Search Results (47)

Accurate separation in microfluidic devices is crucial for biomedical applications; however, enhancing their performance remains challenging due to computational and experimental constraints. This study aims to optimize microfluidic devices by systematically refining spiral microchannel configurations for the segregation of circulating tumor cells (CTCs) and red blood cells (RBCs) through detailed variable analysis and resource-efficient techniques. The spiral design was developed into six variations, considering loop numbers (2, 3, and 4), aspect ratios (2.333, 3.333, and 5), spiral radii (5, 6, and 7 mm), flow rates (1.5, 2, and 3 mL/min), surface roughness levels (0, 0.5, and 1 μm), and particle sizes (12, 18, and 24 μm). Simulations were conducted in COMSOL Multiphysics and evaluated using the Taguchi method to determine the optimal configuration, reducing the analysis set from 216 to 27 through an efficient experimental design approach. The results identified the optimal structure as having an aspect ratio of 3.333, four loops, a spiral radius of 6–7 mm, a flow rate of 3 mL/min, a surface roughness of 1 μm, and a particle diameter of 24 μm. Among the evaluated parameters, aspect ratio (61.2%) had the most significant impact, followed by the number of loops (13.9%) and flow rate (9.4%). The optimized design demonstrated high separation efficiency and purity, achieving 97.5% and 97.6%, respectively. The fabrication process involved 3D-printing the channel mold, followed by polydimethylsiloxane (PDMS) casting, validating the durability and scalability of the proposed design. This study integrates simulation and experimental results, providing a robust framework for developing next-generation microfluidic devices and advancing diagnostic and targeted therapeutic applications. Full article

Background: The metastasis-promoting G-protein-coupled receptor CXC Receptor 4 (CXCR4) is activated by the chemokine CXCL12, also known as stromal cell-derived factor 1 (SDF-1). The CXCL12/CXCR4 pathway in cancer promotes metastasis but the molecular details of how this pathway cross-talks with oncogenes are understudied. An oncogene pathway known to promote breast cancer metastasis in MDA-MB-231 xenografts is that of Mouse Double Minute 2 and 4 (MDM2 and MDM4, also known as MDMX). MDM2 and MDMX promote circulating tumor cell (CTC) formation and metastasis, and positively correlate with a high expression of CXCR4. Interestingly, this MDMX-associated upregulation of CXCR4 is only observed in cells grown in the tumor microenvironment (TME), but not in MDA-MB-231 cells grown in a tissue culture dish. This suggested a cross-talk signaling factor from the TME which was predicted to be CXCL12 and, as such, we asked if the exogenous addition of the cell non-autonomous CXCL12 ligand would recapitulate the MDMX-dependent upregulation of CXCR4. Methods: We used MDA-MB-231 cells and isolated CTCs, with and without MDMX knockdown, plus the exogenous addition of CXCL12 to determine if MDMX-dependent upregulation of CXCR4 could be recapitulated outside of the TME context. We added exogenous CXCL12 to the culture medium used for growth of MDA-MB-231 cells and isogenic cell lines engineered for MDM2 or MDMX depletion. We carried out immunoblotting, and quantitative RT-PCR to compare the expression of CXCR4, MDM2, MDMX, and AKT activation. We carried out Boyden chamber and wound healing assays to assess the influence of MDMX and CXCL12 on the cells’ migration capacity. Results: The addition of the CXCL12 chemokine to the medium increased the CXCR4 cellular protein level and activated the PI3K/AKT signaling pathway. Surprisingly, we observed that the addition of CXCL12 mediated the upregulation of MDM2 and MDMX at the protein, but not at the mRNA, level. A reduction in MDMX, but not MDM2, diminished both the CXCL12-mediated CXCR4 and MDM2 upregulation. Moreover, a reduction in both MDM2 and MDMX hindered the ability of the added CXCL12 to promote Boyden chamber-assessed cell migration. The upregulation of MDMX by CXCL12 was mediated, at least in part, by a step upstream of the proteasome pathway because CXCL12 did not increase protein stability after cycloheximide treatment, or when the proteasome pathway was blocked. Conclusions: These data demonstrate a positive feed-forward activation loop between the CXCL12/CXCR4 pathway and the MDM2/MDMX pathway. As such, MDMX expression in tumor cells may be upregulated in the primary tumor microenvironment by CXCL12 expression. Furthermore, CXCL12/CXCR4 metastatic signaling may be upregulated by the MDM2/MDMX axis. Our findings highlight a novel positive regulatory loop between CXCL12/CXCR4 signaling and MDMX to promote metastasis. Full article

