Shared and distinct biological circuits in effector, memory and exhausted CD8+ T cells revealed by temporal single-cell transcriptomics and epigenetics.

Naïve CD8+ T cells can differentiate into effector (Teff), memory (Tmem) or exhausted (Tex) T cells. These developmental pathways are associated with distinct transcriptional and epigenetic changes that endow cells with different functional capacities and therefore therapeutic potential. The molecular circuitry underlying these developmental trajectories and the extent of heterogeneity within Teff, Tmem and Tex populations remain poorly understood. Here, we used the lymphocytic choriomeningitis virus model of acute-resolving and chronic infection to address these gaps by applying longitudinal single-cell RNA-sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) analyses. These analyses uncovered new subsets, including a subpopulation of Tex cells expressing natural killer cell-associated genes that is dependent on the transcription factor Zeb2, as well as multiple distinct TCF-1+ stem/progenitor-like subsets in acute and chronic infection. These data also revealed insights into the reshaping of Tex subsets following programmed death 1 (PD-1) pathway blockade and identified a key role for the cell stress regulator, Btg1, in establishing the Tex population. Finally, these results highlighted how the same biological circuits such as cytotoxicity or stem/progenitor pathways can be used by CD8+ T cell subsets with highly divergent underlying chromatin landscapes generated during different infections.

Stat5 opposes the transcription factor Tox and rewires exhausted CD8+ T cells toward durable effector-like states during chronic antigen exposure.

Rewiring exhausted CD8+ T (Tex) cells toward functional states remains a therapeutic challenge. Tex cells are epigenetically programmed by the transcription factor Tox. However, epigenetic remodeling occurs as Tex cells transition from progenitor (Texprog) to intermediate (Texint) and terminal (Texterm) subsets, suggesting development flexibility. We examined epigenetic transitions between Tex cell subsets and revealed a reciprocally antagonistic circuit between Stat5a and Tox. Stat5 directed Texint cell formation and re-instigated partial effector biology during this Texprog-to-Texint cell transition. Constitutive Stat5a activity antagonized Tox and rewired CD8+ T cells from exhaustion to a durable effector and/or natural killer (NK)-like state with superior anti-tumor potential. Temporal induction of Stat5 activity in Tex cells using an orthogonal IL-2:IL2Rß-pair fostered Texint cell accumulation, particularly upon PD-L1 blockade. Re-engaging Stat5 also partially reprogrammed the epigenetic landscape of exhaustion and restored polyfunctionality. These data highlight therapeutic opportunities of manipulating the IL-2-Stat5 axis to rewire Tex cells toward more durably protective states.

Developmental Relationships of Four Exhausted CD8+ T Cell Subsets Reveals Underlying Transcriptional and Epigenetic Landscape Control Mechanisms.

CD8+ T cell exhaustion is a major barrier to current anti-cancer immunotherapies. Despite this, the developmental biology of exhausted CD8+ T cells (Tex) remains poorly defined, restraining improvement of strategies aimed at "re-invigorating" Tex cells. Here, we defined a four-cell-stage developmental framework for Tex cells. Two TCF1+ progenitor subsets were identified, one tissue restricted and quiescent and one more blood accessible, that gradually lost TCF1 as it divided and converted to a third intermediate Tex subset. This intermediate subset re-engaged some effector biology and increased upon PD-L1 blockade but ultimately converted into a fourth, terminally exhausted subset. By using transcriptional and epigenetic analyses, we identified the control mechanisms underlying subset transitions and defined a key interplay between TCF1, T-bet, and Tox in the process. These data reveal a four-stage developmental hierarchy for Tex cells and define the molecular, transcriptional, and epigenetic mechanisms that could provide opportunities to improve cancer immunotherapy.

Risk factors and outcomes in patients who switched from peritoneal dialysis to physician-oriented or patient-oriented kidney replacement therapy.

