Regulatory macrophages in solid organ xenotransplantation.

Due to a critical organ shortage, pig organs are being explored for use in transplantation. Differences between species, particularly in cell surface glycans, can trigger elevated immune responses in xenotransplantation. To mitigate the risk of hyperacute rejection, genetically modified pigs have been developed that lack certain glycans and express human complement inhibitors. Nevertheless, organs from these pigs may still provoke stronger inflammatory and innate immune reactions than allotransplants. Dysregulation of coagulation and persistent inflammation remain obstacles in the transplantation of pig organs into primates. Regulatory macrophages (Mregs), known for their anti-inflammatory properties, could offer a potential solution. Mregs secrete interleukin 10 and transforming growth factor beta, thereby suppressing immune responses and promoting the development of regulatory T cells. These Mregs are typically induced via the stimulation of monocytes or macrophages with macrophage colony-stimulating factor and interferon gamma, and they conspicuously express the stable marker dehydrogenase/reductase 9. Consequently, understanding the precise mechanisms governing Mreg generation, stability, and immunomodulation could pave the way for the therapeutic use of Mregs generated in vitro. This approach has the potential to reduce the required dosages and durations of anti-inflammatory and immunosuppressive medications in preclinical and clinical settings.

1,25-Dihydroxy Vitamin D3 Facilitates the M2 Polarization and ß-Amyloid Uptake by Human Microglia in a TREM2-Dependent Manner.

Alzheimer's disease (AD) is a neurodegenerative disorder characterized by dementia as the primary clinical symptom. The production and accumulation of aggregated ß-amyloid (Aß) in patient brain tissues is one of the hallmarks of AD pathogenesis. Microglia, brain-resident macrophages, produce inflammatory cytokines in response to Aß oligomers or fibrils exacerbating Aß pathology in AD. HMO6 cells were treated with Aß42 in the presence or absence of 1,25-dihydroxy vitamin D3 (1,25(OH)2D3) to determine its potential immunomodulatory effects, and the expression of pro-/anti-inflammatory cytokines, M1/M2-associated markers, Toll-like receptors (TLRs), and triggering receptor expressed on myeloid cells 2 (TREM2) was examined. 1,25(OH)2D3 was found to suppress Aß-induced expression of proinflammatory cytokines (TNF-α, IL-1ß, and IL-6), M1 markers (CD86 and iNOS), and TLR2/4, whilst increasing the expression of anti-inflammatory cytokines (IL-4, IL-10, and CCL17) and M2 markers (CD206 and Arg-1). Furthermore, 1,25(OH)2D3 promoted TREM2 expression and Aß uptake by HMO6 cells, and the enhancement of Aß uptake and M2 polarization was revealed to be TREM2-dependent. The findings of this study suggest that 1,25(OH)2D3 facilitates M2 polarization and Aß uptake in a TREM2-dependent manner.

Human Regulatory Macrophages Derived from THP-1 Cells Using Arginylglycylaspartic Acid and Vitamin D3.

Regulatory macrophages (Mregs) are unique in that they have anti-inflammatory and immunosuppressive properties. Thus, treating inflammatory diseases using Mregs is an area of active research. Human Mregs are usually generated by culturing peripheral blood monocytes stimulated using a macrophage colony-stimulating factor with interferon (IFN)-γ. Herein, we generated Mregs with an elongated cell morphology from THP-1 cells that were stimulated with phorbol 12-myristate 13-acetate and cultured with both arginylglycylaspartic acid and vitamin D3. These Mregs regulated macrophage function, and respectively downregulated and upregulated the expression of pro-inflammatory and immunosuppressive mediators. They also expressed Mregs-specific markers, such as dehydrogenase/reductase 9, even when exposed to such inflammatory stimulants as IFN-γ, lipopolysaccharide, purified xenogeneic antigen, and xenogeneic cells. The Mregs also exerted anti-inflammatory and anticoagulatory actions in response to xenogeneic cells, as well as exerting immunosuppressive effects on mitogen-induced Jurkat T-cell proliferation. Our method of generating functional Mregs in vitro without cytokines is simple and cost-effective.

