scholarly journals Effects of Abiotic Elicitors on Expression and Accumulation of Three Candidate Benzophenanthridine Alkaloids in Cultured Greater Celandine Cells

Molecules ◽  
2021 ◽  
Vol 26 (5) ◽  
pp. 1395
Author(s):  
Seyed Mohammad Hashemi ◽  
Mohammad Reza Naghavi ◽  
Mehdi Ghorbani ◽  
Chanditha Priyanatha ◽  
Peiman Zandi
Keyword(s):  
Gene Expression ◽  
Metabolic Regulation ◽  
Gene Expression Analysis ◽  
Significant Enhancement ◽  
Chelidonium Majus ◽  
Benzophenanthridine Alkaloids ◽  
Plant Resource ◽  
Abiotic Elicitors ◽  
Time Courses

Efforts to develop the necessary biotechnologies in Greater Celandine (Chelidonium majus L.), a leading plant resource for the development of plant-derived medicines, have been hampered by the lack of knowledge about transcriptome and metabolome regulations of its medicinal components. Therefore, this study aimed to examine the effect of abiotic elicitors, methyl jasmonate (MJ) and salicylic acid (SA), at different time courses (12, 24, 48, and 72 h), on expression and metabolome of key benzophenanthridine alkaloids (BPAs) in an optimized in vitro culture. Gene expression analysis indicated the upregulation of CFS (cheilanthifoline synthase) to 2.62, 4.85, and 7.28 times higher than the control at 12, 24, and 48 h respectively, under MJ elicitation. Besides, MJ upregulated the expression of TNMT (tetrahydroprotoberberine N-methyltransferase) to 2.79, 4.75, and 7.21 times at 12, 24, and 48 h respectively, compared to the control. Investigation of BPAs revealed a significant enhancement in the chelidonine content (9.86 µg/mg) after 72 h of MJ elicitation. Additionally, sanguinarine content increased to its highest level (3.42 µg/mg) after 24 h of MJ elicitation; however, no significant enhancement was detected in its content in shorter elicitation time courses. Generally, higher gene expression and BPAs’ level was observed through longer elicitation courses (48 and 72 h). Our findings take part in improving the understanding of transcription and metabolic regulation of BPAs in cultured Greater Celandine cells.

scholarly journals Effects of Abiotic Elicitors on Expression and Accumulation of Three Candidate Benzophenanthridine Alkaloids in Cultured Greater Celandine Cells 

2020 ◽  
Author(s):  
Seyed Mohammad Hashemi ◽  
Mohammadreza Naghavi ◽  
Mehdi Ghorbani ◽  
Peiman Zandi
Keyword(s):  
Gene Expression ◽  
Metabolic Regulation ◽  
Gene Expression Analysis ◽  
Significant Enhancement ◽  
Chelidonium Majus ◽  
Benzophenanthridine Alkaloids ◽  
Plant Resource ◽  
Abiotic Elicitors ◽  
Time Courses

Abstract Efforts to develop the necessary biotechnologies in Greater Celandine (Chelidonium majus L.), a leading plant resource for the development of plant-derived medicines, have been hampered by the lack of knowledge about transcriptome and metabolome regulations of its medicinal components. Therefore, this study aimed to examine the effect of abiotic elicitors, methyl jasmonate (MJ) and salicylic acid (SA), at different time courses (12, 24, 48, and 72 h), on expression and metabolome of key benzophenanthridine alkaloids (BPAs) in an optimized in vitro culture. Gene expression analysis indicated the up-regulation of CFS to 2.62, 4.85, and 7.28 times higher than the control at 12, 24, and 48 h, respectively, under MJ elicitation. Besides, MJ up-regulated the expression of TNMT to 2.79, 4.75, and 7.21 times at 12, 24, and 48 h, respectively, compared to the control. Investigation of BPAs revealed a significant enhancement in the chelidonine content (9.86 µg/mg) after 72 h of MJ elicitation. Additionally, sanguinarine content increased to its highest level (3.42 µg/mg) after 24 h of MJ elicitation; however, no significant enhancement was detected in its content in shorter elicitation time courses. Generally, higher gene expression and BPAs’ level was observed through longer elicitation courses (48 and 72 h). Our findings take part in improving the understanding of transcription and metabolic regulation of BPAs in cultured Greater Celandine cells.