BACKGROUND: We aimed to compare the cardiovascular events and mortality in patients who underwent either physician-oriented or patient-oriented kidney replacement therapy (KRT) conversion due to discontinuation of peritoneal dialysis (PD). METHODS: Patients with end-stage kidney disease who were receiving PD and required a switch to an alternative KRT were included. They were divided into physician-oriented group or patient-oriented group based on the decision-making process. Logistic regression analysis was used to explore the influencing factors related to KRT conversion in PD patients. The association of physician-oriented or patient-oriented KRT conversion with outcomes after the conversion was assessed by using Cox proportional hazards models. RESULTS: A total of 257 PD patients were included in the study. The median age at catheterization was 35 years. 69.6% of the participants were male. The median duration of PD was 20 months. 162 participants had patient-oriented KRT conversion, while 95 had physician-oriented KRT conversion. Younger patients, those with higher education levels, higher income, and no diabetes were more likely to have patient-oriented KRT conversion. Over a median follow-up of 39 months, 40 patients experienced cardiovascular events and 16 patients died. Physician-oriented KRT conversion increased nearly 3.8-fold and 4.0-fold risk of cardiovascular events and death, respectively. After adjusting for confounders, physician-oriented KRT conversion remained about a 3-fold risk of cardiovascular events. CONCLUSION: Compared to patient-oriented KRT conversion, PD patients who underwent physician-oriented conversion had higher risks of cardiovascular events and all-cause mortality. Factors included age at catheterization, education level, annual household income, and history of diabetes mellitus.

Cobaltosic oxide-polyethylene glycol-triphenylphosphine nanoparticles ameliorate the acute-to-chronic kidney disease transition by inducing BNIP3-mediated mitophagy.

Accumulating evidence highlights mitochondrial dysfunction as a crucial factor in the pathogenesis of acute kidney injury (AKI); thus, novel therapeutic strategies maintaining mitochondrial homeostasis are highly anticipated. Recent studies have shown that cobaltosic oxide has peroxidase-like catalytic activities, although its role and mechanism remain elusive in AKI. In the present study, we synthesized and identified cobaltosic oxide-polyethylene glycol-triphenylphosphine (COPT) nanoparticles by conjugating cobaltosic oxide with polyethylene glycol and triphenylphosphine, to improve its biocompatibility and mitochondria-targeting property. We found that COPT preferentially accumulated in the kidney proximal tubule cells, and significantly alleviated ischemic AKI in mouse models and gentamicin induced-AKI in the zebrafish model. COPT also inhibited the transition from AKI to chronic kidney disease (CKD), with few side effects. Further studies demonstrated that COPT localized in the mitochondria, and ameliorated hypoxia-reoxygenation-mediated mitochondrial damage through enhancing mitophagy in vitro and in vivo. Mechanistically, COPT dose-dependently induced the expression of Bcl-2/adenovirus E1B 19-kDa interacting protein (BNIP3), while knockdown of BNIP3 attenuated COPT-induced mitophagic flux and mitochondrial protection. Thus, our findings suggest that COPT nanoparticles ameliorate AKI and its progression to CKD through inducing BNIP3-mediated mitophagy, indicating that COPT may serve as a promising mitochondria-targeting therapeutic agent against AKI.

Imbalanced lipid homeostasis caused by membrane αKlotho deficiency contributes to the acute kidney injury to chronic kidney disease transition.

