The Mediator Med23 controls a transcriptional switch for muscle stem cell proliferation and differentiation in muscle regeneration.

Muscle stem cells (MuSCs) contribute to a robust muscle regeneration process after injury, which is highly orchestrated by the sequential expression of multiple key transcription factors. However, it remains unclear how key transcription factors and cofactors such as the Mediator complex cooperate to regulate myogenesis. Here, we show that the Mediator Med23 is critically important for MuSC-mediated muscle regeneration. Med23 is increasingly expressed in activated/proliferating MuSCs on isolated myofibers or in response to muscle injury. Med23 deficiency reduced MuSC proliferation and enhanced its precocious differentiation, ultimately compromising muscle regeneration. Integrative analysis revealed that Med23 oppositely impacts Ternary complex factor (TCF)-targeted MuSC proliferation genes and myocardin-related transcription factor (MRTF)-targeted myogenic differentiation genes. Consistently, Med23 deficiency decreases the ETS-like transcription factor 1 (Elk1)/serum response factor (SRF) binding at proliferation gene promoters but promotes MRTF-A/SRF binding at myogenic gene promoters. Overall, our study reveals the important transcriptional control mechanism of Med23 in balancing MuSC proliferation and differentiation in muscle regeneration.

Synthesis of tunable thickness-to-diameter ratio microcapsules via a diffusion-controlled process for temperature-responsive release.

Designing microcapsules with a complicated functionalized shell to respond to an external stimulus has attracted much attention for triggered release; however, simplifying the synthesis process remains a significant challenge. Herein, we initially propose a novel, simple synthesis strategy that utilizes a mixed solvent as the organic phase to control the diffusion of common monomers during interfacial polymerization, resulting in the successful preparation of microcapsules with tunable thickness-to-diameter ratios (T/D). The morphology of microcapsules is confirmed by scanning electron microscopy. We also observe that the T/D of the designed microcapsules progressively increases as the diffusion of monomers occurs, and the glass transition temperature of microcapsules is controlled. Furthermore, microcapsule-based crosslinking agents are applied to investigate the crosslinking reaction of poly(vinyl chloride). Rotational rheometer results indicate that the microcapsules exhibit an excellent external stimulus response, precisely triggering release at the predetermined temperature. This simple approach for the preparation of microcapsules with tunable physical properties has great potential for triggered release in diverse applications.

Smart windows based on ultraviolet-B persistent luminescence phosphors for bacterial inhibition and food preservation.

Herein, ultraviolet B (UVB) persistent luminescence phosphors containing SrAl12O19: Ce3+, Sc3+ nanoparticles were reported. Thermoluminescence (TL) spectrum analysis reveals that the shallow trap induced by Sc3+ co-doping plays an important role in photoluminescence persistent luminescence (PersL) development, while the deep trap dominates the generation of optical stimulated luminescence (OSL). Owing the appearance of deep trap, the OSL is observed under light (700 nm - 900 nm) excitation. UVB luminescence exerts good bactericidal effects on pathogenic bacteria involved in the process of food spoilage. Thus, the smart window with SrAl12O19: Ce3+, Sc3+/PDMS produces UVB PersL to efficiently inactivate Escherichia coli and Staphylococcus aureus. In addition, the presence of the smart window delays the critical point of pork decay, and greatly reduces the time of pork spoilage. It maximizes the convenience of eradicating bacteria and preserving food, thus offering a fresh perspective on the use of UV light for food sterilization and preservation.

Med23 deficiency reprograms the tumor microenvironment to promote lung tumorigenesis.

