Disseminated Talaromyces marneffei infection initially presenting as cutaneous and subcutaneous lesion in an HIV-Negative renal transplant recipient: a case report and literature review.

BACKGROUND: The incidence of Talaromyces marneffei (T. marneffei) infection has increased in recent years with the development of organ transplantation and the widespread use of immunosuppressive agents. However, the lack of clinical suspicion leading to delay or misdiagnosis is an important reason for the high mortality rate in non-human immunodeficiency virus (HIV) and non-endemic population. Herein, we report a case of disseminated T. marneffei infection in a non-HIV and non-endemic recipient after renal transplant, who initially presented with skin rashes and subcutaneous nodules and developed gastrointestinal bleeding. CASE PRESENTATION: We describe a 54-year-old renal transplantation recipient presented with scattered rashes, subcutaneous nodules and ulcerations on the head, face, abdomen, and right upper limb. The HIV antibody test was negative. The patient had no obvious symptoms such as fever, cough, etc. Histopathological result of the skin lesion sites showed chronic suppurative inflammation with a large number of fungal spores. Subsequent fungal culture suggested T. marneffei infection. Amphotericin B deoxycholate was given for antifungal treatment, and there was no deterioration in the parameters of liver and kidney function. Unfortunately, the patient was soon diagnosed with gastrointestinal bleeding, gastrointestinal perforation and acute peritonitis. Then he rapidly developed multiple organ dysfunction syndrome and abandoned treatment. CONCLUSIONS: The risk of fatal gastrointestinal bleeding can be significantly increased in kidney transplant patients with T. marneffei infection because of the long-term side effects of post-transplant medications. Strengthening clinical awareness and using mNGS or mass spectrometry technologies to improve the detection rate and early diagnosis of T. marneffei are crucial for clinical treatment in non-HIV and non-endemic population.

Crispr-Based Editing of Human Pluripotent Stem Cells for Disease Modeling.

The CRISPR system, as an effective genome editing technology, has been extensively utilized for the construction of disease models in human pluripotent stem cells. Establishment of a gene mutant or knockout stem cell line typically relies on Cas nuclease-generated double-stranded DNA breaks and exogenous templates, which can produce uncontrollable editing byproducts and toxicity. The recently developed adenine base editors (ABE) have greatly facilitated related research by introducing A/T > G/C mutations in the coding regions or splitting sites (AG-GT) of genes, enabling mutant gene knock-in or knock-out without introducing DNA breaks. In this study, we edit the AG bases in exons anterior to achieve gene knockout via the ABE8e-SpRY, which recognizes most expanded protospacer adjacent motif to target the genome. Except for gene-knockout, ABE8e-SpRY can also efficiently establish disease-related A/T-to-G/C variation cell lines by targeting coding sequences. The method we generated is simple and time-saving, and it only takes two weeks to obtain the desired cell line. This protocol provides operating instructions step-by-step for constructing knockout and point mutation cell lines.

Ultrafast energy transfer dynamics in CsPbBr3 nanoplatelets-BODIPY heterostructure.

Understanding and directing the energy transfer in nanocrystals-chromophore heterostructure is critical to improve the efficiency of their photocatalytic and optoelectronic applications. In this work, we studied the energy transfer process between inorganic-organic molecular complexes composed of cesium halide perovskite nanoplatelets (CsPbBr3 NPLs) and boron dipyrromethene (BODIPY) by photoluminescence spectroscopy (PL), time-correlated single photon-counting (TCSPC) and femtosecond transient absorption spectroscopy. The quenching of PL in CsPbBr3 NPLs occurred simultaneously with the PL enhancement of BODIPY implied the singlet energy transfer process. The rate of energy transfer has been determined by transient absorption spectrum as kET = 3.8 × 109 s-1. The efficiency of Förster energy transfer (FRET) has been quantitatively calculated up to 70%. Our work advances the understanding of the interaction between BODIPY and perovskite nanoplatelets, providing a new solution based on their optoelectronic and photocatalytic applications.

In Vivo Base Editing of Scn5a Rescues Type 3 Long QT Syndrome in Mice.

