Sequencing Proteomics Research Articles

Unveiling immunogenic characteristics and neoantigens in endometrial cancer with POLE hotspot mutations for improved immunotherapy

BackgroundImmunotherapy, especially with the use of immune checkpoint inhibitors, has demonstrated efficacy for a variety of malignant tumors. However, the potential of immunotherapy for endometrial cancer (EC) with POLE mutations remains underexplored.MethodsWe utilized multiple databases and clinical specimens to investigate the immunogenicity profiles of EC patients carrying POLE mutations. One particular hotspot mutation POLEP286R was identified and further studied. Consequently, by constructing human leukocyte antigen (HLA) tetramers and incubating them with patients’ peripheral blood mononuclear cells (PBMCs), T cells capable of recognizing the POLEP286R mutation were sorted for further transcriptomic, proteomic and T-cell receptor (TCR) sequencing analyses and for an organoid EC model.ResultsTumor- and immune-related pathways were shown to be activated in the POLEP286R mutant group. Importantly, by using an organoid model of EC, we further confirmed the antitumor potential of T cells that were specific to the POLEP286R mutation.ConclusionsOur study uncovers the pronounced immunogenicity of POLE-mutant EC and characterizes neoantigens that are unique to the POLEP286R mutation, thus providing a promising new immunotherapeutic strategy for EC.

Piplartine alleviates sepsis-induced acute kidney injury by inhibiting TSPO-mediated macrophage pyroptosis.

Sepsis-induced acute kidney injury (SI-AKI) is the most common organ dysfunction of sepsis, characterized with prolonged hospitalization periods and significantly elevated mortality rates. Piplartine (PLG), an alkaloid extracted from Piper longum within the Piperaceae family, has exhibited diverse pharmacological activities, including anti-inflammatory, anti-atherosclerotic, and anti-tumor effects. Herein, we investigated whether the PLG could reverse SI-AKI and explore its possible anti-inflammatory mechanisms. We constructed an SI-AKI model using cecal ligation and puncture (CLP) and systematically evaluated the protective effect of PLG administered by gavage in the SI-AKI mice. Subsequently, we performed proteomic sequencing of the kidney and integrated data from the GeneCards and SwissTargetPrediction databases to identify potential targets and mechanisms. Immunofluorescence and western blotting were used to examine the expression of relevant targets and pathways in vivo and in vitro. The influence of PLG on the predicted target and pathway was verified using an agonist of the target protein and a series of biochemical experiments. PLG exhibited significant efficacy against pathological damage, neutrophil and macrophage infiltration, and macrophage pyroptosis in kidneys at 30 mg/kg. An integrated analysis of proteomic data identified the translocator protein (TSPO) as a potential target for the renoprotective effects of PLG. Moreover, a TSPO agonist (RO5-4864) prominently reversed the protective effect of PLG in SI-AKI mice, as manifested by a deterioration in renal function, histopathological lesions and macrophage pyroptosis in the kidneys. Our results suggest that PLG may ameliorate SI-AKI, potentially through partial inhibition of the TSPO-macrophage pyroptosis pathway.

Unveiling the role of microRNAs in nonhost resistance to Sclerotinia sclerotiorum: Rice-specific microRNAs attack the pathogen via cross-kingdom RNAi.

The development of rapeseed with high resistance against the pathogen Sclerotinia sclerotiorum is impeded by the lack of effective resistance resources within host species. Unraveling the molecular basis of nonhost resistance (NHR) holds substantial value for resistance improvement in crops. In the present study, small RNA sequencing and transcriptome sequencing were carried out between rice (a nonhost species of S. sclerotiorum) and rapeseed during infection, revealing the involvement of rice miRNAs on translation-related processes in both rice and the pathogen. Specifically, rice-specific miRNAs with potential capability for cross-kingdom RNAi against S. sclerotiorum were explored, of which Os-miR169y was selected as a representative case to elucidate its role in resistance to S. sclerotiorum. The silence of Os-miR169y decreased the resistance level of rice to S. sclerotiorum, and heterologous expression of Os-miR169y in Arabidopsis and rapeseed significantly enhanced the host resistance. The dual-luciferase reporter assay indicates that Os-miR169y targets S. sclerotiorum 60S ribosomal protein L19 (SsRPL19). Overexpressing Os-miR169y (OEss-miR169y) and RNAi of SsRPL19 (RNAiss-RPL19) in S. sclerotiorum significantly impaired the growth and pathogenicity of the pathogen, while overexpressing SsRPL19 exhibited a contrast effect. Yeast-two-hybridization revealed an interlinking role of SsRPL19 with multiple large and small ribosomal subunits, indicating its important role in translation. Proteome sequencing detected a decreased amount of proteins in transformants OEss-miR169y and RNAiss-RPL19 and significant suppression on key metabolic pathways such as carbon and nitrogen metabolisms. Collectively, this study suggests that rice can secrete specific miRNAs to suppress genes essential for S. sclerotiorum, such as Os-miR169y, which targets and suppresses SsRPL19 and thus impairs protein synthesis in the pathogen. This study sheds light on the intrinsic mechanisms of rice NHR against S. sclerotiorum, and further demonstrates the potential of using nonhost-specific "pathogen-attacking" miRNAs in improving resistance in host species.