PDMS Micropatterns Coated with PDA and RGD Induce a Regulatory Macrophage-like Phenotype.

Regulatory macrophages (Mreg) are a special cell type that present a potential therapeutic strategy for various inflammatory diseases. In vitro, Mreg generation mainly takes 7-10 days of treatment with chemicals, including cytokines. In the present study, we established a new approach for Mreg generation using a three-dimensional (3D) micropatterned polydimethylsiloxane (PDMS) surface coated with a natural biopolymer adhesive polydopamine (PDA) and the common cell adhesion peptide motif arginylglycylaspartic acid (RGD). The 3D PDMS surfaces were fabricated by photolithography and soft lithography techniques and were subsequently coated with an RGD+PDA mixture to form a surface that facilitates cell adhesion. Human monocytes (THP-1 cells) were cultured on different types of 2D or 3D micropatterns for four days, and the cell morphology, elongation, and Mreg marker expression were assessed using microscopic and flow cytometric analyses. The cells grown on the PDA+RGD-coated 3D micropatterns (20-µm width/20-µm space) exhibited the most elongated morphology and strongest expression levels of Mreg markers, such as CD163, CD206, CD209, CD274, MER-TK, TREM2, and DHRS9. The present study demonstrated that PDA+RGD-coated 3D PDMS micropatterns successfully induced Mreg-like cells from THP-1 cells within four days without the use of cytokines, suggesting a time- and cost-effective method to generate Mreg-like cells in vitro.

Cell Surface Hsp90- and αMß2 Integrin-Mediated Uptake of Bacterial Flagellins to Activate Inflammasomes by Human Macrophages.

All-trans retinoic acid (ATRA) is an active metabolite of vitamin A, which plays an important role in the immune function. Here, we demonstrated that ATRA induces the heat shock protein (Hsp) 90 complex on the surface of THP-1 macrophages, which facilitates the internalization of exogenous bacterial flagellins to activate the inflammasome response. Mass spectrometric protein identification and co-immunoprecipitation revealed that the Hsp90 homodimer interacts with both Hsp70 and αMß2 integrin. ATRA-induced complex formation was dependent on the retinoic acid receptor (RAR)/retinoid X receptor (RXR) pathway and intracellular calcium level and was essential for triggering the internalization of bacterial flagellin, which was clathrin dependent. Notably, in this process, αMß2 integrin was found to act as a carrier to deliver flagellin to the cytosol to activate the inflammasome, leading to caspase-1 activity and secretion of interleukin (IL)-1ß. Our study provides new insights into the underlying molecular mechanism by which exogenous bacterial flagellins are delivered into host cells without a bacterial transport system, as well as the mechanism by which vitamin A contributes to enhancing the human macrophage function to detect and respond to bacterial infection.

Fluorescent Carbon Dots for Sensitive and Rapid Monitoring of Intracellular Ferrous Ion.

Although iron is an essential constituent for almost all living organisms, iron dyshomeostasis at a cellular level may trigger oxidative stress and neuronal damage. Hence, there are numerous reported carbon dots (CDs) that have been synthesized and applied to determine intracellular iron ions. However, among reported CDs focused to detect Fe3+ ions, only a few CDs have been designed to specifically determine Fe2+ ions over Fe3+ ions for monitoring of intracellular Fe2+ ions. We have developed the nitrogen-doped CDs (NCDs) for fluorescence turn-off detection of Fe2+ at cellular level. The as-synthesized NCDs exhibit a strong blue fluorescence and low cytotoxicity, acting as fluorescence probes to detect Fe2+ as low as 0.702 µM in aqueous solution within 2 min and visualize intracellular Fe2+ in the concentration range from 0 to 500 µM within 20 min. The as-prepared NCDs possess some advantages such as high biocompatibility, strong fluorescence properties, selectivity, and rapidity for intracellular Fe2+ monitoring, making NCDs an excellent nanoprobe for biosensing of intracellular ferrous ions.

