Integrative single-cell transcriptomic analyses reveal the cellular ontological and functional heterogeneities of primary and metastatic liver tumors.

BACKGROUND: The global cellular landscape of the tumor microenvironment (TME) combining primary and metastatic liver tumors has not been comprehensively characterized. METHODS: Based on the scRNA-seq and spatial transcriptomic data of non-tumor liver tissues (NTs), primary liver tumors (PTs) and metastatic liver tumors (MTs), we performed the tissue preference, trajectory reconstruction, transcription factor activity inference, cell-cell interaction and cellular deconvolution analyses to construct a comprehensive cellular landscape of liver tumors. RESULTS: Our analyses depicted the heterogeneous cellular ecosystems in NTs, PTs and MTs. The activated memory B cells and effector T cells were shown to gradually shift to inhibitory B cells, regulatory or exhausted T cells in liver tumors, especially in MTs. Among them, we characterized a unique group of TCF7+ CD8+ memory T cells specifically enriched in MTs that could differentiate into exhausted T cells likely driven by the p38 MAPK signaling. With regard to myeloid cells, the liver-resident macrophages and inflammatory monocyte/macrophages were markedly replaced by tumor-associated macrophages (TAMs), with TREM2+ and UBE2C+ TAMs enriched in PTs, while SPP1+ and WDR45B+ TAMs in MTs. We further showed that the newly identified WDR45B+ TAMs exhibit an M2-like polarization and are associated with adverse prognosis in patients with liver metastases. Additionally, we addressed that endothelial cells display higher immune tolerance and angiogenesis capacity, and provided evidence for the source of the mesenchymal transformation of fibroblasts in tumors. Finally, the malignant hepatocytes and fibroblasts were prioritized as the pivotal cell populations in shaping the microenvironments of PTs and MTs, respectively. Notably, validation analyses by using spatial or bulk transcriptomic data in clinical cohorts concordantly emphasized the clinical significance of these findings. CONCLUSIONS: This study defines the ontological and functional heterogeneities in cellular ecosystems of primary and metastatic liver tumors, providing a foundation for future investigation of the underlying cellular mechanisms.

Modulation of gut microbiota by crude mulberry polysaccharide attenuates knee osteoarthritis progression in rats.

Mulberry (Morus alba L.), a kind of common fruits widely cultivated worldwide, has been proven various biological activities. However, its potential role in the progression of knee osteoarthritis (KOA) remains unclear. This study aims to investigate the potential protective effects of crude polysaccharide extracted from mulberry fruit, referred to as a complex blend of polysaccharides and other unidentified extracted impurities, on KOA progression. The KOA rats were established by injection of 1 mg sodium monoiodoacetate into knee, and administrated with crude mulberry polysaccharide (Mup) by gastric gavage for 4 weeks. Furthermore, intestinal bacteria clearance assay (IBCA) and fecal microbiota transplantation were conducted for the evaluation of the effect of gut microbiota (GM) on KOA. Our findings demonstrated that Mup, particularly at a dosage of 200 mg/kg, effectively improved abnormal gait patterns, reduced the level of inflammation, mitigated subchondral bone loss, restored compromised joint surfaces, alleviated cartilage destruction, and positively modulated the dysregulated profile of GM in KOA rats. Moreover, IBCA compromised the protective effects of Mup, while transplantation of fecal bacteria from Mup-treated rats facilitated KOA recovery. Collectively, our study suggested that Mup had the potential to ameliorate the progression of KOA, potentially through its modulation of GM profile.

Three-dimensional Ordered Macroporous Flexible Electrode Design toward High-Performance Zinc-Ion Batteries.

