Computational Identification of Migrating T cells in Spatial Transcriptomics Data.

T cells are the central players in antitumor immunity, and effective tumor killing depends on their ability to infiltrate into the tumor microenvironment (TME) while maintaining normal cytotoxicity. However, late-stage tumors develop immunosuppressive mechanisms that impede T cell movement and induce exhaustion. Investigating T cell migration in human tumors in vivo could provide novel insights into tumor immune escape, although it remains a challenging task. In this study, we developed ReMiTT, a computational method that leverages spatial transcriptomics data to track T cell migration patterns within tumor tissue. Applying ReMiTT to ovarian tumor samples, we identified key genes and pathways that enriched on algorithm-identified T cell migration trails, including leukocyte chemotaxis, cell-cell adhesion, and ECM remodeling. We also characterized the phenotypes of T cells on the migrating trails, suggesting that regulatory T cells may accompany cytotoxic T cells during migration. Our findings provide a novel approach to studying T cell migration and interactions within the TME, offering new insights into tumor-immune dynamics.

Rastermap: a discovery method for neural population recordings.

Neurophysiology has long progressed through exploratory experiments and chance discoveries. Anecdotes abound of researchers listening to spikes in real time and noticing patterns of activity related to ongoing stimuli or behaviors. With the advent of large-scale recordings, such close observation of data has become difficult. To find patterns in large-scale neural data, we developed 'Rastermap', a visualization method that displays neurons as a raster plot after sorting them along a one-dimensional axis based on their activity patterns. We benchmarked Rastermap on realistic simulations and then used it to explore recordings of tens of thousands of neurons from mouse cortex during spontaneous, stimulus-evoked and task-evoked epochs. We also applied Rastermap to whole-brain zebrafish recordings; to wide-field imaging data; to electrophysiological recordings in rat hippocampus, monkey frontal cortex and various cortical and subcortical regions in mice; and to artificial neural networks. Finally, we illustrate high-dimensional scenarios where Rastermap and similar algorithms cannot be used effectively.

Bevacizumab combined with chemotherapy could be superior to chemotherapy alone in relapsed ovarian cancer after PARPi: evidence from a multi-center propensity score-matched analysis.

OBJECTIVE: A retrospective, multi-center propensity score-matched (PMS) analysis was conducted to investigate the efficacy and safety of the treatment strategy that combines bevacizumab and chemotherapy for patients with relapsed epithelial ovarian cancer (EOC) who previously received poly ADP-ribose polymerase inhibitors (PARPis). METHODS: A total of 250 ovarian cancer (OC) patients relapsed after PARPi received chemotherapy with or without bevacizumab at 4 medical centers were enrolled in the study. For both treatments, Kaplan-Meier analysis and Cox regression were used to compare PFS. RESULTS: In the multivariable analysis of 250 patients, the incorporation of bevacizumab into chemotherapy demonstrated a significant enhancement in PFS (hazard ratio [HR]=0.49; 95% confidence interval [CI]=0.34-0.72; p<0.001). Fifty-five patients were enrolled in Group A (bevacizumab combined with chemotherapy) and 55 were enrolled in Group B (chemotherapy alone regime) after PSM analysis. A statistically significant difference in PFS was observed between the 2 regimens (HR=0.62; 95% CI=0.40-0.97; p=0.036), suggesting that the bevacizumab combined with chemotherapy regimen confers superior clinical benefits. The median PFS was 11 months in Group A and 9 months in Group B. A significant variation was noted in PFS between patients without RCRS (HR=0.50; 95% CI=0.30-0.82) and the platinum-resistant subgroup (HR=0.31; 95% CI=0.14-0.68). Adverse effects of Grade 3-4 were more prevalent in Group A than in Group B. Additionally, instances of severe hypertension and bowel perforation were reported solely within Group A. CONCLUSION: In patients diagnosed with EOC relapsed after PARPi, the regime of chemotherapy combined with bevacizumab is associated with better PFS.

A Highly-Sensitive Omnidirectional Acoustic Sensor for Enhanced Human-Machine Interaction.