After acute kidney injury (AKI), renal tubular epithelial cells (RTECs) are pathologically characterized by intracellular lipid droplet (LD) accumulation, which are involved in RTEC injury and kidney fibrosis. However, its pathogenesis remains incompletely understood. The protein, αKlotho, primarily expressed in RTECs, is well known as an anti-aging hormone wielding versatile functions, and its membrane form predominantly acts as a co-receptor for fibroblast growth factor 23. Here, we discovered a connection between membrane αKlotho and intracellular LDs in RTECs. Fluorescent fatty acid (FA) pulse-chase assays showed that membrane αKlotho deficiency in RTECs, as seen in αKlotho homozygous mutated (kl/kl) mice or in mice with ischemia-reperfusion injury (IRI)-induced AKI, inhibited FA mobilization from LDs by impairing adipose triglyceride lipase (ATGL)-mediated lipolysis and lipophagy. This resulted in LD accumulation and FA underutilization. IRI-induced alterations were more striking in αKlotho deficiency. Mechanistically, membrane αKlotho deficiency promoted E3 ligase peroxin2 binding to ubiquitin-conjugating enzyme E2 D2, resulting in ubiquitin-mediated degradation of ATGL which is a common molecular basis for lipolysis and lipophagy. Overexpression of αKlotho rescued FA mobilization by preventing ATGL ubiquitination, thereby lessening LD accumulation and fibrosis after AKI. This suggests that membrane αKlotho is indispensable for the maintenance of lipid homeostasis in RTECs. Thus, our study identified αKlotho as a critical regulator of lipid turnover and homeostasis in AKI, providing a viable strategy for preventing tubular injury and the AKI-to-chronic kidney disease transition.

A large Stokes shift NIR fluorescent probe for visual monitoring of mitochondrial peroxynitrite during inflammation and ferroptosis and in an Alzheimer's disease model.

The excessive formation of peroxynitrite (ONOO-) in mitochondria has been implicated in various pathophysiological processes and diseases. However, owing to short emission wavelengths and small Stokes shifts, previously reported fluorescent probes pose significant challenges for mitochondrial ONOO- imaging in biological systems. In this study, a near-infrared (NIR) fluorescent probe, denoted as DCO-POT, is designed for the visual monitoring of mitochondrial ONOO-, displaying a remarkable Stokes shift of 170 nm. The NIR fluorophore of DCO-CHO is released by DCO-POT upon the addition of ONOO-, resulting in off-on NIR fluorescence at 670 nm. This phenomenon facilitates the high-resolution confocal laser scanning imaging of ONOO- generated in biological systems. The practical applications of DCO-POT as an efficient fluorescence imaging tool are verified in this study. DCO-POT enables the fluorometric visualization of ONOO- in organelles, cells, and organisms. In particular, ONOO- generation is analyzed during cellular and organism-level (zebrafish) inflammation during ferroptosis and in an Alzheimer's disease mouse model. The excellent visual monitoring performance of DCO-POT in vivo makes it a promising tool for exploring the pathophysiological effects of ONOO-.

Geriatric Nutritional Risk Index and Risk of Mortality in Critically Ill Patients With Acute Kidney Injury: A Multicenter Cohort Study.

OBJECTIVES: Malnutrition is associated with adverse outcomes in acute or chronic diseases. However, the prediction value of the Geriatric Nutritional Risk Index (GNRI) in critically ill patients with acute kidney injury (AKI) has not been well studied. METHODS: Data was extracted from the Medical Information Mart for Intensive Care III (MIMIC-III) and the electronic intensive care unit database. We used two nutritional indicators, the GNRI and the modified Nutrition Risk in Critically ill (NUTRIC) score, to evaluate the relationship between the nutritional status of patients with AKI and prognosis. The outcome is in-hospital mortality and 90-day mortality. The prediction accuracy of GNRI was compared with the NUTRIC score. RESULTS: A total of 4,575 participants with AKI were enrolled in this study. The median age of 68 (interquartile range, 56-79) years, and 1,142 (25.0%) patients experienced in-hospital mortality, and 1,238 (27.1%) patients experienced 90-day mortality. Kaplan-Meier survival analysis indicated that lower GNRI levels and high NUTRIC score are associated with lower in-hospital and 90-day survival of patients with AKI (P < .001 by log-rank test). After multivariate adjustment, Cox regression analysis demonstrated a 2-fold increased risk of in-hospital (hazard ratio = 2.019, 95% confidence interval: 1.699-2.400, P < .001) and 90-day (hazard ratio = 2.023, 95% confidence interval: 1.715-2.387, P < .001) mortality in the low GNRI group. Moreover, the multivariate-adjusted Cox model containing GNRI had higher prediction accuracy for the prognosis of patients with AKI than that with NUTRIC score (AUCGNRI model vs. AUCNUTRIC model for in-hospital mortality = 0.738 vs. 0.726, AUCGNRI model vs. AUCNUTRIC model for 90-day mortality = 0.748 vs. 0.726). In addition, the prediction value of GNRI was validated by the electronic intensive care unit database (7,881 patients with AKI) with satisfying performance (AUCGNRI model = 0.680). CONCLUSIONS: Our results demonstrated that GNRI is strongly associated with survival in patients in the intensive care unit coexisting with AKI, and the GNRI has a superior predictive value than the NUTRIC score.

