Metabolomics and transcriptomics reveal the toxic mechanism of Cd and nano TiO2 coexposure on rice (Oryza sativa L.).

Engineered nanoparticles (ENPs) can resist heavy metal toxicity in plants, but their coexposure still exhibits toxicity to plants compared to plants without exposure to ENPs and heavy metals. There have been few studies on the toxic mechanism of nano TiO2-heavy metal coexposure and the effect mechanism of nano TiO2 in plants. Thus, transcriptomics and metabolomics were used to study the toxic mechanism of rutile nano TiO2 or TiO2-Cd (rutile nano TiO2 and CdCl2 mixture) on rice (Oryza sativa L.). After 40 days of exposure, the plant height and root dry weight of rice were significantly decreased in the nano TiO2-Cd group compared to the blank group (nano TiO2 and CdCl2 free). After Cd treatment, 423 differentially expressed genes (DEGs) and 16 differential metabolites were identified. Nano TiO2 exposure induced significant regulation of 299 DEGs and 6 metabolites. After nano TiO2-Cd coexposure, 1660 DEGs and 181 differential metabolites were identified. Notably, the EDGs (e.g., chalcone isomerase and hydroxycinnamoyl transferase) and differential metabolites (e.g., chrysin and galangin) demonstrated the disruption of flavonoid biosynthesis in Cd-treated rice. After rice was exposed to nano TiO2, the DEGs were related to ribosome, whereas the differential metabolites were associated with pyruvate metabolism and valine, leucine, and isoleucine biosynthesis. Furthermore, 14 DEGs (e.g., asparaginyl-tRNA synthetase and methionyl-tRNA formyltransferase) involved in aminoacyl-tRNA biosynthetic pathways were significantly upregulated in rice treated with nano TiO2-Cd, in line with the changes in related metabolites (e.g., L-asparagine and 10-formyltetrahydrofolate). Our results show that it is necessary to pay close attention to the toxicity of nano TiO2-Cd coexposure in paddy ecosystems and use ENPs with caution to combat the phytotoxicity of heavy metals.

Glucose transporter GLUT1 expression is important for oriental river prawn (Macrobrachium nipponense) hemocyte adaptation to hypoxic conditions.

Crustaceans have an open vascular system in which hemocytes freely circulate in hemolymph. Hemocytes are rich in hemocyanin, a specific oxygen-transport protein in crustaceans; therefore, understanding the response of hemocytes to hypoxia is crucial. Although hemocytes take up glucose during hypoxia, the molecular mechanism of glucose uptake in crustaceans remains unclear. Herein, we identified two highly conserved glucose transporters (GLUT1 and GLUT2) in Macrobrachium nipponense (oriental river prawn) and analyzed their tissue-specific expression patterns. Our immunofluorescence assays showed that GLUT1 and GLUT2 are located on the cell membrane, with a strong GLUT1 signal in primary hemocytes under hypoxia. We found that during acute hypoxia, hypoxia-inducible factor-1α-related metabolic alterations result in decreased mitochondrial cytochrome c oxidase activity, implying a classic glycolytic mechanism. As a proof of concept, we replicated these findings in insect S2 cells. Acute hypoxia significantly induced hypoxia-inducible factor-1α, GLUT1, and pyruvate dehydrogenase kinase isozyme 1 expression in primary hemocytes, and hypoxia-induced increases in glucose uptake and lactate secretion were observed. GLUT1 knockdown induced intracellular reactive oxygen species generation and apoptosis in vitro and in vivo, resulting in increased prawn mortality and more apoptotic cells in their brains, implying a vital function of GLUT1 in hypoxia adaptation. Taken together, our results suggest a close relationship between hypoxia-mediated glycolysis and GLUT1 in hemocytes. These results demonstrated that in crustaceans, adaptation to hypoxia involves glucose metabolic plasticity.

The evolutionarily conserved hif-1/bnip3 pathway promotes mitophagy and mitochondrial fission in crustacean testes during hypoxia.