Flexible zinc-ion batteries (ZIBs) have been considered to have huge potential in portable and wearable electronics due to their high safety, cost efficiency, and considerable energy density. Therein, the design and construction of flexible electrodes significantly determine the performance and lifespan of flexible battery devices. In this work, an ultrathin flexible three-dimensional ordered macroporous (3DOM) Sn@Zn anode (60 µm in thickness) is presented to relieve dendrite growth and expand the lifespan of flexible ZIBs. The 3DOM structure can ensure uniform electric field distribution, guide oriented zinc plating/stripping, and extend the lifespan of anodes. The rich zincophilic Sn sites on the electrode surface significantly facilitate Zn nucleation. Accordingly, a lowered nucleation overpotential of 8.9 mV and an ultralong cycling performance of 2400 h at 0.1 mA cm-2 and 0.1 mAh cm-2 are achieved in symmetric cells, and the 3DOM Sn@Zn anode can also operate in deep cycling for over 200 h at 10 mA cm-2 and 5 mAh cm-2. A flexible 3DOM MnO2/Ni cathode with a high structural stability and a high mass-specific capacity is fabricated to match with the anode to form a flexible ZIB with a total thickness of 200 µm. The flexible device delivers a high volumetric energy density of 11.76 mWh cm-3 at 100 mA gMnO2-1 and a high average open-circuit voltage of 1.5 V and exhibits high-performance power supply under deformation in practical application scenarios. This work may shed some light on the design and fabrication of flexible energy-storage devices.

A Comparative Visual Analytics Framework for Evaluating Evolutionary Processes in Multi-Objective Optimization.

Evolutionary multi-objective optimization (EMO) algorithms have been demonstrated to be effective in solving multi-criteria decision-making problems. In real-world applications, analysts often employ several algorithms concurrently and compare their solution sets to gain insight into the characteristics of different algorithms and explore a broader range of feasible solutions. However, EMO algorithms are typically treated as black boxes, leading to difficulties in performing detailed analysis and comparisons between the internal evolutionary processes. Inspired by the successful application of visual analytics tools in explainable AI, we argue that interactive visualization can significantly enhance the comparative analysis between multiple EMO algorithms. In this paper, we present a visual analytics framework that enables the exploration and comparison of evolutionary processes in EMO algorithms. Guided by a literature review and expert interviews, the proposed framework addresses various analytical tasks and establishes a multi-faceted visualization design to support the comparative analysis of intermediate generations in the evolution as well as solution sets. We demonstrate the effectiveness of our framework through case studies on benchmarking and real-world multi-objective optimization problems to elucidate how analysts can leverage our framework to inspect and compare diverse algorithms.

Thoracic Endovascular Aortic Repair Versus Open Surgery for Stanford Type B Aortic Dissection: A Meta-Analysis and Systematic Review.

BACKGROUND: Thoracic endovascular aortic repair is a relatively new technique relative to open surgery, and our aim was to assess whether there is a difference in the risk of common postoperative complications between thoracic endovascular aortic repair and open surgery. METHODS: The PubMed, Web of Science, and Cochrane library were systematically searched for trials comparing thoracic endovascular aortic repair and open surgical repair from January 2000 to September 2022. Primary outcome was death, other outcomes included common associated complications. Data were combined using risk ratio or standardized mean difference with 95% confidence interval. Funnel plot and egger's test were used for assessing publication bias. The study protocol was registered prospectively with PROSPERO (CRD42022372324). RESULTS: This trial included 11 controlled clinical studies with 3667 patients. Thoracic endovascular aortic repair had lower risk of death (risk ratio [RR], 0.59; 95% CI, 0.49 to 0.73; p < 0.00001; I2 = 0), dialysis (RR, 0.55; 95% CI, 0.47 to 0.65; p < 0.00001; I2 = 37%), stroke (RR, 0.71; 95% CI, 0.51 to 0.98; p = 0.03; I2 = 40%), bleeding (RR, 0.44; 95% CI, 0.23 to 0.83; p = 0.01; I2 = 56%), and respiratory complications (RR, 0.67; 95% CI, 0.60 to 0.76; p < 0.00001; I2 = 37%) compared with open surgical repair. In addition, the length of hospital stay was shorter in the thoracic endovascular aortic repair group (SMD, -0.84; 95% CI, -1.30 to -0.38; p = 0.0003; I2 = 80%). CONCLUSIONS: Thoracic endovascular aortic repair has significant advantages over open surgical repair, in terms of postoperative complications and survival in Stanford type B aortic dissection patients.

Heterologous expression of the Leymus chinensis metallothionein gene LcMT3 confers enhanced tolerance to salt stress in Escherichia coli, yeast, and Arabidopsis thaliana.

