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Distant metastasis is the primary cause of unsuccessful treatment in nasopharyngeal carcinoma (NPC), suggesting the crucial need to comprehend this process. A tumor related to NPC does not have flat surfaces, but consists of confined microenvironments, proteins, and surface topography. To mimic the complex microenvironment, three-dimensional platforms with microwells and connecting channels were designed and developed with a fibronectin (FN) coating or nanohole topography. The potential of the transforming growth factor-β (TGF-β) inhibitor (galunisertib) for treating NPC was also investigated using the proposed platform. Our results demonstrated an increased traversing probability of NPC43 cells through channels with an FN coating, which correlated with enhanced cell motility and dispersion. Conversely, the presence of nanohole topography patterned on the platform bottom and the TGF-β inhibitor led to a reduced cell traversing probability and decreased cell motility, likely due to the decrease in the F-actin concentration in NPC43 cells. This study highlights the significant impact of confinement levels, surface proteins, nanotopography, and the TGF-β inhibitor on the metastatic probability of cancer cells, providing valuable insights for the development of novel treatment therapies for NPC. The developed platforms proved to be useful tools for evaluating the metastatic potential of cells and are applicable for drug screening.

This study investigates the oxygen (O 2 ) consumption of single cells during changes in their migration direction. This is the first integration of nanotopographies with an O 2 biosensor in a platform, allowing the real-time monitoring of O 2 consumption in cells and the ability to distinguish cells migrating in the same direction from those migrating in the opposite direction. Advanced nanofabrication technologies were used to pattern nanoholes or nanopillars on grating ridges, and their effects were evaluated using fluorescence microscopy, cell migration assays, and O 2 consumption analysis. The results revealed that cells on the nanopillars over grating ridges exhibited an enhanced migration motility and more frequent directional changes. Additionally, these cells showed an increased number of protrusions and filopodia with denser F-actin areas and an increased number of dotted F-actin structures around the nanopillars. Dynamic metabolic responses were also evident, as indicated by the fluorescence intensity peaks of platinum octaethylporphyrin ketone dye, reflecting an increased O 2 consumption and higher mitochondria activities, due to the higher energy required in response to directional changes. The study emphasizes the complex interplay between O 2 consumption and cell migration directional changes, providing insights into biomaterial science and regenerative medicine. It suggests innovative designs for biomaterials that guide cell migration and metabolism, advocating nanoengineered platforms to harness the intricate relationships between cells and their microenvironments for therapeutic applications.

One third of people with psychosis become antipsychotic treatment-resistant and the underlying mechanisms remain unclear. We investigated whether altered cognitive control function is a factor underlying development of treatment resistance. We studied 50 people with early psychosis at a baseline visit (mean < 2 years illness duration) and follow-up visit (1 year later), when 35 were categorized at treatment-responsive and 15 as treatment-resistant. Participants completed an emotion-yoked reward learning task that requires cognitive control whilst undergoing fMRI and MR spectroscopy to measure glutamate levels from Anterior Cingulate Cortex (ACC). Changes in cognitive control related activity (in prefrontal cortex and ACC) over time were compared between treatment-resistant and treatment-responsive groups and related to glutamate. Compared to treatment-responsive, treatment-resistant participants showed blunted activity in right amygdala (decision phase) and left pallidum (feedback phase) at baseline which increased over time and was accompanied by a decrease in medial Prefrontal Cortex (mPFC) activity (feedback phase) over time. Treatment-responsive participants showed a negative relationship between mPFC activity and glutamate levels at follow-up, no such relationship existed in treatment-resistant participants. Reduced activity in right amygdala and left pallidum at baseline was predictive of treatment resistance at follow-up (67% sensitivity, 94% specificity). The findings suggest that deterioration in mPFC function over time, a key cognitive control region needed to compensate for an initial dysfunction within a social-emotional network, is a factor underlying development of treatment resistance in early psychosis. An uncoupling between glutamate and cognitive control related mPFC function requires further investigation that may present a future target for interventions., (© 2024. The Author(s).)

Background: Several observational studies have documented a potential link between obesity and peripheral artery disease (PAD), although conflicting findings exist. The causal relationship between obesity and PAD continues to be a subject of ongoing debate in the medical community., Objectives: In this study, we employed a bidirectional Mendelian randomization (MR) analysis to explore the potential causal relationship between obesity and the risk of PAD., Methods: To investigate these causal relationships, we conducted bidirectional MR analysis using publicly available genome-wide association study (GWAS) data. Effect estimates were calculated using the random-effects inverse variance-weighted (IVW) method., Results: We identified eight independent single nucleotide polymorphisms (SNPs) associated with obesity in 218,735 samples involving 16,380,465 SNPs, all of which met the genome-wide significance threshold ( p  < 5 × 10 - ⁸). The IVW analysis indicates a significant positive association between genetic obesity and multiple datasets with PAD as the outcome: Queue-1 (GWAS ID: finn-b-I9&#95;PAD) (OR = 1.138, 95% CI: 1.027-1.261, p  = 0.013), Queue-2 (GWAS ID: bbj-a-144) (OR = 1.190, 95% CI: 1.019-1.390, p  = 0.028), Queue-3 (GWAS ID: ebi-a-GCST90018670) (OR = 1.174, 95% CI: 1.014-1.360, p  = 0.032), and Queue-4 (GWAS ID: ebi-a-GCST90018890) (OR = 1.194, 95% CI: 1.099-1.296, p  < 0.001). However, we did not observe a significant genetic-level association between obesity and PAD for Queue-5 (GWAS ID: ukb-d-I9&#95;PAD) (OR = 1.001, 95% CI: 1.000-1.002, p  = 0.071). Furthermore, we conducted a reverse causal MR analysis to explore the potential reverse causal relationship between obesity and PAD. This comprehensive analysis did not provide evidence of a reverse causal association between these two factors., Conclusions: In summary, our study offers genetic evidence suggesting a possible causal link between obesity and PAD. While we did not find evidence supporting the "obesity paradox", prudent weight management remains crucial, as lower weight does not necessarily guarantee better outcomes. As with any study, caution is required in interpreting the findings. Further research is essential to assess the clinical relevance of weight in preventing PAD, which could inform the development of more precise intervention strategies., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (© 2024 Huang, Pang, Zhang and Huang.)

In the context of wound healing and tissue regeneration, precise control of cell migration direction is deemed crucial. To address this challenge, polydimethylsiloxane (PDMS) platforms with patterned 10 nm thick TiO x in arrowhead shape were designed and fabricated. Remarkably, without tall sidewall constraints, MC3T3-E1 cells seeded on these platforms were constrained to migrate along the tips of the arrowheads, as the cells were guided by the asymmetrical arrowhead tips which provided large contact areas. To the best of our knowledge, this is the first study demonstrating the use of thin TiO x arrowhead pattern in combination with a cell-repellent PDMS surface to provide guided cell migration unidirectionally without tall sidewall constraints. Additionally, high-resolution fluorescence imaging revealed that the asymmetrical distribution of focal adhesions, triggered by the patterned TiO x arrowheads with arm lengths of 10, 20, and 35 μm, promoted cell adhesion and protrusion along the arrowhead tip direction, resulting in unidirectional cell migration. These findings have important implications for the design of biointerfaces with ultrathin patterns to precisely control cell migration. Furthermore, microelectrodes were integrated with the patterned TiO x arrowheads to enable dynamic monitoring of cell migration using impedance measurement. This microfluidic device integrated with thin layer of guiding pattern and microelectrodes allows simultaneous control of directional cell migration and characterization of the cell movement of individual MC3T3-E1 cells, offering great potential for the development of biosensors for single-cell monitoring., (© 2024. The Author(s).)

