Mitochondrial perturbation in the intestine causes microbiota-dependent injury and gene signatures discriminative of inflammatory disease.

Mitochondrial dysfunction is associated with inflammatory bowel diseases (IBDs). To understand how microbial-metabolic circuits contribute to intestinal injury, we disrupt mitochondrial function in the epithelium by deleting the mitochondrial chaperone, heat shock protein 60 (Hsp60Δ/ΔIEC). This metabolic perturbation causes self-resolving tissue injury. Regeneration is disrupted in the absence of the aryl hydrocarbon receptor (Hsp60Δ/ΔIEC;AhR-/-) involved in intestinal homeostasis or inflammatory regulator interleukin (IL)-10 (Hsp60Δ/ΔIEC;Il10-/-), causing IBD-like pathology. Injury is absent in the distal colon of germ-free (GF) Hsp60Δ/ΔIEC mice, highlighting bacterial control of metabolic injury. Colonizing GF Hsp60Δ/ΔIEC mice with the synthetic community OMM12 reveals expansion of metabolically flexible Bacteroides, and B. caecimuris mono-colonization recapitulates the injury. Transcriptional profiling of the metabolically impaired epithelium reveals gene signatures involved in oxidative stress (Ido1, Nos2, Duox2). These signatures are observed in samples from Crohn's disease patients, distinguishing active from inactive inflammation. Thus, mitochondrial perturbation of the epithelium causes microbiota-dependent injury with discriminative inflammatory gene profiles relevant for IBD.

lncRNA PAARH impacts liver cancer cell proliferation by engaging miR­6512­3p to target LASP1.

Long non-coding (lnc)RNAs serve a pivotal role as regulatory factors in carcinogenesis. The present study aimed to assess the involvement of the lncRNA progression and angiogenesis-associated RNA in hepatocellular carcinoma (PAARH) in liver cancer, along with the associated underlying mechanism. Through the use of reverse transcription-quantitative (RT-q)PCR, differences in the expression levels of PAARH in HepG2, HEP3B2.1.7, HCCLM3, Huh-7 and MHCC97-H liver cancer cell lines and THLE-2 epithelial cell lines were evaluated. The liver cancer cell line with the greatest, significantly different, level of expression relative to the normal liver cell line was selected for subsequent experiments. Using ENCORI database, the putative target genes of the microRNA (miR) miR-6512-3p were predicted. Cells were then transfected with lentiviruses carrying short-hairpin-PAARH to interfere with PAARH expression. Subsequently, HepG2 liver cancer cells were transfected with a miR-6512-3p mimic and an inhibitor, and the expression levels of miR-6512-3p and the LIM and SH3 domain protein 1 (LASP1) in cells were assessed using RT-qPCR analysis. Cell proliferation was subsequently evaluated using colony formation assays, and immunofluorescence and western blotting were used to assess the expression level of LASP1 in transfected cells. The binding interaction between miR-6512-3p and LASP1 was further evaluated using a dual-luciferase reporter gene assay. Liver cancer cells were found to exhibit higher expression levels of PAARH compared with normal liver cells. Following PAARH interference, the expression level of miR-6512-3p was significantly increased, whereas that of LASP1 was significantly decreased, resulting in a reduction in cell proliferation. In liver cancer cells, miR-6512-3p overexpression led to a significant reduction in the LASP1 level and reduced proliferation, whereas suppressing miR-6512-3p led to a significant increase in LASP1 levels and increased proliferation. Additionally, the inhibition of miR-6512-3p caused the states of low LASP1 expression and reduced cell proliferation to be reversed. LASP1, a recently identified target gene of miR-6512-3p, was demonstrated to be suppressed by miR-6512-3p overexpression, thereby inhibiting liver cancer cell proliferation. Taken together, the findings of the present study demonstrate that the lncRNA PAARH may enhance liver cancer cell proliferation by engaging miR-6512-3p to target LASP1.

METTL3/16-mediated m6A modification of ZNNT1 promotes hepatocellular carcinoma progression by activating ZNNT1/osteopontin/S100A9 positive feedback loop-mediated crosstalk between macrophages and tumour cells.