BACKGROUND: Pathogenic variants in SCN5A can result in long QT syndrome type 3, a life-threatening genetic disease. Adenine base editors can convert targeted A T base pairs to G C base pairs, offering a promising tool to correct pathogenic variants. METHODS: We generated a long QT syndrome type 3 mouse model by introducing the T1307M pathogenic variant into the Scn5a gene. The adenine base editor was split into 2 smaller parts and delivered into the heart by adeno-associated virus serotype 9 (AAV9-ABEmax) to correct the T1307M pathogenic variant. RESULTS: Both homozygous and heterozygous T1307M mice showed significant QT prolongation. Carbachol administration induced Torsades de Pointes or ventricular tachycardia for homozygous T1307M mice (20%) but not for heterozygous or wild-type mice. A single intraperitoneal injection of AAV9-ABEmax at postnatal day 14 resulted in up to 99.20% Scn5a transcripts corrected in T1307M mice. Scn5a mRNA correction rate >60% eliminated QT prolongation; Scn5a mRNA correction rate <60% alleviated QT prolongation. Partial Scn5a correction resulted in cardiomyocytes heterogeneity, which did not induce severe arrhythmias. We did not detect off-target DNA or RNA editing events in ABEmax-treated mouse hearts. CONCLUSIONS: These findings show that in vivo AAV9-ABEmax editing can correct the variant Scn5a allele, effectively ameliorating arrhythmia phenotypes. Our results offer a proof of concept for the treatment of hereditary arrhythmias.

The pseudoenzyme ADPRHL1 affects cardiac function by regulating the ROCK pathway.

BACKGROUND: Pseudoenzymes, catalytically deficient variants of active enzymes, have a wide range of regulatory functions. ADP-ribosylhydrolase-like 1 (ADPRHL1), a pseudoenzyme belonging to a small group of ADP-ribosylhydrolase enzymes that lacks the amino acid residues necessary for catalytic activity, may have a significant role in heart development based on accumulating evidence. However, the specific function of ADPRHL1 in this process has not been elucidated. To investigate the role of ADPRHL1 in the heart, we generated the first in vitro human embryonic stem cell model with an ADPRHL1 knockout. METHOD: Using the CRISPR/Cas9 system, we generated ADPRHL1 knockout in the human embryonic stem cell (hESC) H9 line. The cells were differentiated into cardiomyocytes using a chemically defined and xeno-free method. We employed confocal laser microscopy to detect calcium transients and microelectrode array (MEA) to assess the electrophysiological activity of ADPRHL1 deficiency cardiomyocytes. Additionally, we investigated the cellular mechanism of ADPRHL1 by Bulk RNA sequencing and western blot. RESULTS: The results indicate that the absence of ADPRHL1 in cardiomyocytes led to adhered abnormally, as well as perturbations in calcium transients and electrophysiological activity. We also revealed that disruption of focal adhesion formation in these cardiomyocytes was due to an excessive upregulation of the ROCK-myosin II pathway. Notably, inhibition of ROCK and myosin II effectively restores focal adhesions in ADPRHL1-deficient cardiomyocytes and improved electrical conduction and calcium activity. CONCLUSIONS: Our findings demonstrate that ADPRHL1 plays a critical role in maintaining the proper function of cardiomyocytes by regulating the ROCK-myosin II pathway, suggesting that it may serve as a potential drug target for the treatment of ADPRHL1-related diseases.

The Efficacy of Morodan in Combination with Rabeprazole for the Treatment of Chronic Gastritis and its Impact on Gastric Mucosal Repair.

Objective: This study aimed to investigate the application and effectiveness of Morodan combined with rabeprazole in patients with chronic gastritis, focusing on its impact on gastric mucosa repair. Methods: A cohort of 109 patients diagnosed with chronic gastritis, who received treatment at our hospital between January 2020 and January 2021, were included in this study. Among them, 56 patients were assigned to the control group and received treatment with rabeprazole alone, while 53 were assigned to the research group and received a combination therapy of Morodan and rabeprazole. A comparative analysis was conducted between the two groups, assessing clinical efficacy, gastric mucosa repair effects, serum-related factors, and the incidence of adverse reactions. Results: The research group exhibited a higher total effective rate of treatment (94.64%) compared to the control group (79.25%) (P < .05). Following treatment, the research group showed lower levels of pepsinogen II, serum transforming growth factor α, serum epidermal growth factor, tumor necrosis factor-α, interleukin 6, and C-reactive protein compared to the control group (P < .05). Additionally, the research group displayed higher levels of pepsinogen I compared to the control group (P < .05). There was no significant difference in the incidence of adverse reactions between the research group and the control group (P > .05). Conclusions: The combination therapy of Morodan and rabeprazole demonstrates efficacy in the treatment of chronic gastritis. It promotes gastric mucosa repair, reduces inflammatory damage, and exhibits a higher safety profile with no significant increase in adverse reactions. This treatment approach holds a higher clinical application value.