Background: Colorectal cancer (CRC) is a global health problem. The gut microbiome is now recognized as an important underlying factor to the initiation and progression of CRC. Fusobacterium nucleatum (FN) is one of the most studied bacteria in the aetiology of CRC. This study provided cohort evidence on the association of FN infection with clinicopathologic features in CRC patients., Methods: We analysed the cancerous and adjacent non-cancerous formalin-fixed paraffin embedded (FFPE) tissue of 83 CRC patients from a single medical centre in Malaysia. TaqMan probe-based qPCR targeting the 16S rRNA gene was used to detect the presence of FN in the extracted FFPE DNA. The differences in FN expression between cancer and non-cancer tissues were evaluated. Association studies between FN infection in the tumour and relative FN abundance with available clinical data were conducted., Results: FN was more abundant in the cancerous tissue compared to non-cancerous tissue (p = 0.0025). FN infection in the tumour was significantly associated with lymph node metastasis (p = 0.047) and cancer staging (p = 0.032), but not with other clinicopathologic variables. In double-positive patients where FN was detected in both cancerous and non-cancerous tissue, the expression fold-change of FN, calculated using 2-ΔΔCT formula, was significantly higher in patients with tumour size equal to or greater than 5 cm (p = 0.033) and in KRAS-mutated patients (p = 0.046)., Conclusions: FN is enriched in CRC tumour tissue and is associated with tumour size, lymph node metastasis, cancer staging, and KRAS mutation in this single-centre small cohort study., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

Objective: While current research suggests potential value for docosahexaenoic acid (DHA) in the prevention and management of atopic dermatitis (AD), the causal relationship between DHA and AD remains unclear, and the underlying mechanisms are not well understood., Materials and Methods: To investigate the potential causal relationship between DHA and AD, as well as to explore potential mediating mechanisms, we employed the Mendelian randomization (MR) methods. To study these potential relationships, we conducted MR analysis using publicly available Genome-Wide Association Studies (GWAS) data. Effect estimates were computed using the random-effects inverse-variance weighted method., Results: Our study demonstrates a negative correlation between DHA levels and AD risk (OR: 0.915, 95% CI: 0.858-0.975, p=0.007). Furthermore, in MR analysis using tumor necrosis factor ligand superfamily member 14 (TNFSF14) levels as an outcome, DHA levels also show a negative association with TNFSF14 levels (OR: 0.933, 95% CI: 0.879-0.990, p=0.022). Subsequently, we performed further analysis to explore the relationship between TNFSF14 and AD risk, revealing a positive correlation (OR: 1.069, 95% CI: 1.005-1.137, p=0.033). This suggests a potential mediating role of TNFSF14 in the impact of DHA on AD risk., Conclusions: In summary, our study employs MR analysis to offer genetic evidence indicating a potential role of DHA in reducing the risk of AD, as well as opening avenues for further in-depth investigation into potential mechanisms. These findings emphasize the importance of ongoing research in this field.

The extracellular matrix (ECM) serves as a complex scaffold with diverse physical dimensions and surface properties influencing NPC cell migration. Polydimethylsiloxane (PDMS), a widely used biocompatible material, is hydrophobic and undesirable for cell seeding. Thus, the establishment of a biomimetic model with varied topographies and surface properties is essential for effective NPC43 cell separation from NP460 cells. This study explored how ECM surface properties influence NP460 and NPC43 cell behaviors via plasma treatments and chemical modifications to alter the platform surface. In addition to the conventional oxygen/nitrogen (O 2 /N 2 ) plasma treatment, O 2 and argon plasma treatments were utilized to modify the platform surface, which increased the hydrophilicity of the PDMS platforms, resulting in enhanced cell adhesion. (3-aminopropyl)triethoxysilane and fibronectin (FN) were used to coat the PDMS platforms uniformly and selectively. The chemical coatings significantly affected cell motility and spreading, as cells exhibited faster migration, elongated cell shapes, and larger spreading areas on FN-coated surfaces. Furthermore, narrower top layer trenches with 5 µm width and a lower concentration of 10 µg/mL FN were coated selectively on the platforms to limit NP460 cell movements and enhance NPC43 cell separation efficiency. A significantly high separation efficiency of 99.4% was achieved on the two-layer scaffold platform with 20/5 µm wide ridge/trench (R/T) as the top layer and 40/10 µm wide R/T as the bottom layer, coupling with 10 µg/mL FN selectively coated on the sidewalls of the top and bottom layers. This work demonstrated an innovative application of selective FN coating to direct cell behavior, offering a new perspective to probe into the subtleties of NPC cell separation efficiency. Moreover, this cost-effective and compact microsystem sets a new benchmark for separating cancer cells.

Investigating the effect of nanomedicines on cancer cell behavior in three-dimensional (3D) platforms is beneficial for evaluating and developing novel antitumor nanomedicines in vitro. While the cytotoxicity of nanomedicines on cancer cells has been widely studied on two-dimensional flat surfaces, there is little work using 3D confinement to assess their effects. This study aims to address this gap by applying PEGylated paclitaxel nanoparticles (PEG-PTX NPs) for the first time to treat nasopharyngeal carcinoma (NPC43) cells in 3D confinement consisting of microwells with different sizes and a glass cover. The cytotoxicity of the small molecule drug paclitaxel (PTX) and PEG-PTX NPs was studied in microwells with sizes of 50 × 50, 100 × 100, and 150 × 150 μm 2 both with and without a concealed top cover. The impact of microwell confinement with varying sizes and concealment on the cytotoxicity of PTX and PEG-PTX NPs was analyzed by assessing NPC43 cell viability, migration speed, and cell morphology following treatment. Overall, microwell isolation was found to suppress drug cytotoxicity, and differences were observed in the time-dependent effects of PTX and PEG-PTX NPs on NPC43 cells in isolated and concealed microenvironments. These results not only demonstrate the effect of 3D confinement on nanomedicine cytotoxicity and cell behaviors but also provide a novel method to screen anticancer drugs and evaluate cell behaviors in vitro.

The three-dimensional (3D) structure of the extracellular matrix and cell-cell contacts are two important cues to altering cell migration behavior and the tumor formation process. In this work, we designed and fabricated microwell arrays with a grating-patterned bottom in polydimethylsiloxane platforms to systematically study the effects of confinement, changes in topography, and cell-cell contacts on the migration behavior of nasopharyngeal carcinoma (NPC43) and immortalized nasopharyngeal epithelial (NP460) cells by time-lapse imaging. When two types of cells were co-cultured in microwells, the migration speed and spreading area of NPC43 cells were significantly increased, which might be attributed to the heterotypic cell-cell contacts with NP460 cells. On a flat surface, NPC43 cells could not form clusters due to the frequent interruptions by the active movements of NP460 cells. However, in 3D microwell arrays, clusters of NPC43 cells formed on the bottom surface while the majority of NP460 cells migrated onto the sidewalls. These cell clusters could be further processed to form spheroids for drug screening. These results also revealed that the 3D microenvironments and cell-cell contacts could have significant implications for NPC cell migration and initiation of tumor formation, which will provide insight for NPC progression and dissemination.

The extracellular matrix serves as structural support for cells and provides biophysical and biochemical cues for cell migration. Topography, material, and surface energy can regulate cell migration behaviors. Here, the responses of MC3T3-E1 cells, including migration speed, morphology, and spreading on various platform surfaces, were investigated. Polydimethylsiloxane (PDMS) micropost sensing platforms with nanopillars, silicon oxide, and titanium oxide on top of the microposts were fabricated, and the dynamic cell traction force during migration was monitored. The relationships between various platform surfaces, migration behaviors, and cell traction forces were studied. Compared with the flat PDMS surface, cells on silicon oxide and titanium oxide surfaces showed reduced mobility and less elongation. On the other hand, cells on the nanopillar surface showed more elongation and a higher migration speed than cells on silicon oxide and titanium oxide surfaces. MC3T3-E1 cells on microposts with nanopillars exerted a larger traction force than those on flat PDMS microposts and had more filopodia and long protrusions. Understanding the relationships between platform surface condition, migration behavior, and cell traction force can potentially lead to better control of cell migration in biomaterials capable of promoting tissue repair and regeneration., Competing Interests: Conflict of interestThe authors declare no competing interests., (© The Author(s) 2023.)

