The U-IMPACT can serve as a multifunctional organ-on-a-chip platform for high-content and high-throughput screening.

Standardized video-assisted retroperitoneal minilaparotomy surgery for 615 living donor nephrectomies.

To increase the rate of living kidney donation, the long-term safety of nephrectomy must be demonstrated to potential donors. We analyzed long-term donor outcomes and evaluated the standardization of surgical technique. We evaluated 615 donors who underwent Video-assisted minilaparotomy living donor nephrectomy (VLDN) at Yonsei Severance Hospital between 2003 and 2009. Perioperative data and predictors of outcomes were prospectively analyzed. The mean operative time and mean warm ischemia time were 192.7 and 2.2 min, respectively. Mean estimated blood loss was 195.3 ml. The mean post-transplant serum creatinine levels and Modification of Diet in Renal Disease study equation for estimating glomerular filtration rate were 1.1 mg/dl and 68 ml/min/1.73 m(2) , respectively at 5 years after VLDN. The intra-operative and postoperative complication rate were 3.1% and 6.3%, respectively. Delayed renal function, 5-year graft survival, and complication rates of recipients were 1.1%, 98.4%, and 0.4%, respectively. Predictors of operative time were medical history, vessel anomaly, and surgeon experience (>50 cases). The single predictor of intra-operative complications was vessel anomaly. Standardized VLDN is feasible and safe. Our data on long-term outcomes can assist in demonstrating the long-term safety of donor nephrectomy to potential donors. To compare VLDN to other types of donor nephrectomy, a prospective multicenter study must be performed.

High-throughput injection molded microfluidic device for single-cell analysis of spatiotemporal dynamics.

Single-cell level analysis of various cellular behaviors has been aided by recent developments in microfluidic technology. Polydimethylsiloxane (PDMS)-based microfluidic devices have been widely used to elucidate cell differentiation and migration under spatiotemporal stimulation. However, microfluidic devices fabricated with PDMS have inherent limitations due to material issues and non-scalable fabrication process. In this study, we designed and fabricated an injection molded microfluidic device that enables real-time chemical profile control. This device is made of polystyrene (PS), engineered with channel dimensions optimized for injection molding to achieve functionality and compatibility with single cell observation. We demonstrated the spatiotemporal dynamics in the device with computational simulation and experiments. In temporal dynamics, we observed extracellular signal-regulated kinase (ERK) activation of PC12 cells by stimulating the cells with growth factors (GFs). Also, we confirmed yes-associated protein (YAP) phase separation of HEK293 cells under stimulation using sorbitol. In spatial dynamics, we observed the migration of NIH 3T3 cells (transfected with Lifeact-GFP) under different spatiotemporal stimulations of PDGF. Using the injection molded plastic devices, we obtained comprehensive data more easily than before while using less time compared to previous PDMS models. This easy-to-use plastic microfluidic device promises to open a new approach for investigating the mechanisms of cell behavior at the single-cell level.