It is shown how the Correlated Traits Correlated Methods Minus One (CTC(M − 1)) Multitrait-Multimethod model for cross-classified data can be modified and applied to divergent thinking (DT)-task responses scored for miscellaneous aspects of creative quality by several raters. In contrast to previous Confirmatory Factor Analysis approaches to analyzing DT-tasks, this model explicitly takes the cross-classified data structure resulting from the employment of raters into account and decomposes the true score variance into target-specific, DT-task object-specific, rater-specific, and rater–target interaction-specific components. This enables the computation of meaningful measurement error-free relative variance-parameters such as trait-consistency, object–method specificity, rater specificity, rater–target interaction specificity, and model-implied intra-class correlations. In the empirical application with alternate uses tasks as DT-measures, the model is estimated using Bayesian statistics. The results are compared to the results yielded with a simplified version of the model, once estimated with Bayesian statistics and once estimated with the maximum likelihood method. The results show high trait-correlations and low consistency across DT-measures which indicates more heterogeneity across the DT-measurement instruments than across different creativity aspects. Substantive deliberations and further modifications, extensions, useful applications, and limitations of the model are discussed. Full article

Respiratory Syncytial Virus (RSV) is a significant cause of lower respiratory tract infections (LRTI) across all demographics, with increasing mortality and morbidity among high-risk groups such as infants under two years old, the elderly, and immunocompromised individuals. Although newly approved vaccines and treatments have substantially reduced RSV hospitalizations, accessibility remains limited, and response to treatment varies. This underscores the importance of comprehensive studies on host–RSV interactions. tRNA-derived RNA fragments (tRFs) are recently discovered non-coding RNAs, notable for their regulatory roles in diseases, including viral infections. Our prior work demonstrated that RSV infection induces tRFs, primarily derived from the 5′-end of a limited subset of tRNAs (tRF5), to promote RSV replication by partially targeting the mRNA of antiviral genes. This study found that tRFs could also use their bound proteins to regulate replication. Our proteomics data identified that PABPC1 (poly(A)-binding protein cytoplasmic 1) is associated with tRF5-GluCTC, an RSV-induced tRF. Western blot experimentally confirmed the presence of PABPC1 in the tRF5-GluCTC complex. In addition, tRF5-GluCTC is in the anti-PABPC1-precipitated immune complex. This study also discovered that suppressing PABPC1 with its specific siRNA increased RSV (-) genome copies without impacting viral gene transcription, but led to less infectious progeny viruses, suggesting the importance of PABPC1 in virus assembly, which was supported by its interaction with the RSV matrix protein. Additionally, PABPC1 knockdown decreased the production of the cytokines MIP-1α, MIP-1β, MCP-1, and TNF-α. This is the first observation suggesting that tRFs may regulate viral infection via their bound proteins. Full article

