Yingying Le

Intestinal stem cells (ISCs) play critical roles in the self-renewal and regeneration of the intestinal epithelium under physiological conditions and after injury, respectively. However, the underlying mechanisms are not fully understood. In this study, we investigate the role of the G protein-coupled receptor formyl peptide receptor 2 (FPR2) in intestinal epithelium homeostasis and regeneration. In mice, knocking out Fpr2 in either intestinal epithelial cells (IECs) or ISCs significantly reduces villus height and crypt depth by impairing ISC and transit-amplifying (TA) cell proliferation and differentiation, primarily TA cell differentiation. Mechanistic studies using intestinal organoid culture and bulk and single-cell RNA sequencing revealed that activation of FPR2 promotes proliferation and differentiation of ISCs and TA cells by activating the wingless/integrated (Wnt), Notch, and Hippo signaling pathways via protein kinase C (PKC)−extracellular signal-regulated kinase (ERK). Under physiological conditions, the Wnt and Notch signaling pathways mediate the regulation of ISC proliferation and differentiation by FPR2. Fpr2 deficiency in mouse IECs exacerbates X-ray- and 5-fluorouracil-induced villus and crypt injury, and delays intestinal epithelium regeneration by reducing ISC and TA cell proliferation. Administering an FPR2 agonist to mice significantly increases survival rates and accelerates intestinal epithelium regeneration after irradiation. Taken together, these results demonstrate that intestinal epithelial FPR2 plays a key role in intestinal epithelium homeostasis and regeneration by promoting ISC and TA cell proliferation and differentiation. FPR2 is a potential therapeutic target against chemotherapy- and radiotherapy-induced intestinal injury.

Glycerol kinase (GK) participates in triglyceride (TG) synthesis by catalyzing glycerol metabolism. Whether GK contributes to nonalcoholic fatty liver (NAFL) is unclear. The expression of hepatic Gk is found to be increased in diet‐induced and genetic mouse models of NAFL and is positively associated with hepatic SREBP‐1c expression and TG levels. Cholesterol and fatty acids stimulate GK expression in hepatocytes. In HFD‐induced NAFL mice, knockdown of hepatic Gk decreases expression of SREBP‐1c and its target lipogenic genes as well as DGAT1/2, increases serum glycerol levels, decreases serum TG levels, and attenuates hepatic TG accumulation. Overexpression of GK in hepatocytes in mice or in culture produces opposite results. Mechanistic studies reveal that GK stimulates SREBP‐1c transcription directly by binding to its gene promoter and indirectly by binding to SREBP‐1c protein, thereby increasing lipogenic gene expression and de novo lipogenesis. Studies with truncated GK and mutant GKs indicate that GK induces SREBP‐1c transcription independently of its enzyme activity. GK contributes to lipid homeostasis under physiological conditions by catalyzing glycerol metabolism rather than by regulating SREBP‐1c transcription. Collectively, these results demonstrate that increased hepatic GK promotes de novo lipogenesis and TG synthesis in NAFL by stimulating SREBP‐1c transcription and DGAT1/2 expression and catalyzing glycerol metabolism.

Diabetic retinopathy (DR) is characterized by chronic, low‐grade inflammation. This state may be related to the heightened production of neutrophil extracellular traps (NETs) induced by high glucose (HG). Human cathelicidin antimicrobial peptide (LL37) is an endogenous ligand of G protein‐coupled chemoattractant receptor formyl peptide receptor 2 (FPR2), expressed on neutrophils and facilitating the formation and stabilization of the structure of NETs. In this study, we detected neutrophils cultured under different conditions, the retinal tissue of diabetic mice, and fibrovascular epiretinal membranes (FVM) samples of patients with proliferative diabetic retinopathy (PDR) to explore the regulating effect of LL37/FPR2 on neutrophil in the development of NETs during the process of DR. Specifically, HG or NG with LL37 upregulates the expression of FPR2 in neutrophils, induces the opening of mitochondrial permeability transition pore (mPTP), promotes the increase of reactive oxygen species and mitochondrial ROS, and then leads to the rise of NET production, which is mainly manifested by the release of DNA reticular structure and the increased expression of NETs‐related markers. The PI3K/AKT signaling pathway was activated in neutrophils, and the phosphorylation level was enhanced by FPR2 agonists in vitro. In vivo, increased expression of NETs markers was detected in the retina of diabetic mice and in FVM, vitreous fluid, and serum of PDR patients. Transgenic FPR2 deletion led to decreased NETs in the retina of diabetic mice. Furthermore, in vitro, inhibition of the LL37/FPR2/mPTP axis and PI3K/AKT signaling pathway decreased NET production induced by high glucose. These results suggested that FPR2 plays an essential role in regulating the production of NETs induced by HG, thus may be considered as one of the potential therapeutic targets.