Macrophages are the major components of tumour microenvironment, which play critical roles in tumour development. N6-methyladenosine (m6A) also contributes to tumour progression. However, the potential roles of m6A in modulating macrophages in hepatocellular carcinoma (HCC) are poorly understood. Here, we identified ZNNT1 as an HCC-related m6A modification target, which was upregulated and associated with poor prognosis of HCC. METTL3 and METTL16-mediated m6A modification contributed to ZNNT1 upregulation through stabilizing ZNNT1 transcript. ZNNT1 exerted oncogenic roles in HCC. Furthermore, ZNNT1 recruited and induced M2 polarization of macrophages via up-regulating osteopontin (OPN) expression and secretion. M2 Macrophages-recruited by ZNNT1-overexpressed HCC cells secreted S100A9, which further upregulated ZNNT1 expression in HCC cells via AGER/NF-κB signaling. Thus, this study demonstrates that m6A modification activated the ZNNT1/OPN/S100A9 positive feedback loop, which promoted macrophages recruitment and M2 polarization, and enhanced malignant features of HCC cells. m6A modification-triggered ZNNT1/OPN/S100A9 feedback loop represents potential therapeutic target for HCC.

N6 -methyladenosine-modified FAM111A-DT promotes hepatocellular carcinoma growth via epigenetically activating FAM111A.

As an epitranscriptomic modulation manner, N6 -methyladenosine (m6 A) modification plays important roles in various diseases, including hepatocellular carcinoma (HCC). m6 A modification affects the fate of RNAs. The potential contributions of m6 A to the functions of RNA still need further investigation. In this study, we identified long noncoding RNA FAM111A-DT as an m6 A-modified RNA and confirmed three m6 A sites on FAM111A-DT. The m6 A modification level of FAM111A-DT was increased in HCC tissues and cell lines, and increased m6 A level was correlated with poor survival of HCC patients. m6 A modification increased the stability of FAM111A-DT transcript, whose expression level showed similar clinical relevance to that of the m6 A level of FAM111A-DT. Functional assays found that only m6 A-modified FAM111A-DT promoted HCC cellular proliferation, DNA replication, and HCC tumor growth. Mutation of m6 A sites on FAM111A-DT abolished the roles of FAM111A-DT. Mechanistic investigations found that m6 A-modified FAM111A-DT bound to FAM111A promoter and also interacted with m6 A reader YTHDC1, which further bound and recruited histone demethylase KDM3B to FAM111A promoter, leading to the reduction of the repressive histone mark H3K9me2 and transcriptional activation of FAM111A. The expression of FAM111A was positively correlated with the m6 A level of FAM111A-DT, and the expression of methyltransferase complex, YTHDC1, and KDM3B in HCC tissues. Depletion of FAM111A largely attenuated the roles of m6 A-modified FAM111A-DT in HCC. In summary, the m6 A-modified FAM111A-DT/YTHDC1/KDM3B/FAM111A regulatory axis promoted HCC growth and represented a candidate therapeutic target for HCC.

LncRNA MEG3 Inhibits Tumor Progression by Modulating Macrophage Phenotypic Polarization via miR-145-5p/DAB2 Axis in Hepatocellular Carcinoma.

