Spatial Transcriptomics: A Powerful Tool in Disease Understanding and Drug Discovery.

Recent advancements in modern science have provided robust tools for drug discovery. The rapid development of transcriptome sequencing technologies has given rise to single-cell transcriptomics and single-nucleus transcriptomics, increasing the accuracy of sequencing and accelerating the drug discovery process. With the evolution of single-cell transcriptomics, spatial transcriptomics (ST) technology has emerged as a derivative approach. Spatial transcriptomics has emerged as a hot topic in the field of omics research in recent years; it not only provides information on gene expression levels but also offers spatial information on gene expression. This technology has shown tremendous potential in research on disease understanding and drug discovery. In this article, we introduce the analytical strategies of spatial transcriptomics and review its applications in novel target discovery and drug mechanism unravelling. Moreover, we discuss the current challenges and issues in this research field that need to be addressed. In conclusion, spatial transcriptomics offers a new perspective for drug discovery.

Scutellarin activates IDH1 to exert antitumor effects in hepatocellular carcinoma progression.

Isochlorate dehydrogenase 1 (IDH1) is an important metabolic enzyme for the production of α-ketoglutarate (α-KG), which has antitumor effects and is considered to have potential antitumor effects. The activation of IDH1 as a pathway for the development of anticancer drugs has not been attempted. We demonstrated that IDH1 can limit glycolysis in hepatocellular carcinoma (HCC) cells to activate the tumor immune microenvironment. In addition, through proteomic microarray analysis, we identified a natural small molecule, scutellarin (Scu), which activates IDH1 and inhibits the growth of HCC cells. By selectively modifying Cys297, Scu promotes IDH1 active dimer formation and increases α-KG production, leading to ubiquitination and degradation of HIF1a. The loss of HIF1a further leads to the inhibition of glycolysis in HCC cells. The activation of IDH1 by Scu can significantly increase the level of α-KG in tumor tissue, downregulate the HIF1a signaling pathway, and activate the tumor immune microenvironment in vivo. This study demonstrated the inhibitory effect of IDH1-α-KG-HIF1a on the growth of HCC cells and evaluated the inhibitory effect of Scu, the first IDH1 small molecule agonist, which provides a reference for cancer immunotherapy involving activated IDH1.

Programmed Cell Death Protein 2-like Promotes Inflammation and Oxidative Stress in Vascular Endothelial Cells.

Programmed cell death protein 2-like (PDCD2L) is a shuttle protein of the nucleus and cytoplasm and is related to the ribosome biogenesis. However, there are few reports on the relationship between PDCD2L and inflammation, and the exact relationship between PDCD2L and inflammation has not been determined in vascular endothelial cells yet. Accordingly, we focus on exploring the relationship between PDCD2L and inflammation and its potential mechanisms. Our research findings suggested that PDCD2L is a proinflammatory target. The result showed that, by interfering with the expression of PDCD2L, LPS-induced inflammation of vascular endothelial cells can be reduced, such as IL-6 and IL-1ß, as well as the adhesion factor ICAM1. Meanwhile, overexpression of PDCD2L can further increase LPS-induced inflammation levels, ICAM1, and ROS production, reduce CAT, GSH/GSSG levels, and increase SOD levels. Therefore, we determined that PDCD2L has a regulatory effect on inflammation and oxidative stress of vascular endothelial cells, and its regulatory mechanism may be related to inflammatory transcription factors STAT1, NF-κB regulation, transport of inflammatory messenger mRNA, and ribosome biogenesis. Then, we screened that andrographolide (Andro) can bind to PDCD2L, thus inhibiting inflammation and endothelial cell adhesion caused by the overexpression of PDCD2L. This study reveals that PDCD2L is a potential anti-inflammatory therapeutic target, providing new exploration for the development of anti-inflammatory drugs.

Effects of triclosan exposure on stem cells from human exfoliated deciduous teeth (SHED) fate.