Two Filters for Acquiring the Profiles from Images Obtained from Weak-Light Background, Fluorescence Microscope, Transmission Electron Microscope, and Near-Infrared Camera.

Extracting the profiles of images is critical because it can bring simplified description and draw special attention to particular areas in the images. In our work, we designed two filters via the exponential and hypotenuse functions for profile extraction. Their ability to extract the profiles from the images obtained from weak-light conditions, fluorescence microscopes, transmission electron microscopes, and near-infrared cameras is proven. Moreover, they can be used to extract the nesting structures in the images. Furthermore, their performance in extracting images degraded by Gaussian noise is evaluated. We used Gaussian white noise with a mean value of 0.9 to create very noisy images. These filters are effective for extracting the edge morphology in the noisy images. For the purpose of a comparative study, we used several well-known filters to process these noisy images, including the filter based on Gabor wavelet, the filter based on the watershed algorithm, and the matched filter, the performances of which in profile extraction are either comparable or not effective when dealing with extensively noisy images. Our filters have shown the potential for use in the field of pattern recognition and object tracking.

[Trimethylamine N-Oxide Induces Renal Fibrosis Through the PI3K/AKT/SREBP1 Pathway]. / 氧化三甲胺通过PI3K/AKT/SREBP1通路诱导肾纤维化.

Objective: To investigate the role and mechanism of trimethylamine N-oxide (TMAO), a uremic toxin, in renal fibrosis. Methods: A total of 20 male BALB/c mice were randomly and evenly assigned to a Control group and a TMAO group. Mice in the Control group received intraperitoneal injection of normal saline, while mice in the TMAO group received intraperitoneal injection of TMAO (20 mg/[kg·d]). The injection was given once a day for 8 weeks. Histopathology and fibrosis of kidney were observed by H&E staining and Masson staining. Immunohistochemistry was performed to determine the levels of alpha smooth muscle actin (α-SMA), recombinant human fibronectin fragment (Fibronectin), and sterol-regulatory element binding protein 1 (SREBP1). Western blot was performed to determine α-SMA, SREBP1, phosphatidylinositol 3 kinase (PI3K), phospho-phosphatidylinositol 3 kinase (p-PI3K), protein kinase B (PKB, also known as AKT), and phospho-AKT (p-AKT) protein levels. HK2 cells were treated with SREBP1 small interfering RNA (siRNA) and PI3K/AKT inhibitor, respectively, and the reversal of the effects of TMAO was examined. Results: Animal experiments showed that, compared with the Control group, the mice treated with TMAO experienced pathological damage and fibrosis of the kidney tissue and the expression levels of fibrosis markers, α-SMA and Fibronectin, in the kidney were increased (all P<0.05). According to the findings from further investigation, the TMAO-treatment group showed increased expression of SREBP1 and an up-regulation of PI3K phosphorylation ratio and AKT phosphorylation ratio compared with those of the Control group (all P<0.05). Cell experiments produced results similar to those of the animal experiment. After siRNA interference with SREBP1 expression, the expression levels of fibrosis marker proteins decreased (P<0.05). Besides, the high expression of SREBP1 caused by TMAO was inhibited after HK2 cells were incubated with LY294002, a PI3K-AKT pathway inhibitor (P<0.05). Conclusion: TMAO may induce renal fibrosis by promoting the PI3K/AKT/SREBP1 pathway.

Circular RNA hsa_circ_0018189 drives non-small cell lung cancer growth by sequestering miR-656-3p and enhancing xCT expression.

