Mogroside V alleviates inflammation response by modulating miR-21-5P/SPRY1 axis.

Mogroside V (MV) is a natural sweetener extracted from the edible plant Siraitia grosvenorii that possesses anti-inflammatory bioactivity. It has been reported that microRNAs (miRNAs) play an important role in the inflammation response suppression by natural agents. However, whether the anti-inflammation effect of mogroside V is related to miRNAs and the underlying mechanism remains unclear. Our study aimed to identify the key miRNAs important for the anti-inflammation effect of MV and reveal its underlying mechanisms. Our results showed that MV effectively alleviated lung inflammation in ovalbumin-induced (OVA-induced) asthmatic mice. miRNA-seq and mRNA-seq combined analysis identified miR-21-5p as an important miRNA for the inflammation inhibition effect of MV and it predicted SPRY1 to be a target gene of miR-21-5p. We found that MV significantly inhibited the production of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), interleukin-2 (IL-2), interleukin-6 (IL-6), and nitric oxide (NO), as well as the protein expression of p-P65/P65, cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) in OVA-induced asthmatic mice and LPS-treated RAW 264.7 cells. Moreover, the release of ROS increased in LPS-stimulated RAW 264.7 cells but was mitigated by MV pretreatment. In the meantime, the expression of miR-21-5p was decreased by MV, leading to an increase in the expression of SPRY1 in RAW 264.7 cells. Furthermore, miR-21-5p overexpression or SPRY1 knockdown reversed MV's protective effect on inflammatory responses. Conversely, miR-21-5p inhibition or SPRY1 overexpression enhanced MV's effect on inflammatory responses in LPS-exposed RAW 264.7 cells. Therefore, the significant protective effect of mogroside V on inflammation response is related to the downregulation of miR-21-5p and upregulation of SPRY1 in vitro and in vivo, MiR-21-5p/SPRY1 may be novel therapeutic targets of MV for anti-inflammation treatment.

ANP32A Knockdown Attenuates the Malignant Biological Behavior of Colorectal Cancer Cells by Suppressing Epithelial-mesenchymal Transition and ERK Activation.

Acidic leucine rich nuclear phosphoprotein-32A (ANP32A) protein has a variety of functions, such as regulating cell differentiation, influencing cell apoptosis and cell cycle progression. Our previous study demonstrated that high expression of ANP32A was found in the tumor tissues of colorectal cancer (CRC) patients and was positively associated with tumor grading. However, the function and underlying mechanisms of ANP32A in CRC metastasis have not been fully explored. In this study, we found that ANP32A knockdown significantly attenuated the migration and invasion, and epithelial-mesenchymal transition (EMT) in cells. Further mechanistic studies revealed that ANP32A knockdown inhibited the expression of ß-catenin and phosphorylated-ERK. The immunofluorescent staining experiment has revealed that ANP32A was expressed in the cell membrane, cytosol and nucleus, and its expression was positively associated with ß-catenin expression levels. Moreover, the ability of cell migration and invasion was inhibited, the expression of E-cadherin was enhanced following ANP32A knockdown, and these affects were abolished by an ERK activator PMA, enhanced by an ERK inhibitor PD98059. Moreover, our animal experiment also demonstrated that silenced ANP32A inhibited CRC cell growth, multi-organ metastasis, ERK activation and EMT progression in vivo. Collectively, these findings demonstrated that ANP32A promotes CRC progression and that may be a promising target for the anti-metastasis treatment of CRC.

RNA-binding domain 2 of nucleolin is important for the autophagy induction of curcumol in nasopharyngeal carcinoma cells.