Ocular inoculation of toad venom: toxic cataract and proteomic profiling.

To report a singular case of cataract caused by toad venom inoculation and to scrutinize the pathological mechanisms through proteomic sequencing of the lens specimen. A young Chinese male presented with progressively deteriorating vision in his right eye subsequent to a history of toad venom inoculation. He was diagnosed with a toxic cataract, and underwent phacoemulsification cataract surgery. Anterior capsule, nucleus, and cortex specimens from the patient (designated as PT_CAP, PT_PHACO, and PT_CTX, respectively) and age-related cataract controls (C_CAP, C_PHACO, and C_CTX, respectively) were collected and subjected to 4D label-free quantitative proteomics. A multitude of differentially expressed proteins (DEPs) were identified in the patient's lens compared to those in the controls. Specifically, a total of 204 DEPs were identified in PT_CAP compared to C_CAP, with MYH6, MYL2, MYL3, STAT1, and ANK1 among the foremost regulated DEPs. The DEPs of PT_CAP were principally affiliated with functions including "transportation of small molecules," "regulation of metal ion transport," and "import into cell." A sum of 109 DEPs were delineated in PT_CTX compared to C_CTX, with TPM1 among the top-10 downregulated DEPs. Ninety-five DEPs were pinpointed in PT_PHACO compared to C_PHACO, with hexokinase among the top 10 downregulated DEPs. These proteins were ascertained to be linked with Na+/K+-ATPase activity. This study introduced the first documented case of toxic cataract caused by toad venom inoculation. Proteomic sequencing indicated a correlation between cataract and alterations in Na+/K+-ATPase activity, providing insights for the clinical management of ocular toad venom inoculation in subsequent cases.

Conservative mechanism through various rapeseed (Brassica napus L.) varieties respond to heavy metal (Cadmium, Lead, Arsenic) stress.

Heavy metal soil pollution is a global issue that can be efficiently tackled through the process of phytoremediation. The use of rapeseed in the phytoremediation of heavy metal-contaminated agricultural land shows great potential. Nevertheless, its ability to tolerate heavy metal stress at the molecular level remains unclear. Here, with 7-day seedlings as raw materials, we investigated physiological and biochemical indexes, analyzed the transcriptome sequencing for different treated materials (control, 50×, and 100×), combined with the results of transcriptome and proteome sequencing of the near-isogenic lines (F338 and F335) to reveal the response mechanism to heavy metal stress. Due to oxidative stress response caused by heavy metal stress, there are heavy effects on the emergence of rapeseeds and the growth of seedlings. Although rapeseed can alleviate oxidative stress by enhancing the enzyme activity, especially peroxidase in the oxidation system, this process has its limits. Rapeseed plants activate antioxidase, transport enzymes, and biological regulation to cope with heavy metal stress. Among these responses, peroxidase, ABC transporters, and abscisic acid are particularly significant in this process. Based on this study, we identified a breeding material with high adsorption capacity for heavy metals, which contributed to the research on resistance breeding in rapeseed. The results of this study may be useful to alleviate heavy metal soil pollution and tackle edible oil shortages in China.

