Cytological Observation and RNA-Seq Analyses Reveal miR9564 and Its Target Associated with Pollen Sterility in Autotetraploid Rice.

Understanding the regulation of autotetraploid sterility is essential for harnessing the strong advantages in genomic buffer capacity, biodiversity, and heterosis of autotetraploid rice. miRNAs play crucial roles in fertility regulation, yet information about their reproductive roles and target genes in tetraploid rice remains limited. Here, we used three tetraploid lines, H1 (fertile), HF (fertile), and LF (sterile), to investigate cytological features and identify factors associated with autotetraploid sterility. LF showed abnormal meiosis, resulting in low pollen fertility and viability, ultimately leading to scarce fertilization and a low-seed setting compared to H1 and HF. RNA-seq revealed 30 miRNA-candidate target pairs related to autotetraploid pollen sterility. These pairs showed opposite expression patterns, with differential expression between fertile lines (H1 and HF) and the sterile line (LF). qRT-PCR confirmed that miR9564, miR528, and miR27874 were highly expressed in the anthers of H1 and HF but not in LF, while opposite results were obtained in their targets (ARPS, M2T, and OsRPC53). Haplotype and expression pattern analyses revealed that ARPS was specifically expressed in lines with the same haplotype of MIR9564 (the precursor of miR9564) as LF. Furthermore, the Dual-GFP assay verified that miR9564 inhibited the fluorescence signal of ARPS-GFP. The over-expression of ARPS significantly decreased the seed setting rate (59.10%) and pollen fertility (50.44%) of neo-tetraploid rice, suggesting that ARPS plays important roles in autotetraploid pollen sterility. This study provides insights into the cytological characteristic and miRNA expression profiles of tetraploid lines with different fertility, shedding light on the role of miRNAs in polyploid rice.

Seed development-related genes contribute to high yield heterosis in integrated utilization of elite autotetraploid and neo-tetraploid rice.

Introduction: Autotetraploid rice holds high resistance to abiotic stress and substantial promise for yield increase, but it could not be commercially used because of low fertility. Thus, our team developed neo-tetraploid rice with high fertility and hybrid vigor when crossed with indica autotetraploid rice. Despite these advances, the molecular mechanisms underlying this heterosis remain poorly understood. Methods: An elite indica autotetraploid rice line (HD11) was used to cross with neo-tetraploid rice, and 34 hybrids were obtained to evaluate agronomic traits related to yield. WE-CLSM, RNA-seq, and CRISPR/Cas9 were employed to observe endosperm structure and identify candidate genes from two represent hybrids. Results and discussion: These hybrids showed high seed setting and an approximately 55% increase in 1000-grain weight, some of which achieved grain yields comparable to those of the diploid rice variety. The endosperm observations indicated that the starch grains in the hybrids were more compact than those in paternal lines. A total of 119 seed heterosis related genes (SHRGs) with different expressions were identified, which might contribute to high 1000-grain weight heterosis in neo-tetraploid hybrids. Among them, 12 genes had been found to regulate grain weight formation, including OsFl3, ONAC023, OsNAC024, ONAC025, ONAC026, RAG2, FLO4, FLO11, OsISA1, OsNF-YB1, NF-YC12, and OsYUC9. Haplotype analyses of these 12 genes revealed the various effects on grain weight among different haplotypes. The hybrids could polymerize more dominant haplotypes of above grain weight regulators than any homozygous cultivar. Moreover, two SHRGs (OsFl3 and SHRG2) mutants displayed a significant reduction in 1000-grain weight and an increase in grain chalkiness, indicating that OsFl3 and SHRG2 positively regulate grain weight. Our research has identified a valuable indica autotetraploid germplasm for generating strong yield heterosis in combination with neo-tetraploid lines and gaining molecular insights into the regulatory processes of heterosis in tetraploid rice.

Dechlorination of wastewater from shell-based glucosamine processing by mangrove wetland-derived fungi.