Salinity is poisonous to various plant physiological processes and poses an increasingly severe threat to agricultural productivity worldwide. As a tactic to mitigate this issue, the hunt for salt-tolerance genes and pathways is intensifying. The low-molecular-weight proteins known as metallothioneins (MTs) can effectively reduce salt toxicity in plants. In seeking concrete evidence of its function under salt stress conditions, a unique salt-responsive metallothionein gene, LcMT3, was isolated from the extremely salt-enduring Leymus chinensis and heterologously characterized in Escherichia coli (E. coli), yeast (Saccharomyces cerevisiae), as well as Arabidopsis thaliana. Overexpression of LcMT3 imparted resistance to salt in E. coli cells and yeast, while the development of control cells was completely inhibited. Besides, transgenic plants expressing LcMT3 exhibited significantly enhanced salinity tolerance. They had higher germination rates and longer roots than their nontransgenic counterparts during NaCl tolerance. For several physiological indices of salt tolerance, transgenic lines reduced the accumulation of malondialdehyde (MDA), relative conductivity, and reactive oxygen species (ROS) in comparison to nontransgenic Arabidopsis. They also possessed increased concentrations of proline (Pro), relative water content, chlorophyll content, coupled with three more active antioxidant enzymes (superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT)). Transgenic plants also accumulated less Na+ and maintained a lower Na+/K+ ratio than control, which can be attributable to the transgene's regulatory effect on transporter proteins such as salt overly sensitive (SOS) and Na+/H+ antiporter (NHX1), as demonstrated by qPCR experiments. Collectively, LcMT3 could have a vital function in salinity resistance and be an essential candidate protein for abiotic stress.

Petite Integration Factor 1 knockdown enhances gemcitabine sensitivity in pancreatic cancer cells via increasing DNA damage.

Chemoresistance is still a vital obstacle in various tumors chemotherapy. This study aimed to explore the role of Petite Integration Factor 1 (PIF1) in the sensitivity of gemcitabine response to pancreatic cancer cells. Gene Expression Profiling Interactive Analysis (GEPIA) database was employed for evaluating the level of PIF1 in pancreatic cancer tissues and normal tissues. The mRNA level of PIF1 was detected via reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis. The relative protein expression of PIF1, cleaved caspase-3, and phosphorylated histone H2Ax (γH2Ax) was assessed through western blot. Cell viability and apoptosis were assessed via Cell Counting Kit-8 (CCK-8) assay and flow cytometry, respectively. Moreover, lactate dehydrogenase (LDH) release and caspase-3 activity were determined via the corresponding LDH Cytotoxicity Assay Kit and caspase-3 colorimetric assay kit. PIF1 expression was upregulated in pancreatic cancer tissues and cells. Knockdown of PIF1 exhibited the repressive impact on the viability of AsPC-1 and PANC-1 cells. PIF1 knockdown enhanced LDH release and apoptosis in both AsPC-1 and PANC-1 cells. PIF1 downregulation could augment the sensitivity of gemcitabine in pancreatic cancer cells, as evidenced by lower cell viability and higher LDH release and apoptosis rate after knocking down PIF1 in gemcitabine-treated pancreatic cancer cells relative to pancreatic cancer cells treated with gemcitabine alone. Moreover, PIF1 knockdown increased γH2Ax protein expression and DNA damage, and gemcitabine treatment-induced DNA damage in AsPC-1 and PANC-1 cells was exacerbated by PIF1 silencing. Furthermore, gemcitabine treatment-caused increase of DNA damage was alleviated by PIF1 overexpression; whereas, this effect of PIF1 upregulation was reversed by thymidine, a DNA synthesis inhibitor. In addition, the decreased gemcitabine sensitivity response to pancreatic cancer cells caused by PIF1 upregulation was also hindered by thymidine treatment. In conclusion, PIF1 silencing enhanced gemcitabine sensitivity response to pancreatic cancer cells through aggrandizing DNA damage.

Life Table Construction under Different Temperatures and Insecticide Susceptibility Analysis of Uroleucon formosanum (Hemiptera: Aphididae).