Distribution characteristics and proteomic analysis of glioma-associated oncogene homolog 1 positive cells during mouse orthodontic tooth movement

Objective: To explore the distribution characteristics of glioma-associated oncogene homolog 1 (Gli1) positive cells during orthodontic tooth movement process and conduct a proteomic analysis of these cells. Methods: Forty Gli1-LacZ transgenic mice were used to establish an in vivo orthodontic tooth movement (OTM) model for labeling Gli1 positive cells in Gli1-LacZ transgenic mice (OTM group) and an unforced control group, with tooth movement distance measured using micro-CT. The spatial relationship and distribution characteristics of Gli1 positive cells and H-type vessels of CD31 and endomucin (EMCN) in periodontal tissues were detected by immunofluorescence staining. Twenty Gli1-membrane-targeted tandem dimer Tomato (mT)/membrane-targeted green fluorescent protein (mG) double-genotype mice were bred and Gli1 positive cells were sorted for proteomic sequencing after tamoxifen induction. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) databases were used for enrichment analysis. Results: The micro-CT three-dimensional reconstruction results showed that the mesial movement of the maxillary first molar in mice after 7 days of force application was (69±15) μm, indicating the successful establishment of the Gli1-LacZ transgenic mouse OTM model. Immunofluorescence staining showed that the blood vessels in periodontal tissue were mostly H-type vessels of CD31 and EMCN. The blood vessels in the periodontal tissues are predominantly H-type vessels positive for both CD31 and EMCN. The percentage of Gli1 positive cells in the OTM group, expressed as (54.5±13.2)%, and the relative fluorescence intensity, expressed as 2.6±0.9, were both significantly greater than those in the control group, which had a Gli1 positive cell percentage of (36.3±9.1)% (t=3.60, P=0.002) and a relative fluorescence intensity of 1.0±0.3 (t=5.20, P<0.001). In contrast to the control group where only a small number of Gli1 positive cells were consistent with the distribution of H-type vessels, in the OTM group the number of Gli1 positive cells increased on the tension side were closely associated with the spatial distribution of H-type vessels. GO enrichment analysis of biological processes found that a large number of proteins in Gli1 positive cells were enriched in pathways such as angiogenesis and tissue remodeling. KEGG enrichment analysis found that related proteins were mainly enriched in pathways related to angiogenesis and Gli1, such as hypoxia-inducing factor 1 signaling pathway, vascular endothelial growth factor signaling pathway and hedgehog signaling pathway. Conclusions: The number of Gli1 positive cells increased on tension side and were closely related to H-type blood vessels in response to mechanical force during orthodontic tooth movement. This may be related to profile of inducing blood vessel formation and tissue remodeling.

Therapeutic effects of platelet-derived extracellular vesicles on viral myocarditis correlate with biomolecular content.

Extracellular vesicles (EVs) can potently inhibit inflammation yet there is a lack of understanding about the impact of donor characteristics on the efficacy of EVs. The goal of this study was to determine whether the sex and age of donor platelet-derived EVs (PEV) affected their ability to inhibit viral myocarditis. PEV, isolated from men and women of all ages, was compared to PEV obtained from women under 50 years of age, which we termed premenopausal PEV (pmPEV). Because of the protective effect of estrogen against myocardial inflammation, we hypothesized that pmPEV would be more effective than PEV at inhibiting myocarditis. We injected PEV, pmPEV, or vehicle control in a mouse model of viral myocarditis and examined histology, gene expression, protein profiles, and performed proteome and microRNA (miR) sequencing of EVs. We found that both PEV and pmPEV significantly inhibited myocarditis; however, PEV was more effective, which was confirmed by a greater reduction of inflammatory cells and proinflammatory and profibrotic markers determined using gene expression and immunohistochemistry. Proteome and miR sequencing of EVs revealed that PEV miRs specifically targeted antiviral, Toll-like receptor (TLR)4, and inflammasome pathways known to contribute to myocarditis while pmPEV contained general immunoregulatory miRs. These differences in EV content corresponded to the differing anti-inflammatory effects of the two types of EVs on viral myocarditis.

Effect of the S100A9/AMPK pathway on PM2.5-mediated mouse lung injury.

