Glucose Medium Research Articles

Sodium-Glucose Cotransporter 2 (SGLT2) Inhibition Induces DNA Damage and Radiosensitization in Glioblastoma Cell Lines

Glioblastomas (GBM) are highly dependent on glucose for energy to promote cell proliferation and tumor growth. Preclinical studies have shown that sodium-glucose cotransporter 2 (SGLT2) inhibitors, drugs that are FDA approved for diabetes, reduce the viability and invasiveness of GBM cells. We hypothesize that SGLT2 inhibitors decrease GBM cell growth and viability by promoting DNA damage. The SGLT2 inhibitors canagliflozin, dapagliflozin and empagliflozin were used and cell studies were performed under low glucose (0.6 mM) media conditions. In the cell proliferation assays, cells were counted in triplicate after 24-, 48- and 72-hour treatment with SGLT2 inhibitors. Glucose transport assays were performed using a 6-fluorescent iodinated glucose analog (6FIGA) which is selectively transported by SGLTs and not by glucose transporters (GLUTs). Immunoblots were used to measure protein expression of SGLT2 and markers of DNA damage (phospho-ATM and gamma-H2AX). Cellular radiosensitization was evaluated by clonogenic survival assays. SGLT2 inhibition with canagliflozin, dapagliflozin and empagliflozin decreased cell proliferation in LN229, U118MG and LN18 GBM cells lines. In LN229 cells, 72-hour treatment with canagliflozin (30 µM) decreased cell proliferation by 60% compared to vehicle control cells (p<0.0001) whereas dapagliflozin (30 µM) and empagliflozin (30 µM) decreased cell proliferation compared to vehicle control cells by 17% (p<0.0001) and 19% (p<0.001), respectively. Glucose transport assays show that 6FIGA uptake is reduced by canagliflozin and dapagliflozin in LN229 and U87 GBM cells. Canagliflozin treatment (50 µM) for 24 hours in LN229 cells results in increased phosphorylated ATM protein and increased gamma-H2AX protein, markers of DNA damage, by immunoblot. SGLT2 protein expression was not affected by canagliflozin. Treatment with canagliflozin (50 µM) 48 hours prior to 2, 4 and 6 Gy of ionizing radiation reduced clonogenic survival of LN229, LN18 and U118MG GBM cells. SGLT2 inhibitors canagliflozin, dapagliflozin and empagliflozin decrease GBM cell proliferation, with the highest inhibitory effect seen with canagliflozin. Canagliflozin treatment alone increases DNA damage signaling in GBM cells. Canagliflozin radiosensitizes GBM cells. This work provides preliminary preclinical rationale for canagliflozin as a radiosensitizer in GBM.

Promoting the Anti-Tumor Activity of Radiotherapy on Lung Cancer through a Modified Ketogenic Diet and the AMPK Signaling Pathway