Triclosan (TCS), a widely used broad-spectrum antibacterial agent and preservative, is commonly found in products and environments. Widespread human exposure to TCS has drawn increasing attention from researchers concerning its toxicological effect. However, minimal studies have focused on the impact of TCS exposure on human stem cells. Therefore, the aim of the present study was to evaluate the effects of TCS exposure on stem cells from human exfoliated deciduous teeth (SHED) and its molecular mechanisms. A series of experimental methods were conducted to assess cell viability, morphology, proliferation, differentiation, senescence, apoptosis, mitochondrial function, and oxidative stress after SHED exposure to TCS. Furthermore, transcriptome analysis was applied to investigate the response of SHED to different concentrations of TCS exposure and to explore the molecular mechanisms. We demonstrated that TCS has a dose-dependent proliferation and differentiation inhibition of SHED, while promoting cellular senescence, mitochondrial dysfunction, endoplasmic reticulum (ER) stress, and oxidative stress, as well as significantly induces apoptosis and autophagy flux inhibition at high concentrations. Interestingly, no significant morphological changes in SHED were observed after TCS exposure. Transcriptome analysis of normal and TCS-induced SHED suggested that SHED may use different strategies to counteract stress from different concentrations of TCS and showed significant differences. We discovered that TCS mediates cellular injury of SHED by enhancing the expression of PTEN, thereby inhibiting the phosphorylation levels of PI3K and AKT as well as mTOR expression. Collectively, our findings provide a new understanding of the toxic effects of TCS on human stem cell fate, which is important for determining the risk posed by TCS to human health.

Organ-on-a-chip meets artificial intelligence in drug evaluation.

Drug evaluation has always been an important area of research in the pharmaceutical industry. However, animal welfare protection and other shortcomings of traditional drug development models pose obstacles and challenges to drug evaluation. Organ-on-a-chip (OoC) technology, which simulates human organs on a chip of the physiological environment and functionality, and with high fidelity reproduction organ-level of physiology or pathophysiology, exhibits great promise for innovating the drug development pipeline. Meanwhile, the advancement in artificial intelligence (AI) provides more improvements for the design and data processing of OoCs. Here, we review the current progress that has been made to generate OoC platforms, and how human single and multi-OoCs have been used in applications, including drug testing, disease modeling, and personalized medicine. Moreover, we discuss issues facing the field, such as large data processing and reproducibility, and point to the integration of OoCs and AI in data analysis and automation, which is of great benefit in future drug evaluation. Finally, we look forward to the opportunities and challenges faced by the coupling of OoCs and AI. In summary, advancements in OoCs development, and future combinations with AI, will eventually break the current state of drug evaluation.

Influence of organic rice production mode on weed composition in the soil seed bank of paddy fields.

Introduction: It is of great significance to determine the composition of the soil weed seed bank under different organic rice production modes to provide decision making support for rational integrated weed management in organic rice production. Methods: The soil weed seed bank of the four dominant organic production modes, namely, rice-green manure rotation (RG), rice monoculture (RM), rice-crayfish coculture (RC) and rice-duck coculture (RD), with different numbers of consecutive planting years (3 to 10 years) in different sites in Jiangsu Province were investigated to determine the influence of organic rice production mode on weed composition. Results and Discussion: There were significant differences in the weed composition in the soil seed bank among the four organic rice production modes. The most dominant weed group was broadleaf weeds in the soil seed bank under the RG and RM modes; however, under the RM mode, the most dominant weed species were sedge and grass weeds. Sedge and grass weeds dominated the soil seed bank of the RC and RD modes, respectively. Therefore, specific weed management strategies could be formulated based on the differences in weed composition under different organic rice production modes. The application of organic fertilizer and irrigation were identified as primary factors associated with the differences in weed composition in the soil seed banks, which had higher effects on the weed composition than hand weeding. Consequently, fertilization and irrigation strategies that alter weed composition could be used as improved weed management program components in organic rice production systems. Long-term organic rice planting is beneficial for increasing weed diversity in paddy fields. Our results indicated that weed species diversity increased and weed community evenness and dominance decreased with the increase in the number of consecutive planting years under all four organic rice production modes.

Metformin pre-treatment of stem cells from human exfoliated deciduous teeth promotes migration and angiogenesis of human umbilical vein endothelial cells for tissue engineering.