BACKGROUND: Lung cancer is the leading cause of cancer-related death worldwide. We previously found that Mediator complex subunit 23 (MED23) is important for the tumourigenicity of lung cancer cells with hyperactive Ras activity in vitro, although the in vivo function of MED23 in lung tumourigenesis remains to be explored. METHODS: In this study, we utilized well-characterized KrasG12D-driven non-small cell lung cancer mouse model to investigate the role of MED23 in lung cancer. The lung tumour progression was evaluated by H&E and IHC analysis. Western blotting and qRT-PCR assays were performed to detect changes in gene expression. Immune cells were analyzed by FACS technology. RNA-seq and reporter assays were conducted to explore the mechanism. RESULTS: We observed that lung epithelial Med23 deletion by adeno-Cre resulted in a significant increase in KrasG12D tumour number and size, which was further verified with another mouse model with Med23 specifically deleted in alveolar type II cells. Mice with lung-specific Med23 deficiency also exhibited accelerated tumourigenesis, and a higher proliferation rate for tumour cells, along with increased ERK phosphorylation. Notably, the numbers of infiltrating CD4+ T cells and CD8+ T cells were significantly reduced in the lungs of Med23-deficient mice, while the numbers of myeloid-derived suppressor cells (MDSCs) and Treg cells were significantly increased, suggesting the enhanced immune escape capability of the Med23-deficient lung tumours. Transcriptomic analysis revealed that the downregulated genes in Med23-deficient lung tumour tissues were associated with the immune response. Specifically, Med23 deficiency may compromise the MHC-I complex formation, partially through down-regulating B2m expression. CONCLUSIONS: Collectively, these findings revealed that MED23 may negatively regulate Kras-induced lung tumourigenesis in vivo, which would improve the precise classification of KRAS-mutant lung cancer patients and provide new insights for clinical interventions.

CCNI2 promotes pancreatic cancer through PI3K/AKT signaling pathway.

Globally, pancreatic cancer is recognized as one of the deadliest malignancies that lacks effective targeted therapies. This study aims to explore the role of cyclin I-like protein (CCNI2), a homolog of cyclin I (CCNI), in the progression of pancreatic cancer, thereby providing a theoretical basis for its treatment. Firstly, the expression of CCNI2 in pancreatic cancer tissues was determined through immunohistochemical staining. The biological role of CCNI2 in pancreatic cancer cells was further assessed using both in vitro and in vivo loss/gain-of-function assays. Our data revealed that CCNI2 expression was abnormally elevated in pancreatic cancer, and clinically, increased CCNI2 expression generally correlated with reduced overall survival. Functionally, CCNI2 contributed to the malignant progression of pancreatic cancer by promoting the proliferation and migration of tumor cells. Consistently, in vivo experiments verified that CCNI2 knockdown impaired the tumorigenic ability of pancreatic cancer cells. Moreover, the addition of phosphatidylinositol 3-kinase (PI3K) inhibitors could partially reverse the promoting effect of CCNI2 on the malignant phenotypes of pancreatic cancer cells. CCNI2 promoted pancreatic cancer through PI3K/protein kinase B (AKT) signaling pathway, indicating its potential as a prognostic marker and therapeutic target for pancreatic cancer.

Three-Dimensional MXene-AgNP Hollow Spheres for In Situ Surface-Enhanced Raman Scattering Detection of Catalysis Reactions.

MXenes are attractive candidates in the fields of surface-enhanced Raman scattering (SERS) and catalysis. However, most of the current studies on MXenes are based on blocks and nanosheets, limiting their SERS and catalytic properties. Herein, we have prepared 3D MXene hollow spheres wrapped with silver nanoparticles (Ti3C2-AgNP HSs) using a sacrificial template method, which exhibits excellent sensitivity with a low detection limit due to good light-trapping capability of the hollow sphere and strong localized surface plasmon resonance (LSPR) effect of AgNPs. Furthermore, it shows outstanding photocatalytic performance and realizes in situ SERS monitoring of the 4-nitrobenzenethiol (4-NTP) to 4-aminothiophenol (4-ATP) catalysis reaction. The finite-difference time-domain (FDTD) simulations confirm that 3D Ti3C2-AgNP hollow structures have stronger hot spots than 3D solid structures and higher SERS sensitivity for molecule detection. Therefore, it promises to be an excellent bifunctional material for highly sensitive SERS detection and the in situ monitoring of catalytic reactions.

Bile Acid Overload Induced by Bile Duct and Portal Vein Ligation Improves Survival after Staged Hepatectomy in Rats.