Direct Observation of Ultrafast Relaxation Dynamics of a Mixed Excimer State in Perylene Monoimide Dimer by Femtosecond Transient Absorption.

A J-type dimer PMI-2, two perylene monoimides linked by butadiynylene bridger was prepared, and its excited-state dynamics was studied using ultrafast femtosecond transient absorption spectroscopy, along with steady-state spectroscopy and quantum chemical calculations. It is evidently demonstrated that the symmetry-breaking charge separation (SB-CS) process in PMI-2 is positively mediated by an excimer, which is mixed by localized Frenkel excitation (LE) and an interunit charge transfer (CT) state. Kinetic studies show that, with the polarity increasing of the solvent, the transformation of excimer from a mixture to the CT state (SB-CS) is accelerated, and the recombination time of the CT state is reduced obviously. Theoretical calculations indicate that these are due to PMI-2 obtaining more negative free energy (ΔGcs) and lower CT state energy levels in highly polar solvents. Our work suggests that the mixed excimer can be formed in a J-type dimer with suitable structure, in which the charge separation the process is sensitive to the solvent environment.

Observation of carrier transfer in a vertical 0D-CsPbBr3/2D-MoS2 mixed-dimensional van der Waals heterojunction.

Two-dimensional transition metal dichalcogenides with outstanding properties open up a new way to develop optoelectronic devices such as phototransistors and light-emitting diodes. Heterostructure with light-harvesting materials can produce many photogenerated carriers via charge and/or energy transfer. In this paper, the ultrafast dynamics of charge transfer in zero-dimensional CsPbBr3 quantum dot/two-dimensional MoS2 van der Waals heterostructures are investigated through femtosecond time-resolved transient absorption spectroscopy. Hole and electron transfers in the ps and fs magnitude at the interfaces between MoS2 and CsPbBr3 are observed by modulating pump wavelengths of the pump-probe configurations. Our study highlights the opportunities for realizing the exciton devices based on quantum dot/two-dimensional semiconductor heterostructures.

The Molecular Network behind Volatile Aroma Formation in Pear (Pyrus spp. Panguxiang) Revealed by Transcriptome Profiling via Fatty Acid Metabolic Pathways.

Pears are popular table fruits, grown and consumed worldwide for their excellent color, aroma, and taste. Volatile aroma is an important factor affecting fruit quality, and the fatty acid metabolism pathway is important in synthesizing volatile aromas. Most of the white pear varieties cultivated in China are not strongly scented, which significantly affects their overall quality. Panguxiang is a white pear cultivar, but its aroma has unique components and is strong. The study of the mechanisms by which aroma is formed in Panguxiang is, therefore, essential to improving the quality of the fruit. The study analyzed physiological and transcriptome factors to reveal the molecular network behind volatile aroma formation in Panguxiang. The samples of Panguxiang fruit were collected in two (fruit development at 60, 90, 120, and 147 days, and fruit storage at 0, 7, 14, 21, and 28 days) periods. A total of nine sample stages were used for RNA extraction and paired-end sequencing. In addition, RNA quantification and qualification, library preparation and sequencing, data analysis and gene annotation, gene co-expression network analysis, and validation of DEGs through quantitative real-time PCR (qRT-;PCR) were performed in this study. The WGCNA identified yellow functional modules and several biological and metabolic pathways related to fatty acid formation. Finally, we identified seven and eight hub genes in the fatty acid synthesis and fatty acid metabolism pathways, respectively. Further analysis of the co-expression network allowed us to identify several key transcription factors related to the volatile aroma, including AP2/ERF-ERF, C3H, MYB, NAC, C2H2, GRAS, and Trihelix, which may also be involved in the fatty acid synthesis. This study lays a theoretical foundation for studying volatile compounds in pear fruits and provides a theoretical basis for related research in other fruits.