Taking advantage of the characteristics of high metabolic heterogeneity of tumor cells, the modified ketogenic diet (KD) combined with radiotherapy was used to investigate and analyze the radiosensitivity of a lung cancer model from the perspective of energy metabolism. Different concentrations of glucose and βhydroxybutyrate (βHB) were used at the cellular level to simulate the level of ketone bodies. A cell counting kit was used to detect the effect of different concentrations of glucose (2.78mM, 5.56mM, 12.5mM, and 25mM) and βHB (0mM, 5mM, and 10mM) combined with radiotherapy on the proliferation of LLC cells. Flow cytometry was used to detect tumor cell cycle and apoptosis. Immunofluorescence was used to detect the expression of γH2AX, a DNA damage marker, and western blot was used to detect the expression of AMPK and ρ-AMPK. At the animal level, C57BL/6J female mice were used to establish a transplanted tumor model of lung cancer, and fed with different fat ratio diets combined with radiotherapy. The volume, tumor size, blood glucose level, blood ketone level, survival time and safety of the mice were monitored and observed. The LLC cells were treated with different concentrations of glucose and βHB. The results showed that the survival rate of LLC cells decreased significantly with the increase of irradiation dose when the glucose concentration was 5.56mM and 2.78mM; However, the survival rate of cells in low glucose medium added with βHB was significantly lower than that of the control group, and the survival rate of LLC decreased significantly with the extension of culture time after irradiation (p < 0.001). After irradiation, LG (low glucose) group, LG+βHB 5mM group and LG+βHB 10mM group had a significantly higher proportion of G2 phase, and a significantly higher proportion of early and late phase than the control group. γH2AX foci were detected in LG group, LG +βHB 5mM group and LG +βHB 10mM group at 2h and 24h after radiotherapy, which were significantly higher than those in the control group (p < 0.05). The median survival time was 38 days in the PT group, 55 days in the PT+RT group, 41 days in the 45F group, and not reached in the 45F+RT group. HE staining showed no tumor metastasis and toxic side effects in liver and kidney. The expression of ρ-AMPK/AMPK in the combined treatment group was higher than that in the other groups. The expression of ρ-AMPK/AMPK in RT, 45F and combined treatment group was higher than that in PT group. The expression of ρ-AMPK/AMPK in RT group was higher than that in 45F group, and the difference was statistically significant (p < 0.01). Modified ketogenic diet can enhance the anti-tumor effect of radiotherapy in LLC tumor-bearing mice by reducing glucose and increasing the energy supply ratio from fat.

ВЛИЯНИЕ РАЗЛИЧНЫХ КОНЦЕНТРАЦИЙ ГЛЮКОЗЫ В ПИТАТЕЛЬНОЙ СРЕДЕ НА МОРФОГЕНЕЗ РАСТЕНИЙ ВИНОГРАДА IN VITRO

To study the organic nutrition of grape plants in in vitro culture, we conducted research on the effect of glucose introduced into the nutrient medium on the morphogenesis of test-tube grape plants. The experiments were carried out on two model varieties (Sibirkovyi and Krasnostop Karpi) selected by us from the laboratory collection of in vitro healthy grape plants. These varieties, in addition to differences in genotypes, most noticeably differ in the features of development in in vitro culture, which we noted earlier when they were cultivated and deposited in the collection. The Sibirkovyi variety is one of the fastest growing, and Krasnostop Karpi is one of the slowest developing varieties in in vitro culture. In the experiment, glucose concentrations from 0.0 (sucrose control 10.0 g/l) to 40.0 g/l were used. The varietal response of the model varieties in the experiment with different concentrations of glucose in the nutrient medium was different. The variety Sibirkovyi was already one and a half times ahead of the variety Krasnostop Karpi in development in the first terms of accounting. At the same time, for the Sibirkovyi variety, the optimal range of concentrations for the development and preservation of microplants according to most indicators was the range from 5 to 20 g/l, and for the Krasnostop Karpi variety, the optimal range was narrower – 0-20 g/l. At high glucose concentrations of 30,0-40,0 g/l, the plants of both varieties experienced stress, which was manifested in the intensive release of phenols into the nutrient medium (this was most clearly manifested in the Sibirkovyi variety), and often the plants did not reach the top of the cultivation vessel during ontogenesis. besides, they showed earlier aging processes. At the same time, at high concentrations of glucose, a slowdown in growth processes and its ripening of shoots in individual microplants was noted.

Mechanism of Action of Streptococcus downii, a New Bacterial Species with Probiotic Potential.