BACKGROUND: Non-small cell lung cancer (NSCLC) is one of the cancers with a high mortality rate. CircRNAs have emerged as an important regulatory factor in tumorigenesis in recent years. However, the detailed regulatory mechanism of a circular RNA cullin 2 (hsa_circ_0018189; hsa_circ_0018189) is still unclear in NSCLC. METHODS: RNA levels of hsa_circ_0018189, microRNA (miR)-656-3p, and Solute carrier family seven member 11 (SLC7A11, xCT) were analyzed by real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR), and protein level was assessed by Western blot and immunohistochemical assay. Enzyme-linked immunosorbent assay was conducted to detect cell glutamine metabolism. Effects of hsa_circ_0018189 on cell proliferation, apoptosis, migration, and invasion were analyzed by corresponding assays. Luciferase reporter assay and RNA-immunoprecipitation assay confirmed the target relationship between miR-656-3p and hsa_circ_0018189 or xCT. The in vivo function of hsa_circ_0018189 was verified by xenograft mouse models. RESULTS: Hsa_circ_0018189 abundance was overexpressed in NSCLC cells and samples. Deficiency of hsa_circ_0018189 lowered NSCLC cell proliferative, migrating, invading, and glutamine metabolism capacities, and hsa_circ_0018189 silencing inhibited the growth of tumors in vivo. Hsa_circ_0018189 could up-regulate xCT by sponging miR-656-3p. And miR-656-3p downregulation or xCT overexpression partly overturned hsa_circ_0018189 knockdown or miR-656-3p mimic-mediated repression of NSCLC cell malignancy. CONCLUSION: Hsa_circ_0018189 drove NSCLC growth by interacting with miR-656-3p and upregulating xCT.

Arteriolar hyalinosis and renal outcomes in patients with immunoglobulin A nephropathy.

BACKGROUND: The relationship between arteriolar hyalinosis and renal progression in immunoglobulin A nephropathy (IgAN) is not fully understood. We aimed to investigate the clinicopathological features and outcomes of IgAN with or without arteriolar hyalinosis. METHODS: A total of 762 diagnosed with IgAN patients were retrospectively analyzed. We classified IgAN patients into two groups with or without arteriolar hyalinosis. Then, the clinicopathological characteristics of the two groups were compared. We used Kaplan-Meier survival analysis to compare the composite kidney outcome of the two groups and applied multivariate Cox regression analyses to test the association between arteriolar hyalinosis and composite kidney outcome. RESULTS: Overall, 412 (54.1%) patients had arteriolar hyalinosis, including 173 patients diagnosed with hypertension. IgAN patients with arteriolar hyalinosis were older and had higher proteinuria, urea, uric acid, and blood pressure, while lower eGFR than those without arteriolar hyalinosis. Subgroup analysis showed similar results in IgAN patients with hypertension. Kaplan-Meier survival analysis showed that IgAN patients with arteriolar hyalinosis had worse composite kidney outcome than those without arteriolar hyalinosis. In addition, subgroup analysis revealed that patients with hypertension have worse composite kidney outcome than those without hypertension. Multivariate Cox regression analyses confirm that arteriolar hyalinosis (HR 2.57; 95% CI 1.41-4.69; p = 0.002) is an independent risk factor for renal prognosis in IgAN patients. CONCLUSIONS: Our study demonstrated that arteriolar hyalinosis is a common vascular lesion in IgAN patients. Arteriolar hyalinosis connects closely with hypertension, and arteriolar hyalinosis is an independent risk factor for renal prognosis in patients with IgAN.

IRF-1 promotes renal fibrosis by downregulation of Klotho.