The hypoxia-inducible factor 1 (HIF-1) cascade is an ancient and strongly evolutionarily conserved signaling pathway that is involved in the hypoxic responses of most metazoans. Despite immense advances in the understanding of the HIF-1-mediated regulation of hypoxic responses in mammals, the contribution of the hif-1 cascade in the hypoxic adaptation of nonmodel invertebrates remains unclear. In this study, we used the oriental river prawn Macrobrachium nipponense for investigating the roles of hif-1-regulated mitophagy in crustacean testes under hypoxic conditions. We identified that the Bcl-2/adenovirus E1B 19-kDa interacting protein (bnip3) functions as a regulator of mitophagy in M. nipponense and demonstrated that hif-1α activates bnip3 by binding to the bnip3 promoter. Hif-1α knockdown suppressed the expression of multiple mitophagy-related genes, and prawns with hif-1α knockdown exhibited higher mortality under hypoxic conditions. We observed that the levels of BNIP3 were induced under hypoxic conditions and detected that bnip3 knockdown inhibited the mitochondrial translocation of dynamin-related protein 1 (drp1), which is associated with mitochondrial fission. Notably, bnip3 knockdown inhibited hypoxia-induced mitophagy and aggravated the deleterious effects of hypoxia-induced reactive oxygen species (ROS) production and apoptosis. The experimental studies demonstrated that hypoxia induced mitochondrial fission in M. nipponense via drp1. Altogether, the study elucidated the mechanism underlying hif-1/bnip3-mediated mitochondrial fission and mitophagy and demonstrated that this pathway protects crustaceans against ROS production and apoptosis induced by acute hypoxia.

Expert consensus on thermal ablation therapy of pulmonary subsolid nodules (2021 Edition).

The Expert Consensus reviews current literatures and provides clinical practice guidelines for thermal ablation of pulmonary subsolid nodules or ground-glass nodule (GGN). The main contents include the following: (1) clinical evaluation of GGN; (2) procedures, indications, contraindications, outcomes evaluation, and related complications of thermal ablation for GGN; and (3) future development directions.

A practical screening strategy for Lynch syndrome and Lynch syndrome mimics in colorectal cancer.

OBJECTIVES: The objective of the study is to provide an efficient and practical screening strategy to distinguish a broader spectrum of Lynch syndrome (LS) and LS mimics-associated colorectal cancer (CRC), including Lynch-like syndrome (LLS), constitutional mismatch repair-deficiency, familial CRC type X (FCCTX), and polymerase proofreading-associated polyposis syndrome. MATERIALS AND METHODS: 1294 cases of CRC samples were detected mismatch repair (MMR) status using immunohistochemistry (IHC) staining, in which the cases with MLH1-deficient CRC underwent BRAF mutation analysis by IHC. Following the personal and/or family history survey, next-generation sequencing (NGS) was used to detect gene variants. RESULTS: 1294 CRC patients were dichotomized into tumors caused by a deficient MMR (dMMR) system and a proficient MMR (pMMR) system after MMR status analysis. 45 patients with suspected sporadic dMMR CRC were then separated from MLH1-deficient CRC though BRAF mutation status analysis by IHC. Following the personal and/or family history survey for 1294 patients, as well as germline genetic testing by NGS, 34 patients were diagnosed as LS (8 cases), SLS (13 cases), LLS ( 6 cases), FCCTX (3 cases), and sporadic CRC (4 cases). CONCLUSIONS: Our screening strategy, which consists of clinical and molecular analyses, is expected to improve the screening efficiency and management for the LS and LS mimics.

[Expert Consensus for Thermal Ablation of Pulmonary Subsolid Nodules (2021 Edition)].

"The Expert Group on Tumor Ablation Therapy of Chinese Medical Doctor Association, The Tumor Ablation Committee of Chinese College of Interventionalists, The Society of Tumor Ablation Therapy of Chinese Anti-Cancer Association and The Ablation Expert Committee of the Chinese Society of Clinical Oncology" have organized multidisciplinary experts to formulate the consensus for thermal ablation of pulmonary subsolid nodules or ground-glass nodule (GGN). The expert consensus reviews current literatures and provides clinical practices for thermal ablation of GGN. The main contents include: (1) clinical evaluation of GGN, (2) procedures, indications, contraindications, outcomes evaluation and related complications of thermal ablation for GGN and (3) future development directions.
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Comparison of Multi-Gene Testing Data Between Fresh and Formalin-Fixed Specimens From Core Needle Biopsy in Patients With NSCLC.

