Functionalized ZnMnFe2O4-PEG-FA nanoenzymes integrating diagnosis and therapy for targeting hepatic carcinoma guided by multi-modality imaging.

With its insidious onset and atypical early symptoms, hepatic carcinoma is one of the most common and malignant tumors in the world. Therefore, it is necessary to actively pursue efficient diagnostic and treatment modalities for this malignancy. Photothermal therapy (PTT) is a non-invasive treatment technique that can generate high temperatures locally to induce tumor cell death, but its effectiveness is limited by the tissue-penetration depth of infrared light. Enzyme-catalyzed therapy promotes the production of toxic hydroxyl groups (˙OH) from hydrogen peroxide in tumor cells in situ, but its efficacy is also affected by the catalytic efficiency of ˙OH. Thus, given the complexity of tumors, multimodal therapy is critical for cancer treatment. Herein, we report a novel biomimetic nanoparticle (NP) platform (ZnMnFe2O4-PEG-FA) that enables combined PTT and nanozyme-catalyzed therapy. Due to the excellent photothermal effect of ZnMnFe2O4-PEG-FA, these NPs can reach an ideal temperature and damage tumor cells under lower near-infrared laser power irradiation, while exhibiting enhanced catalytic ability, largely alleviating the limitations of conventional PTT and catalytic therapy. Hence, the combination of these two treatments can provide significantly greater cytotoxicity. Additionally, ZnMnFe2O4-PEG-FA NPs have excellent photoacoustic imaging and magnetic resonance imaging capabilities, which enable monitoring and can guide cancer treatment. Therefore, ZnMnFe2O4-PEG-FA NPs integrate the diagnosis and treatment of tumors. Hence, this study provides a potential model of combined cancer diagnosis and treatment, which could be applied as a multimodal antitumor strategy in clinical settings in the future.

A peroxidase-like magneto-gold nanozyme AuNC@Fe3O4 with photothermal effect for induced cell apoptosis of hepatocellular carcinoma cells in vitro.

Hepatocellular carcinoma (HCC) is one of the most commonly diagnosed and malignant cancers worldwide. Conventional therapy strategies may not completely eradicate the tumor and may cause side effects during treatment. Nano-catalytic therapy, as a novel strategy, has attracted a great deal of attention. This study aimed to synthesize a multifunctional magneto-gold nanozyme AuNC@Fe3O4 and evaluate its anti-cancer potential in HepG2 cells in vitro. The characteristics of AuNC@Fe3O4 were assessed using a transmission electron microscope, dynamic light scattering, and energy-dispersive X-ray. The photothermal performance and peroxidase (POD)-like activity of AuNC@Fe3O4 were detected, using thermal camera and ultraviolet-visible spectrophotometer, respectively. The anti-cancer potential of AuNC@Fe3O4 was examined using cell counting kit-8, live/dead cell staining, and apoptosis analysis. Further research on HepG2 cells included the detection of intracellular reactive oxygen species (ROS) and lysosomal impairment. We observed that the AuNC@Fe3O4 had a small size, good photothermal conversion efficiency and high POD-like activity, and also inhibited cell proliferation and enhanced cell apoptotic ability in HepG2 cells. Furthermore, the AuNC@Fe3O4 enhanced ROS production and lysosomal impairment via the synergistic effect of photothermal and nano-catalytic therapies, which induced cell death or apoptosis. Thus, the magneto-gold nanozyme AuNC@Fe3O4 may offer a potential anti-cancer strategy for HCC.

Suppression of CPSF6 Enhances Apoptosis Through Alternative Polyadenylation-Mediated Shortening of the VHL 3'UTR in Gastric Cancer Cells.

Alternative polyadenylation (APA) is an important RNA post-transcriptional process, which can generate diverse mRNA isoforms. Increasing evidence shows that APA is involved in cell self-renewal, development, immunity, and cancer. CPSF6 is one of the core proteins of CFIm complex and can modulate the APA process. Although it has been reported to play oncogenic roles in cancer, the underlying mechanisms remain unclear. The aim of the present study was to characterize CPSF6 in human gastric cancer (GC). We observed that CPSF6 was upregulated in GC. Knockdown of CPSF6 inhibited proliferation and enhanced apoptosis of GC cells both in vitro and in vivo. Global APA site profiling analysis revealed that knockdown of CPSF6 induced widespread 3'UTR shortening of genes in GC cells, including VHL. We also found CPSF6 negatively regulated the expression of VHL through APA and VHL short-3'UTR isoform enhanced apoptosis and inhibited cell growth in GC cells. Our data suggested that CPSF6-induced cell proliferation and inhibition of apoptosis were mediated by the preferential usage of poly(A) in VHL. Our data provide insights into the function of CPSF6 and may imply potential therapeutic targets against GC.