Colorectal cancer (CRC) remains among the most commonly diagnosed cancers and has been on the rise. It is also one of the most lethal diseases globally, being the third leading cause of cancerrelated death. Depending on the stages and disease conditions, CRC is treated by surgery, chemo-, radio-therapy, immunotherapy or in combination. However, these therapies have subpar results with unwanted side effects, hence continuous effort is ongoing to explore new type of therapeutic modalities. Among the sub-types of CRC, KRAS, BRAF and NRAS mutated CRC comprise approximately 43%, 10% and 3% of the total cases, respectively. These mutations are associated with tumour progression and anti-epidermal growth factor receptor (EGFR) treatment resistance. Due to their important role in CRC, these genes have thus become targets in the development of novel treatments. In this paper, we discuss the current and upcoming treatment on CRC by targeting these mutated genes, with more focus on KRAS and BRAF due to the higher occurrence of mutations in CRC.

Immunoassay for detailed analysis of immune-cancer intercellular interactions can achieve more promising diagnosis and treatment strategies for cancers including nasopharyngeal cancer (NPC). In this study, we report a microfluidic live-cell immunoassay integrated with a microtopographic environment to meet the rising demand for monitoring intercellular interactions in different tumor microenvironments. The developed assay allows: (1) coculture of immune cells and cancer cells on tunable (flat or micrograting) substrates, (2) simultaneous detection of different cytokines in a wide working range of 5-5000 pg/mL, and (3) investigation of migration behaviors of mono- and co-cultured cells on flat/grating platforms for revealing the topography-induced intercellular and cytokine responses. Cytokine monitoring was achieved on-chip by implementing a sensitive and selective microbead-based sandwich assay with an antibody on microbeads, target cytokines, and the matching fluorescent-conjugated detection antibody in an array of active peristaltic mixer-assisted cytokine detection microchambers. Moreover, this immunoassay requires a low sample volume down to 0.5 μL and short assay time (30 min) for on-chip cytokine quantifications. We validated the biocompatibility of the co-culture strategy between immune cells and NPC cells and compared the different immunological states of undifferentiated THP-1 monocytic cells or PMA-differentiated THP-1 macrophages co-culturing with NP460 and NPC43 on topographical and planar substrates, respectively. Hence, the integrated microfluidic platform provides an efficient, broad-range and precise on-chip cytokine detection approach, eliminates the manual sampling procedures and allows on-chip continuous cytokine monitoring without perturbing intercellular microenvironments on different topographical ECM substrates, which has the potential of providing clinical significance in early immune diagnosis, personalized immunotherapy, and precision medicine.

Considering cell migration is essential for understanding physiological processes and diseases. The vertical migration of cells in three dimensions is vital, but most previous studies on cell migration have only focused on two-dimensional horizontal migration. In this paper, cell migration in the vertical direction was studied. Barriers with a height of 1, 5, 10, and 25 µm with grating and arrows in channels as guiding patterns were fabricated. The effects of barrier height and guiding patterns on the vertical migration of MC3T3 cells were explored. The study revealed that taller barriers hinder vertical migration of MC3T3 cells, whereas grating and arrows in channels promote it. The time-lapse and micrograph images showed that as the barrier height increased, the cell climbing angle along the barrier sidewall decreased, and the time taken to climb over the barrier increased. These results indicate that taller barriers increase the difficulty of vertical migration by MC3T3 cells. To promote the vertical migration of MC3T3 cells, 10 µm tall barriers with 18° and 40° sloped sidewalls were fabricated. For barriers with 18° sloped sidewalls, the probability for MC3T3 cells to climb up and down the 10 µm tall barriers was 40.6% and 20.3%, respectively; this is much higher than the migration probability over vertical barriers. This study shows topographic guidance on the vertical migration of MC3T3 cells and broadens the understanding of cell migration in the vertical direction., (© 2022. The Author(s).)

Background: Positive symptoms of psychosis (e.g., hallucinations) often limit everyday functioning and can persist despite adequate antipsychotic treatment. We investigated whether poor cognitive control is a mechanism underlying these symptoms., Methods: 97 patients with early psychosis (30 with high positive symptoms (HS) and 67 with low positive symptoms (LS)) and 40 healthy controls (HC) underwent fMRI whilst performing a reward learning task with two conditions; low cognitive demand (choosing between neutral faces) and high cognitive demand (choosing between angry and happy faces - shown to induce an emotional bias). Decision and feedback phases were examined., Results: Both patient groups showed suboptimal learning behaviour compared to HC and altered activity within a core reward network including occipital/lingual gyrus (decision), rostral Anterior Cingulate Cortex, left pre-central gyrus and Supplementary Motor Cortex (feedback). In the low cognitive demand condition, HS group showed significantly reduced activity in Supplementary Motor Area (SMA)/pre-SMA during the decision phase whilst activity was increased in LS group compared to HC. Recruitment of this region suggests a top-down compensatory mechanism important for control of positive symptoms. With additional cognitive demand (emotional vs. neutral contrast), HS patients showed further alterations within a subcortical network (increased left amygdala activity during decisions and reduced left pallidum and thalamus activity during feedback) compared to LS patients., Conclusions: The findings suggest a core reward system deficit may be present in both patient groups, but persistent positive symptoms are associated with a specific dysfunction within a network needed to integrate social-emotional information with reward feedback., (Copyright © 2022. Published by Elsevier Inc.)

One of the greatest unmet needs hindering the successful treatment of nasopharyngeal carcinomas (NPCs) is for representative physiological and cost-effective models. Although Epstein-Barr virus (EBV) infection is consistently present in NPCs, most studies have focused on EBV-negative NPCs. For the first time, we established and analyzed three-dimensional (3D) spheroid models of EBV-positive and EBV-negative NPC cells and compared these to classical two-dimensional (2D) cultures in various aspects of tumor phenotype and drug responses. Compared to 2D monolayers, the 3D spheroids showed significant increases in migration capacity, stemness characteristics, hypoxia and drug resistance. Co-culture with endothelial cells, which mimics essential interactions in the tumor microenvironment, effectively enhanced spheroid dissemination. Furthermore, RNA sequencing revealed significant changes at the transcriptional level in 3D spheroids compared to expression in 2D monolayers. In particular, we identified known (VEGF, AKT and mTOR) and novel (Wnt-β-catenin and Eph-ephrin) cell signaling pathways that are activated in NPC spheroids. Targeting these pathways in 3D spheroids using FDA-approved drugs was effective in monoculture and co-culture. These findings provide the first demonstration of the establishment of EBV-positive and EBV-negative NPC 3D spheroids with features that resemble advanced and metastatic NPCs. Furthermore, we show that NPC spheroids have potential use in identifying new drug targets., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2021. Published by The Company of Biologists Ltd.)

To realize highly directional guidance for cell migration, both micro- and nano-scale topographies were studied to better understand and mimic the complex extracellular matrix environment. Polydimethylsiloxane-based platforms with micro- and nano-topographies were developed to systematically study their guidance effects on cell migration behaviors. Compared to microtopography such as flat surface or grating, nanotopographies such as nanoholes and nanopillars could promote the generation of filopodia and extension of long protrusions with increased migration speed for MC3T3-E1 cells. Although cells on the grating structures showed lower migration speed, more directional cell migration was achieved due to their anisotropic topography compared to nanohole or nanopillar arrays with isotropic structures. To further enhance the cell migration directionality, the nanotopographies were patterned in grating arrangements and the results showed that both nanoholes and nanopillars in grating arrangements introduced more directional cell migration compared to gratings. The effects of physical dimensions of the nanotopographies on cell migration were studied and the results showed that there was less cell elongation and less directional migration of the nanoholes in grating arrangements with increasing depth of nanoholes. However, the nanopillars in grating arrangements showed more cell elongation, more directional migration, and higher migration speed with increasing height of the nanopillars. Platforms with nanopillars in grating arrangements and large height could be used to control cell migration speed and directionality, which could potentially lead to cell sorting and screening.