Particulate matter 2.5 (PM2.5), particles with an aerodynamic diameter less than 2.5 µm, affect lung function and increase respiratory disease incidence and mortality rate. The molecular mechanism of lung injury and epithelial damage after PM2.5 exposure is not completely clear. Mouth-nose exposure of mice was performed with PM2.5 or neutral saline. In vitro experiments were conducted to investigate the role of the S100A9/AMPK pathway in PM2.5-mediated lung injury. PM2.5 exposure in mice caused lung epithelial damage, alveolar wall thickening, and alveolar wall structure destruction. The 16S rRNA sequencing results suggested that the microecology structure of lung tissue was altered after PM2.5 exposure. Proteomic sequencing was performed to explore the underlying mechanism, and 71 differentially expressed proteins were identified. KEGG database analysis of the up-regulated differential proteins revealed regulatory networks, including fat digestion and absorption, the AMPK signaling pathway, and the PPAR signaling pathway. Moreover, PM2.5 exposure in mice increased the level of S100A9 and ROS, leading to reduction of the ATP level. To achieve a sufficient energy supply by increasing fatty acid transfer and oxidation, activated AMPK up-regulates CD36 and CPT1, which leads to mitochondrial damage of PM2.5-exposed cells and injury or death of lung epithelial cells. siRNA-S100A9 and AMPK inhibitors significantly reduced the occurrence of cell damage. These results may help to clarify biomarkers and specific mechanisms of lung tissue injury induced by PM2.5 exposure.

METTL3-Mediated m6A Modification of ISG15 mRNA Regulates Doxorubicin-Induced Endothelial Cell Apoptosis.

N6-adenosine methylation (m6A) of RNA is involved in the regulation of various diseases. However, its role in chemotherapy-related vascular endothelial injury has not yet been elucidated. We found that methyltransferase-like 3 (METTL3) expression was significantly reduced during doxorubicin (DOX)-induced apoptosis of vascular endothelial cells both invivo and invitro, and that silencing of METTL3 further intensified this process. Combined transcriptome and proteome sequencing analyses revealed that the expression levels of interferon-stimulated gene 15 (ISG15) mRNA and protein significantly increased after METTL3 silencing. Methylated RNA immunoprecipitation (meRIP)-quantitative polymerase chain reaction (qPCR) and mRNA stability assays confirmed that METTL3 regulates the expression of ISG15 by methylating the 1,014,147 site on ISG15 RNA, thereby decreasing ISG15 mRNA levels. Silencing ISG15 significantly suppressed DOX-induced endothelial cell apoptosis and dysfunction caused by METTL3 silencing. In summary, our study revealed that METTL3-mediated methylation of ISG15 mRNA is involved in DOX-induced endothelial cell apoptosis and explored potential therapeutic targets for alleviating chemotherapy-associated vascular injury.

Integrated Proteomic and Metabolomic Analysis of Muscle Atrophy Induced by Hindlimb Unloading.

Skeletal muscle atrophy, which is induced by factors such as disuse, spaceflight, certain medications, neurological disorders, and malnutrition, is a global health issue that lacks effective treatment. Hindlimb unloading is a commonly used model of muscle atrophy. However, the underlying mechanism of muscle atrophy induced by hindlimb unloading remains unclear, particularly from the perspective of the myocyte proteome and metabolism. We first used mass spectrometry for proteomic sequencing and untargeted metabolomics to analyze soleus muscle changes in rats with hindlimb unloading. The study found 1052 proteins and 377 metabolites (with the MS2 name) that were differentially expressed between the hindlimb unloading group and the control group. Proteins like ACTN3, MYH4, MYBPC2, and MYOZ1, typically found in fast-twitch muscles, were upregulated, along with metabolism-related proteins GLUL, GSTM4, and NDUFS4. Metabolites arachidylcarnitine and 7,8-dihydrobiopterin, as well as pathways like histidine, taurine, and hypotaurine metabolism, were linked to muscle atrophy. Protein and metabolism joint analyses revealed that some pathways, such as glutathione metabolism, ferroptosis, and lysosome pathways, were likely to be involved in soleus atrophy. In this study, we have applied integrated deep proteomic and metabolomic analyses. The upregulation of proteins that are expressed in fast-twitch fibers indicates the conversion of slow-twitch fibers to fast-twitch fibers under hindlimb unloading. In addition, some differentially abundant metabolites and pathways revealed the important role of metabolism in muscle atrophy of the soleus. As shown in the graphical abstract, our study provides insights into the pathogenesis and treatment of muscle atrophy that results from unloading by integrating proteomics and metabolomics of the soleus muscles.