Acoustic sensor-based human-machine interaction (HMI) plays a crucial role in natural and efficient communication in intelligent robots. However, accurately identifying and tracking omnidirectional sound sources, especially in noisy environments still remains a notable challenge. Here, a self-powered triboelectric stereo acoustic sensor (SAS) with omnidirectional sound recognition and tracking capabilities by a 3D structure configuration is presented. The SAS incorporates a porous vibrating film with high electron affinity and low Young's modulus, resulting in high sensitivity (3172.9 mVpp Pa-1) and a wide frequency response range (100-20 000 Hz). By utilizing its omnidirectional sound recognition capability and adjustable resonant frequency feature, the SAS can precisely identify the desired audio signal with an average deep learning accuracy of 98%, even in noisy environments. Moreover, the SAS can simultaneously recognize multiple individuals in the auxiliary conference system and the driving commands under background music in self-driving vehicles, which marks a notable advance in voice-based HMI systems.

Quantifying the Functional Relationship Between Visual Acuity and Contrast Sensitivity Function.

Purpose: Studies have reported that individuals with certain ocular disorders may have significant decreases in contrast sensitivity function (CSF) despite having normal or near normal visual acuity (VA). This study seeks to elucidate this phenomenon by investigating the relationship between VA and CSF. Methods: We analyzed data from 14 eyes tested with Electronic Early Treatment Diabetic Retinopathy and quantitative CSF under four Bangerter foil conditions (n = 56). From the CSF data, we estimated peak gain, peak frequency, and contrast sensitivity acuity (CSA). We explored the correlations between VA and various CSF parameters and evaluated five predictive models of VA using CSA alone and in combination with additional CSF parameters through ridge regression. Results: We found that similar VA scores can correspond with markedly different CSFs and observed significant correlations among all CSF parameters and between VA and each CSF parameter (all P < 0.001). The most effective predictive model, incorporating CSA and peak gain, explained 90.97% of the variance with a root mean squared error of 0.0676 logMAR, which is comparable with the average standard deviation of the VA scores (0.0627 logMAR) and accounted for 38.6% of the residual variance not explained by the CSA-alone model. Conclusions: This study offers the first empirical inference of the quantitative relationship between VA and CSF, suggesting that various CSF parameter combinations can yield identical VA. This might help to explain why some clinical populations with normal or near-normal VA exhibit significant CSF deficits and calls for further research in different clinical settings.

Predicting contrast sensitivity functions with digital twins.

We developed and validated digital twins (DTs) for contrast sensitivity function (CSF) across 12 prediction tasks using a data-driven, generative model approach based on a hierarchical Bayesian model (HBM). For each prediction task, we utilized the HBM to compute the joint distribution of CSF hyperparameters and parameters at the population, subject, and test levels. This computation was based on a combination of historical data (N = 56), any new data from additional subjects (N = 56), and "missing data" from unmeasured conditions. The posterior distributions of the parameters in the unmeasured conditions were used as input for the CSF generative model to generate predicted CSFs. In addition to their accuracy and precision, these predictions were evaluated for their potential as informative priors that enable generation of synthetic quantitative contrast sensitivity function (qCSF) data or rescore existing qCSF data. The DTs demonstrated high accuracy in group level predictions across all tasks and maintained accuracy at the individual subject level when new data were available, with accuracy comparable to and precision lower than the observed data. DT predictions could reduce the data collection burden by more than 50% in qCSF testing when using 25 trials. Although further research is necessary, this study demonstrates the potential of DTs in vision assessment. Predictions from DTs could improve the accuracy, precision, and efficiency of vision assessment and enable personalized medicine, offering more efficient and effective patient care solutions.

High-Performance Rotating Structure Triboelectric-Electromagnetic Hybrid Nanogenerator for Environmental Wind Energy Harvesting.

As environmental energy harvesting gains increasing importance in self-powered systems and large-scale energy demands, wind energy, as a clean, pollution-free, and renewable source, has garnered widespread attention. However, achieving efficient wind energy collection remains challenging. This study proposes a high-performance rotating structure triboelectric-electromagnetic hybrid nanogenerator designed for environmental wind energy harvesting. By optimizing the magnetic circuit design of the electromagnetic generator, the dispersed radial magnetic field is converted into a unified axial magnetic field, enabling efficient power generation with only a single annular coil, thereby simplifying the generator design and reducing manufacturing and maintenance costs. Additionally, a triboelectric nanogenerator design with soft contact friction between polycarbonate (PC) fur and fluorinated ethylene propylene (FEP) film was implemented, optimizing the spacing between the electrode and friction layers, thus enhancing output performance and device durability. Furthermore, we simulated and experimentally tested the output waveform of the designed hybrid generator structure, with the results showing a high degree of similarity, further validating the rationality of the device design and providing guidance for structural optimization. Subsequently, we achieved efficient energy storage using an energy management circuit (EMC). With the integration of the EMC, the generator successfully powered a Bluetooth temperature and humidity sensor at a wind speed of 10 m/s, achieving wireless transmission, and demonstrating its potential application in traffic signal systems and other natural environmental systems. This research provides an important reference for further exploration of novel wind energy harvesting technologies.