Synergistic Antiproliferative Effects of All-Trans Retinoic Acid and Paclitaxel on Autosomal Dominant Polycystic Kidney Disease Epithelial Cells.

Autosomal dominant polycystic kidney disease (ADPKD) is a genetic disorder characterized by uncontrollable epithelial cell growth, cyst formation, and kidney malfunction. In the present study, we investigated the antiproliferative effects of the treatment with the combination of paclitaxel (PAC) and all-trans retinoic acid (ATRA) on ADPKD epithelial cells. Our results show that the combined treatment with 1 nM PAC and 10 nM ATRA significantly suppressed ADPKD cell proliferation (20%), while the treatment with ATRA or PAC alone had no such effect. Treatment with PAC and ATRA induced cell cycle arrest at the G2/M phase and apoptosis by upregulating p53 and caspase-8 expression and increased the intracellular calcium (Ca2+) level possibly by enhancing Ca2+ uptake via plasma membrane channels. In addition, this treatment suppressed extracellular signal-regulated kinase signaling possibly through mitogen-activated protein kinase phosphatase-1 activation. Thus, the combination of PAC and ATRA can be explored as a potential treatment regimen for ADPKD.

Advanced Signal-Amplification Strategies for Paper-Based Analytical Devices: A Comprehensive Review.

Paper-based analytical devices (PADs) have emerged as a promising approach to point-of-care (POC) detection applications in biomedical and clinical diagnosis owing to their advantages, including cost-effectiveness, ease of use, and rapid responses as well as for being equipment-free, disposable, and user-friendly. However, the overall sensitivity of PADs still remains weak, posing a challenge for biosensing scientists exploiting them in clinical applications. This review comprehensively summarizes the current applicable potential of PADs, focusing on total signal-amplification strategies that have been applied widely in PADs involving colorimetry, luminescence, surface-enhanced Raman scattering, photoacoustic, photothermal, and photoelectrochemical methods as well as nucleic acid-mediated PAD modifications. The advances in signal-amplification strategies in terms of signal-enhancing principles, sensitivity, and time reactions are discussed in detail to provide an overview of these approaches to using PADs in biosensing applications. Furthermore, a comparison of these methods summarizes the potential for scientists to develop superior PADs. This review serves as a useful inside look at the current progress and prospective directions in using PADs for clinical diagnostics and provides a better source of reference for further investigations, as well as innovations, in the POC diagnostics field.

Trending Technology of Glucose Monitoring during COVID-19 Pandemic: Challenges in Personalized Healthcare.

The COVID-19 pandemic has continued to spread rapidly, and patients with diabetes are at risk of experiencing rapid progression and poor prognosis for appropriate treatment. Continuous glucose monitoring (CGM), which includes accurately tracking fluctuations in glucose levels without raising the risk of coronavirus exposure, becomes an important strategy for the self-management of diabetes during this pandemic, efficiently contributing to the diabetes care and the fight against COVID-19. Despite being less accurate than direct blood glucose monitoring, wearable noninvasive systems can encourage patient adherence by guaranteeing reliable results through high correlation between blood glucose levels and glucose concentrations in various other biofluids. This review highlights the trending technologies of glucose sensors during the ongoing COVID-19 pandemic (2019-2020) that have been developed to make a significant contribution to effective management of diabetes and prevention of coronavirus spread, from off-body systems to wearable on-body CGM devices, including nanostructure and sensor performance in various biofluids. The advantages and disadvantages of various human biofluids for use in glucose sensors are also discussed. Furthermore, the challenges faced by wearable CGM sensors with respect to personalized healthcare during and after the pandemic are deliberated to emphasize the potential future directions of CGM devices for diabetes management.

Metal-enhanced sensing platform for the highly sensitive detection of C-reactive protein antibody and rhodamine B with applications in cardiovascular diseases and food safety.