Cell spreading and migration play a pivotal role in many diseases such as tumor metastasis. In particular, nasopharyngeal tumor cells have known of their tendency of migration to pterygoid muscles and further distant metastasis. Although existing studies revealed key characteristics of the nasopharyngeal tumor cells, their migration preference is yet to be thoroughly understood, especially in the physical aspects including the microtopographical factors. Researchers have developed techniques in recent years to study microtopography-related cell behaviors but they are not yet applied in investigating the nasopharyngeal tumor cells. In this work, we elaborate the spreading and migration characteristics of normal and cancerous nasopharyngeal cells on micrograting substrates mimicking the microtopography of myotubes of the pterygoid muscles. We further apply interference reflection microscopy (IRM) to visualize the cell-substrate adhesion dynamics. We are interested in examining the microtopography-related cell spreading and migration behaviors and their correlations, providing insights for deeper understanding and more promising prediction on the nasopharyngeal tumor metastasis.

Scaffold microstructures were developed to mimic a three-dimensional extracellular matrix in studying cell migration and invasion. The multiple-layer scaffold platforms were designed to investigate cell migration and separation from top to bottom layer. Two cell lines including immortalized nasopharyngeal epithelial (NP460) cells and nasopharyngeal carcinoma (NPC43) cells with Epstein-Barr virus were compared in this study. On one-layer platforms with trench depth of 15 µm, both NP460 and NPC43 cells were guided to migrate along the 18-µm-wide trenches, and exhibited random migration directions when the trench width was 10 or 50 µm. Nearly no cell was found to migrate in the 10-µm-wide trenches on one-layer platforms. However, the NP460 and NPC43 cells showed very different probability in the narrow trenches on two-layer platforms, making it possible to separate the nasopharyngeal epithelial cells from the carcinoma cells. Moreover, 1-µm deep grating topography on the top layer inhibited NP460 cells to migrate from top ridges to the 10-µm-wide trenches, but promoted such behavior for NPC43 cells. The results demonstrated in This study suggest that the engineered multiple-layer scaffold platforms could be used to separate carcinoma cells in NPC tumor as a potential treatment of NPC., (© 2020 Wiley Periodicals LLC.)

Purpose: T4-binding globulin (TBG) is the main thyroid hormone (TH) transporter present in human serum. Inherited thyroxine-binding globulin (TBG) deficiency is caused by mutations in the TBG (SERPINA7) gene, which is located on the X chromosome. This study was performed to report and evaluate coding region mutations in TBG gene for partial thyroxine-binding globulin deficiency., Methods: A pedigree spanning four generations is described in this study. The proband is a female with partial TBG deficiency. All members of this pedigree underwent thyroid function tests, while Sanger sequencing was used to identify the TBG gene mutations. Bioinformatics databases were used to evaluate the deleterious effects of the mutation(s). Two hundred and seven unrelated individuals were used to evaluate the thyroid function of individuals with different TBG mutations. A one-way ANOVA was used to analyze the impact of the TBG mutations on thyroid function., Results: TBG gene sequencing results revealed that the proband had a novel mutation in codon 27 leading to alanine to valine substitution (p.A27V). This mutation was associated with lower serum T4 levels (p < 0.0001) when compared to the groups that did not carry the mutation. The previously reported p.L283F mutation was also found in the proband. The hemizygous p.L283F individuals presenting with lower T4 serum and TBG levels (p < 0.001) when compared to wildtype males and females. Both mutations were deleterious upon SIFT and PolyPhen-2 evaluation., Conclusion: Associated with partial thyroxine-binding globulin deficiency, this study reports a novel p.A27V mutation in the TBG gene.

One of the most crucial components of regenerative medicine is the controlled differentiation of embryonic or adult stem cells into the desired cell lineage. Although most of the reported protocols of stem cell differentiation involve the use of soluble growth factors, it is increasingly evident that stem cells also undergo differentiation when cultured in the appropriate microenvironment. When cultured in decellularized tissues, for instance, stem cells can recapitulate the morphogenesis and functional specialization of differentiated cell types with speed and efficiency that often surpass the traditional growth factor-driven protocols. This suggests that the tissue microenvironment (TME) provides stem cells with a holistic "instructive niche" that harbors signals for cellular reprogramming. The translation of this into medical applications requires the decoding of these signals, but this has been hampered by the complexity of TME. This problem is often addressed by a reductionist approach, in which cells are exposed to substrates decorated with simple, empirically designed geometries, textures, and chemical compositions ("bottom-up" approach). Although these studies are invaluable in revealing the basic principles of mechanotransduction mechanisms, their physiological relevance is often uncertain. This review examines the recent progress of an alternative, "top-down" approach, in which the TME is treated as a holistic biological entity. This approach is made possible by recent advances in systems biology and fabrication technologies that enable the isolation, characterization, and reconstitution of TME. It is hoped that these new techniques will elucidate the nature of niche signals so that they can be extracted, replicated, and controlled. This review summarizes these emerging techniques and how the data they generated are changing our view on TME. Impact statement This review summarizes the current state of art of the understanding of instructive niche in the field of tissue microenvironment. Not only did we survey the use of different biochemical preparations as stimuli of stem cell differentiation and summarize the recent effort in dissecting the biochemical composition of these preparations, through the application of extracellular matrix (ECM) arrays and proteomics, but we also introduce the use of open-source, high-content immunohistochemistry projects in contributing to the understanding of tissue-specific composition of ECM. We believe this review would be highly useful for our peer researching in the same field. "Mr. Tulkinghorn is always the same… so oddly out of place and yet so perfectly at home." -Charles Dickens, Bleak House .

Cell migration is a fundamental process that is crucial for many biological functions in the body such as immune responses and tissue regeneration. Dysregulation of this process is associated with cancer metastasis. In this study, polydimethylsiloxane platforms with various topographical features were engineered to explore the influence of guiding patterns on MC3T3-E1 osteoblast cell migration. Focusing on the guiding effects of grating patterns, variations such as etch depth, pattern discontinuity, and bending angles were investigated. In all experiments, MC3T3-E1 cells on patterned surfaces demonstrated a higher migration speed and alignment when compared to flat surfaces. The study revealed that an increase in etch depth from 150 nm to 4.5 μm enhanced cell alignment and elongation along the grating patterns. In the presence of discontinuous elements, cell migration speed was accelerated when compared to gratings of the same etch depth. These results indicated that cell directionality preference was influenced by a high level of pattern discontinuity. On patterns with bends, cells were more inclined to reverse on 45° bends, with 69% of cells reversing at least once, compared to 54% on 135° bends. These results are attributed to cell morphology and motility mechanisms that are associated with surface topography, where actin filament structures such as filopodia and lamellipodia are essential in sensing the surrounding environment and controlling cell displacement. Knowledge of geometric guidance cues could provide a better understanding on how cell migration is influenced by extracellular matrix topography in vivo.