Low amplified spontaneous emission threshold from 2-thiophenemethylammonium quasi-2D perovskites via phase engineering.

Quasi-2D Ruddlesden-Popper perovskites attract great attention as an optical gain media in lasing applications due to their excellent optoelectronic properties. Herein, a novel quasi-2D Ruddlesden-Popper perovskite based on 2-thiophenemethylammonium (ThMA) is synthesized by a facile solution-processed method. In addition, an anti-solvent treatment method is proposed to tune the phase distribution, and preferential orientation of quasi-2D (ThMA)2Csn-1PbnBr3n+1 thin films. The large-n-dominated narrow domain distribution improves the energy transfer efficiency from small-n to large-n phases. Also, the highly oriented nanocrystals facilitate the efficient Förster energy transfer, beneficial for the carrier population transfer. Furthermore, a green amplified spontaneous emission with a low threshold of 13.92 µJ/cm2 is obtained and a single-mode vertical-cavity laser with an 0.4 nm linewidth emission is fabricated. These findings provide insights into the design of the domain distribution to realize low-threshold multicolor continuous-wave or electrically driven quasi-2D perovskites laser.

Chemically defined and small molecules-based generation of sinoatrial node-like cells.

BACKGROUND: Existing methods for in vitro differentiation of human pluripotent stem cells (hPSCs) into sinoatrial node-like cells (SANLCs) require complex and undefined medium constituents. This might hinder the elucidation of the molecular mechanisms involved in cardiac subtype specification and prevent translational application. In our study, we aimed to establish a chemically defined differentiation methods to generate SANLCs effectively and stably. METHODS: We induced human embryonic stem cells (hESCs)/induced PSCs (hiPSCs) to pan-cardiomyocytes by temporal modulation of the WNT/ß-catenin (WNT) signaling pathway with GSK3 inhibitor and WNT inhibitor. During cardiac mesoderm stage of the differentiation process, signaling of WNT, retinoid acid (RA), and fibroblast growth factor (FGF) was manipulated by three specific molecules. Moreover, metabolic selection was designed to improve the enrichment of SANLCs. Finally, RT-PCR, immunofluorescence, flow cytometry, and whole cell patch clamp were used to identify the SANLCs. RESULTS: WNT, RA, and FGF signaling promote the differentiation of hPSCs into SANLCs in a concentration- and time window-sensitive manner, respectively. Synergetic modulation of WNT, FGF, and RA signaling pathways enhance the pacemaker phenotype and improve the differentiation efficiency of SANLCs (up to 45%). Moreover, the purification based on lactate metabolism and glucose starvation further reached approximately 50% of SANLCs. Finally, the electrophysiological data demonstrate that cells differentiated with the proposed protocol produce a considerable number of SANLCs that display typical electrophysiological characteristics of pacemaker cells in vitro. CONCLUSION: We provide an optimized and chemically defined protocol to generate SANLCs by combined modulation of WNT, RA, and FGF signaling pathways and metabolic selection by lactate enrichment and glucose starvation. This chemically defined method for generating SANLCs might provide a platform for disease modeling, drug discovery, predictive toxicology, and biological pacemaker construction.

Losartan protects human stem cell-derived cardiomyocytes from angiotensin II-induced alcoholic cardiotoxicity.

Alcoholic cardiomyopathy (ACM) is a myocardial injury caused by long-term heavy drinking. Existing evidence indicates that high levels of oxidative stress are the key to pathological cardiomyopathy caused by long-term exposure to high concentrations of alcohol, while angiotensin II (AngII) and its type 1 receptor (AT1R) play an important role in excessive drinking. Whether oxidative stress-induced damage in ACM is related to AngII and AT1R is unclear, and the effects of alcohol on the electrophysiology of myocardial cells have not been reported. Most existing studies have used animal models. This study established an in vitro model of ACM based on human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs). The transcriptional profiling of alcohol treatment was performed by RNA-seq analysis. The role of oxidative stress, the expression of nicotinamide adenine dinucleotide phosphate oxidase (NOX), and the role of AngII and AT1R in the overactivation of oxidative stress were studied using fluorescent labeling, Western blotting, and high-content quantitative analysis. Real-time cell analysis(RTCA) and microelectrode array (MEA) were used to continuously monitor myocardial beating, observe the effects of alcohol on myocardial electrophysiological activity, and clarify the protective effects of the AT1R blocker losartan on ACM. We found that AngII and AT1R contribute to the effects of alcohol on the myocardium through oxidative stress damage, the mechanism of which may be achieved by regulating NOX.