Uroleucon formosanum is an important aphid pest of lettuce, but basic information on its biology is scarce. In this study, effects of three constant temperatures (17, 21, and 25 °C, simulating the mean temperature range in greenhouses) on the development and fecundity of U. formosanum were analyzed by constructing a life table. U. formosanum could develop and reproduce under all three temperatures, but the survival rate, development, and fecundity of U. formosanum were affected by temperature. The intrinsic rate of increase was lowest at 17 °C (0.17) and it was significantly less than at 21 °C (0.20) and 25 °C (0.23). Furthermore, U. formosanum had the lowest finite rate of increase (1.19) and the largest mean generation time (20.21) at 17 °C. These results mean that U. formosanum is less adapted to the lower temperatures (17 °C) among these three set temperatures. To screen insecticides for control, susceptibility of U. formosanum to six insecticides including chlorpyrifos, abamectin, beta-cypermethrin, imidacloprid, nitenpyram, and thiamethoxam was evaluated. U. formosanum was relatively sensitive to all six test insecticides. Chlorpyrifos had the highest toxicity to U. formosanum (LC50 = 3.08 mg/L). These data may help to develop integrated management strategies for better population control of U. formosanum.

The membrane associated NAC transcription factors ANAC060 and ANAC040 are functionally redundant in the inhibition of seed dormancy in Arabidopsis thaliana.

The NAC family of transcription factors is involved in plant development and various biotic and abiotic stresses. The Arabidopsis thaliana ANAC genes ANAC060, ANAC040, and ANAC089 are highly homologous based on protein and nucleotide sequence similarity. These three genes are predicted to be membrane bound transcription factors (MTFs) containing a conserved NAC domain, but divergent C-terminal regions. The anac060 mutant shows increased dormancy when compared with the wild type. Mutations in ANAC040 lead to higher seed germination under salt stress, and a premature stop codon in ANAC089 Cvi allele results in seeds exhibiting insensitivity to high concentrations of fructose. Thus, these three homologous MTFs confer distinct functions, although all related to germination. To investigate whether the differences in function are caused by a differential spatial or temporal regulation, or by differences in the coding sequence (CDS), we performed swapping experiments in which the promoter and CDS of the three MTFs were exchanged. Seed dormancy and salt and fructose sensitivity analyses of transgenic swapping lines in mutant backgrounds showed that there is functional redundancy between ANAC060 and ANAC040, but not between ANAC060 and ANAC089.

Involvement of elevated ASF1B in the poor prognosis and tumorigenesis in pancreatic cancer.

Anti-silencing function 1B (ASF1B) has been reported to be associated with the occurrence of many kinds of tumors. However, the biological effect and action mechanism of ASF1B in pancreatic cancer (PC) tumorigenesis remain unclear. The expression and prognosis value of ASF1B in PC were analyzed using GEPIA, GEO, and Kaplan-Meier plotter databases. The diagnostic value of ASF1B in PC was determined by receiver operating characteristic curve. The relationship between ASF1B expression and the clinical feathers in PC was investigated based on TCGA. qRT-PCR and western blot analyses were used to measure ASF1B expression in PC cells. Cell proliferation was evaluated by MTT and EdU assays, and apoptosis was examined by TUNEL and caspase-3 activity assays. Western blot analysis was utilized to detect the expression of proliferating cell nuclear antigen (PCNA), cyclin D1, Bax, Bcl-2, and phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) signaling proteins. ASF1B was overexpressed in several digestive cancers, including PC. Upregulated ASF1B was correlated with the poor prognosis and clinical features in PC patients. The area under the curve (AUC) value of ASF1B was 0.990. ASF1B was also overexpressed in PC cells. ASF1B silencing inhibited PC cell proliferation, promoted apoptosis, and increased caspase-3 activity, which were accompanied by the reduction of PCNA and cyclin D1 expression and increase of the ratio of Bax/Bcl-2 expression. Additionally, ASF1B silencing suppressed the PI3K/Akt pathway and 740Y-P treatment partially abolished the effects of ASF1B knockdown on PC cells. In conclusion, ASF1B silencing retarded proliferation and promoted apoptosis in PC cells by inactivation of the PI3K/Akt pathway.