Purpose: Currently, formalin-fixed paraffin-embedded (FFPE) tissue specimens are the conventional material for gene testing for non-small cell lung cancer (NSCLC) patients. In our study, we aimed to develop a quick gene testing procedure using fresh core needle biopsy samples from NSCLC patients. Methods: In total, 77 fresh NSCLC samples obtained from core needle biopsy were evaluated by frozen section examination. If the NSCLC diagnosis and adequate tumor cell counts were confirmed by histopathology, the fresh tissues were used to extract DNA and subsequent gene testing by ARMS-PCR. Meanwhile, the paired FFPE core needle biopsy samples from 30 NSCLC patients also underwent gene testing. Results: In total, 77 fresh samples showed an EGFR mutation rate of 61.0%, higher than the levels in the Asian. Following a comparison of gene testing results with fresh tissues and paired FFPE tissues from the 30 patients, no significant difference in the DNA concentration extracted from fresh tissues and FFPE tissues was found. However, DNA purity was significantly higher in fresh tissues than that in FFPE tissues. Gene testing detected the same gene mutations in 93.3% of cases in fresh tissues and paired FFPE tissues. The gene testing procedure using fresh biopsy samples greatly shortens the waiting time of patients. Conclusion: The multi-gene mutation testing using fresh core needle biopsy samples from NSCLC patients is a reasonable, achievable, and quick approach. Fresh tissues may serve as a potential alternative to FFPE tissues for gene testing in NSCLC patients.

Low nitrogen induces root elongation via auxin-induced acid growth and auxin-regulated target of rapamycin (TOR) pathway in maize.

Under low nitrogen (N) supply, an important adaption of the maize root system is to promote the root elongation so as to increase N uptake from a larger soil space. The underlying physiological mechanism is largely unknown. In the present study, two maize inbred lines (Ye478 and Wu312) were used to study the possible involvement of the auxin and target of rapamycin (TOR) pathway in low-N-induced root elongation. Compared to Wu312, primary root elongation of Ye478 was more sensitive to low nitrate supply. Correspondingly, more auxin was accumulated in the root tip, and more protons were secreted, increasing the acidity of the apoplast space. On the other hand, low-N-induced root elongation was greatly reduced when shoot-to-root auxin transport was inhibited by applying N-1-naphthylphthalamic acid (NPA) at the plant base or by pruning the top leaf where auxin is mostly synthesized. Furthermore, exogenous application of TOR inhibitor also eliminated the response of root elongation under low N. The content of TOR kinase and the expression of TOR pathway-related genes were significantly changed when shoot-to-root auxin transport was reduced by NPA treatment. Taken together, it is concluded that low-N stress increases shoot-to-root auxin transport which enhances root elongation via auxin-dependent acid growth and the auxin-regulated TOR pathway in maize.

The lncRNA XIST promotes the progression of breast cancer by sponging miR-125b-5p to modulate NLRC5.

X-inactivation-specific transcript (XIST) is a long noncoding RNA (lncRNA) that functions as an indicator of various human tumors, including those of breast cancer. This study was conducted to characterize a novel regulatory network involving XIST in breast cancer cells. The mRNAs of XIST, miR-125b-5p, and NOD-like receptor family CARD domain containing 5 (NLRC5) in breast cancer cells and tissues were analyzed using quantitative real-time polymerase chain reaction. Cell proliferation, apoptosis, migration, and invasion were separately detected via cell counting kit-8, flow cytometry, and Transwell assays. The relationships between XIST, miR-125b-5p, and NLRC5 were predicted and then confirmed using the dual-luciferase reporter assay. NLRC5 protein expression was quantitated using western blot assays. XIST was found to be overexpressed in breast cancer tissues and cells, which was accompanied by miR-125b-5p downregulation and NLRC5 upregulation. XIST knockdown significantly repressed cell proliferation, anti-apoptosis, migration, and invasion activities in breast cancer cells, and the loss of miR-125b-5p had a similar effect. XIST was shown to sponge miR-125b-5p, which in turn targeted NLRC5. NLRC5, a breast cancer promotor, is negatively regulated by miR-125b-5p. Moreover, the downregulation of NLRC5 induced by the loss of XIST was significantly reversed by miR-125b-5p knockdown. In conclusion, the lncRNA XIST promotes the malignancy of breast cancer cells partly by competitively binding to miR-125b-5p, which then led to increased NLRC5 expression. Our study suggests that targeting XIST may be a possible treatment for breast cancer.