BACKGROUND AIMS: Stem cells from human exfoliated deciduous teeth (SHED) play a significant role in tissue engineering and regenerative medicine. Angiogenesis is crucial in tissue regeneration and a primary target of regenerative medicine. As a first-line anti-diabetic drug, metformin demonstrates numerous valuable impacts on stem cells. This study aimed to explore metformin's impact and mechanism of action on SHED-mediated angiogenesis. METHODS: First, cell proliferation; flow cytometry; osteogenic, adipogenic and chondrogenic induction; and proteomics analyses were conducted to explore the role of metformin in SHED. Subsequently, migration and tube formation assays were used to evaluate chemotaxis and angiogenesis enhancement by SHED pre-treated with metformin under co-culture conditions in vitro, and relative messenger RNA expression levels were determined by quantitative reverse transcription polymerase chain reaction. Finally, nude mice were used for in vivo tube formation assay, and sections were analyzed through immunohistochemistry staining with anti-human CD31 antibody. RESULTS: Metformin significantly promoted SHED proliferation as well as osteogenic, adipogenic and chondrogenic differentiation. Proteomics showed that metformin significantly upregulated 124 differentially abundant proteins involved in intracellular processes, including various proteins involved in cell migration and angiogenesis, such as MAPK1. The co-culture system demonstrated that SHED pre-treated with metformin significantly improved the migration and angiogenesis of human umbilical vein endothelial cells. In addition, SHED pre-treated with metformin possessed greater ability to promote angiogenesis in vivo. CONCLUSIONS: In summary, the authors' findings illustrate metformin's mechanism of action on SHED and confirm that SHED pre-treated with metformin exhibits a strong capacity for promoting angiogenesis. This helps in promoting the application of dental pulp-derived stem cells pre-treated with metformin in regeneration engineering.

The potential of plant extracts in cell therapy.

Cell therapy is the frontier technology of biotechnology innovation and the most promising method for the treatment of refractory diseases such as tumours. However, cell therapy has disadvantages, such as toxicity and poor therapeutic effects. Plant extracts are natural, widely available, and contain active small molecule ingredients that are widely used in the treatment of various diseases. By studying the effect of plant extracts on cell therapy, active plant extracts that have positive significance in cell therapy can be discovered, and certain contributions to solving the current problems of attenuation and adjuvant therapy in cell therapy can be made. Therefore, this article reviews the currently reported effects of plant extracts in stem cell therapy and immune cell therapy, especially the effects of plant extracts on the proliferation and differentiation of mesenchymal stem cells and nerve stem cells and the potential role of plant extracts in chimeric antigen receptor T-cell immunotherapy (CAR-T) and T-cell receptor modified T-cell immunotherapy (TCR-T), in the hope of encouraging further research and clinical application of plant extracts in cell therapy.

Chip-DSF: A rapid screening strategy for drug protein targets.

Identification of the drug target of lead compounds is an important means for rapid and efficient drug discovery. Protein chips are a high-throughput protein function analysis technology that has been widely used in screening drug protein targets in recent years. However, the verification of the results after high-throughput protein chip screening is still cumbersome. Based on our mature protein chip preparation platform, we prepared a protein chip containing 150 important high-frequency protein targets and used antibodies to prove the availability of the protein chip. To improve the accuracy of target screening, we combined the label-free differential scanning fluorimetry (DSF) with the protein chip, proposing the Chip-DSF strategy. Subsequently, we tested the method with small molecular ginsenoside-Rg2 (Rg2). The Chip-DSF strategy was used to successfully screen the potential target protein KRAS(G12C) of Rg2. Consistently, we found that Rg2 could inhibit NCI-H23 cell proliferation by inducing cell cycle arrest. Also, we found that Rg2 could reduce the amount of KRAS protein and inhibit the phosphorylation of KRAS downstream key signaling protein ERK1, RPS6, and P70S6K in NCI-H23 cells. Collectively, our Chip-DSF strategy could achieve rapid target verification which improved the accuracy and efficiency of target screening of protein chips.

Identification of PDCD2 as a Candidate Target of Andrographolide That Arrests the Tumor Cell Cycle by Human Proteome-Scale Screening.