OBJECTIVE: Compared to portal vein ligation (PVL), simultaneous bile duct and portal vein ligation (BPL) can significantly enhance hypertrophy of the intact liver. This study aimed to investigate whether BPL could improve survival after extended hepatectomy independently of an increased remnant liver. METHODS: We adopted rat models of 90% BPL or 90% PVL. To investigate the role of bile acids (BAs) the BA pools in the PVL and BPL groups were altered by the diet. Staged resection preserving 10% of the estimated liver weight was performed 3 days after BPL; PVL; or sham operation. Histology, canalicular network (CN) continuity; and hepatocyte polarity were evaluated. RESULTS: At 3 days after BPL; PVL; or sham operation when the volumetric difference of the intended liver remained insignificant, the survival rates after extended hepatectomy were 86.7%, 47%, and 23.3%, respectively (P<0.01). BPL induced faster restoration of canalicular integrity along with an intensive but transient BA overload. Staged hepatectomy after BPL shortened the duration of the bile CN disturbance and limited BA retention. Decreasing the BA pools in the rats that underwent BPL could compromise these effects, whereas increasing the BA pools of rats that underwent PVL could induce similar effects. The changes in CN restoration were associated with activation of LKB1. CONCLUSION: In addition to increasing the future remnant liver, BPL shortened the duration of the spatial disturbance of the CN and could significantly improve the tolerance of the hypertrophied liver to staged resection. BPL may be a safe and efficient future option for patients with an insufficient remnant liver.

Macrophage-Derived Cathepsin S Remodels the Extracellular Matrix to Promote Liver Fibrogenesis.

BACKGROUND & AIMS: Liver fibrosis is an intrinsic wound-healing response to chronic injury and the major cause of liver-related morbidity and mortality worldwide. However, no effective diagnostic or therapeutic strategies are available, owing to its poorly characterized molecular etiology. We aimed to elucidate the mechanisms underlying liver fibrogenesis. METHODS: We performed a quantitative proteomic analysis of clinical fibrotic liver samples to identify dysregulated proteins. Further analyses were performed on the sera of 164 patients with liver fibrosis. Two fibrosis mouse models and several biochemical experiments were used to elucidate liver fibrogenesis. RESULTS: We identified cathepsin S (CTSS) up-regulation as a central node for extracellular matrix remodeling in the human fibrotic liver by proteomic screening. Increased serum CTSS levels efficiently predicted liver fibrosis, even at an early stage. Secreted CTSS cleaved collagen 18A1 at its C-terminus, releasing endostatin peptide, which directly bound to and activated hepatic stellate cells via integrin α5ß1 signaling, whereas genetic ablation of Ctss remarkably suppressed liver fibrogenesis via endostatin reduction in vivo. Further studies identified macrophages as the main source of hepatic CTSS, and splenectomy effectively attenuated macrophage infiltration and CTSS expression in the fibrotic liver. Pharmacologic inhibition of CTSS ameliorated liver fibrosis progression in the mouse models. CONCLUSIONS: CTSS functions as a novel profibrotic factor by remodeling extracellular matrix proteins and may represent a promising target for the diagnosis and treatment of liver fibrosis.

Overcoming Debye length limitations: Three-dimensional wrinkled graphene field-effect transistor for ultra-sensitive adenosine triphosphate detection.

Adenosine triphosphate (ATP) is closely related to the pathogenesis of certain diseases, so the detection of trace ATP is of great significance to disease diagnosis and drug development. Graphene field-effect transistors (GFETs) have been proven to be a promising platform for the rapid and accurate detection of small molecules, while the Debye shielding limits the sensitive detection in real samples. Here, a three-dimensional wrinkled graphene field-effect transistor (3D WG-FET) biosensor for ultra-sensitive detection of ATP is demonstrated. The lowest detection limit of 3D WG-FET for analyzing ATP is down to 3.01 aM, which is much lower than the reported results. In addition, the 3D WG-FET biosensor shows a good linear electrical response to ATP concentrations in a broad range of detection from 10 aM to 10 pM. Meanwhile, we achieved ultra-sensitive (LOD: 10 aM) and quantitative (range from 10 aM to 100 fM) measurements of ATP in human serum. The 3D WG-FET also exhibits high specificity. This work may provide a novel approach to improve the sensitivity for the detection of ATP in complex biological matrix, showing a broad application value for early clinical diagnosis and food health monitoring. Electronic supplementary materials: The online version contains supplementary material available at 10.1007/s11467-023-1281-7 and https://journal.hep.com.cn/fop/EN/10.1007/s11467-023-1281-7.

Establishment and validation of exhausted CD8+ T cell feature as a prognostic model of HCC.