Ranolazine rescues the heart failure phenotype of PLN-deficient human pluripotent stem cell-derived cardiomyocytes.

Phospholamban (PLN) is a key regulator that controls the function of the sarcoplasmic reticulum (SR) and is required for the regulation of cardiac contractile function. Although PLN-deficient mice demonstrated improved cardiac function, PLN loss in humans can result in dilated cardiomyopathy (DCM) or heart failure (HF). The CRISPR-Cas9 technology was used to create a PLN knockout human induced pluripotent stem cell (hiPSC) line in this study. PLN deletion hiPSCs-CMs had enhanced contractility at day 30, but proceeded to a cardiac failure phenotype at day 60, with decreased contractility, mitochondrial damage, increased ROS production, cellular energy metabolism imbalance, and poor Ca2+ handling. Furthermore, adding ranolazine to PLN knockout hiPSCs-CMs at day 60 can partially restore Ca2+ handling disorders and cellular energy metabolism, alleviating the PLN knockout phenotype of HF, implying that the disorder of intracellular Ca2+ transport and the imbalance of cellular energy metabolism are the primary mechanisms for PLN deficiency pathogenesis.

Generation of a COL1A2 homozygous knockout stem cell line via CRISPR/Cas9 system.

The loss of function of the COL1A2 gene can result in osteogenesis imperfecta (OI) types I, II, III, and IV and Ehlers-Danlos syndrome (cardiac valvular and arthrochalasia type).To further investigate the significance of COL1A2 in osteogenesis imperfecta and cardiac valve disease, we created a homozygous COL1A2-/- human embryonic stem cell line (WAe009-A-72) using CRISPR/Cas9. In vivo, the WAe009-A-72 cell line retained typical colony form, a normal karyotype, and robustly expressed pluripotency markers while differentiating into all three germ layers. This cell line is a potential tool for investigating the role of the COL1A2 gene in associated disorders.

Generation of a homozygous MYH7 gene knockout human embryonic stem cell line (WAe009-A-69) using an episomal vector-based CRISPR/Cas9 system.

Myosin heavy chain 7 (MYH7) encodes the human heart myosin heavy chain subunit, which plays an important role in myocardial contraction. MYH7 is the main pathogenic gene that causes Hypertrophic cardiomyopathy (HCM) and Dilated cardiomyopathy (DCM). In this experiment, a MYH7 homozygous knockout human embryonic stem cell (hESC) line, WAe009-A-69, was generated using an episomal vector-based CRISPR/Cas9 system. It can be an ideal tool to further study the function of MYH7. The cell line was confirmed with pluripotency, normal karyotype and trilineage differentiation potential.

Efficient Correction of a Hypertrophic Cardiomyopathy Mutation by ABEmax-NG.

[Figure: see text].

Generation of a COL4A5 heterozygous mutation human embryonic stem cell line (WAe009-A-58) using an episomal vector-based CRISPR/Cas9 system.

X-linked Alport syndrome (XLAS) is the second most common inherited kidney disease which pathogenic variants related to a mutation in the COL4A5 gene encoding the type IV collagen α5 chain. Here, we have generated a COL4A5 heterozygous mutant human embryonic stem cell (hESC) line (H9-COL4A5+/-) by an episomal vector-based CRISPR/Cas9 system. The generated H9-COL4A5+/- maintained a normal stem cell morphology, stably expressed pluripotent markers, and could differentiate into all three germ layers in vivo. This cell line offers an in vitro efficient platform to explore pathogenic mechanisms in XLAS and provides a cell-based disease model for drug testing.

Generation of a TPM1 homozygous knockout embryonic stem cell line by CRISPR/Cas9 editing.

Alpha-Tropomyosin (TPM1) plays a crucial role in actin regulation and stability and contributes fundamental functions to heart development: without TPM1 expressing, mice embryos will die early in embryogenesis. To further identify the role of TPM1 in human cardiac development, here we generated a homozygous TPM1 knockout (TPM1-/-) human embryonic stem cell (hESC) line using CRISPR/Cas9-based genome editing system. The generated TPM1-/- hESC line maintained normal karyotype, highly expressed pluripotency markers and was able to differentiate into all three germ layers in vivo. This cell line provides a powerful tool to investigate the role of TPM1 in heart development in future.

hERG-deficient human embryonic stem cell-derived cardiomyocytes for modelling QT prolongation.