Retinoic Acid Receptor Agonists Suppress Muscle Fatty Infiltration in Mice

2021 ◽  
Vol 49 (2) ◽  
pp. 332-339
Author(s):  
Hideyuki Shirasawa ◽  
Noboru Matsumura ◽  
Masaki Yoda ◽  
Kazumasa Okubo ◽  
Masayuki Shimoda ◽  
...  
Keyword(s):  
Gene Expression ◽  
Skeletal Muscle ◽  
Rotator Cuff ◽  
Rotator Cuff Tear ◽  
Gene Expression Analysis ◽  
Retinoic Acid Receptor ◽  
Adipogenic Differentiation ◽  
Fatty Infiltration ◽  
Cuff Tear

Background: The infiltration of fat tissue into skeletal muscle, a condition referred to as muscle fatty infiltration or fatty degeneration, is regarded as an irreversible event that significantly compromises the motor function of skeletal muscle. Purpose: To investigate the effect of retinoic acid receptor (RAR) agonists in suppressing the adipogenic differentiation of fibroadipogenic progenitors (FAPs) in vitro and fatty infiltration after rotator cuff tear in mice. Study Design: Controlled laboratory study. Methods: FAPs isolated from mouse skeletal muscle were cultured in adipogenic differentiation medium in the presence or absence of an RAR agonist. At the end of cell culture, adipogenic differentiation was evaluated by gene expression analysis and oil red O staining. A mouse model of fatty infiltration—which includes the resection of the rotator cuff, removal of the humeral head, and denervation the supraspinatus muscle—was used to induce fatty infiltration in the supraspinatus muscle. The mice were orally or intramuscularly administered with an RAR agonist after the surgery. Muscle fatty infiltration was evaluated by histology and gene expression analysis. Results: RAR agonists effectively inhibited the adipogenic differentiation of FAPs in vitro. Oral and intramuscular administration of RAR agonists suppressed the development of muscle fatty infiltration in the mice after rotator cuff tear. In accordance, we found a significant decrease in the number of intramuscular fat cells and suppressed expression in adipogenic markers. RAR agonists also increased the expression of the transcripts for collagens; however, an accumulation of collagenous tissues was not histologically evident in the present model. Conclusion: Muscle fatty infiltration can be alleviated by RAR agonists through suppressing the adipogenic differentiation of FAPs. The results also suggest that RAR agonists are potential therapeutic agents for treating patients who are at risk of developing muscle fatty infiltration. The consequence of the increased expression of collagen transcripts by RAR agonists needs to be clarified. Clinical Relevance: RAR agonists can be used to prevent the development of muscle fatty infiltration after rotator cuff tear. Nevertheless, further studies are mandatory in a large animal model to examine the safety and efficacy of intramuscular injection of RAR agonists.

scholarly journals Effects of Vitrification on the Blastocyst Gene Expression Profile in a Porcine Model

2021 ◽  
Vol 22 (3) ◽  
pp. 1222
Author(s):  
Cristina Cuello ◽  
Cristina A. Martinez ◽  
Josep M. Cambra ◽  
Inmaculada Parrilla ◽  
Heriberto Rodriguez-Martinez ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Analysis ◽  
Survival Rates ◽  
Control Group ◽  
P Value ◽  
Transcriptome Profile ◽  
Embryo Viability ◽  
The Impact

This study was designed to investigate the impact of vitrification on the transcriptome profile of blastocysts using a porcine (Sus scrofa) model and a microarray approach. Blastocysts were collected from weaned sows (n = 13). A total of 60 blastocysts were vitrified (treatment group). After warming, vitrified embryos were cultured in vitro for 24 h. Non-vitrified blastocysts (n = 40) were used as controls. After the in vitro culture period, the embryo viability was morphologically assessed. A total of 30 viable embryos per group (three pools of 10 from 4 different donors each) were subjected to gene expression analysis. A fold change cut-off of ±1.5 and a restrictive threshold at p-value < 0.05 were used to distinguish differentially expressed genes (DEGs). The survival rates of vitrified/warmed blastocysts were similar to those of the control (nearly 100%, n.s.). A total of 205 (112 upregulated and 93 downregulated) were identified in the vitrified blastocysts compared to the control group. The vitrification/warming impact was moderate, and it was mainly related to the pathways of cell cycle, cellular senescence, gap junction, and signaling for TFGβ, p53, Fox, and MAPK. In conclusion, vitrification modified the transcriptome of in vivo-derived porcine blastocysts, resulting in minor gene expression changes.

scholarly journals Insulin-Induced Gene 2 Involvement in Human Adipocyte Metabolism and Body Weight Regulation

2008 ◽  
Vol 93 (5) ◽  
pp. 1995-2001 ◽  
Author(s):  
Sergey Krapivner ◽  
Sergej Popov ◽  
Ekaterina Chernogubova ◽  
Mai-Lis Hellénius ◽  
Rachel M. Fisher ◽  
...  
Keyword(s):  
Gene Expression ◽  
Gene Expression Analysis ◽  
Adipocyte Differentiation ◽  
Regulatory Element ◽  
Human Adipose Tissue ◽  
Human Tissues ◽  
Weight Regulation ◽  
Body Weight Regulation ◽  
Sterol Regulatory Element