BACKGROUND & AIMS: Excessive autophagy induces cell death and is regarded as the treatment of cancer therapy. We have confirmed that the anti-cancer mechanism of curcumol is related to autophagy induction. As the main target protein of curcumol, RNA binding protein nucleolin (NCL) interacted with many tumor promoters accelerating tumor progression. However, the role of NCL in cancer autophagy and in curcumol's anti-tumor effects haven't elucidated. The purpose of the study is to identify the role of NCL in nasopharyngeal carcinoma autophagy and reveal the immanent mechanisms of NCL played in cell autophagy. METHODS & RESULTS: In the current study, we have found that NCL was markedly upregulated in nasopharyngeal carcinoma (NPC) cells. NCL overexpression effectively attenuated the level of autophagy in NPC cells, and NCL silence or curcumol treatment obviously aggravated the autophagy of NPC cells. Moreover, the attenuation of NCL by curcumol lead a significant suppression on PI3K/AKT/mTOR signaling pathway in NPC cells. Mechanistically, NCL was found to be directly interact with AKT and accelerate AKT phosphorylation, which caused the activation of the PI3K/AKT/mTOR pathway. Meanwhile, the RNA Binding Domain (RBD) 2 of NCL interacts with Akt, which was also influenced by curcumol. Notably, the RBDs of NCL delivered AKT expression was related with cell autophagy in the NPC. CONCLUSION: The results demonstrated that NCL regulated cell autophagy was related with interaction of NCL and Akt in NPC cells. The expression of NCL play an important role in autophagy induction and further found that was associated with its effect on NCL RNA-binding domain 2. This study may provide a new perspective on the target protein studies for natural medicines and confirm the effect of curcumol not only regulating the expression of its target protein, but also influencing the function domain of its target protein.

Instrumental and transcriptome analysis reveals the chemotherapeutic effects of doxorubicin-loaded black phosphate nanosheets on abiraterone-resistant prostate cancer.

Prostate cancer is the second most common cause of cancer-related deaths in men and is common in most developed countries. Androgen deprivation therapy (ADT) that uses abiraterone acetate (AA) is an effective second-line treatment for prostate cancer. However, approximately 20-40% of patients develop primary resistance to abiraterone post-treatment. In this study, we aimed to understand the molecular mechanisms underlying the development of abiraterone resistance in prostate cancer cells and the potential use of black phosphorus nanosheets (BPNS) for treating abiraterone-resistant prostate cancer. We first established abiraterone-resistant prostate cancer PC-3 cells and found that these cells have higher migration ability than normal prostate cancer cells. Using comparative transcriptomic and bioinformatics analyses between abiraterone-sensitive PC-3 and abiraterone-resistant PC-3 cells, we highlighted the differentially expressed genes (DEGs) involved in the biological processes related to prostate gland morphogenesis, drug response, immune response, angiogenesis. We further studied the therapeutic effects of BPNS. Our results show that BPNS reduced the proliferation and migration of abiraterone-resistant PC-3 cells. Bioinformatics analysis, including gene ontology, Kyoto encyclopedia of genes and genomes enrichment analysis, and ingenuity pathway analysis (IPA) of the DEGs, suggested that BPNS treatment controlled cancer cell proliferation, metastasis, and oncogenic signaling pathways. Furthermore, the IPA gene network highlighted the involvement of the MMP family, ATF, and notch families in the anti-prostate cancer function of BPNS. Our findings suggest that BPNS may have a chemotherapeutic function in treating abiraterone-resistant prostate cancer.

Correction: Curcumol inhibits breast cancer growth via NCL/ERα36 and the PI3K/AKT pathway.

Correction for 'Curcumol inhibits breast cancer growth via NCL/ERα36 and the PI3K/AKT pathway' by Zhou Lu Wei et al., Food Funct., 2023, https://doi.org/10.1039/d2fo02387c.

Osthole Induces Apoptosis and Caspase-3/GSDME-Dependent Pyroptosis via NQO1-Mediated ROS Generation in HeLa Cells.