The physiological mechanism underlying root elongation in response to nitrogen deficiency in crop plants.

MAIN CONCLUSION: In response to low nitrogen stress, multiple hormones together with nitric oxide signaling pathways work synergistically and antagonistically in crop root elongation. Changing root morphology allows plants to adapt to soil nutrient availability. Nitrogen is the most important essential nutrient for plant growth. An important adaptive strategy for crops responding to nitrogen deficiency is root elongation, thereby accessing increased soil space and nitrogen resources. Multiple signaling pathways are involved in this regulatory network, working together to fine-tune root elongation in response to soil nitrogen availability. Based on existing research, we propose a model to explain how different signaling pathways interact to regulate root elongation in response to low nitrogen stress. In response to a low shoot nitrogen status signal, auxin transport from the shoot to the root increases. High auxin levels in the root tip stimulate the production of nitric oxide, which promotes the synthesis of strigolactones to accelerate cell division. In this process, cytokinin, ethylene, and abscisic acid play an antagonistic role, while brassinosteroids and auxin play a synergistic role in regulating root elongation. Further study is required to identify the QTLs, genes, and favorable alleles which control the root elongation response to low nitrogen stress in crops.

Increased biomass accumulation in maize grown in mixed nitrogen supply is mediated by auxin synthesis.

The use of mixed nitrate and ammonium as a nitrogen source can improve plant growth. Here, we used metabolomics and transcriptomics to study the underlying mechanisms. Maize plants were grown hydroponically in the presence of three forms of nitrogen (nitrate alone, 75%/25% nitrate/ammonium, and ammonium alone). Plants grown with mixed nitrogen had a higher photosynthetic rate than those supplied only with nitrate, and had the highest leaf area and shoot and root biomass among the three nitrogen treatments. In shoot and root, the concentration of nitrogenous compounds (ammonium, glutamine, and asparagine) and carbohydrates (sucrose, glucose, and fructose) in plants with a mixed nitrogen supply was higher than that with nitrate supply, but lower than that with ammonium supply. The activity of the related enzymes (glutamate synthase, asparagine synthase, phosphoenolpyruvate carboxylase, invertase, and ADP-glucose pyrophosphorylase) changed accordingly. Specifically, the mixed nitrogen source enhanced auxin synthesis via the shikimic acid pathway, as indicated by the higher levels of phosphoenolpyruvate and tryptophan compared with the other two treatments. The expression of corresponding genes involving auxin synthesis and response was up-regulated. Supply of only ammonium resulted in high levels of glutamine and asparagine, starch, and trehalose hexaphosphate. We conclude that, in addition to increased photosynthesis, mixed nitrogen supply enhances leaf growth via increasing auxin synthesis to build a large sink for carbon and nitrogen utilization, which, in turn, facilitates further carbon assimilation and nitrogen uptake.

miR-221/222 promote tumor growth and suppress apoptosis by targeting lncRNA GAS5 in breast cancer.

MicroRNAs (miRNAs) are 21-23-nucleotide, short, non-coding RNAs that play important roles in virtually all biological pathways in mammals and other multicellular organisms. The association of miR-221 and miR-222 (miR-221/222) for breast cancer is critical, but their detailed roles in its development and progression remain unclear. In the present study, we found that miR-221/222 were consistently up-regulated in breast cancer tissues. We then investigated the molecular mechanisms by which miR-221/222 contributed to breast cancer and identified growth arrest-specific transcript 5 (GAS5) as a direct target gene of miR-221/222. In contrast with the up-regulated expression levels of miR-221/222, GAS5 levels were significantly down-regulated and negatively correlated with miR-221/222 in breast cancer tissues. In addition, we showed that miR-221/222 inhibitors increased cellular apoptosis, miR-221/222 mimics decreased the cell apoptosis in breast cancer cells, and restoration of GAS5 expression attenuated the anti-apoptotic effects of miR-221/222 in breast cancer cells, indicating that GAS5 was a direct mediator of miR-221/222 function. Finally, we showed that miR-221/222 suppressed GAS5 expression significantly and enhanced tumor growth in a mouse model of breast cancer xenografts. The present study highlighted the important role of miR-221/222 as oncomiRs in breast cancer, which inhibited GAS5 translation. These findings may provide a new perspective for the molecular mechanism of breast carcinogenesis and provide a novel approach to the treatment of breast cancer.