Objectives: The exhausted CD8+T (Tex) cells are a unique cell population of activated T cells that emerges in response to persistent viral infection or tumor antigens. Tex cells showed the characteristics of aging cells, including weakened self-renewal ability, effector function inhibition, sustained high expression of inhibitory receptors including PD-1, TIGIT, TIM-3, and LAG-3, and always accompanied by metabolic and epigenetic reprogramming. Tex cells are getting more and more attention in researching immune-related diseases and tumor immunotherapy. However, studies on Tex-related models for tumor prognosis are still lacking. We hope to establish a risk model based on Tex-related genes for HCC prognosis. Methods: Tex-related GEO datasets from different pathologic factors (chronic HBV, chronic HCV, and telomere shortening) were analyzed respectively to acquire differentially expressed genes (DEGs) by the 'limma' package of R. Genes with at least one intersection were incorporated into Tex-related gene set. GO, KEGG, and GSEA enrichment analyses were produced. Hub genes and the PPI network were established and visualized by the STRING website and Cytoscape software. Transcription factors and targeting small molecules were predicted by the TRUST and CLUE websites. The Tex-related HCC prognostic model was built by Cox regression and verified based on different datasets. Tumor immune dysfunction and exclusion (TIDE) and SubMap algorithms tested immunotherapy sensitivity. Finally, qRT-PCR and Flow Cytometry was used to confirm the bioinformatic results. Results: Hub genes such as AKT1, CDC6, TNF and their upstream transcription factor ILF3, Regulatory factor X-associated protein, STAT3, JUN, and RELA/NFKB1 were identified as potential motivators for Tex. Tex-related genes SLC16A11, CACYBP, HSF2, and ATG10 built the HCC prognostic model and helped with Immunotherapy sensitivity prediction. Conclusion: Our study demonstrated that Tex-related genes might provide accurate prediction for HCC patients in clinical decision-making, prognostic assessment, and immunotherapy. In addition, targeting the hub genes or transcription factors may help to reverse T cell function and enhance the effect of tumor immunotherapy.

Bone mesenchymal stromal cell-derived small extracellular vesicles inhibit inflammation and ameliorate sepsis via delivery of microRNA-21a-5p.

BACKGROUND AIMS: Sepsis is a potentially life-threatening disease that results from a severe systemic inflammatory response due to infection. Mesenchymal stromal cell-derived small extracellular vesicles (MSC sEVs) are able to transfer bioactive molecules and have been demonstrated to play an important role in the pathophysiological process of sepsis. Herein the authors aimed to investigate the potential role and downstream molecular mechanism of MSC sEVs in sepsis. METHODS: MSC sEVs were acquired by ultracentrifugation and then injected into a cecal ligation and puncture mouse model. The efficacy of MSC sEVs in both in vitro and in vivo models of sepsis was evaluated. RESULTS: MSC sEV therapy improved survival, reduced sepsis-induced inflammation, attenuated pulmonary capillary permeability and improved liver and kidney function in septic mice. In addition, the authors found that microRNA-21a-5p (miR-21a-5p) was highly enriched in MSC sEVs, could be transferred to recipient cells, inhibited inflammation and increased survival in septic mice. Furthermore, the authors demonstrated that MSC sEV miR-21a-5p suppressed inflammation by targeting toll-like receptor 4 and programmed cell death 4. The therapeutic efficacy of MSC sEVs was partially abrogated by transfection with miR-21a-5p inhibitors. CONCLUSIONS: Collectively, the authors' data suggest that miR-21a-5p-bearing MSC sEVs may be a prospective and effective sepsis therapeutic strategy.

Exploration and verification of COVID-19-related hub genes in liver physiological and pathological regeneration.