Regulating Contact-Electro-Catalysis Using Polymer/Metal Janus Composite Catalysts.

Interfacial contact electrification can catalyze redox reactions through a process called contact-electro-catalysis (CEC). The two main reaction paths for producing reactive oxygen species via CEC are the water oxidation reaction (WOR) and the oxygen reduction reaction (ORR). Herein, we designed a polymer/metal Janus composite catalyst that regulated the reaction rates of the WOR and ORR based on the catalyst composition. The ORR was preferentially enhanced when the polymer was negatively charged during contact electrification, while the WOR was preferentially enhanced when the polymer was positively charged. This phenomenon was observed for various conductive materials. The increase in the enhancement of the reaction rates depended on the conductivity and work function of the metal. We expect that this efficient CEC method can form a universal strategy for improving the performance of existing catalysts, as contact electrification is common in nature.

Efficacy of oral treprostinil for treating pulmonary arterial hypertension: a systematic review and meta-analysis.

Introduction: Pulmonary arterial hypertension (PAH) is a progressive condition characterized by elevated blood pressure in the pulmonary arteries, leading to significant morbidity and mortality. Treprostinil, a synthetic analogue of prostacyclin, has emerged as a potential treatment option. Aim: The efficacy and safety of oral treprostinil in PAH patients remain subjects of ongoing research. Methods: Comprehensive literature searches were performed across multiple databases for studies assessing the efficacy and/or safety of treprostinil in PAH patients. Quality assessment and statistical analysis were performed using the Cochrane Collaboration's tool and Comprehensive Meta-Analysis software, respectively. Results: From 680 records identified, four studies met the inclusion criteria. These studies demonstrated that oral treprostinil significantly improved exercise capacity as measured by the 6-minute walk distance (6MWD), with a mean difference of 13.13 m in favor of treprostinil, despite high heterogeneity. The analysis also showed an increased risk of adverse events leading to discontinuation in the treprostinil group (OR = 4.39) but a protective effect against clinical worsening (OR = 0.554). No significant impact on mortality was observed. Conclusions: Oral treprostinil offers a significant benefit in improving exercise capacity in PAH patients, with a potential role in delaying clinical worsening. However, the increased risk of adverse events necessitates careful patient management. These findings support the inclusion of oral treprostinil in the treatment strategy for PAH, underscoring the need for further research to optimize its use and understand its long-term impact on patient outcomes.Level of evidence: I.

Self-powered visualized tactile-acoustic sensor for accurate artificial perception with high brightness and record-low detection limit.

The growth of the Internet of Things has focused attention on visualized sensors as a key technology. However, it remains challenging to achieve high sensing accuracy and self-power ability. Here, we propose a self-powered visualized tactile-acoustic sensor (SVTAS) based on an elaborated triboelectrification-induced electroluminescence (TIEL) unit. To date, it features a high brightness of 0.5 mW cm-2 (32 cd m-2) and a record-low detection limit of 0.5 kPa in horizontal-sliding mode. Meanwhile, the SVTAS is applicable to convert acoustic waves into TIEL signals in contact-separation mode, showing the highest response to the 44.07 Hz sound, a high signal-to-noise ratio of 8.7 dB-1, and an ultrafast response time of 0.8 ms. Furthermore, advanced artificial visualized perception systems are constructed with excellent performance in recognizing motion trajectories and human speech with different words/sentences. This work paves the way for the highly efficient and sustainable development of new-generation self-powered visualized perception systems, contributing a solution to wireless communication free from electromagnetic interference.

RECONSTRUCTING RETINAL VISUAL IMAGES FROM 3T FMRI DATA ENHANCED BY UNSUPERVISED LEARNING.