Long non-coding RNA CATIP antisense RNA 1 (lncRNA CATIP-AS1) downregulation contributes to the progression and metastasis of thyroid cancer via epithelial-mesenchymal transition (EMT) pathway.

Thyroid cancer (THCA) is the most common cancer of the endocrine system across the globe. To date, the mechanism of development of THCA remains scarcely known. In this study, we aim to elucidate the long non-coding RNA CATIP antisense RNA 1 (lncRNA CATIP-AS1/CATIP-AS1) role in the pathogenesis of THCA and its regulatory mechanism. The result shows that the CATIP-AS1 was significantly downregulated in THCA tissues and cells and was associated with a poor prognosis of patients diagnosed with THCA. The overexpression of CATIP-AS1 significantly inhibited THCA cell proliferation, migration, and epithelial-mesenchymal transition (EMT) but increased the THCA cell apoptosis. We found that CATIP-AS1 endogenously sponges miR-515-5p and its overexpression could inhibit miR-515-5p regulatory effect. Moreover, the overexpression of miR-515-5p repressed the Smad4 expression level, consequently reversed the inhibiting effect of overexpressed CATIP-AS1 on the proliferation, and migration of THCA cell. It also reversed the increased THCA cell apoptosis and the downregulated-CATIP-AS1-induced cell EMT inhibition. Summarily, we demonstrated that the CATIP-AS1 promotes the progression and metastasis of THCA via EMT pathway partly through regulating the miR-515-5p and Smad4 expression in THCA cell. The CATIP-AS1 could be a promising biomarker for early THCA detection and prognosis and a possible therapeutic target for its treatment.

Terpene synthases in cucumber (Cucumis sativus) and their contribution to herbivore-induced volatile terpenoid emission.

Terpenoids play important roles in flavour, pollinator attraction and defence of plants. In cucumber (Cucumis sativus) they are important components of the herbivore-induced plant volatile blend that attracts natural enemies of herbivores. We annotated the cucumber TERPENE SYNTHASE gene (CsTPS) family and characterized their involvement in the response towards herbivores with different feeding guilds using a combined molecular and biochemical approach. Transcripts of multiple CsTPS genes were upregulated in leaves upon herbivory and the products generated by the expressed proteins match the terpenoids recorded in the volatile blend released by herbivore-damaged leaves. Spatial and temporal analysis of the promoter activity of CsTPS genes showed that cell content-feeding spider mites (Tetranychus urticae) and thrips (Frankliniella occidentalis) induced promoter activity of CsTPS9 and CsTPS19 within hours after initiation of infestation, while phloem-feeding aphids (Myzus persicae) induced CsTPS2 promoter activity. Our findings offer detailed insights into the involvement of the TPS gene family in the dynamics and fine-tuning of the emission of herbivore-induced plant volatiles in cucumber, and open a new avenue to understand molecular mechanisms that affect plant-herbivore interactions.

Screening of CXC chemokines in the microenvironment of ovarian cancer and the biological function of CXCL10.

BACKGROUND: This study aims to screen and identify the biological functions and prognostic value of CXC chemokines in ovarian cancer (OC) through bioinformatics and molecular biology methods, and to provide data support for the selection of biomarkers and prognostic analysis of OC. METHODS: In this study, GEO, ONCOMINE, GEPIA, cBioPortal, GeneMANIA, Metascape, STRING, TRRUST, and TIMER databases were used to study CXC chemokines. Angiogenesis and T cell killing assay were used to detect the effect of CXCL10 on tumor cell immunity and angiogenesis. Real-time quantitative PCR (qRT-PCR), immunoblotting, and ectopic tumor formation experiments were used to verify the effect of CXCL10 on ovarian cancer tumors. RESULTS: We found that CXCL1, CXCL10, CXCL11, CXCL13, and CXCL14 were significantly upregulated in OC samples compared with normal tissues. Our data showed that there was a relationship between the expression of CXC chemokines and the infiltration of six types of immune cells significant correlation. In vitro assay confirmed that overexpression of CXCL10 could enhance the killing effect of T cells and inhibit angiogenesis. Further in vivo assay had shown that CXCL10 could affect the progression of ovarian cancer by increasing the expression of cytotoxic T cells and inhibiting angiogenesis. CONCLUSION: In conclusion, we hope that our data will provide new insights into the development of immunotherapy and the selection of prognostic markers for patients with OC.