Tumour metastasis is a complex process that strongly influences the prognosis and treatment of cancer. Apart from intracellular factors, recent studies have indicated that metastasis also depends on microenvironmental factors such as the biochemical, mechanical and topographical properties of the surrounding extracellular matrix (ECM) of tumours. In this study, as a proof of concept, we conducted tumour spheroid dissemination assay on engineered surfaces with micrograting patterns. Nasopharyngeal spheroids were generated by the 3D culture of nasopharyngeal carcinoma (NPC43) cells, a newly established cell line that maintains a high level of Epstein-Barr virus, a hallmark of NPC. Three types of collagen I-coated polydimethylsiloxane (PDMS) substrates were used, with 15 μm deep "trenches" that grated the surfaces: (a) 40/10 μm ridges (R)/trenches (T), (b) 18/18 μm (R/T) and (c) 50/50 μm (R/T). The dimensions of these patterns were designed to test how various topographical cues, different with respect to the size of tumour spheroids and individual NPC43 cells, might affect dissemination behaviours. Spreading efficiencies on all three patterned surfaces, especially 18/18 μm (R/T), were lower than that on flat PDMS surface. The outspreading cell sheets on flat and 40/10 μm (R/T) surfaces were relatively symmetrical but appeared ellipsoid and aligned with the main axes of the 18/18 μm (R/T) and 50/50 μm (R/T) grating platforms. Focal adhesions (FAs) were found to preferentially formed on the ridges of all patterns. The number of FAs per spheroid was strongly influenced by the grating pattern, with the least FAs on the 40/10 μm (R/T) and the most on the 50/50 μm (R/T) substrate. Taken together, these data indicate a previously unknown effect of surface topography on the efficiency and directionality of cancer cell spreading from tumour spheroids, suggesting that topography, like ECM biochemistry and stiffness, can influence the migration dynamics in 3D cell culture models., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Purpose: Although important for apoptosis, the signaling pathway involving MOAP-1(Modulator of Apoptosis 1), RASSF1A (RAS association domain family 1A), and Bax (Bcl-2 associated X protein) is likely to be dysfunctional in many types of human cancers due to mechanisms associated with gene mutation and DNA hyper-methylation. The purpose of the present study was to assess the potential impact of generating physiologically relevant signaling pathway mediated by MOAP-1, Bax, and RASSF1A (MBR) in cancer cells and chemo-drug resistant cancer cells., Methods: The tricistronic expression construct that encodes MOAP-1, Bax, and RASSF1A (MBR) or its mutant, MOAP-1∆BH3L, Bax and RASSF1A (MBRX) was expressed from an IRES (Internal Ribosome Entry Site)-based tricistronic expression vector in human breast cancer cells, including MCF-7, MCF-7-CR (cisplatin resistant) and triple negative breast cancer cells, BMET05, for functional characterization through in vitro and in vivo models., Results: Transient expression of MBR potently promoted dose-dependent apoptotic signaling and chemo-sensitization in the cancer cells, as evidenced by loss of cell viability, nuclei condensation and Annexin-V positive staining while stable expression of MBR in MCF-7 cells significantly reduced the number of MBR stable clone by 86% and the stable clone exhibited robust chemo-drug sensitivity. In contrast, MBRX stable clone exhibited chemo-drug resistance while transiently over-expressed MOAP-1ΔBH3L inhibited the apoptotic activity of MBR. Moreover, the spheroids derived from the MBR stable clone displayed enhanced chemo-sensitivity and apoptotic activity. In mouse xenograft model, the tumors derived from MBR stable clone showed relatively high level of tumor growth retardation associated with the increase in apoptotic activity, leading to the decreases in both tumor weight and volume., Conclusions: Expression of MBR in cancer cells induces apoptotic cell death with enhanced chemo-sensitization requiring the BH3L domain of MOAP-1. In animal model, the expression of MBR significantly reduces the growth of tumors, suggesting that MBR is a potent apoptotic sensitizer with potential therapeutic benefits for cancer treatment.

In the present study, 3D biomimetic platforms were fabricated with guiding grating to mimic extracellular matrix topography, porous membrane to resemble the epithelial porous interface and trenches below to represent blood vessels as an in vitro tissue microenvironment. Fabrication technologies were developed to integrate the transparent biocompatible polydimethylsiloxane platforms with preciously controlled dimensions. Cell migration behaviors of an immortalized nasopharyngeal epithelial cell line (NP460) and a nasopharyngeal carcinoma cell line (NPC43) were studied on the 2D and 3D platforms. The NP460 and NPC43 cells traversing through the porous membrane and migrating in the trenches below were studied by time-lapse imaging. Before traversing through the pores, NP460 and NPC43 cells migrated around the pores but NPC43 cells had a lower migration speed with less lamellipodia spreading. After traversing to trenches below, NPC43 cells moved faster with an alternated elongated morphology (mesenchymal migration mode) and round morphology (amoeboid migration mode) compared with only mesenchymal migration mode for NP460 cells. The cell traversing probability through porous membrane on platforms with 30 μm wide trenches below was found to be the highest when the guiding grating was perpendicular to the trenches below and the lowest when the guiding grating was parallel to the trenches below. The present study shows important information on cell migration in complex 3D microenvironment with various dimensions and could provide insight for pathology and treatment of nasopharyngeal carcinoma., (© 2020 The Author(s).)

Filopodia are thin finger-like protrusions from cells and they are hard to detect using electrical, mechanical, or optical sensors because of their nanometer scale features. Besides, the signals from filopodia and the cell membrane are often mixed together which makes the detection of filopodia challenging. Here, a 3D nanoplasmonic biosensor with microposts is proposed to overcome these limitations. By using suitable chemical coating and physical dimensions, the signals from filopodia and the cell membrane were separated by having the microposts keep the cell membrane from making contact with the nanoplasmonic biosensor. The filopodia were detected by the 3D asymmetrical nanopillars with sharp Fano resonance. The sensitivity and figure of merit of the nanoplasmonic biosensor were 650 nm per refractive index unit and 28.3, respectively. A large peak shift of 6 nm was observed for the detection of MC3T3 osteoblastic cell filopodia at a concentration of 1300 cells per mm 2 . To the best of our knowledge, this is the first demonstration of filopodia detection using nanoplasmonic biosensors, where microposts were used to separate the cell membrane from filopodia and the 3D nanoplasmonic biosensors were used to monitor filopodia on the nanometer scale. These combined 3D micro- and nano-structures allow filopodia to be detected using different sensors without interference from the cell membrane.

Three-dimensional polydimethylsiloxane platforms were developed to mimic the extracellular matrix with blood vessels by having scaffolds with micropatterns, porous membrane and trenches. Precisely controlled physical dimensions, layouts, and topography as well as different surface chemical treatments were applied to study their influences on nasopharyngeal carcinoma cell (10-15 μm in diameter) migration in mimicked platforms over 15-hour of time-lapse imaging. By placing the pores at different distance from the edges of the trenches, pores with different trench sidewall exposures and effective sizes were generated. Pores right next to the trench sidewalls showed the highest cell traversing probability, most likely related to the larger surface contact area with cells along the sidewalls. Straight grating oriented perpendicular to trenches below the top layer increased cell traversing probability. Pore shape as well as pore size influenced the cell traversing probability and cells could not traverse through pores that were 6 μm or less in diameter, which is much smaller than the cell size. Trench depth of 15 μm could induce more cells to traverse through the porous membrane, while shallower trenches impeded cell traversing and longer time was needed for cells to traverse because 3 and 6 μm deep trenches were much smaller than cell size which required large cell deformation. Hydrophobic surface coating on the top layer and fibronectin in pores and trenches increased the cell traversing probability and reduced the pore size that cells could traverse from 8 to 6 μm, which indicated that cells could have larger deformation with certain surface coatings., Competing Interests: The authors have declared that no competing interests exist.

Cells in tissues are enveloped by an instructive niche made of the extracellular matrix. These instructive niches contain three general types of information: topographical, biochemical and mechanical. While the combined effects of these three factors are widely studied, the functions of each individual one has not been systematically characterised, because it is impossible to alter a single factor in a tissue microenvironment without simultaneously affecting the other two. Silica BioReplication (SBR) is a process that converts biological samples into silica, faithfully preserving the original topography at the nano-scale. We explored the use of this technique to generate inorganic replicas of intact mammalian tissues, including tendon, cartilage, skeletal muscle and spinal cord. Scanning electron and atomic force microscopy showed that the resulting replicas accurately preserved the three-dimensional ultrastructure of each tissue, while all biochemical components were eradicated by calcination. Such properties allowed the uncoupling the topographical information of a tissue microenvironment from its biochemical and mechanical components. Here, we showed that human mesenchymal stem cells (MSC) cultured on the replicas of different tissues displayed vastly different morphology and focal adhesions, suggesting that the topography of the tissue microenvironment captured by SBR could profoundly affect MSC biology. MSC cultured on tendon replica elongated and expressed tenocytes marker, while MSC on the spinal cord replica developed into spheroids that resembled neurospheres, in morphology and in the expression of neurosphere markers, and could be further differentiated into neuron-like cells. This study reveals the significance of topographical cues in a cell niche, as tissue-specific topography was sufficient in initiating and directing differentiation of MSC, despite the absence of any biochemical signals. SBR is a convenient and versatile method for capturing this topographical information, facilitating the functional characterisation of cell niches. STATEMENT OF SIGNIFICANCE: Various studies have shown that three major factors, topographical, biochemical and mechanical, in a tissue microenvironment (TME) are essential for cellular homeostasis and functions. Current experimental models are too simplistic to represent the complexity of the TME, hindering the detailed understanding of its functions. In particular, the importance each factor in a tissue microenvironment have not been individually characterised, because it is challenging to alter one of these factors without simultaneously affecting the other two. Silica bioreplication (SBR) is a process that converts biological samples into silica replicas with high structural fidelity. SBR is a convenient and versatile method for capturing this topographical information on to a biologically inert material, allowing the functional characterisation of the architecture of a TME., (Copyright © 2019. Published by Elsevier Ltd.)