Streptococcus downii is a recently reported bacterial species of oral origin, with inhibitory capacity against Streptococcus mutans, Actinomyces naeslundii, Veillonella parvula and Aggregatibacter actinomycetemcomitans, which confers upon it the potential of being an oral probiotic. The aim of the present study was to identify the potential mechanisms by which S. downii exerts its inhibitory effect on S. mutans. To this end, the study assessed the consumption of glucose and proteins available in the culture medium, the modification of the pH, the production of short-chain fatty acids, the changes in the protein panel of the inhibition halo, the production of hydrogen peroxide and the effect of proteinase K. There were no differences in the glucose values or in the protein content of the medium, but there was a reduction in pH (with no effect on the growth of S. mutans). Significant increases were detected in the levels of lactic and formic acid (with no effect on the growth of S. mutans), as well as changes in the peptide panel (with no effect on the growth of S. mutans). The inhibitory effect was maintained in the presence of peroxidase but disappeared after adding proteinase K. Based on these results, it is suggested that the main mechanism of inhibition of S. downii against S. mutans is the production of bacteriocins.

Differential Gene Expression Analysis in Candida albicans: A Comparative Study Between Malic Acid and Glucose Media for Insights into Pathogenicity and Metabolic Pathway Interactions

Differential Gene Expression Analysis in Candida albicans: A Comparative Study Between Malic Acid and Glucose Media for Insights into Pathogenicity and Metabolic Pathway Interactions

Exosomal circHIPK3 derived from umbilical cord-derived mesenchymal stem cells enhances skin fibroblast autophagy by blocking miR-20b-5p/ULK1/Atg13 axis.

Umbilical cord-derived mesenchymal stem cells (UCMSCs) could alleviate diabetes-induced injury. Hence, this investigation aimed to explore the role and mechanism of UCMSCs-derived exosomal circHIPK3 (exo-circHIPK3) in diabetes mellitus (DM). HFF-1 cells were cultured in high glucose (HG) medium or normal medium, and treated with UCMSCs-derived exo-circHIPK3 or miR-20b-5p mimics or Unc-51-like autophagy activating kinase 1 (ULK1) overexpression vector. The surface markers of UCMSCs were analyzed using a flow cytometer. The differentiation potential of UCMSCs was evaluated using oil red O staining, alizarin red staining and alkaline phosphatase (ALP) staining. Cell proliferation was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The miRNA expressions were analyzed by reverse transcription-quantitative polymerase chain reaction (qRT-PCR). Protein levels were quantified by western blot. An immunofluorescence staining was used for observing LC3 expression. The interaction between miR-20b-5p and circHIPK3, and between miR-20b-5b and ULK1 were identified by a RNA immunoprecipitation (RIP) assay and a luciferase reporter assay. Up-regulation of circHIPK3 was found in UCMSCs-derived exosomes. Exo-circHIPK3 decreased the miR-20b-5p level while increasing the contents of ULK1 and autophagy-related gene 13 (Atg13) in HG-induced fibroblasts. In addition, exo-circHIPK3 activated HG-induced fibroblast autophagy and proliferation. Overexpressed miR-20b-5p promoted fibroblast injury by inhibiting cell autophagy via the ULK1/Atg13 axis in HG conditions of high glucose. Moreover, exo-circHIPK3 enhanced autophagy and cell viability in HG-induced fibroblasts through the miR-20b-5p/ULK1/Atg13 axis. UCMSCs-derived exosomal circHIPK3 promoted cell autophagy and proliferation and accelerated the fibroblast injury repair by the miR-20b-5p/ULK1/Atg13 axis.

Lachancea quebecensis a Novel Isolate for the Production of Craft Beer.

Yeasts are ubiquitously present in different natural sources. Some of these yeasts have interesting characteristics for the production of fermented food products. This study characterized Lachancea thermotolerans and L. quebecensis isolated from insects to determine their brewing potential. The yeasts were evaluated according to their fermentative potential in glucose and maltose-defined media and their resistance to ethanol and hop. Finally, craft beer was elaborated at a laboratory scale (10 L). The yeasts utilized glucose as the only carbon source and produced 3.25 ± 1.77, and 4.25 ± 1.06% (v/v), of ethanol for L. thermotolerans and quebecensis, respectively. While in the maltose-defined medium, ethanol content reached 3.25 ± 0.45, and 3.92 ± 0.36, respectively. The presence of alpha acids and ethanol affected the growth of L. quebecensis, which showed lower growth at 90 IBU and 8 ethanol% (v/v) mixtures. The craft beer brewed with L. quebecensis in monoculture experiments showed fruity flavors associated with ethyl acetate and isoamyl acetate. The ethanol content reached 3.50 ± 0.46% (v/v). The beer pH was 4.06 ± 0.20, with a lactic acid concentration of 1.21 ± 0.05 g/L. The sensory panel identified the beer as "fruity", "floral", "hoppy", "sweet", and "sour". To our knowledge, this is the first time L. quebecensis was reported as a potential candidate for sour beer production with reduced ethanol content.