Although the key role of renal fibrosis in the progression of chronic kidney disease (CKD) is well known, the causes of renal fibrosis are not fully clarified. In this study, interferon regulatory factor 1 (IRF-1), a mammalian transcription factor, was highly expressed in fibrotic kidney of CKD patients. Concordantly, the expression level of IRF-1 was significantly elevated in the kidney of unilateral ureteral obstruction (UUO) and Adriamycin nephropathy (ADR) mice. In tubular epithelial cells, overexpression of IRF-1 could induce profibrotic markers expression, which accompanied by dramatic downregulation of Klotho, an important inhibitor of renal fibrosis. Luciferase reporter analysis and ChIP assay revealed that IRF-1 repressed Klotho expression by downregulation of C/EBP-ß, which regulates Klotho gene transcription via directly binding to its promoter. Further investigation showed that tumor necrosis factor-alpha may be an important inducement for the increase of IRF-1 in tubular epithelial cells after UUO and genetic deletion of IRF-1 attenuated renal fibrosis in UUO mice. Hence, these findings demonstrate that IRF-1 contributes to the pathogenesis of renal fibrosis by downregulation of Klotho, and suppresses IRF-1 may be a potential therapeutic target for CKD.

Analysis of neurite length of hippocampal neurons cultured into 3D artificial network patterned microfluidic chips.

Purpose: The study aims to lay a foundational probe for the thorough application microfluidic chips in brain function research with microfluidic chips. Neuron slide culture is a common culture method in vitro, and the microfluidic chip with the artificial network pattern not only can realize neuron cells 3 D culture in vitro, but also limit the extension space of neurite outgrow.Materials and Methods: In order to analyze the differences of hippocampal cells neurite growth length between the 3 D chips and the common 2 D culture, the experiments utilized statistical analysis method analyzing the length of the hippocampus neuron neurite of 3 days, 5 days and 7 days, respectively, with the common glass slide 2 D culture method and the microfluidic chip 3 D culture in vitro.Results: The results showed that there was no significant difference in the neurite length after 3 days. However, there was a significant difference after 5 days and 7 days. It can be seen that the microfluidic chip with artificial network pattern has limitations to the growth of neurite after 5 days.Conclusions: We concluded that the growth state of hippocampal cells in the restricted 3 D space is different from that of conventional 2 D culture.It showed that the artificial network pattern design has limited the growth space of the dendrites but also affected its growth.

Putative promoters within gene bodies control exon expression via TET1-mediated H3K36 methylation.

Hypermethylation of gene promoter has been indicated for the contribution of gene silencing, and DNA demethylating drugs, such as 5-aza-2'-deoxycytidine (DAC), has been used clinically for cancer treatment. However, the reason why a proportion of genes with hypermethylated promoter exhibit high expression levels remains unclear and this drug is not much successful as expected in use. Furthermore, CpG islands (CGIs) are found to be located in not only promotors, but also in gene bodies. By RNA-seq and reduced representation bisulfite sequencing, we found the mismatch between the level of promoter methylation and gene expression. By chromatin Immunoprecipitation-quantitative polymerase chain reaction and luciferase reporter assay, we identified putative promoters in gene body, and proved the activities of putative promoters were affected by the methylation level of the CGI nearby. DAC can reverse the DNA hypermethylation at promoter CGIs effectively but not the CGIs in gene body. We also found that TET1 could demethylate CGIs both in promoter and gene body. Furthermore, we revealed a novel mechanism that H3K36me3 could affect the activity of putative promoter, and 5hmC recruited MeCP2 and CREB1 as a coactivator to SETD2 promoter, to enhance its gene expression and result in increased H3K36me3 in gene body. Our results concluded that putative promoters existed in the gene bodies, and TET1 could influence the transcriptional activity of putative promoters by intragenic demethylation.

ROS/p38MAPK-induced lamin B1 accumulation promotes chronic kidney disease-associated vascular smooth muscle cells senescence.