As a special type of clay, expansive clay is widely distributed in China. Its characteristics of swelling and softening when meeting water and shrinking and cracking when losing water bring many hidden dangers to engineering construction. Expansive clay is known as “engineering cancer”, and in-depth research on the unloading mechanical response characteristics and the unloading constitutive relationships of expansive clay is a prerequisite for conducting geotechnical engineering design and safety analysis in expansive-soil areas. In order to obtain the unloading mechanical response characteristics and the expression of the unloading tangent modulus of expansive clay, typical expansive clay in the Hefei area was taken as the research object, and triaxial unloading stress path tests were conducted. The stress–strain properties, microstructures, macro failure modes, and strength indexes of the expansive clay were analyzed under unloading stress paths. Through an applicability analysis of several classical soil strength criteria, an unloading constitutive model and the unloading tangent modulus expression of the expansive clay were constructed based on the Mohr–Coulomb (hereinafter referred to as “M-C”) criterion, the Drucker–Prager (hereinafter referred to as “D-P”) criterion, and the extended Spatial Mobilized Plane (hereinafter referred to as “SMP”) criterion theoretical frameworks. The following research results were obtained: (1) The stress–strain curves of the three stress paths of the expansive clay were hyperbolic. The expansive clay showed typical strain-hardening characteristics and belonged to work-hardening soil. (2) Under the unloading stress paths, the soil particles were involved in the unloading process of stress release, and the failure samples showed obvious stretching, curling, and slipping phenomena in their soil sheet elements. (3) Under both unloading stress paths, the strength of the expansive clay was significantly weakened and reduced. Under the lateral unloading paths, the cohesive force (c) of the expansive clay was reduced by 32.7% and the internal friction angle (φ) was increased by 19% compared with those under conventional loading, while under the axial unloading path, c was reduced by 63.5% and φ was reduced by 28.7%. (4) For typical expansive clay in Hefei, the conventional triaxial compression (hereinafter referred to as “CTC”) test, the reduced triaxial compression (hereinafter referred to as “RTC”) test, and the reduced triaxial extension (hereinafter referred to as “RTE”) test stress paths were suitable for characterization and deformation prediction using the M-C strength criterion, D-P strength criterion, and extended SMP strength criterion, respectively. (5) The derived unloading constitutive model and the unified tangent modulus formula of the expansive clay could accurately predict the deformation characteristics of the unloading stress path of the expansive clay. These research results will provide an important reference for future engineering construction in expansive-clay areas. Full article

Background and Objectives: Congenital thyroid dyshormonogenesis is caused by alterations in the synthesis of thyroid hormones in a newborn. Additionally, 10 to 20% of these cases are hereditary, caused by defects in proteins involved in hormonal synthesis. One of the most common causes is mutations in the thyroid peroxidase (TPO) enzyme gene, an autosomal recessive disease. We aimed to detect mutations of the TPO gene in 12 Chilean patients with congenital hypothyroidism due to dyshormonogenesis (CHD) and to characterize these patients clinically and molecularly. Materials and Methods: Twelve patients under 20 years of age with CHD, controlled at San Juan de Dios Hospital in Santiago, Chile, were selected according to the inclusion criteria: elevated neonatal TSH, persistent hypothyroidism, and thyroid normotopic by imaging study. Those with deafness, Down syndrome, and central or transient congenital hypothyroidism were excluded. Blood samples were taken for DNA extraction, and the 17 exons and exon–intron junctions of the TPO gene were amplified by PCR. The PCR products were sequenced by Sanger. Results: Two possibly pathogenic mutations of the TPO gene were detected: c.2242G>A (p.Val748Met) and c.1103C>T (p.Pro368Leu). These mutations were detected in 2 of 12 patients (16.6%): 1 was compound heterozygous c.1103C>T/c.2242G>A, and the other was heterozygous for c.2242G>A. In the diagnostic confirmation test, both patients presented diffuse hyper-uptake goiter on thyroid scintigraphy and high TSH in venous blood (>190 uIU/mL). Conclusions: The frequency of patients with possibly pathogenic mutations in TPO with CHD was 16.6%. Its study would allow for genetic counseling to be offered to the families of affected patients. Full article

Both the soybean variety and processing method affect the end soybean product’s characteristics. This study’s objective was to characterize the effects of four extraction methods (variations of soaking and grinding) combined with cooking on the content and composition of phenolic substances and the antioxidant and anti-DU145 prostate cancer cell proliferation properties of soymilks prepared from a yellow soybean of the ProSoy variety, which is a high-protein variety. The results showed that the soymilk processing yield was the greatest using method 4, although method 2 gave the highest solid and protein yields by about 14 and 12%, respectively. Method 4, a two-step grinding method, also gave increased yields (8 and 7% for solids and proteins, respectively), and in all but one instance produced higher total phenolic content (TPC), total flavonoid content (TFC), condensed tannin content (CTC), and total isoflavone content values in both raw and cooked soymilks as compared to method 1. Cooking the soymilks reduced 14–17% of their total phenolic substances. Cooking reduced the anti-cancer capacity of the phenolic extracts from the soymilk prepared using method 4 by increasing the IC50 value from about 4.9 mg/mL to 6.8 mg/mL. The increases in phenolic compounds and antioxidants produced in the Prosoy soymilks using methods 2 and 4, with simultaneous increases in product and solid yields, are of significant benefit to the soymilk industry and consumer health. Full article