Glucose Starvation Stimulates the Promoting Strength of a Novel Evolved Suc2 Promoter.

Promoters are essential for designing Saccharomyces cerevisiae cell factories. Identifying novel promoters tuned to produce specific metabolites under increasingly diverse industrial stresses is required to improve the economic feasibility of whole fermentation processes. In this study, a positively evolved Suc2 promoter (SUC 2p) with promoter activity stronger than that of the wild-type Suc2 promoter (SUC 2wtp) was obtained. Quantitative real-time PCR, fluorescence analysis, Western blotting, and a β-galactosidase activity assay revealed that SUC 2p is a medium-strength promoter compared with eight reported promoters at a medium glucose concentration (2% (w/v)). Different glucose concentrations modulated the strength of SUC 2p. Low glucose concentrations (0.05 and 0.5% (w/v)) enhanced the promoter strength of SUC 2p dramatically, with promoter activity higher than that of reported strong promoters. Glucose starvation resulted in the formation of a new Msn2/4 binding site on SUC 2p. Our work should facilitate the development of promoters with novel fine-tuning properties and the construction of S. cerevisiae cell factories suitable for the industrial production of essential chemicals under glucose-deprived conditions.

PLA-Mg film degradation under in vitro environments supplemented with glucose and/or ketone bodies

Materials used for the manufacture of implants may suffer alterations in their surface properties as a result of continuous contact with physiological fluids. Moreover, in biodegradable and bioabsorbable materials, such as polylactic acid (PLA), these changes may be more accentuated, and their biological response may be affected by the presence of proteins, enzymes or other compounds of an oxidizing character. This research proposes to study the degradation of PLA in a close-to-reality environment by supplementing the physiological buffer m-SBF with concentrations of glucose and ketone bodies of healthy individuals. To this end, parameters such as hydrophobicity, surface tension, topography and surface chemical composition of PLA films and PLA films doped with magnesium particles after degradation were evaluated to determine how these components influence these properties compared to degradation in standard buffer. The presence of glucose and/or ketone bodies in the degradation medium of PLA doped with magnesium particles altered the composition of the salt layers absorbed on the surface of the material due to the action of gluconate and/or hydroxybutyrate anions, which were able to coordinate with ions from the solution as well as Mg2+ cations from polymer degradation. The salt accumulation on polymeric films changes the surface mechanical response increasing the Young's modulus after degradation.

Production of surfactin from novel Bacillus spp. isolated from soil and its antifungal properties

Aim: This research aimed to screen the biosurfactant-producing bacteria from soil, especially for the betterment of the environment in terms of bioremediation and biodegradation of lipids and hydrocarbons. Methodology: Bacillus spp. isolated from the soil was cultured and inoculated into different mineral media such as Glucose, Sawdust, and Bagasse mineral media. The surfactin was extracted and the activity was studied using oil emulsification test, oil spreading assay, and drop collapse assay. Bacillus spp. that provided better results were sequenced and submitted to NCBI. The surfactin of the specific bacteria was characterized using TLC, UV-vis spectra, and FTIR analysis. Finally, the antifungal activity assay was performed against Aspergillus flavus, Rhizopus spp., and Aspergillus niger. Results: Bacillus species subjected to gene sequencing was identified as Bacillus mycoids strain ARJCBNR (On464184). UV-vis spectra and FTIR analyses revealed various functional groups such as 3356.14 cm-1 (N-H stretch), 1635.64 cm-1 (C=O amide), and 1211 cm-1 (C-O stretch) representing the surfactin compound between 4000 and 500 cm-1. Upon antifungal activity, sawdust-extracted surfactin had a higher zone of inhibition against all the tested fungi. Interpretation: This study revealed that Bacillus spp. can utilize agro-industrial waste as substrate to produce surfactin. Also, the surfactin extracted from sawdust mineral media showed higher antifungal activity. Key words: Bacillus spp., Biosurfactant, Mineral media, Soil, Surfactin