Distinct patterns of cell adhesion, migration, and morphology in olfactory neuroepithelium cells of bipolar disorder patients

Abstract Background Bipolar disorder (BD) is a severe, chronic mental illness that remains difficult to diagnose due to the lack of specific biomarkers, relying primarily on clinical assessments. Early diagnosis and treatment are essential for improving prognosis and lowering suicide risk. This study aimed to identify biomarkers and therapeutic targets by utilizing olfactory neuroepithelium (ONE) cells from patients with BD and controls. Methods Immunofluorescence of ONE cells, along with proteomic and RNA sequencing analyses, was performed to investigate cytoskeletal changes and pathways involved in cell adhesion, movement, and morphology. Additionally, potential biomarkers were investigated in blood samples to improve clinical accessibility. Results Thus, according to functional assays, ONE cells derived from BD patients exhibited decreased substrate adhesion, reduced cell migration, and morphological changes compared to control cells. In addition, proteomic and RNAseq analyses in ONE cells and peripheral blood mononuclear cells (PBMCs) revealed alterations in pathways such as RhoA/PAK/Integrin and Actin Cytoskeleton Signaling, as well as significant changes in inflammatory and immunological pathways. AUROC analysis identified proteins like PTK2 as potential diagnostic biomarkers, showing altered expression in both ONE cells and PBMCs. PTK2 RNA expression correlated with distinct morphological traits in BD ONE cells. Conclusions In summary, this study identified cytoskeletal alterations, reduced adhesion, and disrupted migration patterns in BD ONE cells, highlighting molecular mechanisms underlying these changes and emphasizing PTK2’s role as a potential diagnostic biomarker for BD.

Revealing the key role of the folate synthesis regulatory gene DHNA in tobacco leaf yellowing based on BSA-seq, RNA-seq, and proteomic sequencing

BackgroundThe degree of yellowing in tobacco leaves is an important indicator for determining the maturity and harvesting time of tobacco leaves. Decreasing chlorophyll levels helps speed up the ripening process of tobacco leaves for easier mechanical harvesting. Identifying and utilizing genes that regulate yellowing in tobacco leaves are crucial for developing tobacco varieties suitable for mechanized harvesting and understanding the molecular processes that control leaf color changes.ResultsIn this study, the phenotypes of the yellow-leaf K326 and K326 varieties were analysed, and it was observed that the yellow-leaf K326 variety exhibited a distinct yellow leaf phenotype with a significant reduction in chlorophyll content. Subsequently, using a combination of BSA-seq, transcriptomic sequencing (RNA-seq), and proteomic sequencing approaches, we identified the candidate gene Nitab4.5_0008674g0010 that encodes dihydroneopterin aldolase as a factor associated with tobacco leaf yellowing. Finally, by measuring the folate content in K326 and Huangye K326, the folate content in Huangye K326 was observed to be significantly lower than that in K326, thus indicating that folate synthesis plays a crucial role in phenotypic changes in tobacco yellow leaves.ConclusionsThis study is the first to use BSA-seq combined with RNA-seq and proteomic sequencing to identify candidate genes in tobacco yellow leaves. The results provide a theoretical basis for the analysis of the mechanism of tobacco yellow leaf mutations.

Single-cell RNA sequencing reveals the contribution of smooth muscle cells and endothelial cells to fibrosis in human atrial tissue with atrial fibrillation

AimsAtrial fibrillation (AF) has high mortality and morbidity rates. However, the intracellular molecular complexity of the atrial tissue of patients with AF has not been adequately assessed.Methods and resultsWe investigated the cellular heterogeneity of human atrial tissue and changes in differentially expressed genes between cells using single-cell RNA sequencing, fluorescence in situ hybridization, intercellular communication, and cell trajectory analysis. Using genome-wide association studies (GWAS) and proteomics, we discovered cell types enriched for AF susceptibility genes. We discovered eight different cell types, which were further subdivided into 23 subpopulations. In AF, the communication strength between smooth muscle cells (SMCs) and fibroblast (FB) 3 cells increased and the relevant signaling pathways were quite similar. Subpopulations of endothelial cells (ECs) are mainly involved in fibrosis through TXNDC5 and POSTN. AF susceptibility genes revealed by GWAS were especially enriched in neuronal and epicardial cells, FB3, and lymphoid (Lys) cells, whereas proteomic sequencing differential proteins were concentrated in FB3 cells and SMCs.ConclusionsThis study provides a cellular landscape based on the atrial tissue of patients with AF and highlights intercellular changes and differentially expressed genes that occur during the disease process. A thorough description of the cellular populations involved in AF will facilitate the identification of new cell-based interventional targets with direct functional significance for the treatment of human disease.