The spleen and small intestines are the primary immune organs that provide important immunity against various diseases. Artemisia ordosica polysaccharide (AOP) could be used as an immunologic enhancer to boost immunity in response to infection. This study was performed to explore the effects of the dietary supplementation of AOP on the growth performance and spleen and small intestine immune function in broilers. A total of 288 AA broilers (1 day old) were randomly assigned into six dietary groups. Each group included six replicates of eight broilers per cage. The broilers were fed with a basal diet supplemented with 0 mg/kg (CON), 50 mg/kg chlortetracycline (CTC), 250, 500, 750, and 1000 mg/kg AOP for 42 d. The results showed that dietary AOP supplementation affected broiler growth performance, with 750 and 1000 mg/kg of AOP being able to significantly improve broiler BWG, and 750 mg/kg of AOP was able to significantly reduce the FCR. The dietary AOP supplementation increased the levels of IgA, IgG, IgM, IL-1β, IL-2, and IL-4 in the spleen and small intestine in a dose-dependent manner (p < 0.05). Meanwhile, we found that AOP can promote the mRNA expression of TLR4/MAPK/NF-κB signaling-pathway-related factors (TLR4, MyD88, P38 MAPK, JNK, NF-κB p50, and IL-1β). In addition, the dietary supplementation of 750 mg/kg AOP provides better immunity in the tissue than the CON group but showed no significant difference from the CTC group. Therefore, AOP has an immunoregulatory action and can modulate the immune function of broilers via the TLR4/ NF-ΚB/MAPK signal pathway. In conclusion, dietary supplementation with 750 mg/kg AOP may be alternatives to antibiotics for enhancing broilers’ health, immunity, and growth performance. Full article

Rational: Circulating tumor cells (CTCs) appear to be a promising tool for predicting the clinical outcome and monitoring the response to treatment in patients with solid tumors. The current study assessed the clinical relevance of monitoring CTCs in patients with metastatic castration resistant prostate cancer (mCRPC) treated with cabazitaxel. Patients and Methods: Patients with histologically confirmed mCRPC who were previously treated with a docetaxel-containing regimen and experienced disease progression were enrolled in this multicenter prospective study. CTC counts were enumerated using the CellSearch system at baseline (before cabazitaxel initiation), after one cabazitaxel cycle (post 1st cycle) and at disease progression (PD). Patients were stratified into predetermined CTC-positive and CTC-negative groups. The phenotypic characterization was performed using double immunofluorescence staining with anti-CKs and anti-Ki67, anti-M30 or anti-vimentin antibodies. Results: The median PFS and OS were 4.0 (range, 1.0–17.9) and 14.5 (range, 1.2–33.9) months, respectively. At baseline, 48 out of 57 (84.2%) patients had ≥1 CTCs/7.5 mL of peripheral blood (PB) and 37 (64.9%) had ≥5 CTCs/7.5 mL of PB. After one treatment cycle, 30 (75%) out of the 40 patients with available measurements had ≥1 detectable CTC/7.5 mL of PB and 24 (60%) ≥ 5CTCs/7.5 mL of PB; 12.5% of the patients with detectable CTCs at the baseline sample had no detectable CTCs after one treatment cycle. The detection of ≥5CTCs/7.5 mL of PB at baseline and post-cycle 1 was associated with shorter PFS and OS (p = 0.002), whereas a positive CTC status post-cycle 1 strongly correlated with poorer OS irrespective of the CTC cut-off used. Multivariate analysis revealed that the detection of non-apoptotic (CK⁺/M30⁻) CTCs at baseline is an independent predictor of shorter OS (p = 0.005). Conclusions: In patients with mCRPC treated with cabazitaxel, CTC counts both at baseline and after the first cycle retain their prognostic significance, implying that liquid biopsy monitoring might serve as a valuable tool for predicting treatment efficacy and survival outcomes. Full article