Background: Hepatocellular carcinoma (HCC) is the predominant histological type of primary liver cancer, which ranks sixth among the most common human tumors. Tumor-associated macrophages (TAMs) are an important component of tumor microenvironment (TME) and the M2 macrophage polarization substantially contributes to tumor growth and metastasis. Long non-coding RNA (lncRNA) MEG3 was reported to restrain HCC development. However, whether MEG3 regulates macrophage phenotypic polarization in HCC remains unclear. Methods: Bone marrow derived macrophages (BMDMs) were treated with LPS/IFNγ and IL4/IL13 to induce the M1 and M2 macrophage polarization, respectively. M2-polarized BMDMs were simultaneously transfected with adenovirus vector overexpressing MEG3 (Adv-MEG3). Subsequently, M2-polarized BMDMs were cultured for 24 h with serum-free medium, the supernatants of which were harvested as conditioned medium (CM). HCC cell line Huh7 was cultured with CM for 24 h. F4/80+CD68+ and F4/80+CD206+ cell percentages in M1-and M2-polarized BMDMs were calculated using flow cytometry. Huh7 cell migration, invasion and angiogenesis were determined via Transwell assay and tube formation experiment. Nude mice were implanted with Huh7 cells and Adv-MEG3-transfected M2-polarizd BMDMs, and tumor growth and M2 macrophage polarization markers were assessed. The binding between miR-145-5p and MEG3 or disabled-2 (DAB2) was verified by luciferase reporter assay. Results: MEG3 presented lower expression in HCC tissues than in normal controls, and low expression of MEG3 was correlated to poorer prognosis of HCC patients. MEG3 expression was enhanced during LPS/IFNγ-induced M1 polarization, but was reduced during IL4/IL13-induced M2 polarization. MEG3 overexpression inhibited the expression of M2 polarization markers in both M2-polarized BMDMs and mice. Mechanically, MEG3 bound with miR-145-5p to regulate DAB2 expression. Overexpressing MEG3 suppressed M2 polarization-induced HCC cell metastasis and angiogenesis by upregulating DAB2 and inhibited in vivo tumor growth. Conclusion: LncRNA MEG3 curbs HCC development by repressing M2 macrophage polarization via miR-145-5p/DAB2 axis.

E3 ubiquitin ligase HECTD3 is a tumor suppressor and mediates the polyubiquitination of SLC7A11 to promote ferroptosis in colon cancer.

Homologous to the E6-associated protein carboxyl terminus domain containing 3 (HECTD3) has been reported to play an essential role in biological processes, including drug resistance, metastasis or apoptosis. However, the relationships between HECTD3 and Colorectal cancer (CRC) remain to be unclear. In this study, we discovered that HECTD3 expressed lowly in CRC compared with normal tissues and patients with low HECTD3 suffered from poorer survival outcomes relative to those with high HECTD3 levels. HECTD3 inhibition could significantly enhance proliferative, clone abilities and self-renewal capacities of CRC cells in vitro and in vivo. Mechanistically, our findings revealed that HECTD3 had endogenous interactions with SLC7A11 proteins. HECTD3 promoted the polyubiquitination of SLC7A11 to trigger the degradation of SLC7A11 proteins. Targeting HECTD3 could notably prolong the half-life period of SLC7A11 proteins, thereby promoting its stability. However, the cysteine mutation at amino acid 823 (ubiquitinase active site) of HECTD3 impaired the polyubiquitination of SLC7A11. HECTD3 deficiency depended on accumulated SLC7A11 proteins to accelerate malignant progression of CRC in vitro and in vivo. Thus, HECTD3 could suppress SLC7A11 levels to attenuate the SLC7A11-mediated cystine uptake, leading to enhanced CRC ferroptosis. SLC7A11 inhibition through polyubiquitination by HECTD3 increased ferroptosis, thereby inhibiting CRC tumor growth. Taken together, these results showed that HECTD3 controlled the stability of SLC7A11 and uncovered the function of HECTD3/SLC7A11 axis in regulating CRC progression.

Toxicity Analysis of Mesoporous Polydopamine on Intestinal Tissue and Microflora.

As a promising therapy, photothermal therapy (PTT) converts near-infrared (NIR) light into heat through efficient photothermal agents (PTAs), causing a rapid increase in local temperature. Considering the importance of PTAs in the clinical application of PTT, the safety of PTAs should be carefully evaluated before their widespread use. As a promising PTA, mesoporous polydopamine (MPDA) was studied for its clinical applications for tumor photothermal therapy and drug delivery. Given the important role that intestinal microflora plays in health, the impacts of MPDA on the intestine and on intestinal microflora were systematically evaluated in this study. Through biological and animal experiments, it was found that MPDA exhibited excellent biocompatibility, in vitro and in vivo. Moreover, 16S rRNA analysis demonstrated that there was no obvious difference in the composition and classification of intestinal microflora between different drug delivery groups and the control group. The results provided new evidence that MPDA was safe to use in large doses via different drug delivery means, and this lays the foundation for further clinical applications.