Objectives An acute injury is often accompanied by tissue regeneration. In this process, epithelial cells show a tendency of cell proliferation under the induction of injury stress, inflammatory factors, and other factors, accompanied by a temporary decline of cellular function. Regulating this regenerative process and avoiding chronic injury is a concern of regenerative medicine. The severe coronavirus disease 2019 (COVID-19) has posed a significant threat to people's health caused by the coronavirus. Acute liver failure (ALF) is a clinical syndrome resulting from rapid liver dysfunction with a fatal outcome. We hope to analyze the two diseases together to find a way for acute failure treatment. Methods COVID-19 dataset (GSE180226) and ALF dataset (GSE38941) were downloaded from the Gene Expression Omnibus (GEO) database, and the "Deseq2" package and "limma" package were used to identify differentially expressed genes (DEGs). Common DEGs were used for hub genes exploration, Protein-Protein Interaction (PPI) network construction, Gene Ontology (GO) functional enrichment, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment. The real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) was used to verify the role of hub genes in liver regeneration during in vitro expansion of liver cells and a CCl4-induced ALF mice model. Results: The common gene analysis of the COVID-19 and ALF databases revealed 15 hub genes from 418 common DEGs. These hub genes, including CDC20, were related to cell proliferation and mitosis regulation, reflecting the consistent tissue regeneration change after the injury. Furthermore, hub genes were verified in vitro expansion of liver cells and in vivo ALF model. On this basis, the potential therapeutic small molecule of ALF was found by targeting the hub gene CDC20. Conclusion We have identified hub genes for epithelial cell regeneration under acute injury conditions and explored a new small molecule Apcin for liver function maintenance and ALF treatment. These findings may provide new approaches and ideas for treating COVID-19 patients with ALF.

Damaged collagen detected by collagen hybridizing peptide as efficient diagnosis marker for early hepatic fibrosis.

Liver fibrosis is characterized by excessive synthesis and deposition of extracellular matrix (ECM) in liver tissues. However, it still has been lacking of early detection and diagnosis methods. The collagen hybridizing peptide (CHP) is a novel synthetic peptide that enables detection of collagen damage and tissue remodeling. Here, we showed that obvious CHP-positive staining could be detected in the liver while given CCl4 for only 3 days, which was significantly enhanced while given CCl4 for 7 days. However, H&E staining showed no significant changes in fibrous tissue, and sirius red-positive staining could only be observed while given CCl4 for 14 days. Moreover, CHP-positive staining enhanced initially at portal area which further extended into the hepatic lobule, which was increased more significantly than sirius red-positive staining in the model of 10 and 14 days. Further proteomic analysis of CHP-positive staining revealed that pathways associated with ECM remodeling were significantly increased, while retinol metabolism was downregulated. Meanwhile, proteins enriched in cellular gene transcription and signal transduction involved in fibrogenesis were also upregulated, suggesting that fibrosis occurred in CHP-positive staining. Our study provided evidence that CHP could detect the collagen damage in liver, which might be an efficient indicator for the diagnosis of liver fibrosis at a very early stage.

Estrogen-related genes influence immune cell infiltration and immunotherapy response in Hepatocellular Carcinoma.

Background: Immunotherapy has been the first-line treatment option in advanced Hepatocellular Carcinoma(HCC); but now, there are no established molecular markers that can predict immunotherapy response. Estrogen has a crucial role in the development of a variety of liver illnesses, including liver fibrosis, Nonalcoholic fatty liver disease (NAFLD), and HCC. Nonetheless, the significance of estrogen-related genes in HCC immunotherapy and the underlying molecular mechanisms are not yet fully understood. Method: In this study, we constructed a novel estrogen-related gene prognostic signature (ERGPS) by analyzing bulk RNA sequencing data from 365 HCC patients. Based on the median risk score, we divided 365 HCC patients into low- and high-risk groups. Tumor mutation burden (TMB), Microsatellite instability (MSI), T cell receptor (TCR) richness, B cell receptor (BCR) richness, single-nucleotide variants (SNV) Neoantigens, Cancer Testicular Antigens (CTA) scores, and Tumour Immune Dysfunction and Exclusion (TIDE) scores were used to evaluate the magnitude of immunotherapy response. Multiple external datasets validate the validity and robustness of the prognostic signature. Real-time quantitative polymerase chain reaction (qRT-PCR) was used to validate estrogen-related gene overexpression in HCC tissue samples. Results: ERGPS is an independent risk factor affecting the prognosis of HCC patients and is superior to other clinical variables in predicting patient survival and immunotherapy response. Multiple independent external datasets confirmed the superior predictive efficacy of the prognostic signature. The prognostic signature was positively correlated with TMB score, MSI score, TCR richness, BCR richness, SNV Neoantigens score, CTA score, expression levels of immune checkpoint-related genes, and TIDE score. Patients with HCC in the high-risk group identified by the prognostic signature were likely to be more responsive to immunotherapy and more suitable for immunotherapy. qRT-PCR confirmed that estrogen-related genes of the construct signature were highly expressed in HCC tumor tissues. Conclusion: Estrogen-related genes are overexpressed in HCC tissues. Our novel prognostic signature can accurately predict not only the prognosis but also the immunotherapy response of HCC patients. In the future, prognostic signatures will be a useful tool for clinicians to screen patients with HCC who are suitable for immunotherapy.