The reconstruction of human visual inputs from brain activity, particularly through functional Magnetic Resonance Imaging (fMRI), holds promising avenues for unraveling the mechanisms of the human visual system. Despite the significant strides made by deep learning methods in improving the quality and interpretability of visual reconstruction, there remains a substantial demand for high-quality, long-duration, subject-specific 7-Tesla fMRI experiments. The challenge arises in integrating diverse smaller 3-Tesla datasets or accommodating new subjects with brief and low-quality fMRI scans. In response to these constraints, we propose a novel framework that generates enhanced 3T fMRI data through an unsupervised Generative Adversarial Network (GAN), leveraging unpaired training across two distinct fMRI datasets in 7T and 3T, respectively. This approach aims to overcome the limitations of the scarcity of high-quality 7-Tesla data and the challenges associated with brief and low-quality scans in 3-Tesla experiments. In this paper, we demonstrate the reconstruction capabilities of the enhanced 3T fMRI data, highlighting its proficiency in generating superior input visual images compared to data-intensive methods trained and tested on a single subject.

ENHANCING 3T RETINOTOPIC MAPS USING DIFFEOMORPHIC REGISTRATION.

Retinotopic mapping aims to uncover the relationship between visual stimuli on the retina and neural responses on the visual cortical surface. This study advances retinotopic mapping by applying diffeomorphic registration to the 3T NYU retinotopy dataset, encompassing analyze-PRF and mrVista data. Diffeomorphic Registration for Retinotopic Maps (DRRM) quantifies the diffeomorphic condition, ensuring accurate alignment of retinotopic maps without topological violations. Leveraging the Beltrami coefficient and topological condition, DRRM significantly enhances retinotopic map accuracy. Evaluation against existing methods demonstrates DRRM's superiority on various datasets, including 3T and 7T retinotopy data. The application of diffeomorphic registration improves the interpretability of low-quality retinotopic maps, holding promise for clinical applications.

Cardiomyocyte-specific knockout of ADAM17 alleviates doxorubicin-induced cardiomyopathy via inhibiting TNFα-TRAF3-TAK1-MAPK axis.

The pathogenesis of doxorubicin-induced cardiomyopathy remains unclear. This study was carried out to test our hypothesis that ADAM17 aggravates cardiomyocyte apoptosis induced by doxorubicin and inhibition of ADAM17 may ameliorate doxorubicin-induced cardiomyopathy. C57BL/6J mice were intraperitoneally injected with a cumulative dose of doxorubicin to induce cardiomyopathy. Cardiomyocyte-specific ADAM17-knockout (A17α-MHCKO) and ADAM17-overexpressing (AAV9-oeA17) mice were generated. In addition, RNA sequencing of the heart tissues in different mouse groups and in vitro experiments in neonatal rat cardiomyocytes (NRCMs) receiving different treatment were performed. Mouse tumor models were constructed in A17fl/fl and A17α-MHCKO mice. In addition, cardiomyocyte-specific TRAF3-knockdown and TRAF3-overexpressing mice were generated. ADAM17 expression and activity were markedly upregulated in doxorubicin-treated mouse hearts and NRCMs. A17α-MHCKO mice showed less cardiomyocyte apoptosis induced by doxorubicin than A17fl/fl mice, and cardiomyocyte ADAM17 deficiency did not affect the anti-tumor effect of doxorubicin. In contrast, AAV9-oeA17 mice exhibited markedly aggravated cardiomyocyte apoptosis relative to AAV9-oeNC mice after doxorubicin treatment. Mechanistically, doxorubicin enhanced the expression of transcription factor C/EBPß, leading to increased expression and activity of ADAM17 in cardiomyocyte, which enhanced TNF-α shedding and upregulated the expression of TRAF3. Increased TRAF3 promoted TAK1 autophosphorylation, resulting in activated MAPKs pathway and cardiomyocyte apoptosis. ADAM17 acted as a positive regulator of cardiomyocyte apoptosis and cardiac remodeling and dysfunction induced by doxorubicin by upregulating TRAF3/TAK1/MAPKs signaling. Thus, targeting ADAM17/TRAF3/TAK1/MAPKs signaling holds a promising potential for treating doxorubicin-induced cardiotoxicity.

Mapping optimal orthodontic implant sites in the palate using cone-beam computed tomography.