Osthole is a natural coumarin which has been proved to inhibit growth of cancer cells by inducing cell death, while its mechanism was considered to be just caused by apoptosis. In our study, we found that osthole activated not just apoptosis, but also pyroptosis which is a form of regulated cell death accompanied by loss of cell membrane integrity and lactate dehydrogenase (LDH) release. Caspase-3 is a key protein of apoptosis as well as pyroptosis. The apoptosis and pyroptosis induced by osthole were all inhibited by irreversible caspase-3 inhibitor Z-DEVD-FMK. Meanwhile, knockdown of gasdermin E (GSDME) only reduced the osthole-induced pyroptosis but did not affect the occurrence of apoptosis. Our proteomic analysis revealed that the expression of NAD(P)H: quinone oxidoreductase 1 (NQO1) was decreased in osthole-treated cells. Moreover, NQO1 inhibition by osthole induced the overproduction of reactive oxygen species (ROS), as well as apoptosis and pyroptosis. ROS inhibitor N-Acetyl-L-cysteine (NAC) not only reduced osthole-induced apoptosis but also reversed its effect on the pyroptosis. Meanwhile, knockdown of NQO1 by si-NQO1 or its inhibitor dicoumarol (DIC) not only enhanced ROS generation but also strengthened the GSDME-mediated pyroptosis. Finally, we demonstrated that osthole inhibited tumor growth and the expression of NQO1 in a HeLa xenograft mode. Similar to the results in vitro, osthole stimulated the activation of caspase-3, PARP, and GSDME in vivo. Taken together, all these data suggested that osthole induced apoptosis and caspase-3/GSDME-mediated pyroptosis via NQO1-mediated ROS accumulation.

Curcumol inhibits EBV-positive Nasopharyngeal carcinoma migration and invasion by targeting nucleolin.

Metastasis is a major cause of recurrence and death in patients with EBV-positive Nasopharyngeal carcinoma (NPC). Previous reports documented that curcumol has both anti-cancer and anti-viral effects, but there is little literature systematically addressing the mechanism of curcumol in EBV-positive tumors. Previously we found that nucelolin (NCL) is a target protein of curcumol in CNE2 cells, an EBV-negative NPC, and in this experiment, we reported a critical role for NCL in promoting migration and invasion of C666-1 cells, an EBV-positive NPC, and found that the expression of NCL determined the level of curcumol's efficacy. Mechanistically, NCL interacted with Epstein-Barr Virus Nuclear Antigen 1 (EBNA1) to activate VEGFA/VEGFR1/PI3K/AKT signaling pathway, which in turn promoted NPC cell invasion and metastasis. Moreover, further study showed that the differential expression of NCL and curcumol intervention only had a regulatory effect on the nuclear accumulation of VEGFR1, which strengthened the anti-cancer effect of curcumol mediated through NCL. Our findings indicated that curcumol exerted anti EBV-positive NPC invasion and metastasis by downregulating EBNA1 and inhibiting VEGFA/VEGFR1/PI3K/AKT signaling by targeting NCL, which provides a novel pharmacological basis for curcumol's clinical use in treating patients with EBV-positive NPC.

Curcumol inhibits the viability and invasion of colorectal cancer cells via miR-30a-5p and Hippo signaling pathway.

MicroRNA-30a-5p (miR-30a-5p), which functions as a tumor suppressor, has been reported to be downregulated in colorectal cancer (CRC) tissues and to be associated with cancer invasion. However, the detailed regulatory mechanism of curcumol in the malignant progression of CRC remains unknown. MTT, Transwell, scratch, western blotting and reverse transcription-quantitative PCR assays were performed to examine how curcumol inhibited CRC cell viability, invasion and migration, and to detect the role of miR-30a-5p and curcumol in the invasion and Hippo signaling pathways of CRC cells. The present study revealed that miR-30a-5p expression was downregulated in human CRC tissues and cells. The results demonstrated that miR-30a-5p downregulation was accompanied by the inactivation of the Hippo signaling pathway, which was demonstrated to promote CRC cell viability, invasion and migration. Curcumol treatment was identified to increase miR-30a-5p expression and to activate the Hippo signaling pathway, which in turn inhibited the invasion and migration of CRC cells. Overexpression of miR-30a-5p enhanced the effects of curcumol on cell invasion and migration, and the Hippo signaling pathway in CRC cells. Furthermore, downregulation of miR-30a-5p reversed the effects of curcumol on cell invasion and migration, and the Hippo signaling pathway in CRC cells. These findings identified novel signaling pathways associated with miR-30a-5p and revealed the effects of curcumol on miR-30a-5p expression. Therefore, curcumol may serve as a potential therapeutic strategy to delay CRC progression.

Inhibition of human carboxylesterases by magnolol: Kinetic analyses and mechanism.