Medium and process optimizations for Euglena gracilis with high biomass production enriched with protein

Axenic Euglena cultures were grown under different LED conditions, medium compositions, and durations of cultivation to identify the factors involved in tuning Euglena biomass quality, especially protein and paramylon contents. Compositional analysis suggests that paramylon and protein concentrations in biomass can be tuned to different extents depending on medium and process conditions, including medium compositions, glucose concentrations, and illuminations of various LED spectra. Through the optimization of these parameters, the biomass productivities under specified conditions reached ~50–68 g/L. The protein-rich (>50 % dry basis) Euglena cultures can be produced with higher biomass productivities (38 g/L) under mixotrophic conditions with the presence of white and red LED lights and the modified medium composition. The cell morphology studies of Euglena cultures under various cultivation conditions indicate the correlation of slender cell-shape to the cell feature of high-protein contents in the biomass. The fresh cellular volumes and dry weights of paramylon-rich round-shaped cells were also estimated to be 5–9 times higher than those of protein-rich slender-shaped cells. The present study demonstrates an effective method to produce high-protein Euglena biomass with higher productivity in a shorter time.

Integration of systematic review, lipidomics with experiment verification reveals abnormal sphingolipids facilitate diabetic retinopathy by inducing oxidative stress on RMECs

ObjectiveThis study aims to explore the potential biomarkers in the development of diabetes mellitus (DM) into diabetic retinopathy (DR). MethodsSystematic review of diabetic metabolomics was used to screen the differential metabolites and related pathways during the development of DM. Non-targeted lipidomics of rat plasma was performed to explore the differential metabolites in the development of DM into DR in vivo. To verify the effects of differential metabolites in inducing retinal microvascular endothelial cells (RMECs) injury by increasing oxidative stress, high glucose medium containing differential metabolites was used to induce rat RMECs injury and cell viability, malondialdehyde (MDA) contents, superoxide dismutase (SOD) activities, reactive oxygen species (ROS) levels and mitochondrial membrane potential (MMP) were evaluated in vitro. Network pharmacology was performed to explore the potential mechanism of differential metabolites in inducing DR. ResultsThrough the systematic review, 148 differential metabolites were obtained and the sphingolipid metabolic pathway attracted our attention. Plasma non-targeted lipidomics found that sphingolipids were accompanied by the development of DM into DR. In vitro experiments showed sphinganine and sphingosine-1-phosphate aggravated rat RMECs injury induced by high glucose, further increased MDA and ROS levels, and further decreased SOD activities and MMP. Network pharmacology revealed sphinganine and sphingosine-1-phosphate may induce DR by regulating the AGE-RAGE and HIF-1 signaling pathways. ConclusionsIntegrated systematic review, lipidomics and experiment verification reveal that abnormal sphingolipid metabolism facilitates DR by inducing oxidative stress on RMECs. Our study could provide the experimental basis for finding potential biomarkers for the diagnosis and treatment of DR.

Long-read direct RNA sequencing of the mitochondrial transcriptome of Saccharomyces cerevisiae reveals condition-dependent intron abundance.