The incidence of cardiovascular thrombotic events which are highly associated with atherosclerotic plaque vulnerability and its rupture is much higher in chronic kidney disease (CKD) patients than that in the general population. It has been reported that the thinning of fibrous cap in atherosclerotic plaque is a crucial factor in plaque vulnerability and thrombosis. Moreover, vascular smooth muscle cells (VSMCs) senescence play a crucial role in maintaining the thickness of fibrous cap. Lamin B1, one of the members of laminin family, is an important component of the nuclear membrane and it is related to cell senescence. While whether lamin B1 participates CKD-related VSMCs senescence and plaque vulnerability and the underlying mechanism remain unclear. Here, we found that CKD promoted fibrous cap thinning and reduced the stability of atherosclerotic plaque through accelerating VSMCs senescence. VSMCs senescence induced by CKD was related to the increased expression of lamin B1 and abnormality of nuclear membrane structure. Knocking down the expression of lamin B1 with RNA interference prevented CKD-induced aberrant nuclear membrane structure and senescence in VSMCs. Additionally, overproduction of reactive oxidative stress (ROS) and subsequent activation of ROS/p38MAPK under CKD milieus contribute to these series of outcomes, as scavenging ROS with N-acety-l-cysteine (NAC) or inhibiting p38MAPK signal pathway with SB203580 could inhibit CKD-induced activation of ROS/p38MAPK, increased expression of lamin B1, abnormality of nuclear membrane structure and VSMCs senescence. Taken together, these results suggested that ROS/p38MAPK-mediated increased expression of lamin B1 and abnormality of nuclear membrane structure was an important mechanism of CKD-induced VSMCs senescence.

Nrf2 promotes breast cancer cell migration via up-regulation of G6PD/HIF-1α/Notch1 axis.

Abnormal metabolism of tumour cells is closely related to the occurrence and development of breast cancer, during which the expression of NF-E2-related factor 2 (Nrf2) is of great significance. Metastatic breast cancer is one of the most common causes of cancer death worldwide; however, the molecular mechanism underlying breast cancer metastasis remains unknown. In this study, we found that the overexpression of Nrf2 promoted proliferation and migration of breast cancers cells. Inhibition of Nrf2 and overexpression of Kelch-like ECH-associated protein 1 (Keap1) reduced the expression of glucose-6-phosphate dehydrogenase (G6PD) and transketolase of pentose phosphate pathway, and overexpression of Nrf2 and knockdown of Keap1 had opposite effects. Our results further showed that the overexpression of Nrf2 promoted the expression of G6PD and Hypoxia-inducing factor 1α (HIF-1α) in MCF-7 and MDA-MB-231 cells. Overexpression of Nrf2 up-regulated the expression of Notch1 via G6PD/HIF-1α pathway. Notch signalling pathway affected the proliferation of breast cancer by affecting its downstream gene HES-1, and regulated the migration of breast cancer cells by affecting the expression of EMT pathway. The results suggest that Nrf2 is a potential molecular target for the treatment of breast cancer and targeting Notch1 signalling pathway may provide a promising strategy for the treatment of Nrf2-driven breast cancer metastasis.

Loss of G3BP1 suppresses proliferation, migration, and invasion of esophageal cancer cells via Wnt/ß-catenin and PI3K/AKT signaling pathways.

Accumulating evidence suggests that Ras GTPase-activating protein SH3 domain-binding protein 1 (G3BP1) is very crucial to regulate tumorigenesis and metastasis. Recently, many research works have suggested that G3BP1 is overexpressed in many human cancers including esophageal cancer. Nevertheless, the functional roles of G3BP1 in esophageal cancer are still unknown. Here, the results suggested that silencing of G3BP1 inhibited proliferation, migration, and invasion of esophageal cancer cells, whereas overexpression of G3BP1 led to opposite effects on the growth and metastasis. Surprisingly, G3BP1-depletion had no effect on cell death but caused the arrest of cell cycle in the G0 /G1 phase and increased the levels of p53 and p21. In addition, loss of G3BP1 led to a significant elevation of E-cadherin and decrease of N-cadherin, Vimentin, Snail, MMP-9, and MMP-2. Mechanistically, loss of G3BP1 dramatically suppressed Wnt-stimulated T-cell factor/lymphoid enhancer factor (TCF/LEF) transcription factor activity and downregulated its target genes including c-Myc, Axin2, and cyclin D1. Moreover, knockdown of G3BP1 downregulated the expression levels of p-PI3K, p-AKT, and p-GSK-3ß, but the total PI3K, AKT, and GSK-3ß were not changed. Furthermore, our data proved that the promoting effects of G3BP1-overexpression on cell proliferation, migration, and invasion could be rescued by PI3K inhibitor LY294002 treatment. Collectively, our results here elucidate that G3BP1-depletion suppresses proliferation, migration, and invasion capabilities of esophageal cancer cells via the inactivation of Wnt/ß-catenin and PI3K/AKT signaling pathways. Furthermore, our findings imply that G3BP1 can participate in the regulation of esophageal cancer progression, and will be taken as a promising target to treat esophageal cancer.