CPSF6 links alternative polyadenylation to metabolism adaption in hepatocellular carcinoma progression.

BACKGROUND: Alternative polyadenylation (APA) is an important mechanism of gene expression regulation through generation of RNA isoforms with distinct 3' termini. Increasing evidence has revealed that APA is actively involved in development and disease, including hepatocellular carcinoma (HCC). However, how APA functions in tumor formation and progression remains elusive. In this study, we investigated the role of cleavage factor I (CFIm) subunit CPSF6 in human hepatocellular carcinoma (HCC). METHODS: Expression levels of CPSF6 in clinical tissues and cell lines were determined by qRT-PCR and western blot. Functional assays, including the cell number, MTT, colony formation and transwell, were used to determine the oncogenic role of CPSF6 in HCC. Animal experiments were used to determine the role of CPSF6 in HCC tumorigenicity in vivo. Deep sequencing-based 3 T-seq was used to profile the transcriptome-wide APA sites in both HCC cells and CPSF6 knockdown HCC cells. The function of CPSF6-affected target NQO1 with distinct 3'UTRs was characterized by metabolism assays. RESULTS: We observed CPSF6 was upregulated in HCC and the high expression of CPSF6 was associated with poor prognosis in patients. Overexpression of CPSF6 promoted proliferation, migration and invasion of HCC cells in vitro and in vivo. Transcriptome-wide APA profiling analysis indicated that high expression of CPSF6 promoted the favorable usage of the proximal poly(A) site in the 3'UTR of NQO1. We demonstrated CPSF6-induced tumorigenic activities were mediated by the NQO1 isoform with short 3'UTR. Furthermore, we found that CPSF6 induced metabolic alterations in liver cells through NQO1. CONCLUSION: CPSF6 plays a critical role in HCC progression by upregulating NQO1 expression through APA. These findings provide evidence to demonstrate that APA of NQO1 contributes to HCC progression and may have implications for developing new therapeutic strategy against this disease.

CPF impedes cell cycle re-entry of quiescent lung cancer cells through transcriptional suppression of FACT and c-MYC.

Blockade of cell cycle re-entry in quiescent cancer cells is a strategy to prevent cancer progression and recurrence. We investigated the action and mode of action of CPF mixture (Coptis chinensis, Pinellia ternata and Fructus trichosanthis) in impeding a proliferative switch in quiescent lung cancer cells. The results indicated that CPF impeded cell cycle re-entry in quiescent lung cancer cells by reduction of FACT and c-MYC mRNA and protein levels, with concomitant decrease in H3K4 tri-methylation and RNA polymerase II occupancy at FACT and c-MYC promoter regions. Animals implanted with quiescent cancer cells that had been exposed to CPF had reduced tumour volume/weight. Thus, CPF suppresses proliferative switching through transcriptional suppression of FACT and the c-MYC, providing a new insight into therapeutic target and intervention method in impeding cancer recurrence.

PLZF suppresses differentiation of mouse spermatogonial progenitor cells via binding of differentiation associated genes.

Promyelocytic leukaemia zinc finger (PLZF) is a key factor in inhibiting differentiation of spermatogonial progenitor cells (SPCs), but the underlying mechanisms are still largely unknown. In this study, the regulation of PLZF on Kit, Stra8, Sohlh2, and Dmrt1 (SPCs differentiation related genes) was investigated. We found some PLZF potential binding sites existed in the promoters of Kit, Stra8, Sohlh2, and Dmrt1. Additionally, the expressions of KIT, STRA8, SOHLH2, and DMRT1 were upregulated when PLZF was knockdown in SPCs. Furthermore, chromatin immunoprecipitation quantitative polymerase chain reaction revealed PLZF directly bound to the promoters of Kit, Stra8, Sohlh2, and Dmrt1. Besides, dual luciferase assay verified PLZF repressed those gene expressions. Collectively, our finding indicate that PLZF binds to the promoter regions of Kit, Stra8, Sohlh2, and Dmrt1 to regulate SPCs differentiation, which facilitate us to further understand the regulatory mechanism of PLZF in SPCs fates.

Mutant UBQLN2P497H in motor neurons leads to ALS-like phenotypes and defective autophagy in rats.