Mitochondria fulfil many essential roles and have their own genome, which is expressed as polycistronic transcripts that undergo co- or posttranscriptional processing and splicing. Due to the inherent complexity and limited technical accessibility of the mitochondrial transcriptome, fundamental questions regarding mitochondrial gene expression and splicing remain unresolved, even in the model eukaryote Saccharomyces cerevisiae. Long-read sequencing could address these fundamental questions. Therefore, a method for the enrichment of mitochondrial RNA and sequencing using Nanopore technology was developed, enabling the resolution of splicing of polycistronic genes and the quantification of spliced RNA. This method successfully captured the full mitochondrial transcriptome and resolved RNA splicing patterns with single-base resolution and was applied to explore the transcriptome of S. cerevisiae grown with glucose or ethanol as the sole carbon source, revealing the impact of growth conditions on mitochondrial RNA expression and splicing. This study uncovered a remarkable difference in the turnover of Group II introns between yeast grown in either mostly fermentative or fully respiratory conditions. Whether this accumulation of introns in glucose medium has an impact on mitochondrial functions remains to be explored. Combined with the high tractability of the model yeast S. cerevisiae, the developed method enables to monitor mitochondrial transcriptome responses in a broad range of relevant contexts, including oxidative stress, apoptosis and mitochondrial diseases.

Submerged fermentation using Aspergillus tubingensis as an efficient strategy to obtain antioxidant extracts from black cricket proteins

In recent years, there has been growing interest in edible insects as a novel food source due to their high nutritional value and more sustainable cultivation system compared to traditional animal sources. Edible insects are rich in proteins, which can be used to produce bioactive peptides through enzymatic hydrolysis or fermentation. The main objective of this study was to evaluate the production of an antioxidant extract using proteins extracted from the black cricket (Gryllus assimilis) as a substrate for the submerged fermentative process conducted by the filamentous fungus Aspergillus tubingensis. A response surface methodology (CCRD) was employed to investigate the effect of glucose and black cricket protein concentrations in the culture medium on the release of the peptides. The results demonstrated that the maximum values for DPPH (290.79 µmol TE g–1), ABTS (862.82 µmol TE g–1), and FRAP (1020.11 µmol TE g–1) were obtained at 72 or 96 h of fermentation using lower concentrations of both protein and glucose. These findings confirm that black cricket proteins are suitable for producing an antioxidant extract via microbial fermentation.

Resveratrol improves palmitic acid‑induced insulin resistance via the DDIT4/mTOR pathway in C2C12 cells.

The present study aimed to establish a model of palmitic acid (PA)‑induced insulin resistance (IR) in C2C12 cells and to determine the mechanism underlying how resveratrol (RSV) improves IR. C2C12 cells were divided into the control (CON), PA, PA + RSV, PA + RSV + DNA damage‑inducible transcript 4 (DDIT4)‑small interfering (si)RNA and PA + RSV + MHY1485 (mTOR agonist) groups. Glucose contents in culture medium and triglyceride contents in cells were determined. Oil red O staining was performed to observe the pathological changes in the cells. Reverse transcription‑quantitative PCR and western blotting were conducted to evaluate the mRNA and protein expression levels, respectively, of DDIT4, mTOR, p70 ribosomal protein S6 kinase (p70S6K), insulin receptor substrate (IRS)‑1, PI3K, AKT and glucose transporter 4 (GLUT4). Compared with in the CON group, glucose uptake was decreased, cellular lipid deposition was increased, phosphorylated (p)‑IRS‑1, p‑mTOR and p‑p70S6K protein expression levels were increased, and p‑PI3K, p‑AKT, GLUT4 and DDIT4 protein expression levels were decreased in the PA group. By contrast, compared with in the PA group, culture medium glucose content and cellular lipid deposition were decreased, p‑PI3K, p‑AKT, GLUT4 and DDIT4 protein expression levels were increased, p‑IRS‑1 protein expression levels were decreased, and mTOR and p70S6K mRNA and protein expression levels were decreased in the PA + RSV group. Compared with in the PA + RSV group, DDIT4 protein and mRNA expression levels were reduced in the PA + RSV + DDIT4‑siRNA group, but showed no change in the PA + RSV + MHY1485 group. Following transfection with DDIT4‑siRNA or treatment with MHY1485, the effects of RSV on improving IR and lipid metabolism were weakened, mTOR and p70S6K protein expression levels were upregulated, p‑PI3K, p‑AKT and GLUT4 protein expression levels were down‑regulated, p‑IRS‑1 protein expression levels were upregulated, and culture medium glucose content and cellular lipid deposition were increased. In conclusion, RSV may improve PA‑induced IR in C2C12 cells through the DDIT4/mTOR/IRS‑1/PI3K/AKT/GLUT4 signaling pathway, as well as via improvements in glucose and lipid metabolism.