The potential applications of metal-enhanced fluorescence (MEF) devices include biosensors for the detection of trace amounts in biosciences, biotechnology, and pathogens that are relevant to medical diagnostics and food control. In the present study, the silver (Ag) film thickness (56 nm) of an MEF system was calibrated to maximize the depth-to-width ratio (Γ) of the surface plasmon resonance (SPR) active metal from reflectance dip curves. Upon plasmon coupling with thermally evaporated Ag, we demonstrated a 2.21-fold enhancement compared to the pristine flat substrate with the coefficient of variation (CV) ≈0.22% and the limit of detection (LOD) 0.001 mg L-1 of the concentration of an Alexa Fluor 488-labeled anti-C-reactive protein antibody (CRP@Alexa fluor 488). The structure was developed to simplify the in situ generation of biosensors for the surface-enhanced Raman spectroscopy (SERS) to determine Rhodamine B (RhB) with a highly robust performance. The procedure presented a simple and rapid sample pretreatment for the determination of RhB with a limit of quantification of 10-10 M and a satisfactory linear response (0.98). The results showed the excellent performance of the surface plasmon coupled emission (SPCE), which opens up possibilities for the accurate detection of small-volume and low-concentration target analytes due to the improved sensitivity and signal-to-noise ratio (SNR).

Graphene Integrated Hydrogels Based Biomaterials in Photothermal Biomedicine.

Recently, photothermal therapy (PTT) has emerged as one of the most promising biomedical strategies for different areas in the biomedical field owing to its superior advantages, such as being noninvasive, target-specific and having fewer side effects. Graphene-based hydrogels (GGels), which have excellent mechanical and optical properties, high light-to-heat conversion efficiency and good biocompatibility, have been intensively exploited as potential photothermal conversion materials. This comprehensive review summarizes the current development of graphene-integrated hydrogel composites and their application in photothermal biomedicine. The latest advances in the synthesis strategies, unique properties and potential applications of photothermal-responsive GGel nanocomposites in biomedical fields are introduced in detail. This review aims to provide a better understanding of the current progress in GGel material fabrication, photothermal properties and potential PTT-based biomedical applications, thereby aiding in more research efforts to facilitate the further advancement of photothermal biomedicine.

Nanomaterial-based Optical and Electrochemical Biosensors for Amyloid beta and Tau: Potential for early diagnosis of Alzheimer's Disease.

INTRODUCTION: Alzheimer's disease (AD), a heterogeneous pathological process representing the most common causes of dementia worldwide, has required early and accurate diagnostic tools. Neuropathological hallmarks of AD involve the aberrant accumulation of Amyloid beta (Aß) into Amyloid plaques and hyperphosphorylated Tau into neurofibrillary tangles, occurring long before the onset of brain dysfunction.Areas covered:Considering the significance of Aß and Tau in AD pathogenesis, these proteins have been adopted as core biomarkers of AD, and their quantification has provided precise diagnostic information to develop next-generation AD therapeutic approaches. However, conventional diagnostic methods may not suffice to achieve clinical criteria that are acceptable for proper diagnosis and treatment. The advantages of nanomaterial-based biosensors including facile miniaturization, mass fabrication, ultra-sensitivity, make them useful to be promising tools to measure Aß and Tau simultaneously for accurate validation of low-abundance yet potentially informative biomarkers of AD.. EXPERT OPINION: The study has identified the potential application of advanced biosensors as standardized clinical diagnostic tools for AD, evolving the way for new and efficient AD control with minimum economic and social burden. After clinical trial, nanobiosensors for measuring Aß and Tau simultaneously possess innovative diagnosis of AD to provide significant contributions to primary Alzheimer's care intervention.

Nanobiosensors for Non-Amyloidbeta-Tau Biomarkers as Advanced Reporters of Alzheimer's Disease.

Emerging nanomaterials providing benefits in sensitivity, specificity and cost-effectiveness are being widely investigated for biosensors in the application of Alzheimer's disease (AD) diagnosis. Core biomarkers amyloid-beta (Aß) and Tau have been considered as key neuropathological hallmarks of AD. However, they did not sufficiently reflect clinical severity and therapeutic response, proving the difficulty of the Aß- and Tau-targeting therapies in clinical trials. In recent years, there has still been a shortage of sensors for non-Aß-Tau pathophysiological biomarkers that serve as advanced reporters for the early diagnosis of AD, predict AD progression, and monitor the treatment response. Nanomaterial-based sensors measuring multiple non-Aß-Tau biomarkers could improve the capacity of AD progression characterization and supervised treatment, facilitating the comprehensive management of AD. This is the first review to principally represent current nanobiosensors for non-Aß-Tau biomarker and that strategically deliberates future perspectives on the merit of non-Aß-Tau biomarkers, in combination with Aß and Tau, for the accurate diagnosis and prognosis of AD.