Mutations in ubiquilin2 (UBQLN2) have been linked to abnormal protein aggregation in amyotrophic lateral sclerosis (ALS). The mechanisms underlying UBQLN2-related neurodegenerative diseases remain unclear. Using a tetracycline-regulated gene expression system, the ALS-linked UBQLN2P497H mutant was selectively expressed in either the spinal motor neurons or astrocytes in rats. We found that selectively expressing mutant UBQLN2P497H in the spinal motor neurons caused several core features of ALS, including the progressive degeneration of motor neurons, the denervation atrophy of skeletal muscles, and the abnormal protein accumulation. Furthermore, mutant UBQLN2P497H accumulation was associated with an age-dependent decrease in several core autophagy-related proteins. ALS-like phenotypes were not observed when mutant UBQLN2P497H was overexpressed in the astrocytes, however, even though the expression of the mutant UBQLN2P497H protein was higher in these rats. Our results suggest that selectively expressing mutant UBQLN2P497H in motor neurons is sufficient to trigger the development of ALS in rats. Our results further indicate that the compromised autophagy-lysosomal pathway plays a critical role in the pathogenesis of UBQLN2-related neurodegenerative diseases.

STAT3 is required for proliferation and exhibits a cell type-specific binding preference in mouse female germline stem cells.

LIF-mediated STAT3 signaling is critically involved in stem cells and development. However, its function in mouse female germline cells (FGSCs) remains elusive. In this study, we demonstrated that LIF-induced STAT3 activation contributes to the proliferation and undifferentiation maintenance of mouse FGSCs. Characterization of the STAT3-mediated transcriptional network by intersecting ChIP-seq and RNA-seq datasets revealed 405 direct target genes of STAT3, which are primarily involved in proliferation and germline development. In particular, we observed that STAT3 exhibits a FGSC-specific binding pattern when compared with mouse embryonic stem cells. Taken together, our study reported that the LIF-mediated STAT3 activation is actively involved in FGSCs and functions through a distinctive binding pattern across the FGSC genome. This cell-type specific binding preference provides an insight into understanding the genetic base for STAT3-driven cellular functions in germline stem cells.

CHIR99021 enhances Klf4 Expression through ß-Catenin Signaling and miR-7a Regulation in J1 Mouse Embryonic Stem Cells.

Understanding the mechanisms that regulate pluripotency of embryonic stem cells (ESCs) is important to ensure their safe clinical use. CHIR99021 (CHIR)-induced activation of Wnt/ß-catenin signaling promotes self-renewal in mouse ESCs (mESCs). ß-catenin functions individually or cooperates with transcription factors to activate stemness factors such as c-Myc, Esrrb, Pou5f1, and Nanog. However the relationship between the core pluripotent factor, Kruppel-like factor 4 (also known as GKLF or EZF) and Wnt/ß-catenin signaling, remains ambiguous in J1 mESCs. DNA microarray analysis revealed that CHIR-treatment promoted pluripotency-maintaining transcription factors and repressed germ layer specification markers. CHIR also promoted genes related to the development of extracellular regions and the plasma membrane to maintain pluripotency of J1 mESCs. Among the CHIR-regulated genes, Klf4 has not been reported previously. We identified a novel cis element in the Klf4 gene that was activated by ß-catenin in J1 mESCs. We determined that ß-catenin interacted with this cis element, identifying Klf4 as a ß-catenin target gene in this context. Moreover, several microRNAs that targeted the 3'-UTR of Klf4 mRNA were identified, with miR-7a being down-regulated by CHIR in a ß-catenin-independent manner in J1 mESCs. These data collectively suggest that CHIR enhances Klf4 expression by repressing miR-7a expression or canonical Wnt pathway activation.

Maintenance of Self-Renewal and Pluripotency in J1 Mouse Embryonic Stem Cells through Regulating Transcription Factor and MicroRNA Expression Induced by PD0325901.

Embryonic stem cells (ESCs) have the ability to grow indefinitely and retain their pluripotency in culture, and this self-renewal capacity is governed by several crucial molecular pathways controlled by specific regulatory genes and epigenetic modifications. It is reported that multiple epigenetic regulators, such as miRNA and pluripotency factors, can be tightly integrated into molecular pathways and cooperate to maintain self-renewal of ESCs. However, mouse ESCs in serum-containing medium seem to be heterogeneous due to the self-activating differentiation signal of MEK/ERK. Thus, to seek for the crucial miRNA and key regulatory genes that establish ESC properties in MEK/ERK pathway, we performed microarray analysis and small RNA deep-sequencing of J1 mESCs treated with or without PD0325901 (PD), a well-known inhibitor of MEK/ERK signal pathway, followed by verification of western blot analysis and quantitative real-time PCR verification; we found that PD regulated the transcript expressions related to self-renewal and differentiation and antagonized the action of retinoic acid- (RA-) induced differentiation. Moreover, PD can significantly modulate the expressions of multiple miRNAs that have crucial functions in ESC development. Overall, our results demonstrate that PD could enhance ESC self-renewal capacity both by key regulatory genes and ES cell-specific miRNA, which in turn influences ESC self-renewal and cellular differentiation.