Phloretin Modulates Human Th17/Treg Cell Differentiation In Vitro via AMPK Signaling.

OBJECTIVE: We conducted studies to explore the effect of phloretin on glucose uptake, proliferation, and differentiation of human peripheral blood CD4+ T cells and investigated the mechanism of phloretin on inducing Th17/Treg development. METHODS: Naïve CD4+ T cells were purified from peripheral blood of healthy volunteers, stimulated with anti-CD3/CD28 antibodies, and polarized in vitro to generate Th17 or Treg cells. Glucose uptake, proliferation, cell cycle, protein expression (phospho-Stat3, phospho-Stat5), and Th17 and Treg cell numbers were analyzed by flow cytometry. AMP-activated protein kinase (AMPK) signaling was analyzed by western blot. Results and Discussion. Phloretin could inhibit the glucose uptake and proliferation of activated CD4+ T cells. The proliferation inhibition was due to the G0/G1 phase arrest. Phloretin decreased Th17 cell generation and phospho-Stat3 expression as well as increased Treg cell generation and phospho-Stat5 expression in the process of inducing Th17/Treg differentiation. The phosphorylation level of AMPK was significantly enhanced, while the phosphorylation level of mTOR was significantly decreased in activated CD4+ T cells under phloretin treatment. The AMPK signaling inhibitor compound C (Com C) could neutralize the effect of phloretin, while the agonist 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR) could impact the Th17/Treg balance similar to phloretin during Th17/Treg induction. CONCLUSION: Our results suggest that phloretin can mediate the Th17/Treg balance by regulating metabolism via the AMPK signal pathway.

Intraoperative Hepatic Blood Inflow Can Predict Early Acute Kidney Injury following DCD Liver Transplantation: A Retrospective Observational Study.

PURPOSE: Acute kidney injury (AKI) is a major and severe complication following donation-after-circulatory-death (DCD) liver transplantation (LT) and is associated with increased postoperative morbidity and mortality. However, the risk factors and the prognosis factors of AKI still need to be further explored, and the relativity of intraoperative hepatic blood inflow (HBI) and AKI following LT has not been discussed yet. The purpose of this study was to investigate the correlation between HBI and AKI and to construct a prediction model of early acute kidney injury (EAKI) following DCD LT with the combination of HBI and other clinical parameters. METHODS: Clinical data of 132 patients who underwent DCD liver transplantation at the first hospital of China Medical University from April 2005 to March 2017 were analyzed. Data of 105 patients (the first ten years of patients) were used to develop the prediction model. Then we assessed the clinical usefulness of the prediction models in the validation cohort (27 patients). EAKI according to Kidney Disease Improving Global Outcomes (KDIGO) criteria based on serum creatinine increase during 7-day of postoperative follow-up. RESULTS: After Least Absolute Shrinkage and Selection Operator (LASSO) regression and simplification, a simplified prediction model consisting of the Child-Turcotte-Pugh (CTP) score (p=0.033), anhepatic phase (p=0.014), packed red blood cell (pRBC) transfusion (p=0.027), and the HBI indexed by height (HBI/h) (p=0.002) was established. The C-indexes of the model in the development and validation cohort were 0.823 [95% CI, 0.738-0.908] and 0.921 [95% CI, 0.816-1.000], respectively. CONCLUSIONS: In this study, we demonstrated the utility of HBI/h as a predictor for EAKI following DCD LT, as well as the clinical usefulness of the prediction model through the combination of the CTP score, anhepatic phase, pRBC transfusion and HBI/h.

Pentapeptide Protects INS-1 Cells From hIAPP-Mediated Apoptosis by Enhancing Autophagy Through mTOR Pathway.