LBH589 Inhibits Glioblastoma Growth and Angiogenesis Through Suppression of HIF-1α Expression.

Glioblastoma (GBM) is an angiogenic malignancy with a highly unfavorable prognosis. Angiogenesis in GBM represents an adaptation to a hypoxic microenvironment and is correlated with tumor growth, invasion, clinical recurrence, and lethality. LBH589 (also called panobinostat) is a histone deacetylase (HDAC) inhibitor with potent antitumor activity. In the current study, we investigated the mechanism and effects of LBH589 on GBM growth and hypoxia-induced angiogenesis in vitro and in vivo. To determine the antitumor and angiogenesis activity and mechanism of LBH589, we used cell proliferations in vitro and GBM xenografts in vivo. To clarify mechanisms of LBH589 on angiogenesis, HDAC assay, RT-PCR, Western blot, and co-immunoprecipitation assays were performed. We found LBH589 displayed significant antitumor effects on GBM as demonstrated by inhibited cell proliferation, slower tumor growth, and decreased microvessel density of subcutaneous xenografts. These actions of LBH589 resulted from the disruption of heat shock protein 90/HDAC6 complex, increased HIF-1α instability and degradation, and decreased VEGF expression. Our results indicate the potential antiangiogenic activity of LBH589 in human GBM and provide some preclinical data to warrant further exploration of HDAC inhibitors for the treatment of advanced glioma. Moreover, our study supports the role of HDAC inhibitors as a therapeutic strategy to target tumor angiogenesis.

Valproic acid ameliorates olfactory dysfunction in APP/PS1 transgenic mice of Alzheimer's disease: Ameliorations from the olfactory epithelium to the olfactory bulb.

Olfactory dysfunction is a common and early symptom of many neurodegenerative diseases, particularly of Alzheimer's disease (AD) and mild cognitive impairment, pointing to the progression to dementia. Recent studies have revealed that valproic acid (VPA) has neuroprotective effects in rodent models of AD. In this study, we investigated the effects of VPA on olfactory dysfunction of APP/PS1 double transgenic mouse models of AD. After continuous treatment with a 100mg/kg daily dose of VPA for 3 months, APP/PS1 mice showed improved olfactory performances. In agreement with the behavioral findings, VPA treatment reduced amyloid ß (Aß) burden in the olfactory epithelium (OE) of transgenic mice. And, VPA increased epithelial thickness of the olfactory mucosa through decreased cell apoptosis and increased cell proliferation. In the olfactory bulb (OB), VPA administration also reduced senile plaques and levels of soluble and insoluble Aß42 peptides. Besides, VPA promoted the increase of mitral cells and decrease of neurofilament immunostaining. In hence, VPA treatment completely improved the olfactory performances and prevented degenerative changes of the OE and OB. Our study raises the possibility of AD diagnosis by OE biopsy. Moreover, VPA may provide a novel therapeutic strategy for the treatment of olfactory dysfunction in AD patients.

Overexpression of eIF4E in colorectal cancer patients is associated with liver metastasis.

PURPOSE: Liver metastasis is one of the leading causes of death in colorectal cancer (CRC) patients. The present study aimed to evaluate the value of eIF4E as a prognostic marker of colorectal liver metastasis (CLM) and identify the functional role of eIF4E in CRC metastasis. PATIENTS AND METHODS: The expression level of eIF4E in CRC tissues was analyzed by immunohistochemical staining and Western blot. Expression of eIF4E in CRC cell lines was evaluated by reverse transcription quantitative polymerase chain reaction (RT-qPCR) and Western blot. Cell Counting Kit-8 (CCK-8) and Transwell assays were performed to assess the effects of eIF4E on cell proliferation, migration, and invasion. Western blot was further used to investigate the mechanism of eIF4E in tumor metastasis. RESULTS: The upregulation frequency of eIF4E in the CLM group (82.5%) was higher than that in the non-CLM group (65.0%). Of the 80 patients recruited for the follow-up study, 23 were in the low eIF4E group (ratio of tumor to nontumor tissue

Can the signal-to-noise ratio of choline in magnetic resonance spectroscopy reflect the aggressiveness of endometrial cancer?