Metastasis, rather than the growth of the primary tumor, accounts for approximately 90% of breast cancer patient deaths. Microtentacles (McTNs) formation represents an important mechanism of metastasis. Triple-negative breast cancer (TNBC) is the most aggressive subtype with limited targeted therapies. The present study aimed to isolate viable circulating tumor cells (CTCs) and functionally analyze them in response to drug treatment. CTCs from 20 TNBC patients were isolated and maintained in culture for 5 days. Biomarker expression was identified by immunofluorescence staining and VyCap analysis. Vinorelbine-induced apoptosis was evaluated based on the detection of M30-positive cells. Our findings revealed that the CTC absolute number significantly increased using TetherChips analysis compared to the number of CTCs in patients’ cytospins (p = 0.006) providing enough tumor cells for drug evaluation. Vinorelbine treatment (1 h) on live CTCs led to a significant induction of apoptosis (p = 0.010). It also caused a significant reduction in Detyrosinated α-tubulin (GLU), programmed death ligand (PD-L1)-expressing CTCs (p < 0.001), and disruption of McTNs. In conclusion, this pilot study offers a useful protocol using TetherChip technology for functional analysis and evaluation of drug efficacy in live CTCs, providing important information for targeting metastatic dissemination at a patient-individualized level. Full article

Circulating tumor cells (CTCs) and/or circulating tumor microemboli (CTM) from non-small cell lung cancer (NSCLC) patients may be a non-invasive tool for prognosis, acting as liquid biopsy. CTCs interact with platelets through the transforming growth factor-β/transforming growth factor-β receptor type 1 (TGF-β/TGFβRI) forming clusters. CTCs also may express the Cluster of Differentiation 47 (CD47) protein, responsible for the inhibition of phagocytosis, the “don’t eat me” signal to macrophages. Objectives: To isolate, quantify and analyze CTCs/CTMs from metastatic NSCLC patients, identify TGFβRI/CD47 expression in CTCs/CTMs, and correlate with progression-free survival (PFS). Methods: Blood (10 mL) was collected at two time-points: T1 (before the beginning of any line of treatment; T2 (60 days after initial collection). CTCs were isolated using ISET^®. Immunocytochemistry was conducted to evaluate TGFβRI/CD47 expression. Results: 45 patients were evaluated. CTCs were observed in 82.2% of patients at T1 (median: 1 CTC/mL; range: 0.33–11.33 CTCs/mL) and 94.5% at T2 (median: 1.33 CTC/mL; 0.33–9.67). CTMs were observed in 24.5% of patients and significantly associated with poor PFS (10 months vs. 17 months for those without clusters; p = 0.05) and disease progression (p = 0.017). CTMs CD47+ resulted in poor PFS (p = 0.041). TGFβRI expression in CTCs/CTMs was not associated with PFS. Conclusion: In this study, we observed that CTC/CTM from NSCLC patients express the immune evasion markers TGFβRI/CD47. The presence of CTMs CD47+ is associated with poor PFS. This was the first study to investigate CD47 expression in CTCs/CTM of patients with NSCLC and its association with poor PFS. Full article

Stage III melanoma includes nodal metastasis or in-transit disease. Five-year survival rates vary between 32% and 93%. The identification of high-risk patients is important for clinical decision making. We demonstrated previously that ≥1 circulating tumor cells (CTCs) at baseline was associated with recurrence. In this study, we investigated how frequently CTCs were identified prior to radiologically detected recurrence. Stage III patients (n = 325) had imaging at baseline and q 3 months. Baseline and q 6–12 months blood draws (7.5 mL) were performed to identify CTCs up to 3.5 years from diagnosis. CTC assessment was performed using the immunomagnetic capture of CD146-positive cells and anti-MEL-PE. The presence of one or more CTCs was considered positive. We analyzed the cohort of patients with relapse confirmed by radiologic imaging. CTC collection dates were assessed to determine the lead time for CTC detection. CTC-negative patients were significantly less likely to relapse compared to patients positive for CTCs (p-value < 0.001). Within the 325-patient cohort, 143 patients (44%) had recurrence, with a median follow-up of 52 months from diagnosis. The cohort (n = 143) with positive imaging and CTC results revealed 76% of patients (108/143) had CTC+ results before the radiological identification of relapse. The median time between positive CTC and positive imaging was 9 months. CTCs were positive in >75% of patients prior to relapse at a median of 9 months before radiologic detection. Full article