All-trans retinoic acid increases NF-κB activity in PMA-stimulated THP-1 cells upon unmethylated CpG challenge by enhancing cell surface TLR9 expression.

An active metabolite of vitamin A, all-trans retinoic acid (ATRA), is known to exert immunomodulatory functions. This study investigates the possible immune potentiating effect of ATRA on NF-κB activity in human monocytic THP-1 cells after exposure to unmethylated CpG DNA ODN2006. We observed that challenge with ODN2006 significantly enhanced the NF-κB activity of PMA-differentiated THP-1 cells. ATRA synergistically enhanced NF-κB activity of cells, in a concentration- and time-dependent manner. The enhanced NF-κB activity of PMA-differentiated THP-1 cells after ODN2006 challenge was dependent on the RAR/RXR pathway. To determine the mechanism involved in increasing in the NF-κB activity of stimulated THP-1 cells, we examined the effects of PMA and ATRA on the expression of TLR9 (a receptor of ODN2006) in THP-1 cells. PMA treatment significantly enhanced both the intracellular and cell surface expression of TLR9, while ATRA alone showed no effect. However, ATRA synergistically enhanced the cell surface TLR9 expression of PMA-differentiated cells. To determine whether the ATRA-enhanced NF-κB activity is due to the enhanced cell surface TLR9 expression, we examined NF-κB activity after treatment with anti-TLR9 blocking antibody. Results revealed that the anti-TLR9 antibody treatment almost completely reverses the ATRA-enhanced NF-κB activity, suggesting that ATRA enhances NF-κB activity through upregulation of the cell surface TLR9 expression in PMA-differentiated and unmethylated CpG challenged THP-1 cells.

All-Trans Retinoic Acid Enhances Bacterial Flagellin-Stimulated Proinflammatory Responses in Human Monocyte THP-1 Cells by Upregulating CD14.

All-trans retinoic acid (ATRA), an active form of vitamin A, exerts immunomodulatory functions. In this study, we examined the immune potentiating effect of ATRA on bacterial flagellin-induced NF-κB activation and proinflammatory cytokine production in human monocytic cell line THP-1. ATRA treatment significantly enhanced the flagellin-induced NF-κB/AP-1 activity in THP-1 via the RAR/RXR pathway. Similarly, ATRA enhanced the expression and production of TNF-α and IL-1ß in THP-1 cells upon flagellin challenge. The cell surface expression of toll-like receptor 5 (TLR5), which is the receptor for bacterial flagellin, was significantly reduced by ATRA in a concentration- and time-dependent manner. To determine the mechanisms underlying the ATRA-enhanced immune response against bacterial flagellin despite the reduced cell surface expression of TLR5 in ATRA-treated THP-1, we examined the cell surface expression of CD14, which has been proposed to be a TLR co-receptor that enhances the response to microbial components. The cell surface expression of CD14 was significantly enhanced by ATRA treatment, especially in the presence of flagellin. Anti-CD14 antibody treatment prior to ATRA and flagellin treatments completely abolished ATRA-enhanced TNF-α and IL-1ß production. Our results suggest that ATRA enhances flagellin-stimulated proinflammatory responses in human monocyte THP-1 cells by upregulating CD14 in a RAR/RXR-dependent manner.

All-Trans Retinoic Acid Enhances Matrix Metalloproteinase 2 Expression and Secretion in Human Myeloid Leukemia THP-1 Cells.