Transcriptomic and proteomic sequencing unveils the role of vitamin D and metabolic flux shifts in the induction of human hepatic organoids

BackgroundHepatic organoids (HOs), validated through comparative sequencing with human liver tissues, are reliable models for liver research. Comprehensive transcriptomic and proteomic sequencing of HOs throughout their induction period will enhance the platform’s utility, aiding in the elucidation of liver development’s molecular mechanisms.MethodsWe developed hepatic organoids (HOs) from embryonic stem cells (ESCs) through a de novo induction protocol, mimicking the stages of fetal liver development: ESCs to definitive endoderm (DE), then to foregut (FG), hepatoblasts (HB), and finally to HOs stage 1 (HO1), culminating in self-organizing HOs stage 2 (HO2) via dissociation and re-inoculation. The successful establishment of HOs was validated by immunofluorescence staining and RT-qPCR for specific markers. Comprehensive transcriptomic and proteomic sequencing and analysis were conducted on FG, HB, HO1, and HO2.ResultsOur data suggest that several transcription factors (TFs) activated during the HB stage share overlapping target genes with the vitamin D receptor (VDR). Calcitriol, a direct activator of VDR, notably facilitated the FG to HB stage transition by activating VDR and enhancing key TFs, thereby promoting hepatic progenitor cell maturation. Furthermore, our findings revealed a significant transition towards glycolytic energy metabolism at the HO2 stage, characterized by increased glycolytic flux and reduced oxidative phosphorylation. Inhibition of glycolysis using 2-deoxy-D-glucose (2-DG) led to suppressed growth and differentiation at the HO2 stage. Analysis of signaling pathways indicated upregulation of the HIF-1 pathway, which is associated with glycolysis activation, as well as the MAPK and PI3K-AKT pathways, which regulate HIF-1α protein translation.ConclusionsWe elucidated a pivotal role for calcitriol in facilitating the transition from FG to HB by activating VDR and augmenting the expression of critical transcription factors (TFs). Besides, our research underscores a shift in metabolic pathways toward glycolytic energy metabolism in HO2 organoids. Overall, our multiomics approach reveals the intricate molecular regulation during the development of HOs.

A Comparative Transcriptomic and Proteomic Analysis of the Pileus of Agaricus bisporus During Its Different Developmental Phases

The analysis of the developmental stages of Agaricus bisporus, a major edible and medicinal mushroom, has always been an important focus in this research area. The process of the growth and development of edible mushrooms is complex and involves the regulation of multiple genes and metabolic pathways. Less data exist on the mechanism of their growth and development at the overall level. In this study, RNA sequencing analyses (RNA-Seq) and data-independent acquisition (DIA) proteomic analyses were carried out at the button phase (BP), harvesting phase (HP), and opening phase (OP) stages of Agaricus bisporus ‘Shuangbao 106’ to reveal the changes in gene expression during the different growth periods of its substrates. The authors screened and explored 3351 differentially expressed genes (DEGs) with a difference factor of ≥2.0, including 2080 up-regulated and 1271 down-regulated genes. After proteome sequencing, 1156 differentially expressed proteins (DEPs) were screened, including 524 up-regulated and 632 down-regulated genes. The expression in TPM of glycoside hydrolase, catalytic core, and chitinase II decreased during both the HP and OP compared with the BP. This may be because mushrooms require higher levels of glycoside hydrolase, catalytic core, and chitinase II activity during the BP to cope with external threats and the need for cell wall remodeling. Conversely, the growth of mushrooms slowed down and the need for cell wall remodeling decreased during the HP and OP, leading to a decrease in the expression of glycoside hydrolase, catalytic core, and chitinase II. This change is related to the need for environmental adaptation, immune defense, and cell wall remodeling, and may regulate the post-growth process of A. bisporus via the hydrolysis of cell wall chitin and glycoside hydrolase. It may also inhibit the growth of mushroom pilei.