Background: A large amount of evidence from clinical studies has demonstrated that circulating tumor cells are strong predictors of outcomes in many cancers. However, the clinical significance of CTC enumeration in metastatic colorectal cancer is still questioned. The aim of this study was to evaluate the clinical value of CTC dynamics in mCRC patients receiving first-line treatments. Materials and methods: Serial CTC data from 218 patients were used to identify CTC trajectory patterns during the course of treatment. CTCs were evaluated at baseline, at a first-time point check and at the radiological progression of the disease. CTC dynamics were correlated with clinical endpoints. Results: Using a cut-off of ≥1 CTC/7.5 mL, four prognostic trajectories were outlined. The best prognosis was obtained for patients with no evidence of CTCs at any timepoints, with a significant difference compared to all other groups. Lower PFS and OS were recognized in group 4 (CTCs always positive) at 7 and 16 months, respectively. Conclusions: We confirmed the clinical value of CTC positivity, even with only one cell detected. CTC trajectories are better prognostic indicators than CTC enumeration at baseline. The reported prognostic groups might help to improve risk stratification, providing potential biomarkers to monitor first-line treatments. Full article

The co-localization of platelets and tumor cells in hematogenous metastases has long been recognized. Interactions between platelets and circulating tumor cells (CTCs) contribute to tumor cell survival and migration via the vasculature into other tissues. Taking advantage of the interactions between platelets and tumor cells, two schemes, direct and indirect, were proposed to target the modified human serum albumin submicron particles (HSA-MPs) towards tumor cells. HSA-MPs were constructed by the Co-precipitation–Crosslinking–Dissolution (CCD) method. The anti-CD41 antibody or CD62P protein was linked to the HSA-MPs separately via 1-ethyl-3-(-3-dimethyl aminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) EDC/NHS chemistry. The size of modified HSA-MPs was measured at approximately 1 µm, and the zeta potential was around −24 mV. Anti-CD41-HSA-MPs adhered to platelets as shown by flowcytometry and confocal laser scanning microscopy. In vitro, we confirmed the adhesion of platelets to tumor lung carcinoma cells A549 under shearing conditions. Higher cellular uptake of anti-CD41-HSA-MPs in A549 cells was found in the presence of activated platelets, suggesting that activated platelets can mediate the uptake of these particles. RNA-seq data in the Cancer Cell Lineage Encyclopedia (CCLE) and The Cancer Genome Atlas (TCGA) database showed the expression of CD62P ligands in different types of cancers. Compared to the non-targeted system, CD62P-HSA-MPs were found to have higher cellular uptake in A549 cells. Our results suggest that the platelet-based and platelet-mimicking modified HSA-MPs could be promising options for tracking metastatic cancer. Full article

Circulating tumor cells (CTCs) exist in low quantities in the bloodstream in the early stages of cancers. It, therefore, remains a technical challenge to isolate them in large enough quantities for a precise diagnosis and downstream analysis. We introduce the BMProbe™, a minimally invasive device that isolates CTCs during a 30-minute incubation in the median cubital vein. The optimized geometry of the device creates flow conditions for improved cell deposition. The CTCs are isolated using antibodies that are bound to the surface of the BMProbe™. In this study, flow experiments using cell culture cells were conducted. They indicate a 31 times greater cell binding efficiency of the BMProbe™ compared to a flat geometry. Further, the functionality of isolating CTCs from patient blood was verified in a small ex vivo study that compared the cell count from seven non-small-cell lung carcinoma (NSCLC) patients compared to nine healthy controls with 10 mL blood samples. The median cell count was 1 in NSCLC patients and 0 in healthy controls. In conclusion, the BMProbe™ is a promising method to isolate CTCs in large quantities directly from the venous bloodstream without removing blood from a patient. The future step is to verify the functionality in vivo. Full article