Andrographolide (andro) and its derivatives have been reported to have antitumor activity by arresting the cell cycle. However, the more precise mechanism has been controversial. Here, a proteome chip was used to screen drug targets in cells, and we discovered that andro can bind to PDCD2 (PD2), which has been shown to be associated with the cell cycle and mRNA nuclear export. Then, RNA-binding protein immunoprecipitation for PD2 was used to detect the quantity of cell cycle-related mRNAs, and the nuclear distribution difference analyses of these mRNAs in tumor cells after andro intervention, followed by systematic experiments, were performed to assess the downstream effects of this event in vivo and in vitro. Thus, the target spectrum of andro was revealed at the level of the human proteome chip for the first time, and this work demonstrated that andro, through targeting PD2, blocks the nuclear output of CDK mRNAs in the nucleus of tumor cells, further reduces the expression of cell CDK proteins, and finally causes tumor cell cycle arrest in phenotype and tumor tissue growth arrest in vivo.

Hydrogel supplemented with human platelet lysate enhances multi-lineage differentiation of mesenchymal stem cells.

Stem cells from human exfoliated deciduous teeth (SHED) can be used as a potential clinical material. But the use of xenogeneic ingredients will increase the risk of zoonotic disease transmission. Human platelet lysate (HPL) is a potential surrogate and used in human cell expansion with reliability in clinical applications. In this study, we synthesized chitosan/gelatin/gellan gum hydrogel supplemented with HPL and investigated the effect of 3D culture for SHED. TMT-tagged proteomics was used to decipher the secretome protein profiles of SHEDs and a total of 3209 proteins were identified, of which 23 were up-regulated and 192 were down-regulated. The results showed that hydrogel supplemented with HPL promoted SHED proliferation. After induction, the hydrogel coating contributed to osteogenic differentiation, adipogenic differentiation and differentiation into neural-like cells of SHED. SHED encapsulated in a hydrogel promotes migration and angiogenesis of HUVEC. In conclusion, our research found that hydrogel supplemented with HPL can be used as a method for SHED in standardized production and can contribute to the clinical application of SHED in cell therapy.

Chitosan Hydrogel Supplemented with Metformin Promotes Neuron-like Cell Differentiation of Gingival Mesenchymal Stem Cells.

Human gingival mesenchymal stem cells (GMSCs) are derived from migratory neural crest stem cells and have the potential to differentiate into neurons. Metformin can inhibit stem-cell aging and promotes the regeneration and development of neurons. In this study, we investigated the potential of metformin as an enhancer on neuronal differentiation of GMSCs in the growth environment of chitosan hydrogel. The crosslinked chitosan/ß-glycerophosphate hydrogel can form a perforated microporous structure that is suitable for cell growth and channels to transport water and macromolecules. GMSCs have powerful osteogenic, adipogenic and chondrogenic abilities in the induction medium supplemented with metformin. After induction in an induction medium supplemented with metformin, Western blot and immunofluorescence results showed that GMSCs differentiated into neuron-like cells with a significantly enhanced expression of neuro-related markers, including Nestin (NES) and ß-Tubulin (TUJ1). Proteomics was used to construct protein profiles in neural differentiation, and the results showed that chitosan hydrogels containing metformin promoted the upregulation of neural regeneration-related proteins, including ATP5F1, ATP5J, NADH dehydrogenase (ubiquinone) Fe-S protein 3 (NDUFS3), and Glutamate Dehydrogenase 1 (GLUD1). Our results help to promote the clinical application of stem-cell neural regeneration.

Transfer of Tumor-Bearing Mice Intestinal Flora Can Ameliorate Cognition in Alzheimer's Disease Mice.