Wastewater from processing crustacean shell features ultrahigh chloride content. Bioremediation of the wastewater is challenging due to the high chloride ion content, making it inhospitable for most microorganisms to survive and growth. In this study, mangrove wetland-derived fungi were first tested for their salt tolerance, and the highly tolerant isolates were cultured in shrimp processing wastewater and the chloride concentration was monitored. Notably, the filamentous fungal species Aspergillus piperis could remove over 70% of the chloride in the wastewater within 3 days, with the fastest biomass increase (2.01 times heavier) and chloride removal occurring between day one and two. The chloride ions were sequestered into the fungal cells. The genome of this fungal species contained Cl- conversion enzymes, which may have contributed to the ion removal. The fungal strain was found to be of low virulence in larval models and could serve as a starting point for further considerations in bioremediation of shell processing wastewater, promoting the development of green technology in the shell processing industry.

Comparative cytological and transcriptome analyses of ny2 mutant delayed degeneration of tapetal cells and promotes abnormal microspore development in neo-tetraploid rice.

We aimed to investigate the genetic defects related to pollen development and infertility in NY2, a novel tetraploid rice germplasm known as Neo-tetraploid rice. This rice variety was created through the crossbreeding and selective breeding of various autotetraploid rice lines and has previously shown high fertility. Our previous research has revealed that the NY2 gene, encoding a eukaryotic translation initiation factor 3 subunit E, regulates pollen fertility. However, the underlying mechanism behind this fertility is yet to be understood. To shed light on this matter, we performed a combined cytological and transcriptome analysis of the NY2 gene. Cytological analysis indicated that ny2 underwent abnormal tapetal cells, microspore, and middle layer development, which led to pollen abortion and ultimately to male sterility. Genetic analysis revealed that the F1 plants showed normal fertility and an obvious advantage for seed setting compared to ny2. Global gene expression analysis in ny2 revealed a total of 7545 genes were detected at the meiosis stage, and 3925 and 3620 displayed upregulation and downregulation, respectively. The genes were significantly enriched for the gene ontology (GO) term "carbohydrate metabolic process. Moreover, 9 genes related to tapetum or pollen fertility showed down-regulation, such as OsABCG26 (ATP Binding Cassette G26), TMS9-1 (Thermosensitive Male Sterility), EAT1 (Programmed cell death regulatory), KIN14M (Kinesin Motor), OsMT1a (Metallothionein), and OsSTRL2 (Atypical strictosidine synthase), which were validated by qRT-PCR. Further analyses of DEGs identified nine down-regulated transcription factor genes related to pollen development. NY2 is an important regulator of the development of tapetum and microspore. The regulatory gene network described in this study may offer important understandings into the molecular processes that underlie fertility control in tetraploid rice.

Confirmation of the Absence of Somogyi Effect in Patients with Type 2 Diabetes by Retrospective Continuous Glucose Monitoring Systems.

Background: The Somogyi effect is defined as fasting hyperglycemia secondary to nocturnal hypoglycemia. In past decades, this effect proved to be rare or absent. However, many endocrinologists still believe in this phenomenon in clinical practice. Does the Somogyi effect truly exist? We aimed to answer this question with a study based on a larger sample size. Methods: We collected retrospective CGMs data from 2,600 patients with type 2 diabetes with stable treatment of insulin. Nocturnal hypoglycemia was defined as a CGMs sensor glucose of less than 3.9 mmol/L for at least 15 min between 24:00 and 06:00. Morning fasting glucose was compared between people with nocturnal hypoglycemia and without nocturnal hypoglycemia. Results: Valid CGMs data were obtained on 4,705 of 5,200 nights. Morning fasting glucose was observed lower after nights with nocturnal hypoglycemia compared with nights without hypoglycemia (P < 0.001). 84 cases presented fasting glucose of more than 7 mmol/L after nocturnal glucose of less than 3.9 mmol/L. Only 27 cases presented fasting glucose of more than 7 mmol/L after nocturnal glucose of less than 3.0 mmol/L. Fasting glucose values below 3.9 mmol/l in the morning were associated with a 100% risk of nocturnal hypoglycemia, while fasting glucose values over 9.6 mmol/l in the morning were associated with no risk of nocturnal hypoglycemia. Correlation analysis showed that the nocturnal glucose nadir was significantly correlated with fasting glucose levels (r = 0.613, P < 0.001). Conclusions: Our data provided no support for the existence of the Somogyi effect. If fasting glucose exceeds 9.6 mmol/L, we do not have to worry about asymptomatic nocturnal hypoglycemia in patients with type 2 diabetes.