The human islet amyloid polypeptide (hIAPP), the major component of islet amyloid deposition, is one of the amyloidogenic peptides and has been associated with ß cell loss and dysfunction in type 2 diabetes (T2D). Autophagy plays a central role in the clearance of hIAPP aggregates, thereby diminishing the hIAPP-induced cytotoxicity. Conversely, hIAPP has been reported to have interfering effects on the autophagy. The pentapeptide FLPNF developed in our previous study has been shown to have effects on the level of the downstream proteins of mTOR and autophagy-lysosome pathway. In the present study, the peptide FLPNF-mediated increase in autophagy flux and its underlying mechanisms, as well as its protecting effect on INS-1 cells, were investigated. Autophagy flux in INS-1 cells overexpressing hIAPP (hIAPP-INS-1 cells) markedly increased after exposure to peptide FLPNF for 24 h and peaked at a concentration of 200 µM. Peptide FLPNF enhanced the autophagy by inhibiting the mTORC1 activity. Flow cytometry results showed the peptide FLPNF bind to mammalian target of rapamycin (mTOR), and further molecular docking analysis revealed a direct interaction between peptide FLPNF and the FRB domain of mTOR. Meanwhile, both peptide FLPNF and rapamycin significantly decreased the hIAPP-induced apoptosis, whereas 3-MA increased the apoptosis. Furthermore, peptide FLPNF reduced the hIAPP oligomer and improved the hIAPP-INS-1 cells insulin release function at high glucose concentration. Taken together, the peptide FLPNF decreased the hIAPP oligomer via upregulating autophagy by inhibiting mTORC1 activity, thus protecting the INS-1 cells from hIAPP-induced apoptosis and improving the insulin release function of INS-1 cells.

A Novel Pentapeptide Inhibitor Reduces Amyloid Deposit Formation by Direct Interaction with hIAPP.

BACKGROUNDS: The presence of amyloid deposits of human islet amyloid polypeptide (hIAPP) in islet ß-cells has been associated with type 2 diabetes occurrence and islet graft failure. Self-assembly into oligomers and fibrils during the process of aggregation by hIAPP can lead to failure and depletion of ß-cells. Studies have shown that some critical regions of hIAPP might contribute to the aggregation. Thus, many studies focused on finding the effective molecules, especially the short-peptide inhibitors, that bind to these regions and disrupt the aggregation of hIAPP. In the present study, a novel pentapeptide inhibitor Phe-Leu-Pro-Asn-Phe (FLPNF) was designed and its effectiveness on the inhibition of the formation of amyloid deposits was examined. METHODS: The binding mode between FLPNF and hIAPP was performed using molecular docking. The effectiveness of FLPNF on inhibiting hIAPP amyloid aggregation was tested by Thioflavin T (ThT) staining. Furthermore, negative stain electron microscopy was used to observe hIAPP fibrils. A biolayer interferometry analysis was used to identify the interaction between FLPNF and hIAPP. In addition, the cytotoxicity toward INS-1 cells was tested by a cell proliferation assay. RESULTS: FLPNF was predicted to have a compact conformation to bind at the site of hIAPP. FLPNF strongly inhibited the amyloid aggregation of hIAPP at a 10 : 1 molar ratio in vitro. Coincubation of FLPNF with hIAPP decreased the amount of hIAPP fibrils. Furthermore, a direct interaction between FLPNF and hIAPP was confirmed. FLPNF could also decrease the cytotoxic effect of hIAPP. CONCLUSIONS: The novel pentapeptide inhibitor FLPNF was constructed and inhibited the aggregation through direct binding to hIAPP. It is considered a suitable inhibitor for hIAPP amyloid deposit formation.

Resveratrol attenuates hIAPP amyloid formation and restores the insulin secretion ability in hIAPP-INS1 cell line via enhancing autophagy.