Invadosomes are invasive protrusions generated by cells which can secrete matrix metalloproteinases for focal digestion of extracellular matrix. They also aid invasive cancer cells in their transmigration through vascular endothelium. However, how the physical and chemical cues in a three-dimensional (3D) system signal the spatial localization of invadosomes remains largely unknown. Here we study the topographic guidance of invadosome formation in invasive nasopharyngeal cells under the stimulation of an inflammatory cytokine, TGF-β1, using engineered gratings with different width and depth. We first report that TGF-β1 can act as an external signal to upregulate the formation of invadosomes with a random distribution on a plane 2D surface. When the cells were seeded on parallel 3D gratings of 5 µm width and 1 µm depth, most of the invadosomes aligned to the edges of the gratings, indicating a topographic cue to the control of invadosome localization. While the number of invadosomes per cell were not upregulated when the cells were seeded on 3D topography, guidance of invadosomes localization to edges is correlated with cell migration directionality on 1 µm deep gratings. Invadosomes preferentially form at edges when the cells move at a lower speed and are guided along narrow gratings. The invadosomes forming at 3D edges also have a longer half-life than those forming on a plane surface. These data suggest that there are integrated biochemical and 3D geometric cues underlying the spatial regulation of invasive structures so as to elicit efficient invasion or metastasis of cells. STATEMENT OF SIGNIFICANCE: Nasopharyngeal cells were integrated with the biological cues and matrix topography to govern the activity and spatial distribution of invadosomes. The biochemical induction of invadosome formation by TGF-β1 in nasopharyngeal cells was observed. When the cells were seeded on parallel 3D gratings, most of the invadosomes aligned to the edges of the gratings due to topographical induced invadosome localization. While the number of invadosomes per cell were not upregulated, guidance of invadosomes localization to edges is correlated with cell migration directionality on 1 µm deep gratings. Invadosomes preferentially form at edges with a higher stability when the cells are guided along narrow gratings. The integrated biochemical and 3D geometric cues could elicit efficient invasion or metastasis of cells., (Copyright © 2019. Published by Elsevier Ltd.)

Colorectal cancer (CRC) continues to be one of the most common cancers globally. The incidence has increased in developing countries in the past few decades, this could be partly attributed to aging populations and unhealthy lifestyles. While the treatment of CRC has seen significant improvement since the advent of target-specific therapies and personalized medicine, CRC is oftentimes detected at late or advanced stages, thereby reducing the efficacy of treatment. Hence, screening for early detection is still the key to combat CRC and to increase overall survival (OS). Considering that the field of medical diagnostics is moving towards molecular diagnostics, CRC can now be effectively screened and diagnosed with high accuracy and sensitivity. Depending on the tumor genotype and genetic profile of the individual, personalized treatments including tyrosine kinase inhibitor therapy and immunotherapy can be administered. Notably, there can be no one single treatment that is effective for all CRC patients due to the variation in tumor genetics, which highlights the importance of molecular diagnostics. This review provides insights on therapeutic modalities, molecular biomarkers, advancement of diagnostic technologies, and current challenges in managing CRC.

Implantable neural probes are widely used to record and stimulate neural activities. These probes should be stiff enough for insertion. However, it should also be flexible to minimize tissue damage after insertion. Therefore, having dynamic control of the neural probe shank flexibility will be useful. For the first time, we have successfully fabricated flexible neural probes with embedded microfluidic channels for dynamic control of neural probe stiffness by controlling fluidic pressure in the channels. The present hybrid neural probes consisted of polydimethylsiloxane (PDMS) and polyimide (PI) layers could provide the required stiffness for insertion and flexibility during operation. The PDMS channels were fabricated by reversal imprint using a silicon mold and bonded to a PI layer to form the embedded channels in the neural probe. The probe shape was patterned using an oxygen plasma generated by an inductively coupled plasma etching system. The critical buckling force of PDMS/PI neural probes could be tuned from 0.25-1.25 mN depending on the applied fluidic pressure in the microchannels and these probes were successfully inserted into a 0.6% agarose gel that mimicked the stiffness of the brain tissue. Polymer-based neural probes are typically more flexible than conventional metal wire-based probes, and they could potentially provide less tissue damage after implantation., Competing Interests: The authors have declared that no competing interests exist.

Natural killer (NK) cells are lymphocytes which play an important role in the immune system by recognizing and killing potentially malignant cells without antigen sensitization, and could be utilized in cancer therapy. NK cell migration is an essential process to find and kill target cells, which is well known to be driven by the chemotaxis effect. NK cells also experience a topographical effect induced by the extracellular matrix (ECM) during their migration. However, topographical effects on NK cell locomotion in three dimensional (3D) environments are not well studied yet. In this work, polydimethylsiloxane based platforms containing microchannels with different types of perturbations and decorated with various surface patterns were fabricated to systematically study the topographical effect on NK cell migration with and without the chemotaxis effect. The results showed that perturbation sites in channels induced pauses and reversals in chemotaxis driven NK cell migration. Surface topography such as gratings in confined environments could introduce directional preference to NK cell movement even without chemoattractants. These findings showed that NK cell migration could be controlled by contact guidance, which provides future possibility to manipulate NK cell migration in controlled in vitro bioengineering systems. Results in this study showed that the complex topography of 3D microenvironments in the ECM could have significant effects on NK cell migration in different tissues and organs, and provided insight for explaining the dynamics of NK cell activities in clinical experiments.

Three-dimensional (3D) cell migrations are regulated by force interactions between cells and a 3D extracellular matrix (ECM). Mapping the 3D traction force generated by cells on the surrounding ECM with controlled confinement and contact area will be useful in understanding cell migration. In this study, double-sided micropost arrays were fabricated. The cell traction force was mapped by microposts on the top and bottom of opposing surfaces with a controlled separating distance to create different confinements. The density of micropost arrays was modified to investigate the effect of cell contact area on 3D traction force development. Using MC3T3-E1 osteoblastic cells, the leading traction force was found to increase with additional contact surface on the top. Summing force vectors on both surfaces, a large force imbalance was found from the leading to trailing regions for fast migrating cells. With 10 μm separation and densely arranged microposts, the traction force on the top surface was the largest at 28.6 ± 2.5 nN with the highest migration speed of 0.61 ± 0.07 μm min -1 . Decreasing the density of the top micropost arrays resulted in a reduced traction force on the top and lower migration speed. With 15 μm separation, the cell traction force on the top and migration speed further decreased simultaneously. These results revealed traction force development on 3D ECM with varied degrees of confinement and contact area, which is important in regulating 3D cell migration., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

Understanding cell migration in a 3D microenvironment is essential as most cells encounter complex 3D extracellular matrix (ECM) in vivo Although interactions between cells and ECM have been studied previously on 2D surfaces, cell migration studies in 3D environment are still limited. To investigate cell migration under various degrees of confinements and coating conditions, 3D platforms with micropost arrays and controlled fibronectin (FN) protein coating were developed. MC3T3-E1 cells spread and contacted the top surface of microposts if FN was coated on top. When FN was coated all over the microposts, cells were trapped between microposts with 3 μm spacing and barely moved. As the spacing between microposts increased from 3 to 5 μm, cells became elongated with limited cell movement of 0.18 μm/min, slower than the cell migration speed of 0.40 μm/min when cells moved on top. When cells were trapped in between the microposts, cell nuclei were distorted and actin filaments formed along the sidewalls of microposts. With the addition of a top cover to introduce cell confinement, the cell migration speed was 0.23 and 0.84 μm/min when the channel height was reduced from 20 to 10 μm, respectively. Cell traction force was monitored at on the top and bottom microposts with 10 μm channel height. These results show that the MC3T3-E1 cell morphology, migration speed, and movement position were affected by surface coating and physical confinement, which will provide significant insights for in vivo cell migration within a 3D ECM., (© 2019 The Author(s).)