BACKGROUND: Long-QT syndrome type 2 (LQT2) is a common malignant hereditary arrhythmia. Due to the lack of suitable animal and human models, the pathogenesis of LQT2 caused by human ether-a-go-go-related gene (hERG) deficiency is still unclear. In this study, we generated an hERG-deficient human cardiomyocyte (CM) model that simulates 'human homozygous hERG mutations' to explore the underlying impact of hERG dysfunction and the genotype-phenotype relationship of hERG deficiency. METHODS: The KCNH2 was knocked out in the human embryonic stem cell (hESC) H9 line using the CRISPR/Cas9 system. Using a chemically defined differentiation protocol, we obtained and verified hERG-deficient CMs. Subsequently, high-throughput microelectrode array (MEA) assays and drug interventions were performed to characterise the electrophysiological signatures of hERG-deficient cell lines. RESULTS: Our results showed that KCNH2 knockout did not affect the pluripotency or differentiation efficiency of H9 cells. Using high-throughput MEA assays, we found that the electric field potential duration and action potential duration of hERG-deficient CMs were significantly longer than those of normal CMs. The hERG-deficient lines also exhibited irregular rhythm and some early afterdepolarisations. Moreover, we used the hERG-deficient human CM model to evaluate the potency of agents (nifedipine and magnesium chloride) that may ameliorate the phenotype. CONCLUSIONS: We established an hERG-deficient human CM model that exhibited QT prolongation, irregular rhythm and sensitivity to other ion channel blockers. This model serves as an important tool that can aid in understanding the fundamental impact of hERG dysfunction, elucidate the genotype-phenotype relationship of hERG deficiency and facilitate drug development.

Cell sheet formation enhances the therapeutic effects of human umbilical cord mesenchymal stem cells on myocardial infarction as a bioactive material.

Stem cell-based therapy has been used to treat ischaemic heart diseases for two decades. However, optimal cell types and transplantation methods remain unclear. This study evaluated the therapeutic effects of human umbilical cord mesenchymal stem cell (hUCMSC) sheet on myocardial infarction (MI). METHODS: hUCMSCs expressing luciferase were generated by lentiviral transduction for in vivo bio-luminescent imaging tracking of cells. We applied a temperature-responsive cell culture surface-based method to form the hUCMSC sheet. Cell retention was evaluated using an in vivo bio-luminescent imaging tracking system. Unbiased transcriptional profiling of infarcted hearts and further immunohistochemical assessment of monocyte and macrophage subtypes were used to determine the mechanisms underlying the therapeutic effects of the hUCMSC sheet. Echocardiography and pathological analyses of heart sections were performed to evaluate cardiac function, angiogenesis and left ventricular remodelling. RESULTS: When transplanted to the infarcted mouse hearts, hUCMSC sheet significantly improved the retention and survival compared with cell suspension. At the early stage of MI, hUCMSC sheet modulated inflammation by decreasing Mcp1-positive monocytes and CD68-positive macrophages and increasing Cx3cr1-positive non-classical macrophages, preserving the cardiomyocytes from acute injury. Moreover, the extracellular matrix produced by hUCMSC sheet then served as bioactive scaffold for the host cells to graft and generate new epicardial tissue, providing mechanical support and routes for revascularsation. These effects of hUCMSC sheet treatment significantly improved the cardiac function at days 7 and 28 post-MI. CONCLUSIONS: hUCMSC sheet formation dramatically improved the biological functions of hUCMSCs, mitigating adverse post-MI remodelling by modulating the inflammatory response and providing bioactive scaffold upon transplantation into the heart. TRANSLATIONAL PERSPECTIVE: Due to its excellent availability as well as superior local cellular retention and survival, allogenic transplantation of hUCMSC sheets can more effectively acquire the biological functions of hUCMSCs, such as modulating inflammation and enhancing angiogenesis. Moreover, the hUCMSC sheet method allows the transfer of an intact extracellular matrix without introducing exogenous or synthetic biomaterial, further improving its clinical applicability.