All-trans retinoic acid (ATRA) is an effective drug for the induction therapy of acute promyelocytic leukemia. However, the treatment is associated with adverse events such as retinoic acid syndrome (RAS) in some patients, whose histologic characteristics included organ infiltration by leukemic cells. Matrix metalloproteinase 2 (MMP-2) is often upregulated in tumor cells and plays a role in tumor cell migration and invasion by degrading the extracellular matrix. In this study, we examined the possible modulatory effects of ATRA on MMP-2 expression and secretion in human myeloid leukemia cell line THP-1. The cells were treated with various concentrations of ATRA, and MMP-2 expression and secretion were examined. MMP-2 expression and secretion started to increase with ATRA concentration as low as 0.1 nM and gradually increased thereafter. Agonists of retinoic acid receptor (RAR) or retinoid X receptor (RXR) alone could enhance MMP-2 secretion, and RAR or RXR antagonists alone could reverse ATRA-induced MMP-2 secretion. ATRA increased intracellular calcium ion levels, and a calcium-channel blocker inhibited ATRA-induced MMP-2 secretion. Dexamethasone suppressed ATRA-induced MMP-2 secretion. Our results suggest that ATRA enhances MMP-2 expression and secretion in human myeloid leukemia THP-1 cells in a calcium ion dependent manner through RAR/RXR signaling pathways, and this enhanced expression and secretion may be associated with the possible mechanisms of RAS.

Hsp90 Inhibition Reduces TLR5 Surface Expression and NF-κB Activation in Human Myeloid Leukemia THP-1 Cells.

Tumors highly express active heat shock protein 90 (Hsp90), which is involved in tumor survival and progression. Enhanced Toll-like receptor (TLR) 5 expression and signaling were reported to be associated with acute myeloid leukemia. In the present study, we investigated the possible modulatory effects of Hsp90 inhibitors on TLR5 expression and signaling in the human myeloid leukemia cell line THP-1. Cells were pretreated with various concentrations of the Hsp90 inhibitor geldanamycin (GA) or the Hsp70 inhibitor VER155008, followed by stimulation with bacterial flagellin. Flagellin-induced nuclear factor-κB (NF-κB) activation was significantly reduced by treatment with GA or VER155008. To elucidate the underlying mechanism of this effect, mRNA and cell surface expression of TLR5 was examined. TLR5 mRNA expression was enhanced by both GA and VER155008, whereas cell surface expression of TLR5 was reduced by three different Hsp90 inhibitors, including GA, 17-(allylamino)-17-demethoxygeldanamycin, and radicicol, and an Hsp70 inhibitor. The inhibitory effect of Hsp90 inhibitors was much higher than that of Hsp70 inhibitor. Our results suggest that Hsp90 inhibitors suppress TLR5 surface expression and activation of NF-κB in THP-1 cells in response to TLR5 ligand, and these inhibitory effects may be associated with the possible mechanisms by which Hsp90 inhibitors suppress myeloid leukemia.

Identification and Characterization of a Splicing Variant in the 5' UTR of the Human TLR5 Gene.

Toll-like receptors (TLRs) are essential components of the innate immune system. TLR5 is the receptor for flagellin, the principal protein component of bacterial flagella. The TLR5 gene has 6 exons. In an RT-PCR analysis, we found long TLR5 transcripts, in addition to those of the expected size (short TLR5 transcripts). A sequence analysis revealed that the long TLR5 transcripts contain a new exon of 94 nucleotides located between previously reported exons IV and V in the 5' untranslated region (5' UTR). A real-time PCR analysis of the two alternatively spliced variants in various cell lines showed that the long TLR5 transcripts are abundantly expressed in nonimmune cells. The ratios of long/short transcripts in human nonimmune cell lines, such as A549, T98G, HaCaT, H460, HEK-293, and Caco-2 cells, and primary mesenchymal stem cells were in the range of 1.25 to 4.31. In contrast, those of human monocytic THP-1 and U937 cells and E6.1 T cells and Ramos B cells were around 0.9. These ratios in human monocytic THP-1 cells were decreased by treatment with IFN-γ in a concentration-dependent manner. Based on our findings, we suggest that the newly found long TLR5 transcripts may be involved in the negative regulation of TLR5 expression and function.