In this study, two-dimensional (2D), quasi-three-dimensional (3D), and 3D plasmonic photonic crystal (PPC) nanostructures with point-defect cavities were developed and fabricated using direct and reversal nanoimprint lithography. As a result of the hybrid coupling of localized surface plasmon resonance and Fabry-Perot cavity modes, the quasi-3D plasmonic nanoholes showed higher electromagnetic field intensity and sensitivity than the 2D plasmonic nanoholes. Specifically, the sensitivity of the quasi-3D plasmonic nanoholes was 483 nm per refractive index unit (RIU), whereas that of the 2D plasmonic nanoholes was 276 nm RIU-1. In addition, by enhancing electromagnetic field intensity around corners and generating an additional subradiant dark mode, the symmetrical breakage of the quasi-3D plasmonic nanoholes further increased the sensitivity to 946 nm RIU-1. Among all the nanostructures developed in the study, the 3D PPC nanostructures with point-defect cavities showed the highest sensitivity up to 1376 nm RIU-1 and highest figure of merit of 11.6 as the result of the hybrid coupling of plasmonics and photonic crystal modes with multilayered plasmonic nanostructures. The spacing between the 3D PPC nanostructures was comparable with the average size of exosomes derived from fibroblast L cells, which allowed the exosomes to spread around the 3D PPC nanostructures with increased sensing area. This effect further enhanced the detection sensitivity with a large peak shift of 9 nm when using the 3D PPC biosensor to detect exosomes at the concentration of 1 × 104 particles per ml, and the peak shift increased to 102 nm as exosome concentration increased to 1 × 1011 particles per ml.

Three-dimensional (3D) multilayered plasmonic nanostructures consisting of Au nanosquares on top of SU-8 nanopillars, Au asymmetrical nanostructures in the middle, and Au asymmetrical nanoholes at the bottom were fabricated through reversal nanoimprint technology. Compared with two-dimensional and quasi-3D plasmonic nanostructures, the 3D multilayered plasmonic nanostructures showed higher electromagnetic field intensity, longer plasmon decay length and larger plasmon sensing area, which are desirable for highly sensitive localized surface plasmonic resonance biosensors. The sensitivity and resonance peak wavelength of the 3D multilayered plasmonic nanostructures could be adjusted by varying the offset between the top and bottom SU-8 nanopillars from 31% to 56%, and the highest sensitivity of 382 and 442 nm/refractive index unit were observed for resonance peaks at 581 and 805 nm, respectively. Live lung cancer A549 cells with a low concentration of 5 × 10 3 cells ml -1 and a low sample volume of 2 μl could be detected by the 3D multilayered plasmonic nanostructures integrated in a microfluidic system. The 3D plasmonic biosensors also had the advantages of detecting DNA hybridization by capturing the complementary target DNA in the low concentration range of 10 -14 -10 -7 M, and providing a large peak shift of 82 nm for capturing 10 -7 M complementary target DNA without additional signal amplification.

Functional microbeads have been widely applied in molecular identification and other biochemical applications in the past decade, owing to the compatibility with flow cytometry and the commercially available microbeads for a wide range of molecular identification. Nevertheless, there is still a technical hurdle caused by the significant sample volume required (∼50  μ l), limited molecular detection limit (∼20 pg/ml), complicated liquid/microbead handling procedures, and the long reaction time (>2 h). In this work, we optimize the operation of an automated microbead-based microfluidic device for the reagent mixing and the dynamic cytokine detection. In particular, we adopt fluorescence microscopy for quantification of multiple microbeads in each microchamber instead of flow cytometry for a lower detection limit. The operation parameters are then configured for improved measurement performance. As demonstrated, we consider the cytokine secretion of human macrophage-differentiating lymphocytes stimulated by lipopolysaccharides. We examine requirements on the mixing duration, minimal sample volume, and the image analysis scheme for the smaller biosample volume (<5  μ l), the lower cytokine detection limit (∼5 pg/ml), and shorter process time (∼30 min). Importantly, this microfluidic strategy can be further extended in the molecular profiling using other functional microbeads for a broad range of biomedical applications.

The hand is commonly affected by osteoarthritis (OA). The development and progression of OA are believed to involve inflammation, even in the early stages of the disease. Inflammatory and proinflammatory cytokines have also been shown to be elevated in the flexor tenosynovium of idiopathic carpal tunnel syndrome (CTS). A large percentage of patients with hand OA also have a concomitant CTS. This study evaluated the results of a pulsed electrical joint stimulator in patients who had hand OA with or without CTS. Pain, tenderness, and swelling; grip strength and pinch force; and Patient and Physician Global Assessment and Disabilities of the Arm, Shoulder and Hand (DASH) results were evaluated. The primary efficacy outcome was pain due to OA in the study hand in the past 48 hours. Secondary outcomes consisted of OA pain in the study thumb in the past 48 hours, grip strength, pinch force, and Patient and Physician Global Assessment and DASH results. All 7 outcome parameters improved in OA patients. On physical examination, individual finger and wrist joints had also improved regarding pain, swelling, and tenderness. In the subset of patients with CTS, CTS pain, paresthesia, weakness, and all CTS symptoms had significantly improved. Patient and Physician Global Assessment and DASH results and pinch force were also significantly improved. This pulsed electrical joint stimulator is effective in providing clinically relevant and statistically significant reductions in the signs and symptoms of OA of the hand and CTS. It could be a useful modality for the treatment of patients who have one of these conditions or both. [Orthopedics. 2018; 41(4):e550-e556.]., (Copyright 2018, SLACK Incorporated.)

Paraneoplastic Ma Family (PNMA) comprises a growing number of family members which share relatively conserved protein sequences encoded by the human genome and is localized to several human chromosomes, including the X-chromosome. Based on sequence analysis, PNMA family members share sequence homology to the Gag protein of LTR retrotransposon, and several family members with aberrant protein expressions have been reported to be closely associated with the human Paraneoplastic Disorder (PND). In addition, gene mutations of specific members of PNMA family are known to be associated with human mental retardation or 3-M syndrome consisting of restrictive post-natal growth or dwarfism, and development of skeletal abnormalities. Other than sequence homology, the physiological function of many members in this family remains unclear. However, several members of this family have been characterized, including cell signalling events mediated by these proteins that are associated with apoptosis, and cancer in different cell types. Furthermore, while certain PNMA family members show restricted gene expression in the human brain and testis, other PNMA family members exhibit broader gene expression or preferential and selective protein interaction profiles, suggesting functional divergence within the family. Functional analysis of some members of this family have identified protein domains that are required for subcellular localization, protein-protein interactions, and cell signalling events which are the focus of this review paper., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Introduction: Continuous development of cell traction force can regulate cell migration on various extracellular matrixes in vivo . However, the topographical effect on traction force is still not fully understood., Methods: Micropost sensors with parallel guiding gratings were fabricated in polydimethylsiloxane to track the cell traction force during topographical guidance in real time. The force distributions along MC3T3-E1 mouse osteoblasts were captured every minute. The traction force in the leading, middle, and trailing regions was monitored during forward and reversed cell migration., Results: The traction force showed periodic changes during cell migration when the cell changed from elongated to contracted shape. For cell migration without guiding pattern, the leading region showed the largest traction force among the three regions, typically 5.8 ± 0.8 nanonewton (nN) when the cell contracted and 7.1 ± 0.5 nN when it elongated. During guided cell migration, a lower traction force was obtained. When a cell contracted, the trailing traction force was 4.1 ± 0.4 for non-guided migration and 2.2 ± 0.2 nN for guided migration. As a cell became elongated, the trailing traction force was 6.0 ± 0.5 nN during non-guided migration and 4.8 ± 0.3 nN under guidance. When a cell reversed its migration direction, the magnitudes of the traction force from the leading to the trailing regions also flipped., Conclusion: The cell traction force is continuously influenced by topographical guidance, which determines cell migration speed and direction. These results of cell traction force development on various topographies could lead to better cell migration control using topotaxis., (© Biomedical Engineering Society 2017.)