Abstract Background: Insulin-induced genes (INSIGs) encode proteins that block proteolytic activation of sterol regulatory element-binding proteins, transcription factors that regulate lipogenic enzymes, and adipocyte differentiation. Objective: Here, we analyzed the relative significance of INSIG1 and INSIG2 in human liver and adipocyte metabolism, and defined a novel, functional polymorphism in the promoter of INSIG2 associated with body mass index. Research Methods: Variations in gene expression of different human tissues, of hepatoma cells exposed to INSIG1 and INSIG2 gene silencing probes, and of differentiating 3T3-L1 adipocytes were determined by real-time quantitative PCR. The functional significance of a novel polymorphism in the promoter of INSIG2 was analyzed using in vitro methods and gene expression analysis of human adipose tissue, whereas the phenotype associated with this polymorphism was studied in two cohorts of middle-aged men. Results: Gene expression analysis of 17 human tissues demonstrated that INSIG1 is highly expressed in the liver, whereas INSIG2 is ubiquitously expressed. Gene silencing experiments confirmed that INSIG1, but not INSIG2, regulates the expression of sterol regulatory element-binding proteins target genes in human hepatoma cells. In contrast, adipocyte differentiation of 3T3-L1 cells was associated with a 13-fold increase in expression of INSIG2. Significant relationships between the INSIG2–102G/A polymorphism and body mass index were observed in two cohorts of middle-aged men (ANOVA P = 0.017 and 0.044, respectively). In vitro studies and analysis of allele-specific expression in human adipose tissue substantiated the functional significance of the INSIG2–102G/A polymorphism. Conclusion: INSIG2 is involved in adipocyte metabolism and body weight regulation.

212 INTERSPECIFIC CLONING AND EMBRYO AGGREGATION INFLUENCE THE EXPRESSION OF oct4, nanog, sox2, AND cdx2 IN CHEETAH AND DOMESTIC CAT BLASTOCYSTS

2015 ◽  
Vol 27 (1) ◽  
pp. 196
Author(s):  
L. N. Moro ◽  
D. Veraguas ◽  
L. Rodriguez-Alvarez ◽  
M. I. Hiriart ◽  
C. Buemo ◽  
...  
Keyword(s):  
Gene Expression ◽  
Expression Analysis ◽  
Internal Control ◽  
Gene Expression Analysis ◽  
Early Embryo ◽  
Domestic Cat ◽  
Nuclear Reprogramming ◽  
Acinonyx Jubatus ◽  
Standard Curve

The cheetah (Ch, Acinonyx jubatus) is a species considered globally endangered and cloning is one of the assisted reproductive techniques that can help to preserve it and to study early embryo development. However, the production of cloned felid embryos remains inefficient, probably because of the difficulty to control the process of nuclear reprogramming and obtain adequate gene expression. Embryo aggregation has been demonstrated to improve the cloning efficiency in several species and to normalise cdx2 in the mouse by lowering its expression (Balbach et al. 2010), but it has not been evaluated in felids before. To better understand the effect of interspecific somatic-cell nuclear transfer (iSCNT) and embryo aggregation in nuclear reprogramming, we analysed the expression of oct4, sox2, nanog, and cdx2 in cheetah blastocysts generated by iSCNT, domestic cat blastocysts (Dc) generated by SCNT, and IVF blastocysts as control. To achieve this, domestic cat oocytes were in vitro matured and zona-free SCNT or iSCNT was performed, as previously described (Moro et al. 2014, Reprod. Fertil. Dev.). Zona-free reconstructed embryos were then cultured individually (1X) or two embryo were cultured together (2X) in microwells, in synthetic oviductal fluid (SOF) medium. The experimental groups were Dc1X, Dc2X, Ch1X, Ch2X, and IVF. After 8 days of in vitro culture the blastocysts obtained were stored in RNA-later at –20°C. For gene expression analysis, blastocysts were pooled as follows: Dc1X, 4 replicates of 3 blastocysts each; Dc2X, 4 replicates of 3 blastocysts each; Ch1X, 2 replicates of 2 blastocysts and 1 replicate of 1 blastocyst; Ch2X, 4 replicates of 3 blastocysts each; IVF 3 replicates of 3 blastocysts each. Embryos were treated with a Cells-to-cDNA TM II kit (Life Technologies, Carlsbad, CA, USA) lyses buffer and treated with DNase I (0.04 U μL–1) for genomic DNA digestion. Gene expression analysis was performed by real-time qPCR using the standard curve method. In all qPCRs, GAPDH was used as an internal control. The statistical analysis was performed using a non-parametric Kruskal–Wallis test (P < 0.05). We observed that Dc1X blastocysts overexpressed the 4 genes evaluated respect to the IVF control. However, the gene expression of the aggregated group (Dc2X) was lower for all the genes, achieving the same levels of nanog and sox2 as the IVF blastocysts. The expression of oct4 and cdx2 were also closer to the expression levels of the control in the Dc2X group than in the Dc1X group. With respect to interspecific embryos, the amount of oct4 and cdx2 was also significantly reduced in the Ch2X blastocysts respect to Ch1X blastocysts. Both cheetah groups showed significantly lower expression of oct4, cdx2, and nanog than the IVF control. In conclusion, transcription of pluripotent and early differentiation factors in cheetah embryos was not as efficient as in the domestic cat embryos, probably caused by interspecific transfer. Our study demonstrated for the first time that defects in gene expression of domestic cat embryos can be corrected by embryo aggregation, providing a simple strategy to improve felid cloning.