Exosomes derived from minor salivary gland mesenchymal stem cells: a promising novel exosome exhibiting pro-angiogenic and wound healing effects similar to those of adipose-derived stem cell exosomes

BackgroundsMinor salivary gland mesenchymal stem cells (MSGMSCs) can be easily extracted and have a broad range of sources. Applying exosomes to wounds is a highly promising method for promoting wound healing. Exosomes derived from different stem cell types have been proven to enhance wound healing, with adipose-derived stem cell (ADSC)-derived exosomes being the most extensively researched. Considering that MSGMSCs have advantages such as easier extraction compared to ADSCs, MSGMSCs should also be a very promising type of stem cell in exosome therapy. However, whether MSGMSC-derived exosomes (MSGMSC-exos) can promote wound healing and how they compare to ADSC-derived exosomes (ADSC-exos) in the wound healing process remain unclear.MaterialsThe effects of MSGMSC-exos and ADSC-exos on angiogenesis in wound healing were investigated in vitro using CCK-8, scratch assays, and tube formation assays. Subsequently, the promotion of wound healing by MSGMSC-exos and ADSC-exos was evaluated in vivo using a full-thickness wound defect model in mice. Immunohistochemistry was used to verify the effects of MSGMSC-exos and ADSC-exos on promoting collagen deposition, angiogenesis, and cell proliferation in the wound. Immunofluorescence staining was performed to investigate the role of MSGMSC-exos and ADSC-exos in modulating the inflammatory response in the wound. Furthermore, proteomic sequencing was conducted to investigate the functional similarities and differences between the proteomes of MSGMSC-exos and ADSC-exos, with key protein contents verified by ELISA.ResultsMSGMSC-exos exhibited similar effects as ADSC-exos in promoting the migration, proliferation, and tube formation of human umbilical vein endothelial cells (HUVECs) in vitro, with a comparable dose-dependent effect. In vivo experiments confirmed that MSGMSC-exos have similar wound healing-promoting functions as ADSC-exos. MSGMSC-exos promoted the neovascularization and maturation of blood vessels in vivo at a level comparable to ADSC-exos. Despite MSGMSC-exos showing less collagen deposition than ADSC-exos, they exhibited stronger anti-scar formation and anti-inflammatory effects. Proteomic analysis revealed that the proteins promoting wound healing in both MSGMSC-exos and ADSC-exos were relatively conserved, with ITGB1 identified as a critical protein for angiogenesis. Further differential analysis revealed that the functions specifically enriched in MSGMSC-exos and ADSC-exos reflected the functions of their source tissue.ConclusionsOur study confirms that MSGMSC-exos exhibit highly similar wound healing and angiogenesis-promoting functions compared to ADSC-exos, and the proteins involved in promoting wound healing in both are relatively conserved. Moreover, MSGMSC-exos show stronger anti-scar formation and anti-inflammatory effects than ADSC-exos. This suggests that MSGMSCs are a promising stem cell source with broad applications in wound healing treatment.

Endothelial cell Ass1 inhibits arteriosclerotic calcification in diabetes mellitus

Endothelial cell (EC) dysfunction is an important pathological feature of early calcification in diabetic plaques. Argininosuccinic synthase 1 (Ass1) is important in protecting EC activity. Therefore, this study aimed to explore the effect of endothelial Ass1 on calcification in diabetic plaques and its potential regulatory mechanism. In this study, serum Ass1 levels were measured in 84 patients, and the study showed that the serum Ass1 level in patients with diabetes was significantly decreased compared with the non-diabetic group, and the serum Ass1 level in patients with coronary artery calcification was significantly decreased compared with the non-coronary artery calcification group. The ApoE-/- mouse diabetic plaque calcification model and the mouse aortic endothelial cell (MAEC) calcification model were constructed, and the influence of endothelial cell Ass1 on diabetic plaque calcification was further investigated by adeno-associated virus and plasmid intervention. Molecular biology studies have shown that endothelial Ass1 overexpression can reduce plaque calcification and inhibit MAEC osteogenic differentiation in diabetic mice, and Ass1 has protective effects on EC and blood vessels in mice. 4D-label-free proteomic sequencing, bioinformatics analysis, and IP experiments were performed on ApoE-/- mouse aorta after adeno-associated virus intervention. It was found that the differential protein Ptk2b was closely related to vascular calcification (VC) and interacted with the target protein Ass1. The above studies indicate that endothelial Ass1 affects calcification formation in diabetic plaques, and the mechanism may be related to Ptk2b. Ass1 may be a new target for the treatment of diabetic VC.

Mechanism of enhancing chemotherapy efficacy in pancreatic ductal adenocarcinoma with paricalcitol and hydroxychloroquine.