RATIONALE AND OBJECTIVES: To differentiate endometrial cancer (ECa) from benign lesions in endometrial or in submucosa (BLs-ESm), and investigate whether the signal-to-noise ratio (SNR) of choline-containing compounds (Cho) obtained from three-dimensional (1)H magnetic resonance spectroscopy (MRS) is associated with the aggressiveness of ECa. MATERIALS AND METHODS: Thirty-three patients with ECa and 15 patients with BLs-ESm underwent preoperative multivoxel (1)H MRS at 3 T MR. The amplitude of Cho peak of each voxel was recorded, and the corresponding SNR of Cho peak (ChoSNR) was calculated. The maximum ChoSNR (max ChoSNR) for each lesion was identified. The max ChoSNR of ECa and BLs-ESm, as well as type I ECa and type II ECa, was compared. The relationship between max ChoSNR and pathologic characteristics of tumors, including tumor grade, stage, type, and tumor size, was analyzed. RESULTS: The mean max ChoSNR (±standard deviation [SD]) was 30.93 ± 16.89 for ECa and 10.40 ± 3.07 for BLs-ESm (P < .001). The mean max ChoSNR for type II ECa (48.54 ± 21.46) was higher than that for type I ECa (26.19 ± 12.02, P = .001). There were no significant differences among different grades (P = .449). The Spearman coefficient between max ChoSNR and stage was 0.423 (P = .014); the difference existed only between Ia and III ECa (P = .048). The Pearson coefficient between ChoSNR and tumor size was 0.515 (P = .002). CONCLUSIONS: The max ChoSNR obtained from MRS can differentiate ECa from BLs and type I ECa and type II ECa, but cannot differentiate between each grade ECa and each International Federation of Gynecology and Obstetrics stage ECa. However, max ChoSNR increased with the increase in International Federation of Gynecology and Obstetrics stage and size of ECa.

Can magnetic resonance spectroscopy differentiate endometrial cancer?

OBJECTIVES: To investigate whether the choline-containing compounds (Cho) obtained from three-dimensional (1)H magnetic resonance (MR) spectroscopy can differentiate endometrial cancer (ECa) from benign lesions in endometria or in submucosa (BLs-ESm) and is associated with the aggressiveness of ECa. METHODS: Fifty-seven patients (ECa, 38; BLs-ESm, 19) underwent preoperative multi-voxel MR spectroscopy at 3.0 T. The ratio of the sum of the Cho peak integral to the sum of the unsuppressed water peak integral (Cho/water) and the coefficient of variation (CV) used to describe the variability of Cho/water in one lesion were calculated. RESULTS: Mean Cho/water (±standard deviation [SD]) was (3.02 ± 1.43) × 10(-3) for ECa and (1.68 ± 0.33) × 10(-3) for BLs-ESm (p < 0.001). Mean Cho/water was (4.42 ± 1.53) × 10(-3) for type II ECa and (2.65 ± 1.17) × 10(-3) for type I ECa (p = 0.001). There were no significant differences among different stages of ECa (p = 0.107) or different grades of ECa (p = 0.142). The Cho/water was positively correlated with tumour stage (r = 0.386, p = 0.017) and size (r = 0.333, p = 0.041). The CV was also positively correlated with tumour stage (r = 0.537, p = 0.001) and size (r = 0.34, p = 0.037). CONCLUSION: The Cho/water can differentiate ECa from BLs-ESm and differentiate type II from type I ECa, but cannot differentiate different stages of ECa or different grades of ECa. Cho/water increased with the increase of tumour stage and size. KEY POINTS: • First report to attempt to assess ECa aggressiveness with magnetic resonance spectroscopy (MRS). • MRS can differentiate type I from type II ECa. • MRS can differentiate ECa from BLs-ESm. • MRS cannot differentiate different stages of ECa or different grades of ECa. • Cho/water increased with the increase of tumour stage and size.