Objective: To measure the palatal soft tissue thickness and cortical bone density to determine safe regions for the placement of orthodontic mini-implants and to examine the influence of sex and age on soft tissue thickness and cortical bone density. Materials and methods: Cone-beam computed tomography images of 42 patients (22 males and 20 females), including 21 adults and 21 adolescents, were examined in this study. The palatal soft tissue thickness and cortical bone density were measured at the coronal planes between the premolars (P4-5), between the second premolars and first molars (P5-6), and between the first molars and second molars (P6-7). Results: The thickness of the soft tissue revealed similar coronal planes, but the bone density varied. The mean thickness was 3.8 mm at 0°-60° and 1.5 mm at 60°-90°. P4-5 had the highest bone density (>600 HU), decreasing toward P6-7 (<600 HU). Bone density decreased from 90° to 0° coronally, whereas the soft tissue thickness increased. Age, sex, and their interaction affected bone and soft tissues. Conclusions: In general, areas with a high bone density tended to have thin soft tissue coronally, thus the preferred implant site tends to be more anterior to the P4-5 plane and closer to 60°-90°. Considering individual variances, mapping of the recommended regions for palatal mini-implants is suggested.

Ultra-High Sensitivity Real-Time Monitoring of Landslide Surface Deformation via Triboelectric Nanogenerator.

Monitoring surface deformation is crucial for the early warning of landslides, facilitating timely preventive measures. Triboelectric nanogenerator (TENG) demonstrates great potential for self-powered distributed monitoring in remote and power-scarce landslide areas. However, landslides deform typically at a rate of a few millimeters per day (mm d-1), making it challenging for TENG to directly monitor the deformation process. Herein, a method for monitoring surface deformation of landslides by constructing an ultra-low-speed triboelectric displacement sensor (US-TDS) is reported. Utilizing a force storage-release device and an accelerator, the US-TDS can produce obvious sensing signals at a linear input speed of 4.32 mm d-1. The coefficient of determination (R2) for the fitting curve of the pulse signals within the speed range of 21.6 to 129.6 mm d-1 reaches 0.999. Moreover, US-TDS can detect deformation displacement as small as 0.0382 mm. The stability of US-TDS displacement measurements is confirmed at a speed of 108 mm d-1, with relative errors under 1%. Ultimately, a real-time monitoring and early warning system for landslide surface deformation is constructed and verified through a combination of indoor simulations and outdoor experiments. This work provides a feasible solution for the scientific monitoring and early warning of the landslide development.

Comparison of the quantitative contrast sensitivity function between early keratoconus and normal eyes.

PURPOSE: To compare the characteristics of the quantitative contrast sensitivity function (qCSF) in eyes with early keratoconus (Early-KC) and normal control (NC) eyes and investigate the associated factors. DESIGN: A cross-sectional study. METHODS: This study included 43 eyes of 43 patients with Early-KC (including subclinical keratoconus [SKC] and forme fruste keratoconus [FFKC]) and 77 NC eyes of 77 participants with corrected distance visual acuity (CDVA) all ≥ 20/20. Contrast sensitivity (CS) was assessed using the qCSF tests. Subgroup analysis was performed according to keratoconus type(SKC and FFKC) and astigmatism(cylindrical refraction >-1.0D or ≤-1.0D). RESULTS: Sex ratio, spherical refraction, and spherical equivalent (SE) varied significantly between the two groups (all P < 0.01). The area under log CSF (AULCSF), CSF Acuity, and CS at low (1.0 and 1.5 cycles per degree [cpd]) and high (12.0 and 18.0 cpd) spatial frequencies decreased significantly in the Early-KC group than that in the NC group (all P < 0.05). The subgroup analysis revealed a similar decrease in the SKC group (all P < 0.05). AULCSF, CSF Acuity, and CS at high spatial frequencies of patients with cylindrical refraction ≤-1.0D in the Early-KC group decreased significantly (all P < 0.05) than those in the NC group. The index of vertical asymmetry and index of height decentration correlated negatively with CS at 1.5 cpd (r= -0.321 and -0.306; both P < 0.05). CONCLUSIONS: CS decreased significantly at low and high spatial frequencies in Early-KC, though with normal CDVA. The qCSF test can sensitively reflect visual performance in early keratoconus.

Enhancing CAR-T cell therapy against solid tumor by drug-free triboelectric immunotherapy.