Case report of the first cured patient with Mycobacterium Chimaera infection following cardiac valve replacement in the mainland of China.

BACKGROUND: Mycobacterium chimaera infections subsequent to cardiac surgery are related to contaminated heater-cooler devices, with high mortality. Nevertheless, few studies have been reported in Asia. CASE PRESENTATION: We described the case of a 55-year-old man with Mycobacterium chimaera infection following cardiac surgery in the mainland of China. He was diagnosed with endocarditis caused by Mycobacterium chimaera subsequent to open heart surgery. Metagenomic next-generation sequencing (mNGS) and 16S rRNA gene PCR analysis were used to identify potential pathogens. The patient underwent redo valve replacement surgery and received combination therapy with azithromycin, ethambutol, linezolid, and amikacin. No signs of relapse were observed during the 11-month follow-up visit. CONCLUSIONS: This is the first documented case of Mycobacterium chimaera infection following cardiac surgery in the mainland of China and the first documented transnational imported case worldwide. Moreover, mNGS is a novel diagnostic technology that can guide antimicrobial therapy prior to obtaining fluid/tissue culture results for Mycobacterium chimaera, providing a new approach for the detection of potential Mycobacterium chimaera infection.

Designing Organic Semiconductors with Ultrasmall Reorganization Energies: Insights from Molecular Symmetry, Aromaticity and Energy Gap.

An intuitive design strategy for organic semiconductors with ultrasmall reorganization energy (λ) is proposed. Learning from a total of 98 molecules condensed by benzene and/or thiophene rings, we find that linear compounds in D2h symmetry have the smallest λ in each of the three molecular categories (PAHs, thienothiophenes, benzothiophenes). 2D expanded analogues that contain these D2h building blocks also give unusually small λ (<100 meV). λ of 1D elongated polycyclics show an approximate linear correlation with the ring-averaged HOMA indices and the HOMO-LUMO gaps. Compared to the symmetry principle, the HOMA and energy gap, though much less intuitive to design a priori, provide additional quantitative guidelines to further optimize λ through substitutions, for example, when molecules have the same symmetries. Our results indicate that ring-fused π-conjugates that have narrower HOMO-LUMO gaps and are less aromatic are better candidates to achieve ultrasmall λ.

Mechanism of epithelial­mesenchymal transition inhibited by miR­203 in non­small cell lung cancer.