An intellectual disability-related MED23 mutation dysregulates gene expression by altering chromatin conformation and enhancer activities.

Transcriptional Mediator controls diverse gene programs for various developmental and pathological processes. The human Mediator MED23/R617Q mutation was reported in a familial intellectual disability (ID) disorder, although the underlying mechanisms remain poorly understood. Constructed by gene editing, the Med23/R617Q knock-in mutant mice exhibited embryonic lethality due to the largely reduced Med23/R617Q protein level, but the R617Q mutation in HEK293T cells didn't change its expression and incorporation into Mediator Complex. RNA-seq revealed that MED23/R617Q mutation disturbed gene expression, related to neural development, learning and memory. Specifically, R617Q mutation reduced the MED23-dependent activities of ELK1 and E1A, but in contrast, upregulated the MAPK/ELK1-driven early immediate genes (IEGs) JUN and FOS. ChIP-seq and Hi-C revealed that the MED23 R617Q mutation reprogramed a subset of enhancers and local chromatin interactions, which correlated well with the corresponding gene expression. Importantly, the enhancers and chromatin interactions surrounding IEGs were unchanged by the R617Q mutation, but DACH1, an upstream repressor of IEGs, showed reduced enhancer-promoter interactions and decreased expression in mutant cells, thus relieving its inhibition to the intellectual-related IEGs. Overall, unraveling the MED23-DACH1-IEG axis provides a mechanistic explanation for the effects of the MED23/R617Q mutation on gene dysregulation and inherited ID.

Whole-exome sequencing identifies a set of genes as markers of hepatocellular carcinoma early recurrence.

BACKGROUND: Hepatocellular carcinoma (HCC) is characterized by a high recurrence rate and poor prognosis. In recent years, the therapeutic regimen of programmed cell death protein 1 (PD-1) antibody combined with multi-targeted tyrosine kinase inhibitors (mTKIs) has achieved better results in the clinical application of hepatocellular carcinoma. Whole-exome sequencing can reflect the mutational characteristics of patients' exons and guide the clinical selection of molecular targeting drugs more accurately, which is in line with the concept of precision medicine. METHODS: We performed exome sequencing on 63 patients with HCC treated with radical surgery at our hospital and collected their clinical indexes and postoperative follow-up data. Using machine learning, a prediction model for recurrence within 1 year was constructed and the model was presented in a nomogram. Patients treated with PD-1 antibodies in combination with mTKIs after relapse were grouped by prognosis, and the valuable mutated genes were screened according to whole-exome sequencing data. The tumor tissue immune cells were analyzed using the UCSC Xena database. The expressions of target proteins were verified by Polymerase Chain Reaction (PCR) and Immunohistochemistry (IHC), respectively, on commercial HCC cell lines and pathological specimens of hepatocellular carcinoma collected clinically. RESULTS: The proportion of patients who relapsed within a year was 41% and the prognosis of those patients was poor. The characteristic exon mutation profile with a high frequency of variants in multiple mucin genes was present in Chinese HCC patients. Multiple nidi and 30 exon variants were brought into the prediction model with an area under the curve (AUC) = 0.94. MUC6 gene mutation was obvious in patients with an early recurrence, and MUC3A and MUC4 gene mutations were evident in patients with poorer responses to PD-1 antibodies combined with mTKIs. Those three mucins were negatively correlated with immune infiltrating cells. CONCLUSIONS: We depicted the exon characteristics of hepatocellular carcinoma in the Chinese population and established a predictive model for recurrence within 1 year after radical surgical treatment. Moreover, we found that mucins were worthy targets of hepatocellular carcinoma.

Establishment of a mouse model for bile duct repair and tissue engineering.