Magnolol, the most abundant bioactive constituent of the Chinese herb Magnolia officinalis, has been found with multiple biological activities, including anti-oxidative, anti-inflammatory and enzyme-regulatory activities. In this study, the inhibitory effects and inhibition mechanism of magnolol on human carboxylesterases (hCEs), the key enzymes responsible for the hydrolytic metabolism of a variety of endogenous esters as well as ester-bearing drugs, have been well-investigated. The results demonstrate that magnolol strongly inhibits hCE1-mediated hydrolysis of various substrates, whereas the inhibition of hCE2 by magnolol is substrate-dependent, ranging from strong to moderate. Inhibition of intracellular hCE1 and hCE2 by magnolol was also investigated in living HepG2 cells, and the results showed that magnolol could strongly inhibit intracellular hCE1, while the inhibition of intracellular hCE2 was weak. Inhibition kinetic analyses and docking simulations revealed that magnolol inhibited both hCE1 and hCE2 in a mixed manner, which could be partially attributed to its binding at two distinct ligand-binding sites in each carboxylesterase, including the catalytic cavity and the regulatory domain. In addition, the potential risk of the metabolic interactions of magnolol via hCE1 inhibition was predicted on the basis of a series of available pharmacokinetic data and the inhibition constants. All these findings are very helpful in deciphering the metabolic interactions between magnolol and hCEs, and also very useful for avoiding deleterious interactions via inhibition of hCEs.

An Optimized Two-Photon Fluorescent Probe for Biological Sensing and Imaging of Catechol-O-Methyltransferase.

A practical two-photon fluorescent probe was developed for highly sensitive and selective sensing of the activities of catechol-O-methyltransferase (COMT) in complex biological samples. To this end, a series of 3-substituted 7,8-dihydroxycoumarins were designed and synthesized. Among them, 3-BTD displayed the best combination of selectivity, sensitivity, reactivity, and fluorescence response following COMT-catalyzed 8-O-methylation. The newly developed two-photon fluorescent probe 3-BTD can be used for determining the activities of COMT in complex biological samples and bio-imaging of endogenous COMT in living cells and tissue slices with good cell permeability, low cytotoxicity, and high imaging resolution. All these findings suggest that 3-BTD holds great promise for developing therapeutic molecules that target COMT, as well as for exploring COMT-associated biological processes and its biological functions in living systems. Furthermore, the strategy also sheds new light on the development of fluorescent probes for other conjugative enzymes.

Assessment of the inhibitory effects of pyrethroids against human carboxylesterases.

Pyrethroids are broad-spectrum insecticides that widely used in many countries, while humans may be exposed to these toxins by drinking or eating pesticide-contaminated foods. This study aimed to investigate the inhibitory effects of six commonly used pyrethroids against two major human carboxylesterases (CES) including CES1 and CES2. Three optical probe substrates for CES1 (DME, BMBT and DMCB) and a fluorescent probe substrate for CES2 (DDAB) were used to characterize the inhibitory effects of these pyrethroids. The results demonstrated that most of the tested pyrethroids showed moderate to weak inhibitory effects against both CES1 and CES2, but deltamethrin displayed strong inhibition towards CES1. The IC50 values of deltamethrin against CES1-mediated BMBT, DME, and DMCB hydrolysis were determined as 1.58µM, 2.39µM, and 3.3µM, respectively. Moreover, deltamethrin was cell membrane permeable and capable of inhibition endogenous CES1 in living cells. Further investigation revealed that deltamethrin inhibited CES1-mediated BMBT hydrolysis via competitive manner but noncompetitively inhibited DME or DMCB hydrolysis. The inhibition behaviors of deltamethrin against CES1 were also studied by molecular docking simulation. The results demonstrated that CES1 had at least two different ligand-binding sites, one was the DME site and another was the BMBT site which was identical to the binding site of deltamethrin. In summary, deltamethrin was a strong reversible inhibitor against CES1 and it could tightly bind on CES1 at the same ligand-binding site as BMBT. These findings are helpful for the deep understanding of the interactions between xenobiotics and CES1.