The aim of the present study was to investigate whether miR­203 can inhibit transforming growth factor­ß (TGF­ß)­induced epithelial­mesenchymal transition (EMT), and the migration and invasion ability of non­small cell lung cancer (NSCLC) cells by targeting SMAD3. In the present study, the expression levels of miR­203, SMAD3 mRNA and protein in NSCLC tissues were examined, as well as their corresponding paracancerous samples. The miR­203 mimics and miR­203 inhibitor were transfected into the H226 cell line. RT­qPCR was used to assess the expression levels of E­cadherin, Snail, N­cadherin and vimentin mRNA, and western blotting was performed to detect the expression levels of p­SMAD2, SMAD2, p­SMAD3, SMAD3 and SMAD4. The cell migration and invasion abilities were detected by Transwell assays. The target site of SMAD3 was predicted by the combined action between miR­203 and dual luciferase. The results revealed that the RNA levels of miR­203, compared with paracancerous tissues, were decreased in NSCLC tissues, while SMAD3 mRNA and protein levels were upregulated, and miR­203 inhibited SMAD3 expression. Induction of TGF­ß led to decreased E­cadherin mRNA levels, upregulation of Snail, N­cadherin and vimentin mRNA levels (P<0.05), and significant increase in cell migration and invasion, whereas transfection of miR­203 mimics reversed the aforementioned results (P<0.05). Conversely, miR­203 inhibitor could further aggravate the aforementioned results (P<0.05). Western blot results revealed that transfection of miR­203 mimics significantly reduced the protein expression of SMAD3 and p­SMAD3 (P<0.05). Furthermore, the results of the Dual­Luciferase assay revealed that miR­203 inhibited SMAD3 expression by interacting with specific regions of its 3'­UTR. Overall, a novel mechanism is revealed, in which, miR­203 can inhibit SMAD3 by interacting with specific regions of the 3'­UTR of SMAD3, thereby restraining TGF­ß­induced EMT progression and migration and invasion of NSCLC cells.

Efficient travelling-mode quantum key agreement against participant's attacks.

Quantum key agreement (QKA) is to negotiate a final key among several participants fairly and securely. In this paper, we show that some existing travelling-mode multiparty QKA protocols are vulnerable to internal participant's attacks. Dishonest participants can exploit a favorable geographical location or collude with other participants to predetermine the final keys without being discovered. To resist such attacks, we propose a new travelling-mode multiparty QKA protocol based on non-orthogonal Bell states. Theoretical analysis shows that the proposed protocol is secure against both external and internal attacks, and can achieve higher efficiency compared with existing travelling-mode multiparty QKA protocols. Finally we design an optical platform for each participant, and show that our proposed protocol is feasible with current technologies.

The influence of external electric fields on charge reorganization energy in organic semiconductors.

Charge reorganization energies (λ) of inter-ring carbon-carbon (IRCC) bond connected conjugated polycyclics are shown to exhibit an electric-field-driven anisotropic character. An external electric field parallel to the IRCC linker(s) reduces λ while the field vertical to the molecular plane does the opposite. This anisotropic character can be modulated by introducing inter-ring noncovalent locks into appropriate sites.

MiR-125b-1-3p Exerts Antitumor Functions in Lung Carcinoma Cells by Targeting S1PR1.

BACKGROUND: MicroRNAs (miRNAs) have been extensively studied over the decades and have been identified as potential molecular targets for cancer therapy. To date, many miRNAs have been found participating in the tumorigenesis of non-small cell lung cancer (NSCLC). The present study was designed to evaluate the functions of miR-125b-1-3p in NSCLC cells. METHODS: MiR-125b-1-3p expression was detected in tissue samples from 21 NSCLC patients and in NSCLC cell lines using the real-time polymerase chain reaction. A549 cell lines were transfected with a miR-125b-1-3p mimic or miR-125b-1-3p antisense. Cell counting kit-8, wound healing, Matrigel invasion assays, and flow cytometry were used to assess the effects of these transfections on cell growth, migration, invasion, and apoptosis, respectively. Western blotting was used to detect apoptosis-related proteins, expression of S1PR1, and the phosphorylation status of STAT3. Significant differences between groups were estimated using Student's t-test or a one-way analysis of variance. RESULTS: MiR-125b-1-3p was downregulated in NSCLC samples and cell lines. Overexpression of miR-125b-1-3p inhibited NSCLC cell proliferation (37.8 ± 9.1%, t = 3.191, P = 0.013), migration (42.3 ± 6.7%, t = 6.321, P = 0.003), and invasion (57.6 ± 11.3%, t = 4.112, P = 0.001) and simultaneously induced more NSCLC cell apoptosis (2.76 ± 0.78 folds, t = 3.772, P = 0.001). MiR-125b-1-3p antisense resulted in completely opposite results. S1PR1 was found as the target gene of miR-125b-1-3p. Overexpression of miR-125b-1-3p inhibited S1PR1 protein expression (27.4 ± 6.1% of control, t = 4.083, P = 0.007). In addition, S1PR1 siRNA decreased STAT3 phosphorylation (16.4 ± 0.14% of control, t = 3.023, P = 0.015), as in cells overexpressing miR-125b-1-3p (16.7 ± 0.17% of control, t = 4.162, P = 0.026). CONCLUSION: Our results suggest that miR-125b-1-3p exerts antitumor functions in NSCLC cells by targeting S1PR1.