Pancreatic ductal adenocarcinoma (PDAC) has a minimal (<15%) 5-year existence, in part due to resistance to chemoradiotherapy. Previous research reveals the impact of paricalcitol (P)and hydroxychloroquine (H)on altering the lysosomal fusion, decreasing stromal burden, and triggering PDAC to chemotherapies. This investigation aims to elucidate the molecular properties of the H and P combination and their potential in sensitizing PDAC to gemcitabine (G). PH potentiates the effects of G in invitro, orthotopic mouse models, and a patient-derived xenograft model of PDAC. Proteomic and single-cell RNA sequencing (RNA-seq) analyses reveal that GPH treatment upregulates autophagy and endoplasmic reticulum (ER) stress-related transcripts. GPH treatment decreases the number of Ki67, fibroblast-associated protein (FAP), and alpha-smooth muscle actin (SMA)-expressing fibroblasts with a decrease in autophagy-related transcripts. The GPH treatment increases M1 polarization and CD4+ and CD8+ Tcells and reduces CD4+ and CD8+ regulatory Tcells (Tregs). These effects of GPH were confirmed in paired biopsies obtained from patients treated in a clinical trial (NCT04524702).

Oral secretions from striped stem borer (Chilo suppressalis) induce defenses in rice.

The striped stem borer (SSB, Chilo suppressalis) is one of the most destructive insect pests on rice. As a chewing insect, SSB larval feeding causes a dramatic increase in rice defense responses. However, the effects of oral secretions (OSs) during SSB feeding on rice defense remain largely unexplored. In this study, based on transcriptome analysis results, treatment with SSB OSs regulated the expression of genes involved in the plant defense-related pathways of calcium, mitogen-activated protein kinases, reactive oxygen species, jasmonic acid (JA), herbivore-induced plant volatiles (HIPVs), and protease inhibitors. Unsurprisingly, treatment with SSB OSs elicited the accumulation of JA and JA-isoleucine in rice. The defense mechanisms activated by the cascade not only induced the expression of trypsin inhibitors, inhibiting the normal growth of SSB larvae but also induced HIPVs emission, rendering rice attractive to a common larval parasitoid. High-throughput proteome sequencing of SSB OSs led to 534 proteins being identified and 343 proteins with two or more unique peptides being detected. The study demonstrates that SSB OSs trigger both direct and indirect defense mechanisms in rice, akin to the effects of SSB feeding. It identifies specific proteins in SSB OSs that may influence the interactions between SSB and rice during feeding, providing valuable insights for effectors research. © 2024 Society of Chemical Industry.

Analysis of differentially expressed proteins and related metabolic pathways in response to lead stress in the leaves of Pogonatherum crinitum

Proteomics provides an essential means of explaining the mechanisms underlying gene expression regulation. The proteomic mechanisms by which heavy metal hyperaccumulators respond to lead (Pb) stress remain largely unclear. To this end, we examined Pogonatherum crinitum (Thunb.) Kunth and employed proteomic sequencing technology to screen for differential proteins that respond to Pb stress. The connection between Pb-tolerant proteins in metabolic pathways and their functions were analyzed. Differences in the downstream molecules of Pb-resistant proteins in P. crinitum were also assessed. Furthermore, we utilized Parallel Reaction Monitoring (PRM) technology to validate the selected Pb-tolerant differential proteins across various stress concentration gradients. A total of 5275 protein families were identified, and 118 DEPs were observed between the stressed and control groups, including 76 upregulated and 42 downregulated proteins. Functional annotation analysis using Gene Ontology and the Kyoto Encyclopedia of Genes and Genomes revealed that Pb stress led to the upregulation of 16 proteins within P. crinitum leaves. These proteins were primarily involved in the metabolic processes of energy and carbohydrate metabolism (PcCht1, PcSPS5, PcGME-1, and PcPEP4) as well as protein translation and oxidative stress (PcHSP26.7, PcHSP18, PcCAT3, and PcCAT1). Bioinformatic analysis indicated that DEPs responding to Pb stress were primarily related to the MAPK signaling pathway, amino sugar and nucleotide sugar metabolism, and starch and sucrose metabolism. Pathway analysis revealed maltose, acetylcholine, N-acetylglucosamine, and oxalic acid as the downstream products. Moreover, the levels of these indicators all increased with increasing Pb concentrations. PRM of the 16 DEPs revealed that nine proteins were upregulated under different Pb concentrations. PRM and data-independent acquisition results for the upregulation of these nine DEPs were identical, suggesting the reliability of our analytical outcomes. In conclusion, the upregulation of specific proteins in P. crinitum enables the regulation of glucose metabolism and antioxidant balance within the plant and represents a mechanism underlying its Pb stress response.