Optimized Exon-Exon Junction Library and its Application on Rodents' Brain Transcriptome Analysis

ABSTRACT Background: Alternative splicing (AS), which plays an important role in gene expression and functional regulation, has been analyzed on genome-scale by various bioinformatic approaches based on RNA-seq data. Compared with the huge number of studies on mouse, the AS researches approaching the rat, whose genome is intermedia between mouse and human, were still limited. To enrich the knowledge on AS events in rodents' brain, we perfomed a comprehensive analysis on four transcriptome libraries (mouse cerebrum, mouse cerebellum, rat cerebrum, and rat cerebellum), recruiting high-throughput sequencing technology. An optimized exon-exon junction library approach was introduced to adapt the longer RNA-seq reads and to improve mapping efficiency. Results: In total, 7,106 mouse genes and 2,734 rat genes were differentially expressed between cerebrum and cerebellum, while 7,125 mouse genes and 1,795 rat genes exhibited varieties on transcript variant level. Only half of the differentially expressed exon-exon junctions could be reflected at gene expression level. Functional cluster analysis showed that 32 pathways in mouse and 9 pathways in rat were significantly enriched, and 6 of them were in both. Interestingly, some differentially expressed transcript variants did not show difference on gene expression level, such as PLCβ1 and Kcnma1. Conclusion: Our work provided a case study of a novel exon-exon junction strategy to analyze the expression of genes and isoforms, helping us understand which transcript contributes to the overall expression and further functional change.

[The relationship between catechol O-methyltransferase and diseases].

Catechol O-methyltransferase (COMT), one of the endogenous phase II metabolizing enzymes, expressed by chromosome 22. COMT catalyzes the transfer of a methyl group from common methyl donor S-adenosyl-L-methionine(Ado Met or SAM) to one of the catechol hydroxyls. COMT participates in the metabolism of many catechols in vivo, e.g. dopamine, epinephrine, noradrenaline, estradiol. Furthermore COMT also plays important roles in the metabolism of xenobiotic catechols from food and drug. COMT play a critical role in the management of catechols. Metabolism disorders of COMT can cause many diseases or an increased risk of diseases, e.g. Pakinson diseases, schizophrenia, and breast cancer. In this review, we explains the relationship of COMT and related-diseases through expounding disease caused by the COMT metabolic disorders. Finally, we hope that there will be more effective treatments for the COMT metabolism related diseases.

A Highly Selective Ratiometric Two-Photon Fluorescent Probe for Human Cytochrome P450 1A.

Cytochrome P450 1A (CYP1A), one of the most important phase I drug-metabolizing enzymes in humans, plays a crucial role in the metabolic activation of procarcinogenic compounds to their ultimate carcinogens. Herein, we reported the development of a ratiometric two-photon fluorescent probe NCMN that allowed for selective and sensitive detection of CYP1A for the first time. The probe was designed on the basis of substrate preference of CYP1A and its high capacity for O-dealkylation, while 1,8-naphthalimide was selected as fluorophore because of its two-photon absorption properties. To achieve a highly selective probe for CYP1A, a series of 1,8-naphthalimide derivatives were synthesized and used to explore the potential structure-selectivity relationship, by using a panel of human CYP isoforms for selectivity screening. After screening and optimization, NCMN displayed the best combination of selectivity, sensitivity and ratiometric fluorescence response following CYP1A-catalyzed O-demetylation. Furthermore, the probe can be used to real-time monitor the enzyme activity of CYP1A in complex biological systems, and it has the potential for rapid screening of CYP1A modulators using tissue preparation as enzyme sources. NCMN has also been successfully used for two-photon imaging of intracellular CYP1A in living cells and tissues, and showed high ratiometric imaging resolution and deep-tissue imaging depth. In summary, a two-photon excited ratiometric fluorescent probe NCMN has been developed and well-characterized for sensitive and selective detection of CYP1A, which holds great promise for bioimaging of endogenous CYP1A in living cells and for further investigation on CYP1A associated biological functions in complex biological systems.

rs17501976 polymorphism of CLDN1 gene is associated with decreased risk of colorectal cancer in a Chinese population.