Fusion with mesenchymal stem cells differentially affects tumorigenic and metastatic abilities of lung cancer cells.

Cell fusion plays a crucial role in cancer progression and leads to massive aberrant changes in chromosome and gene expression involved in tumor metastasis. Cancer cells can fuse with many cell types, including stromal cells, epithelial cells, macrophages, and endothelial cells. Mesenchymal stem cells (MSCs) have been reported to migrate and incorporate into tumor sites during cancer progression. However, the underlying mechanism of stem cell fusion in tumor metastasis has not been fully deciphered. In this research, we established a cell fusion model between lung cancer cells and MSCs in vitro. We found that the hybrid cells showed enhanced metastatic capacity with increased expression of MMP-2 and MMP-9, whereas the proliferation ability was inhibited and cell cycle was blocked in the G0 /G1 phase with elevated expression of p21, p27, and p53. Moreover, the hybrid cells lost epithelial morphology and exhibited an epithelial-mesenchymal transition (EMT) change with downregulation of E-cadherin and upregulation of N-cadherin, Vimentin, α-SMA and Fibronectin1. Meanwhile, the expressions of EMT transcription factors, including Snail1, Slug, Twist1, Zeb1, and Zeb2, were also increased in hybrid cells. More important, the fusion hybrids acquired stem cell-like properties, which exhibited increased expression stem cell transcription factors Oct4, Sox2, Nanog, Kif4 as well as Bmi1. Taken together, our results suggested that cell fusion between lung cancer cells and MSCs offered enhanced metastatic capacity and characteristics of cancer stem cell by undergoing EMT. This study will contribute to explaning the origin of lung cancer stem cells and to elucidate the role of cell fusion in cancer metastasis.

Tet methylcytosine dioxygenase 1 promotes hypoxic gene induction and cell migration in colon cancer.

Ten-eleven translocation 1 (TET1), a widely reported DNA demethylation protein, has been associated with tumorigenesis and metastasis. However, whether TET1 is an oncogene or tumor suppressor gene has been controversial; the mechanism of how TET1 affects cancer progression remains unclear. The current study aims to investigate how TET1 is changed in the tumor microenvironment and to explore the mechanisms of how TET1 affects colon cancer progression. Because hypoxia prevails on solid tumors, we established an important connection between hypoxia and DNA demethylation in tumorigenesis. By qPCR and RNA interference (RNAi) technology, we found that hypoxia increased TET1 expression with a hypoxia-inducible factor-1-alpha (HIF-1α)-dependent manner. By CHIP-qPCR and pyrosequencing technology, we demonstrated that TET1 regulated the target gene expression of HIF-1α through HIF-1α binding to hypoxia-responsive elements (HREs), and HIF-1α binding to HREs depended on CpG methylation levels. By Cell Counting Kit-8 (CCK-8) and transwell assay, we showed that loss of TET1 did not affect cell proliferation but inhibited migration. We also identified two novel gene mutants of TET1 in 120 paired tumor/normal tissue specimens by DNA sequencing and found that TET1 E2082K mutant blocked the TET1-enhanced cell migration. Our results showed that the downregulation of TET1 rescued the abnormally high levels of gene expression resulting from hypoxia in tumors and reduced the migration activity of tumor cells, suggesting a therapeutic role by interference with TET1 in colon cancer treatment. By demonstrating that hypoxia upregulated TET1 and that TET1 drove HIF-1α-responsive genes, we showed that an epigenetic mechanism and tumor microenvironment-driven models coexisted and mutually affected colon cancer.