To shorten the complex and time-consuming process of the identification method of the traditional food angiotensin-I-converting enzyme (ACE-I) inhibitory peptides, we propose AHTPeptideFusion based on a segmented fusion with the protein language model and deep learning. The statistical analysis found that hydrophobic amino acids, N-terminal valine is a dominant amino acid in the activity of ACE-I inhibitory peptides. In 12 machine learning (ML) algorithms, the transformer outperformed the other 11 models, with the best performance in predicting short and medium peptides. In the external dataset, AHTPeptideFusion fused by transformer and random forest (RF) showed excellent performance (accuracy > 0.9) in predicting ACE-I inhibitory peptides with lengths ranging from 2 to 15 amino acid residues and different activity distributions, and the reliability and accuracy of AHTPeptideFusion was demonstrated by synthetic peptide and ACE-I inhibition experiments. In addition, hydrogen bonding and electrostatic interaction between 4 synthetic peptides and active residues of ACE-I were found by molecular docking. To further explore the ACE-I inhibitory peptides from animal-derived foods, we established an automated pipeline consisting of the trinity of proteomics, virtual enzymatic digestion and AHTPeptideFusion, and tapped the ACE-I inhibitory peptide released from royal jelly after digestion in the gastrointestinal tract. In conclusion, this computational pipeline will become a powerful screening tool for active peptides from animal-derived foods, which can help food scientists accelerate the mining and design of active peptides from animal-derived foods. Overall, AHTPeptideFusion will be a powerful ACE-I inhibitor peptide prediction tool, it can help food scientists accelerate the mining and design of ACE-I inhibitory peptides.

The screening of enzymes for catalyzing specific substrate–product pairs is often constrained in the realms of metabolic engineering and synthetic biology. Existing tools based on substrate and reaction similarity predominantly rely on prior knowledge, demonstrating limited extrapolative capabilities and an inability to incorporate custom candidate-enzyme libraries. Addressing these limitations, we have developed the Substrate–product Pair-based Enzyme Promiscuity Prediction (SPEPP) model. This innovative approach utilizes transfer learning and transformer architecture to predict enzyme promiscuity, thereby elucidating the intricate interplay between enzymes and substrate–product pairs. SPEPP exhibited robust predictive ability, eliminating the need for prior knowledge of reactions and allowing users to define their own candidate-enzyme libraries. It can be seamlessly integrated into various applications, including metabolic engineering, de novo pathway design, and hazardous material degradation. To better assist metabolic engineers in designing and refining biochemical pathways, particularly those without programming skills, we also designed EnzyPick, an easy-to-use web server for enzyme screening based on SPEPP. EnzyPick is accessible at http://www.biosynther.com/enzypick/.