The purpose of this study was to determine the relationship between polymorphisms in Claudin-1 (CLDN1) and the risk of colorectal cancer in a Chinese population. In this study, a case-control study was conducted in which polymorphisms in CLDN1 were analyzed in 50 patients with colorectal cancer (CRC) and 50 healthy individuals as controls. No rs16865344 and rs17429833 polymorphism were found among all analyzed samples. For the rs17501976 polymorphism, the TC genotype (OR = 0. 41, 95% CI = 0.18-0.91, and P = 0.045) was closely associated with the risk of colorectal cancer compared with the more common TT genotype. And the TC + CC genotypes (OR = 0.41, 95% CI = 0.18-0.91, and P = 0.045) were also significantly associated with the risk of CRC compared with the TT genotype. However, a C > T change of the rs17501976 polymorphism did not show a difference in transcription factor binding to the promoter region of CLDN1. For rs12696600 polymorphism, no significant difference was found in colorectal cancer risk between cases and controls in corresponding genotypes. Collectively, our data suggest that rs17501976 polymorphism significantly associated with a decreased susceptibility to CRC in a Chinese population.

Association of a variant in MIR 196A2 with susceptibility to hepatocellular carcinoma in male Chinese patients with chronic hepatitis B virus infection.

MicroRNAs (miRNAs) are small noncoding RNAs with regulatory functions as tumor suppressors and oncogenes. Recent studies have implicated that the rs11614913 SNP in MIR196A2 was associated with susceptibility of lung cancer, congenital heart disease, breast cancer and shortened survival time of nonsmall cell lung cancer. To assess whether this polymorphism is associated with susceptibility to and clinicopathologic characteristics of hepatitis B virus (HBV)-related hepatocellular carcinoma (HCC), a total of 560 patients with chronic HBV infection and 391 healthy volunteers were enrolled, and MIR196A2 polymorphism was genotyped by polymerase chain reaction-ligation detection reaction (PCR-LDR). In our study group, there was no significant association between MIR196A2 polymorphism and the risk of HBV-related HCC in all subjects, however, the risk of HCC was significantly higher with MIR196A2 rs11614913 CC genotype or C allele compared with those with the TT genotype or T allele in male patients. Furthermore, in a subsequent analysis of the association between this polymorphism and clinicopathologic characteristics, there was still no significant difference in both the distribution of genotype or allelic frequency. However, we observed that the T allele was significantly more frequent in male HCC patients with lymphatic metastasis. Our results suggested that MIR196A2 polymorphism was associated with susceptibility to HBV-related HCC in a male Chinese population.

CTLA-4 +49A>G polymorphism is associated with the risk but not with the progression of colorectal cancer in Chinese.

PURPOSE: Colorectal cancer (CRC) is one of the most common malignancies in the world and a multipathway disease. Cytotoxic T-lymphocyte antigen-4 (CTLA-4) is a potent immunoregulatory molecule that suppresses antitumor response by down-regulating T-cell activation. The most studied +49A>G polymorphism of CTLA-4 gene has been associated with several autoimmune or cancer diseases. Our aim was to investigate the association between this genetic variant and the risk as well as progression of colorectal cancer in Chinese. METHODS: We conducted a case-control study of 124 colorectal cancer cases and 407 healthy controls. DNA was extracted from blood specimens, and +49A>G polymorphism in the CTLA-4 gene was genotyped by polymerase chain reaction-ligation detection reaction (PCR-LDR). RESULTS: In our study group, the frequency of AG or GG or carrying at least one G allele at position +49 was significantly different in colorectal cancer patients and the control group, indicating that the risk of CRC was significantly higher among subjects with the AG or GG genotype or carrying at least one G allele at position +49 than among the subjects with the AA genotype. However, we observed no association between CTLA-4 +49A>G polymorphism and the progression of CRC. Interestingly, the CTLA-4 +49A allele was in non-significantly higher numbers in CRC patients with distant metastasis. CONCLUSIONS: Our results suggested that CTLA-4 +49A>G polymorphism was associated with an increased risk of colorectal cancer, but this polymorphism did not play an important role in the progression of CRC in Chinese.