Elabela inhibits TRAF1/NF-κB induced oxidative DNA damage to promote diabetic foot ulcer wound healing

Diabetic foot ulcer (DFU) is a serious complication of diabetes. Elabela (ELA), a ligand of apelin receptor (APJ), was shown to promote angiogenesis and suppress inflammation. This study aimed to illustrate the role of ELA in DFU wound healing. A whole-skin defect model was constructed using db/m and db/db mice to observe the effects of ELA on wound healing. The function of ELA in endothelial cells cultured in high glucose medium was investigated. Administration of ELA in peri-wound area of db/db mice accelerated wound closure and reduced inflammatory infiltration. Indicators of DNA damage, elevated reactive oxygen species (ROS) levels and tail DNA amounts, were downregulated by ELA but compromised after TRAF1 overexpression. ELA-mediated inhibition of NF-κB phosphorylation improved cell migration and angiogenesis, which were blocked by APJ silencing. The findings imply that ELA suppresses TRAF1-mediated NF-κB signal activation, reducing ROS-related oxidative DNA damage and improving protection of endothelial function.

Identification and Validation of Genes Related to RNA Methylation Modification in Diabetic Retinopathy

Purpose To identify and validate the differentially expressed genes related to RNA methylation modification in diabetic retinopathy. Methods The data sets GSE12610 and GSE111465 related to diabetic retinopathy in the Gene Expression Omnibus were selected. The R software package was used to identify differentially expressed genes related to RNA methylation modification in diabetic retinopathy. Protein-protein interaction network was constructed to explore the interactions between proteins and predict proteins. Then, Gene Ontology annotation analysis and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analysis were used to analyze the potential enrichment pathways and clarify the biological functions of these genes. In addition, the correlation between them and immune cells was visualized, and receiver operating characteristic curves were drawn to evaluate the diagnostic performance of each one of them for diabetic retinopathy. To verify the differentially expressed genes, the mRNA expression of rat retinal vascular endothelial cells cultured in low and high glucose medium separately were detected by RT-qPCR. Results The expression of Lrpprc, Nsun4, Nsun6 and Trdmt1 were significantly up-regulated in diabetic retinopathy samples, while the expression of Cbll1, Hnrnpc, Mettl3 and Wtap were significantly down-regulated. Differentially expressed genes were mainly enriched in the RNA-methylation-medication pathways and biological function. The results of immune infiltration analysis proved that eosinophils aggregated more in diabetic group, while T cells follicular helper aggregated more in normal samples. These genes of Cbll1 (AUC = 0.986), Hnrnpc (AUC = 0.819), Lrpprc (AUC = 0.806), Mettl3 (AUC = 0.917), Nsun4 (AUC = 0.819), Nsun6 (AUC = 0.819), Trdmt1 (AUC = 0.972) and Wtap (AUC = 0.972) were respectively used as the diagnostic basis of diabetic retinopathy. According to the RT-qPCR results, the expression of Mettl3 was significantly down-regulated (p < 0.0005) in cells cultured in high glucose, while Trdmt1 (p < 0.05), Nsun4 (p < 0.05) and Nsun6 (p < 0.05) were significantly up-regulated. Conclusion Differentially expressed genes such as Mettl3, Nsun4, Nsun6, and Trdmt1 should be conducted to explore, and the role of RNA methylation in the process of diabetic retinopathy would be revealed in-depth.