MiR-760 enhances TRAIL sensitivity in non-small cell lung cancer via targeting the protein FOXA1.

Non-small cell lung cancer (NSCLC) is one of the leading cause of death worldwide. TNF-related apoptosis-inducing ligand (TRAIL) is a promising anti-tumor agent with the ability to kill tumor cells while spare normal ones. MicroRNAs (miRNAs) are small, non-coding RNAs that play vital roles in carcinogenesis. Although miR-760 has been reported to be dysregulated in a variety of cancers, the role of miR-760 in NSCLC is not fully understood, and the relationship between miR-760 dysregulation and TRAIL sensitivity is still elusive. In the current study, we found that miR-760 is significantly downregulated in NSCLC tissues and cell lines. We also found that ectopic expression of miR-760, by targeting the FOXA1, enhanced TRAIL sensitivity in NSCLC cells. Correspondingly, silencing of FOXA1 also sensitized NSCLC cell to TRAIL-induced apoptosis and proliferation inhibition. In summary, these findings suggest that miR-760 should be considered as a tumor suppressor since it negatively regulates the oncogene protein FOXA1 and regulated TRAIL sensitivity in NSCLC cells.

MicroRNA profiling during rat ventricular maturation: A role for miR-29a in regulating cardiomyocyte cell cycle re-entry.

Recent studies demonstrated that the mammalian heart possesses some capacity to proliferate. We observed cardiomyocyte proliferation within 4 weeks of age (P4W) in rats. We found 95 microRNAs that are differentially expressed in P4W cardiomyocytes. MicroRNA-29a was among the most highly up-regulated microRNAs in P4W cardiomyocytes. Overexpression of microRNA-29a suppressed the proliferation of H9c2 cell line. MicroRNA-29a inhibition induced cardiomyocytes to proliferate, accelerated the G1/S and G2/M transition, and up-regulated the cell cycle gene expression. Cyclin D2 (CCND2) was identified as a direct target of microRNA-29a. These findings indicate that microRNA-29a is involved in cardiomyocyte proliferation during postnatal development.

MicroRNA-193 pro-proliferation effects for bone mesenchymal stem cells after low-level laser irradiation treatment through inhibitor of growth family, member 5.

The enhanced proliferation of mesenchymal stem cells (MSCs) can be helpful for the clinical translation of cell therapy. Low-level laser irradiation (LLLI) has been demonstrated as regulating MSC proliferation. MicroRNAs (miRNAs) are involved in various pathophysiologic processes in stem cells, but the role of miRNAs in the LLLI-based promotion of MSC proliferation remains unclear. We found that the proliferation level and cell cycle-associated genes in MSCs were increased after LLLI treatment in a time-dependent manner. Microarray assays revealed subsets of miRNAs to be differentially regulated, and these dynamic changes were confirmed by quantitative real-time polymerase chain reaction (qRT-PCR) after LLLI. miR-193 was the most highly up-regulated miRNA, and the change in it was related with the proliferation level. Gain-loss function experiments demonstrated that miR-193 could regulate the proliferation of MSCs, including human's and rat's, but could not affect the apoptosis and differentiation level. Blockade of miR-193 repressed the MSC proliferation induced by LLLI. By qRT-PCR, we found that miR-193, in particular, regulated cyclin-dependent kinase 2 (CDK2) expression. Bioinformatic analyses and luciferase reporter assays revealed that inhibitor of growth family, member 5 (ING5) could be the best target of miR-193 to functionally regulate proliferation and CDK2 activity, and the mRNA and protein level of ING5 was regulated by miR-193. Furthermore, the ING5 inhibited by small interfering RNA (siRNA) could up-regulate the proliferation of MSCs and the expression of CDK2. Taken together, these results strongly suggest that miR-193 plays a critical part in MSC proliferation in response to LLLI stimulation, which is potentially amenable to therapeutic manipulation for clinical application.