Due to the lack of a suitable model, research on biliary biology is far behind that on other organs. A mouse model of common bile duct (CBD) dilation (BDD) was first established and compared with CBD ligation mice (BDL). Then, in a transplantation experiment, the dilated CBD of recipient BDD mice was injured by making an elliptical incision and repaired by transplanting a bile duct patch from donor BDD mice. Biochemical and histological changes were analyzed and cell proliferation of the bile duct grafts was determined. Slightly dilated and unblocked CBD with a diameter of 2.89±0.76 mm was obtained in BDD mice, while the CBD diameter was 0.51±0.08 mm in the Sham group and 4.71±0.64 mm in the BDL group on day 14 after surgery. The liver damage was very mild in BDD mice compared with BDL mice, proving that the BDD model could be further used for bile duct transplantation. By cross transplanting the bile duct patch from enhanced green fluorescence protein and wild-type BDD mice, it was found that the CBD injury was well repaired and the cells of the bile duct patch were completely replaced by recipient-derived cells at 12 week after the repair operation. α Smooth muscle actin, Ki67 and cytokeratin 19 immunofluorescence staining showed that the proliferation of bile duct epithelial cells and abundant active fibroblasts were found within the bile duct patch during the regeneration process. Therefore, a reliable new mouse model of bile duct injury and repair was successfully established and can be used in the study of biliary repair mechanisms and tissue engineering of biliary ducts.

DhMYB2 and DhbHLH1 regulates anthocyanin accumulation via activation of late biosynthesis genes in Phalaenopsis-type Dendrobium.

Phalaenopsis-type Dendrobium is a popular orchid with good ornamental and market value. Despite their popularity, molecular regulation of anthocyanin biosynthesis during flower development remains poorly understood. In this study, we systematically investigated the regulatory roles of the transcription factors DhMYB2 and DhbHLH1 in anthocyanins biosynthesis. Gene expression analyses indicated that both DhMYB2 and DhbHLH1 are specifically expressed in flowers and have similar expression patterns, showing high expression in purple floral tissues with anthocyanin accumulation. Transcriptomic analyses showed 29 differentially expressed genes corresponding to eight enzymes in anthocyanin biosynthesis pathway have similar expression patterns to DhMYB2 and DhbHLH1, with higher expression in the purple lips than the yellow petals and sepals of Dendrobium 'Suriya Gold'. Further gene expression analyses and Pearson correlation matrix analyses of Dendrobium hybrid progenies revealed expression profiles of DhMYB2 and DhbHLH1 were positively correlated with the structural genes DhF3'H1, DhF3'5'H2, DhDFR, DhANS, and DhGT4. Yeast one-hybrid and dual-luciferase reporter assays revealed DhMYB2 and DhbHLH1 can bind to promoter regions of DhF3'H1, DhF3'5'H2, DhDFR, DhANS and DhGT4, suggesting a role as transcriptional activators. These results provide new evidence of the molecular mechanisms of DhMYB2 and DhbHLH1 in anthocyanin biosynthesis in Phalaenopsis-type Dendrobium.

Identification of CKS1B as a prognostic indicator and a predictive marker for immunotherapy in pancreatic cancer.

As a regulatory subunit of cyclin kinase, CKS1B promotes cancer development and is associated with poor prognosis in multiple cancer patients. However, the intrinsic role of CKS1B in pancreatic cancer remains elusive. In our research, CKS1B expression in pancreatic tumor tissue was higher than that in normal tissue by TCGA, Oncomine and CPTAC databases analysis. Similar result was verified in our center tissues by qRT-PCR. CKS1B expression was closely relevant to histologic grading, prognosis, and TMB. GSEA showed that CKS1B mainly participated in the regulation of autophagy and T cell receptor signaling pathway. Furthermore, CIBERSORT analysis showed that there was a strong correlation between CKS1B expression and tumor immune cells infiltration. Drug sensitivity analysis showed that patients with high CKS1B expression appeared to be more sensitive to gemcitabine, 5-fluorouracil, and paclitaxel. We then investigated cell viability and migratory ability by CCK8 and transwell assay, respectively. Results indicated that CKS1B knockdown by short hairpin RNA significantly reduced pancreatic cancer cell viability and invasion via regulating PD-L1 expression. In conclusion, our research further demonstrates the role of CKS1B in pancreatic cancer and the signaling pathways involved. The association of CKS1B with immune infiltration and immune checkpoint may provide a new direction for immunotherapy of pancreatic cancer.