175 Nobiletin enhances the quality of in vitro-matured bovine oocytes and blastocysts by altering the transcription of key developmental genes

2019 ◽  
Vol 31 (1) ◽  
pp. 212
Author(s):  
Y. N. Cajas ◽  
K. Cañón-Beltrán ◽  
M. E. González ◽  
P. Ramos-Ibeas ◽  
A. Gutierrez-Adán ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Gene Expression ◽  
Expression Analysis ◽  
Gene Expression Analysis ◽  
Statistical Significance ◽  
Housekeeping Genes ◽  
Cumulus Cells ◽  
Cell Junction ◽  
In Vitro Production

One of the problems associated with in vitro production of embryos in bovine is the increase in reactive oxygen species (ROS), which leads to cell alterations and death. Nobiletin is a polymethoxyflavone isolated from citrus fruits with various beneficial effects on cell cycle regulation and inhibition of ROS production. In a preliminary study, we demonstrated that supplementation of 25 or 50 µM nobiletin to the in vitro maturation (IVM) medium reduces oxidative stress and improves oocyte nuclear and cytoplasmic maturation and embryo development. Thus, in this study, we aimed to evaluate the antioxidant activity of nobiletin during IVM on bovine matured oocytes, their cumulus cells (CC), and blastocysts by quantitative changes of gene expression. Immature cumulus oocytes complexes (COC) were aspirated from ovaries of slaughtered heifers. Selected COC underwent IVM in TCM-199+10% FCS and 10ng mL−1 epidermal growth factor (EGF; Control) supplemented with 25 µM (Nob25) or 50 µM (Nob50) nobiletin (MedChemExpress, Monmouth Junction, NJ, USA) or 0.001% dimethyl sulfoxide (DMSO control), a vehicle for nobiletin dilution, in 5% CO2 in air at 38.5°C. After 24h, 50 matured oocytes/group and their CC were snap-frozen in LN2 for gene expression analysis. The remaining oocytes were fertilized (Day 0) and cultured in vitro. Blastocysts (Day 7; n=50/group) were snap-frozen in LN2 for gene expression analysis (5 replicates). The mRNA abundance of candidate genes related with oxidative stress (SOD2, CYP51); apoptosis (BAX); quality (BMP15, BMP7, CLIC1, MAPK1, ABCB1); and cell junction (GJA1) was measured by quantitative PCR; H2AFZ and 18S rRNA were used as housekeeping genes. Statistical significance was assessed by one-way ANOVA. Supplementation of IVM medium with Nob25 or Nob50 produced changes in the expression levels of genes related to oxidative stress and apoptosis during IVM compared with controls. SOD2 and CYP51 were down-regulated in oocytes and CC (P&lt;0.05) but not in blastocysts, whereas BAX was down-regulated only in CC (P&lt;0.05). Nobiletin supplementation in IVM increased the expression of MAPK1 in oocytes and blastocysts (P&lt;0.05); however, no differences were observed in CC. BMP15 for oocytes and their CC and GJA1 for CC were up-regulated in Nob25 and Nob50 groups compared with controls (P&lt;0.05). The relative abundance of CLIC1 decreased in blastocysts from both nobiletin groups compared with controls (P&lt;0.05). No significant differences in the expression in ABCB1 and BMP7 were detected. In conclusion, our results suggest that supplementation of 25 or 50 µM nobiletin to the IVM medium reduces oxidative stress in oocytes and CC, decreases CC apoptosis, and provokes positive changes in the expression of genes related to oocyte and embryo quality. This research was supported by Spanish MINECO (AGL2015-70140-R and AGL2015-66145-R). Y. N. Cajas was supported by a grant from SENESCYT-Ecuador.

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