BACKGROUND: Fecal microbiota transplant (FMT) is a potential treatment approach for many diseases. Alzheimer's disease (AD) and cancer have been proven to have a specific antagonistic relationship to FMT. OBJECTIVE: This article aims to explore whether intestinal flora transplantation from cancer individuals can ameliorate cognitive impairment. METHODS: Morris water maze and object recognition tests were performed to assess cognitive function after the fecal flora from tumor-bearing and WT mice were transplanted into AD mice by gavage. The effect of flora transplantation on AD was analyzed by thioflavin T staining, western blot, and 16S RNA sequencing. RESULTS: AD mice with FMT significantly improved short-term memory level and cognitive ability compared with Tg + NaCl group. Inflammatory factors in the plasma were regulated, and Aß plaques burden in the hippocampus and cortex were decreased. FMT in the tumor-bearing group showed a higher significant amelioration in symptoms compared to the healthy group. 16S RNA sequencing revealed that FMT treatments could reverse the increased Firmicutes and Prevotella and the decreased Bacteroidetes, Bacteroides, and Sutterella in AD mice. AD mice transplanted with tumor-bearing mice feces additionally increased the density of Oscillospira, Odoribacter, and AF12. Furthermore, the predicted functional analyses showed that the metabolism of inorganic and organic salts in the intestinal flora of AD mice was also reversed by FMT. CONCLUSION: Intestinal flora transplantation from tumor-bearing mice can ameliorate the cognitive impairment of AD mice.

Metformin enhances the osteogenesis and angiogenesis of human umbilical cord mesenchymal stem cells for tissue regeneration engineering.

Human umbilical cord mesenchymal stem cells (hUC-MSCs) are a potential clinical material in regenerative medicine applications. Metformin has shown safety and effectiveness as a clinical drug. However, the effect of metformin as a treatment on hUC-MSCs is unclear. Our research aimed to explore the effects of metformin on the osteogenesis, adipogenesis and angiogenesis of hUC-MSCs, and attempted to explain the molecular fluctuations of metformin through the mapping of protein profiles. Proliferation assay, osteogenic and adipogenic differentiation induction, cell cycle, flow cytometry, quantitative proteomics techniques and bioinformatics analysis were used to detect the influences of metformin treatment on hUC-MSCs. Our results demonstrated that low concentrations of metformin promoted the proliferation of hUC-MSCs, but high concentrations of metformin inhibited it. Metformin exhibited promotion of osteogenesis but inhibition of adipogenesis. Metformin treated hUC-MSCs up-regulated the expression of osteogenic marker ALP, OCN and RUNX2, but down-regulated the expression of adipogenic markers PPARγ and LPL. Proteomics analysis found that up-regulation of differentially expressed proteins in metformin treatment group involved the biological process of cell migration in Gene Ontology analysis. Metformin enhanced cell migration of HUVEC in a co-culture system, and hUC-MSCs treated with metformin exhibited stronger angiogenesis in vitro and in vivo compared to the hUC-MSCs group. The results of RT-qPCR revealed that the SCF and VEGFR2 were raised in metformin treatment. This study can promote the application of hUC-MSCs treated with metformin to tissue engineering for vascular reconstruction and angiogenesis.

End-to-End Automatic Pronunciation Error Detection Based on Improved Hybrid CTC/Attention Architecture.

Advanced automatic pronunciation error detection (APED) algorithms are usually based on state-of-the-art automatic speech recognition (ASR) techniques. With the development of deep learning technology, end-to-end ASR technology has gradually matured and achieved positive practical results, which provides us with a new opportunity to update the APED algorithm. We first constructed an end-to-end ASR system based on the hybrid connectionist temporal classification and attention (CTC/attention) architecture. An adaptive parameter was used to enhance the complementarity of the connectionist temporal classification (CTC) model and the attention-based seq2seq model, further improving the performance of the ASR system. After this, the improved ASR system was used in the APED task of Mandarin, and good results were obtained. This new APED method makes force alignment and segmentation unnecessary, and it does not require multiple complex models, such as an acoustic model or a language model. It is convenient and straightforward, and will be a suitable general solution for L1-independent computer-assisted pronunciation training (CAPT). Furthermore, we find that find that in regards to accuracy metrics, our proposed system based on the improved hybrid CTC/attention architecture is close to the state-of-the-art ASR system based on the deep neural network-deep neural network (DNN-DNN) architecture, and has a stronger effect on the F-measure metrics, which are especially suitable for the requirements of the APED task.