MicroRNA-24 regulates cardiac fibrosis after myocardial infarction.

Cardiac fibrosis after myocardial infarction (MI) has been identified as a key factor in the development of heart failure. Although dysregulation of microRNA (miRNA) is involved in various pathophysiological processes in the heart, the role of miRNA in fibrosis regulation after MI is not clear. Previously we observed the correlation between fibrosis and the miR-24 expression in hypertrophic hearts, herein we assessed how miR-24 regulates fibrosis after MI. Using qRT-PCR, we showed that miR-24 was down-regulated in the MI heart; the change in miR-24 expression was closely related to extracellular matrix (ECM) remodelling. In vivo, miR-24 could improve heart function and attenuate fibrosis in the infarct border zone of the heart two weeks after MI through intramyocardial injection of Lentiviruses. Moreover, in vitro experiments suggested that up-regulation of miR-24 by synthetic miR-24 precursors could reduce fibrosis and also decrease the differentiation and migration of cardiac fibroblasts (CFs). TGF-ß (a pathological mediator of fibrotic disease) increased miR-24 expression, overexpression of miR-24 reduced TGF-ß secretion and Smad2/3 phosphorylation in CFs. By performing microarray analyses and bioinformatics analyses, we found furin to be a potential target for miR-24 in fibrosis (furin is a protease which controls latent TGF-ß activation processing). Finally, we demonstrated that protein and mRNA levels of furin were regulated by miR-24 in CFs. These findings suggest that miR-24 has a critical role in CF function and cardiac fibrosis after MI through a furin-TGF-ß pathway. Thus, miR-24 may be used as a target for treatment of MI and other fibrotic heart diseases.

Central zone of myocardial infarction: a neglected target area for heart cell therapy.

The purpose of this study was to investigate the fate of transplanted cells in the central zone of myocardial infarction (MI), and to clarify the relationship between the injection-site impact and the efficacy of cell therapy. MI was created by coronary ligation in female rats. Three weeks later, 3-million labelled male bone marrow mesenchymal stem cells (BMSCs) were directly injected into the border (BZC group) or central zone (CZC group) of MI area. As a control, culture medium was injected into the same sites. Cell survival was evaluated by quantitative real-time polymerase chain reaction, and apoptosis was assayed with TUNEL and caspase-3 staining. Four weeks after transplantation, heart function and cardiac morphometry were evaluated by echocardiography and Masson's Trichrome staining, respectively. Angiogenesis and myogenesis were detected by immunofluorescence staining. After cell transplantation into the border or central zone, there was no cell migration between the different zones of MI. BMSCs in the CZC group exhibited no difference in apoptotic percentage, in the long-term survival, when compared with those in the BZC group. However, they did effectively promote angiogenesis and cellular myogenic differentiation. Although cell delivery in the central zone of MI had no effect on the recovery of heart function compared with the BZC group, the retained BMSCs could still increase the scar thickness, and subsequently exhibit a trend in the reverse remodelling of ventricular dilation. Hence, we concluded that the central zone of MI should not be ignored during cell-based therapy. Multiple site injection (border+central zone) is strongly recommended during the procedure of cell transplantation.