RNA-Seq Based Toxicity Analysis of Mesoporous Polydopamine Nanoparticles in Mice Following Different Exposure Routes.

Mesoporous polydopamine nanoparticles (MPDA NPs) are promising nanomaterials that have the prospect of clinical application for multi-strategy antitumor therapy, while the biosecurity of MPDA NPs remains indistinct. Here, transcriptome sequencing (RNA-Seq) was performed to systematically reveal the toxicity of MPDA NPs to five categories of organs after three different exposure routes, including intravenous injection, intramuscular injection, and intragastric administration. Our results uncovered that MPDA NPs could be deposited in various organs in small amounts after intravenous administration, not for the other two exposure routes. The number of differentially expressed genes (DEGs) identified in the heart, liver, spleen, lung, and kidney from the intragastric administration group was from 22 to 519. Similarly, the corresponding number was from 23 to 64 for the intramuscular injection group and was from 11 to 153 for the intravenous injection group. Functional enrichment analyses showed 6, 39, and 4 GO terms enriched for DEGs in intragastric administration, intramuscular injection, and intravenous injection groups, respectively. One enriched pathway was revealed in intragastric administration group, while no enriched pathway was found in other groups. Our results indicated that MPDA NPs produced only slight changes at the transcriptome level in mice, which provided new insights for further clinical application of MPDA NPs.

Triggering Immune System With Nanomaterials for Cancer Immunotherapy.

Cancer is a major cause of incidence rate and mortality worldwide. In recent years, cancer immunotherapy has made great progress in the preclinical and clinical treatment of advanced malignant tumors. However, cancer patients will have transient cancer suppression reaction and serious immune related adverse reactions when receiving immunotherapy. In recent years, nanoparticle-based immunotherapy, which can accurately deliver immunogens, activate antigen presenting cells (APCs) and effector cells, provides a new insight to solve the above problems. In this review, we discuss the research progress of nanomaterials in immunotherapy including nanoparticle-based delivery systems, nanoparticle-based photothermal and photodynamic immunotherapy, nanovaccines, nanoparticle-based T cell cancer immunotherapy and nanoparticle-based bacteria cancer immunotherapy. We also put forward the current challenges and prospects of immunomodulatory therapy.

Long non-coding RNA PAARH promotes hepatocellular carcinoma progression and angiogenesis via upregulating HOTTIP and activating HIF-1α/VEGF signaling.

Hepatocellular carcinoma (HCC) is one of the leading lethal malignancies and a hypervascular tumor. Although some long non-coding RNAs (lncRNAs) have been revealed to be involved in HCC. The contributions of lncRNAs to HCC progression and angiogenesis are still largely unknown. In this study, we identified a HCC-related lncRNA, CMB9-22P13.1, which was highly expressed and correlated with advanced stage, vascular invasion, and poor survival in HCC. We named this lncRNA Progression and Angiogenesis Associated RNA in HCC (PAARH). Gain- and loss-of function assays revealed that PAARH facilitated HCC cellular growth, migration, and invasion, repressed HCC cellular apoptosis, and promoted HCC tumor growth and angiogenesis in vivo. PAARH functioned as a competing endogenous RNA to upregulate HOTTIP via sponging miR-6760-5p, miR-6512-3p, miR-1298-5p, miR-6720-5p, miR-4516, and miR-6782-5p. The expression of PAARH was significantly positively associated with HOTTIP in HCC tissues. Functional rescue assays verified that HOTTIP was a critical mediator of the roles of PAARH in modulating HCC cellular growth, apoptosis, migration, and invasion. Furthermore, PAARH was found to physically bind hypoxia inducible factor-1 subunit alpha (HIF-1α), facilitate the recruitment of HIF-1α to VEGF promoter, and activate VEGF expression under hypoxia, which was responsible for the roles of PAARH in promoting angiogenesis. The expression of PAARH was positively associated with VEGF expression and microvessel density in HCC tissues. In conclusion, these findings demonstrated that PAARH promoted HCC progression and angiogenesis via upregulating HOTTIP and activating HIF-1α/VEGF signaling. PAARH represents a potential prognostic biomarker and therapeutic target for HCC.