Chimeric antigen receptor (CAR) T cell therapy is a highly effective immunotherapy for hematological tumors, but its efficacy against most solid tumors remains challenging. Herein, a novel synergistic combination therapy of drug-free triboelectric immunotherapy and CAR-T cell therapy against solid tumor was proposed. A triboelectric nanogenerator (TENG) that can generate pulsed direct-current by coupling triboelectrification effect and electrostatic breakdown effect was fabricated. The TENG can generate up to 30 pulse direct-current peaks with peak current output ≈35 µA in a single sliding to power the triboelectric immunotherapy. The pulsed direct-current stimulation induced immunogenic cell death of tumor cells (survival rate of 35.9 %), which promoted dendritic cells maturation, accelerated the process of antigen presentation to CAR-T cells and enhanced the systemic adaptive immune response. Furthermore, triboelectric immunotherapy promoted M1-like macrophage polarization, reduced regulatory T cells differentiation and reprogrammed the tumor immunosuppressive microenvironment, which ultimately enhanced the efficacy of CAR-T cells to eradicate nearly 60 % of NALM6 solid tumor mass. Notably, considering that triboelectric immunotherapy is a safe and effective drug-free antitumor strategy, the combined therapy did not increase the burden of double-medication on patients.

Key subdomains of mesencephalic astrocyte-derived neurotrophic factor attenuate myocardial ischemia/reperfusion injury by JAK1/STAT1/NF-κB signaling pathway.

BACKGROUND: Myocardial ischemia/reperfusion (I/R) injury is a common pathological process in clinical practice. Developing effective therapeutic strategies to reduce or prevent this injury is crucial. The article aimed to investigate the role and mechanism of mesencephalic astrocyte-derived neurotrophic factor (MANF) and its key subdomains in modulating myocardial I/R-induced cardiomyocyte apoptosis. METHODS: MANF stable knockout cell line and MANF mutant overexpression plasmids were constructed. The effects of MANF and mutants on apoptosis and endoplasmic reticulum (ER) stress related proteins were evaluated in hypoxia/reoxygenation-induced HL-1 cardiomyocytes by western blot, immunofluorescence, Tunel and flow cytometry. Echocardiography, ELISA, TTC and Masson were used to observe the effects of recombinant MANF protein (rMANF) on cardiac function in myocardial I/R mice. RESULTS: This study observed increased expression of MANF in both myocardial infarction patients and I/R mice. MANF overexpression in cardiomyocytes decreased ER stress-induced apoptosis, while MANF knockout exacerbated it. rMANF improved cardiac function in I/R mice by reducing injury and inflammation. This study specifically demonstrates that mutations in the α-helix of MANF were more effective in reducing ER stress and cardiomyocyte apoptosis. Mechanistically, MANF and the α-helix mutant attenuated I/R injury by inhibiting the JAK1/STAT1/NF-κB signaling pathway in addition to reducing ER stress-induced apoptosis. CONCLUSION: These findings highlight MANF and its subdomains as critical regulators of myocardial I/R injury, offering promising therapeutic targets with significant clinical implications for I/R-related diseases.

Biosynthesis of Nonribosomal Peptides Chitinimides Reveal a Special Type of Thioesterase Domains.

Non-ribosomal peptide synthetases (NRPSs) and their tailored enzymes have diverse biological functions. In this study, we investigated the biosynthesis and function of chitinimides, which belong to the non-ribosomal peptide (NRP) subfamily featuring a pyrrolidine-containing part (X part) connected to the polypeptide chain via an ester bond. A conserved gene cassette, chmHIJK, is responsible for oxyacylation of the pyrrolidine moiety in the X part. The thioesterase (TE) domain of ChmC (ChmC-TE) catalyzes transesterification reactions with a free X part or methanol as a nucleophilic reagent to form different chitinimides. The crucial amino acid residues in the ChmC-TE domains responsible for the specific recognition of the X part were identified, and they were conserved in all the biosynthetic pathways of this NRP subfamily to form a signature motif, YNHNR, suggesting a special type of TE domain in NRPSs. Chitinimides demonstrate the biological function of promoting the swarming ability of the native producer. This study provides deep insights into the biosynthesis of this special NRP subfamily, and shows that the special TE domain could be used to generate diverse NRPs by combinatorial biosynthesis.