Gold (Au) resistive temperature sensors were integrated on flexible polyimide-based neural probes to monitor temperature changes during neural probe implantation and stimulation. Temperature changes were measured as neural probes were implanted to infer the positions of the neural probes, and as the retina or the deep brain region was stimulated electrically. The temperature sensor consisted of a serpentine Au resistor and surrounded by four Au electrodes with 200 and [Formula: see text] diameter (dia.). The Au temperature sensors had temperature coefficient of 0.32%, and they were biocompatible and small in size. In vivo measurements of temperature changes during implantation and stimulation were carried out in the retina and deep brain region in rats. The desired implantation position was reached when temperature measured by the sensor increased to the calibrated level and became stable. There was no temperature increase when low level stimulation current of 8 and [Formula: see text] each for the two 200- and 400- [Formula: see text]-dia. electrodes, respectively, were applied. When higher level stimulation current of 100 and [Formula: see text] each were applied to the two 200- and 400- [Formula: see text]-dia. electrodes, respectively, maximum temperature increases of 1.2 °C in retina and 1 °C in deep brain region were found.

In this study, we aimed at studying the effects of engineered and patterned substrates on the migration characteristics of mammalian cancer cell lines. On the shallow topographical patterns, cells from different histological origins showed different migration speed and directionality. We also observed that cells from the same origin showed distinctive behaviour, suggesting these substrate topographies could distinguish cancer subtypes. To eliminate the influence of genetic background, we examined two isogenic subpopulations of ovarian cancer cell lines for their different metastatic activities. While these cell lines showed indistinguishable migration characteristics on a flat substrate, their motilities on the patterned substrates were highly different, suggesting that cancer cells' motilities on these substrates varied in a metastasis-dependent manner. While cells with different metastatic activities showed similar morphology and focal adhesion distribution on flat surface, vinculin aggregated into single cytoplasmic foci in metastatic cells cultured on the engineered substrates. This implies that the topographical patterns on the substrates induced vinculin redistribution in cancer cells with a higher invasive activity. The fabricated platforms with topographical patterns offer a novel in vitro technique for metastasis assessment. Moreover, such platforms could potentially provide the opportunity to sort cells in different metastatic states using advanced pattern designs and features.

Natural killer (NK) cells serve an important role in immune system by recognizing and killing the potentially malignant cells without antigen sensitization, and could be promising in cancer therapy. We have designed and fabricated microwell arrays with microchannel connections in polydimethylsiloxane (PDMS) substrates to study the interaction dynamics of NK-92MI cells with MCF7 breast cancer cells using time-lapse imaging by fluorescence microscopy for 15 h. Although cell seeding density was the same, NK cell cytotoxicity was found to be higher in larger microwells, which is manifested as increased target death ratio from 13.7 ± 3.1 to 46.3 ± 3.3% and shorter triggering time of first target lysis from 502 ± 49 to 391 ± 63 min in 150 μm × 150 μm microwells comparing to 50 μm × 50 μm wells in 15 h. Mirochannel connection between adjacent microwells of the same size increased the overall target death ratio by >10%, while connection between microwells of different sizes led to significantly increased target death ratio and delayed first target lysis in smaller microwells. Our findings reveal unique cell interaction dynamics, such as initiation and stimulation, of NK cell cytotoxicity in a confined microenvironment, which is different from population-based study, and the results could lead to a better understanding of the dynamics of NK cell cytotoxicity.

While terahertz communications are considered to be the future solutions for the increasing demands on bandwidth, terahertz equivalents of radio frequency front-end components have not been realized. It remains challenging to achieve wideband, low profile antenna arrays with highly directive beams of radiation. Here, based on the complementary antenna approach, a wideband 2 × 2 cavity-backed slot antenna array with a corrugated surface is proposed. The approach is based on a unidirectional antenna with a cardiac radiation pattern and stable frequency characteristics that is achieved by integrating a series-resonant electric dipole with a parallel-resonant magnetic dipole. In this design, the slots work as magnetic dipoles while the corrugated surface radiates as an array of electric dipoles. The proposed antenna is realized at 1 THz operating frequency by stacking multiple metallized layers using the microfabrication technology. S-parameter measurements of this terahertz low-profile metallic antenna array demonstrate high efficiency at terahertz frequencies. Fractional bandwidth and gain are measured to be 26% and 14 dBi which are consistent with the simulated results. The proposed antenna can be used as the building block for larger antenna arrays with more directive beams, paving the way to develop high gain low-profile antennas for future communication needs.

Purpose: Members of paraneoplastic Ma (PNMA) family have been identified as onconeuronal antigens, which aberrant expressions in cancer cells of patients with paraneoplastic disorder (PND) are closely linked to manifestation of auto-immunity, neuro-degeneration, and cancer. The purpose of present study was to determine the role of PNMA5 and its functional relationship to MOAP-1 (PNMA4) in human cancer cells., Methods: PNMA5 mutants were generated through deletion or site-directed mutagenesis and transiently expressed in human cancer cell lines to investigate their role in apoptosis, subcellular localization, and potential interaction with MOAP-1 through apoptosis assays, fluorescence microscopy, and co-immunoprecipitation studies, respectively., Results: Over-expressed human PNMA5 exhibited nuclear localization pattern in both MCF-7 and HeLa cells. Deletion mapping and mutagenesis studies showed that C-terminus of PNMA5 is responsible for nuclear localization, while the amino acid residues (391KRRR) within the C-terminus of PNMA5 are required for nuclear targeting. Deletion mapping and co-immunoprecipitation studies showed that PNMA5 interacts with MOAP-1 and N-terminal domain of PNMA5 is required for interaction with MOAP-1. Furthermore, co-expression of PNMA5 and MOAP-1 in MCF-7 cells significantly enhanced chemo-sensitivity of MCF-7 to Etoposide treatment, indicating that PNMA5 and MOAP-1 interact synergistically to promote apoptotic signaling in MCF-7 cells., Conclusions: Our results show that PNMA5 promotes apoptosis signaling in HeLa and MCF-7 cells and interacts synergistically with MOAP-1 through its N-terminal domain to promote apoptosis and chemo-sensitivity in human cancer cells. The C-terminal domain of PNMA5 is required for nuclear localization; however, both N-and C-terminal domains of PNMA5 appear to be required for pro-apoptotic function.

Objectives: To investigate the influence of variations in resting pressure (precompression) on thyroid ultrasound supersonic shear wave elastography (SWE)., Methods: Thirty-five normal thyroid glands (Norm), 55 benign hyperplastic nodules (BHN), and 17 papillary thyroid cancers (PTC) in 96 subjects underwent thyroid SWE. Four precompression levels were applied manually by the operator, ranging from A (baseline, 0 % strain) to D (high, 22-30 % strain). SWE results at each precompression level were compared using ANOVA tests with P < 0.05 indicating significance., Results: SWE indices were highest in PTC, followed by BHN and Norm at each precompression level (P < 0.05). All tissue types showed successive increases in SWE results as precompression increased, although the rate was higher for PTC than BHN and Norm (Ps < 0.05). SWE values (kPa) of Norm, BHN, and PTC at baseline precompression (A) were 10.3 ± 3.3, 17.7 ± 7.6, and 22.2 ± 11.9 compared with 21.1 ± 4.2, 42.3 ± 16.0, and 97.6 ± 46.8 at high precompression (D). SWE index differences between precompression levels A and D were 10.8 kPa for Norm, 24.6 kPa for BHN, and 75.4 kPa for PTC., Conclusion: PTCs show greater SWE stiffening than BHN as precompression rises. Precompression effects on thyroid nodules are not negligible and may account for wide discrepancies in published SWE discriminatory performance results for thyroid malignancy., Key Points: • Increases in resting pressure (precompression) applied by the operator increases thyroid stiffness. • Papillary cancers show greater increases in stiffness (strain hardening) than benign nodules. • Precompression may affect the diagnostic performance of shearwave elastography for thyroid malignancy.