Skin epidermal stem cells (EpSCs) play a critical role in wound healing and are ideal seed cells for skin tissue engineering. Exosomes from human adipose-derived stem cells (ADSC-Exos) promote human EpSC proliferation, but the underlying mechanism remains unclear. Here, we investigated the effect of miR-100-5p, one of the most abundant miRNAs in ADSC-Exos, on the proliferation of human EpSCs and explored the mechanisms involved. MTT and BrdU incorporation assays showed that miR-100-5p mimic transfection promoted EpSC proliferation in a time-dependent manner. Cell cycle analysis showed that miR-100-5p mimic transfection significantly decreased the percentage of cells in the G1 phase and increased the percentage of cells in the G2/M phase. Myotubularin-related protein 3 (MTMR3), a lipid phosphatase, was identified as a direct target of miR-100-5p. Knockdown of MTMR3 in EpSCs by RNA interference significantly enhanced cell proliferation, decreased the percentage of cells in the G1 phase and increased the percentage of cells in the S phase. Overexpression of MTMR3 reversed the proproliferative effect of miR-100-5p on EpSCs, indicating that miR-100-5p promoted EpSC proliferation by downregulating MTMR3. Mechanistic studies showed that transfection of EpSCs with miR-100-5p mimics elevated the intracellular PIP3 level, induced AKT and ERK phosphorylation, and upregulated cyclin D1, E1, and A2 expression, which could be attenuated by MTMR3 overexpression. Consistently, intradermal injection of ADSC-Exos or miR-100-5p-enriched ADSC-Exos into cultured human skin tissues significantly reduced MTMR3 expression and increased the thickness of the epidermis and the number of EpSCs in the basal layer of the epidermis. The aforementioned effect of miR-100-5p-enriched ADSC-Exos was stronger than that of ADSC-Exos and was reversed by MTMR3 overexpression. Collectively, our findings indicate that miR-100-5p promotes EpSC proliferation through MTMR3-mediated elevation of PIP3 and activation of AKT and ERK. miR-100-5p-enriched ADSC-Exos can be used to treat skin wound and expand EpSCs for generating epidermal autografts and engineered skin equivalents.

β₂-agonists are a group of synthetic phenylethanolamine compounds which are traditionally used for treating bronchospasm. These compounds can also increase skeletal muscle mass and decrease body fat. The illegal use of β₂-agonists in food-producing animals results in residue of β₂-agonists in edible tissues and causes adverse health effects in humans. Thus, the detection of β₂-agonists at trace level in complex sample matrices is of great importance for monitoring the abuse of β₂-agonists. Many methods have been developed to detect β₂-agonists. Among them, a variety of antigen–antibody interaction-based techniques have been established to detect β₂-agonists in various samples, including animal feed, urine, serum, milk, tissues and hair. In this review, we summarized current achievement in the extraction of β₂-agonists from testing samples and detection of β₂-agonists using immunological techniques. Future perspectives were briefly discussed.

Scope: Low circulating 25‐hydroxyvitamin D (25(OH)D) levels associate with obesity, diabetes, and hyperlipidemia, but the underlying mechanisms remain uncertain. As energy‐dense diet contributes to these disorders, this study investigates whether diet could impair vitamin D metabolism.Methods and Results: Compared with control chow‐fed mice, high fat diet (HFD)‐fed mice show lower serum 25(OH)D₃ and 1,25(OH)₂D₃ levels, lower hepatic vitamin D 25‐hydroxylase Cyp2r1 expression but comparable renal vitamin D metabolic enzymes expression. Time course studies show that after HFD feeding, the serum concentrations of cholesterol, triglycerides, fatty acids, glucose, and insulin elevate sequentially and before the reduction of hepatic Cyp2r1 expression and serum 25(OH)D₃ levels. Hepatic Cyp2r1 expression also reduces after consuming high fat and high sucrose diet. After high cholesterol diet feeding, serum total cholesterol rises and hepatic Cyp2r1 expression decreases ahead of the reduction of serum 25(OH)D₃. In vitro studies demonstrate that high concentrations of cholesterol, glucose, and insulin significantly inhibit Cyp2r1expression in primary murine hepatocytes. Further studies show that dietary restriction in HFD‐fed mice ameliorates hypercholesterolemia, hyperglycemia, and hypertriglyceridemia, and elevates hepatic Cyp2r1 expression and serum 25(OH)D₃ level.Conclusion: These findings suggest that diet‐induced elevation of circulating cholesterol, glucose, and insulin reduces serum 25(OH)D₃ level through suppressing hepatic Cyp2r1 expression.