Zoledronic acid epigenetically alleviates high-glucose-suppressed osteogenic differentiation of MC3T3-E1 cells.

Due to the impact of excessive glucose on osteogenic differentiation, diabetic osteopathy frequently results in skeletal fragility, osteoporosis, and bone pain. Zoledronic acid, a bisphosphonate (BP) that effectively inhibits osteoclastic bone resorption is given yearly to improve bone mineral density (BMD) in patients with osteoporosis. However, the detailed molecular mechanisms remained unclear. This study investigates the possible pathways by which zoledronic acid regulates osteogenesis when blood glucose levels are high. MC3T3-E1 cells were treated with one mM zoledronic acid or not in a standard or high glucose culture medium. A quantitative polymerase chain reaction (qPCR) assay was utilized to assess the expression of the target candidate genes, including RUNX2, MALAT1, miR-133, miR-20a, and miR-204. In a high-glucose condition, zoledronic acid treatment significantly lowered MALAT1 (p < 0.0001) and miR-20a (p < 0.0001) expression. Conversely, in a high-glucose condition, RUNX2, miR-133, and miR-204 expressions were found to be significantly increased in the zoledronic acid treatment group as compared to no treatment (all p < 0.0001). In conclusion, under a high-glucose environment, zoledronic acid can modulate the expression of the RUNX2 transcription factor through epigenetic regulation.

Development of tissue culture system with automated pulsation and Kalman filter control for an artificial artery model.

Tissue-engineered arterial vessels have been used as substitutes for unnecessary animal experiments to evaluate the pharmacokinetics of drugs targeting various arteriopathies caused by structural or physiological arterial defects. An arterial tissue culture system was established to simulate the mechanical characteristics of a heart-beating pump and to do online feedback control of lactate and glucose concentrations. The mechanically controlled flow pump mimicked the heart pumping inside a tissue-engineered artery composed of muscle and endothelial cells within a nanofibrous scaffold. After monitoring the pH of the culture medium online, lactate and glucose were estimated using the Kalman filter algorithm, and the set-point online control was operated to maintain glucose for artery tissue engineering. The composition of the artificial artery was confirmed by immunofluorescence staining, and its mechanical characteristics were examined. The online automated system successfully demonstrated its applicability as a standardized process for arterial tissue culture to replace animal arterial experiments.

Glucose promoting the early embryonic development by increasing the lipid synthesis at 2-cell stage.

The optimization of culture conditions is one of the main strategies to improve the embryo development competence in in vitro fertilization (IVF). Glucose is an important carbon source while also exists in the oviductal fluid in vivo, the effect of glucose in embryo development microenvironment is still unclear. Here we employed the LC-MS to detect and analyze the metabolites in the culture medium of different cleavage stages including 2-Cell, 4-Cell and 8-Cell mouse embryos, respectively. The effects of the external glucose were estimated by measuring the development rate at different glucose concentrations from 0 to 5mmol/L, and the gene expression changes were detected to explore the potential mechanism after the addition of glucose in the media. Our results indicated the 2-Cell and 8-Cell stages had defined characteristic metabolites, while 4-Cell stage was the transition state. Global and contiguous metabolic characteristics showed the glycometabolism play a critical role at each early cleavage stages during the embryo development. The 8-Cell rates demonstrated the addition of glucose in culture media significantly improve the embryo competence, the highest rate was 87.33% using 3mmol/L glucose in media, in contrast only 9.95% using the media without glucose. Meanwhile, the blocked embryos were mainly enriched at 2-Cell stage. Further transcriptome study found 3mmol/L glucose in media remarkably upregulated the gene expression of lipid biosynthesis at 2-Cell stage, the increased lipid was confirmed by nile red staining. These data indicated the glucose may promote the development competence through increasing the lipid biosynthesis to overcoming the 2-Cell block. Our findings were helpful for the further optimization of IVF culture media, as well as the estimation of embryo quality using metabolites in the culture media.