It has been proved that human islet amyloid polypeptide (hIAPP), the main constituent of islet amyloid deposition, is one of the important factors that can induce type 2 diabetes or graft failure after islet transplantation. As there is no research on whether resveratrol degrading the amyloid deposition by its special chemical structure or enhancing autophagy had been published, we decided to detect the function of resveratrol in degrading the amyloid deposition in pancreatic beta cells. We established stable hIAPP-INS1 cell line via transfecting INS1 cells by lentivirus that overexpresses hIAPP. Our research demonstrates that amyloid deposition existed in hIAPP-INS1 cell by the thioflavin S fluorescent staining, meanwhile the function of insulin secretion of hIAPP-INS1 cells was decreased significantly (p < 0.01). After treatment with resveratrol (20 µM) for 24 h, amyloid deposition in hIAPP-INS1 cells was decreased significantly, and the insulin secretion was restored significantly (p < 0.01). Once inhibited the autophagy of hIAPP-INS1 cells by 3-methyladenine for 24 h, resveratrol does not effectively remove hIAPP deposits again, and cannot improve the function of insulin secretion. These results provide a novel thought that resveratrol can degrade the amyloid deposition in type 2 diabetes and the graft after islet transplantation.

miR-663a inhibits tumor growth and invasion by regulating TGF-ß1 in hepatocellular carcinoma.

BACKGROUND: The dysregulation of miR-663a is frequently observed in many human cancers. However, the functional role and precise mechanism of miR-663a have been controversial in hepatocellular carcinoma (HCC) and need to be studied in depth. METHODS: The expression of miR-663a was detected in human cell lines and HCC tissues by quantitative RT-PCR (qRT-PCR), and data from the Cancer Genome Atlas (TCGA). Cell proliferation was investigated using MTS, EdU, colony formation assays, and xenograft animal experiments, and the cell invasion capacity was evaluated using the transwell assay. The target gene of miR-663a was identified by qRT-PCR, Western blot, and dual-luciferase reporter assays. The clinicopathological features of miR-663a and the correlation between miR-663a and TGF-ß1 expression were also investigated in the clinical samples of HCC. RESULTS: miR-663a was significantly downregulated in HCC cells relative to immortal normal liver cells, as indicated using qRT-PCR, and the lower expression of miR-663a was also confirmed in HCC tissue samples and the data from TCGA. The expression of miR-663a in HCC tissue samples was statistically significantly associated with size and the number of tumors. In addition, the upregulation of miR-663a inhibited the proliferation and invasion of HCC cells in vitro. Further study showed that miR-663a directly targeted transforming growth factor beta 1 (TGF-ß1) to suppress HCC invasion, and that the inhibitory effect of miR-663a on cell invasion could be regulated by TGF-ß1. In vivo studies showed that miR-663a significantly inhibited tumor growth. A negative correlation between miR-663a and TGF-ß1 expression was also confirmed from the clinical samples of HCC. CONCLUSIONS: miR-663a acts as a tumor suppressor and exerts a substantial role in inhibiting the proliferation, invasion, and tumorigenesis of HCC by regulating TGF-ß1 in vitro and in vivo. These observations indicate that miR-663a may be a suitable diagnostic, therapeutic, and prognostic target for the treatment of HCC.

DNMTs inhibitor SGI-1027 induces apoptosis in Huh7 human hepatocellular carcinoma cells.

SGI-1027, a novel class of relatively stable, highly lipophilic quinoline-based small-molecule inhibitors of DNA methyltransferase enzymes (DNMTs), is able to inhibit DNMTs activity, and reactivate tumor suppressor genes. However, the potential anticancer mechanisms of SGI-1027 on human hepatocellular carcinoma (HCC) cells are still not clearly understood. Thus, the objective of the present study was to clarify the inhibitory effect of SGI-1027 on the cell cycle and apoptosis of the Huh7 cell line. The results revealed that treatment with SGI-1027 resulted in a significant dose-dependent decrease in cell viability. Flow cytometric analysis identified that a 24 h treatment of SGI-1027 resulted in cell apoptosis, and typical apoptotic nucleic alterations were observed with fluorescence microscopy following terminal deoxynucleotidyl-transferase-mediated dUTP nick end labeling staining. Immunoblot analysis further demonstrated that SGI-1027 downregulated the expression of B cell lymphoma-2 and upregulated the expression of Bcl-associated X protein. However, no significant alterations of the cell cycle phases were observed. Overall, it is demonstrated that SGI-1027 causes cell apoptosis via the mitochondrial-mediated pathway, which advances current understanding of the molecular mechanisms of SGI-1027 in HCC management.