MARCKS mRNA Research Articles

Dapagliflozin Suppresses High Glucose-Induced Proliferation, Oxidative Stress, and Fibrosis by Reducing Mettl3-Induced m6A Modification in Marcks mRNA.

Diabetic cardiomyopathy (DCM) is a common and severe complication of Diabetes mellitus (DM). Dapagliflozin (DAPA) is an oral anti-diabetic drug worldwide for the treatment of type 2 DM. However, the action and mechanism of DAPA in cardiac fibrosis during DCM remain vague. Primary cardiac fibroblasts (CFs) were incubated with high glucose (HG) in vitro. Cell proliferation was detected by MTT and EdU assays. Oxidative stress was evaluated by determining the production of reactive oxygen species and malondialdehyde. Cell fibrosis was assessed by detecting fibrosis-related proteins by western blotting. Levels of Mettl3 (Methyltransferase 3) and Marcks (myristoylated alanine-rich C kinase substrate) were measured using qRT-PCR and western blotting. The m6A modification profile was determined by methylated RNA immunoprecipitation assay and the interaction between Mettl3 and Marcks was verified using dual-luciferase reporter and RIP assays. DAPA treatment alleviated HG-induced proliferation, oxidative stress, and fibrosis in CFs. HG promoted the expression of Mettl3 in CFs. Knockdown of Mettl3 reversed HG-induced proliferation, oxidative stress, and fibrosis in CFs; moreover, forced expression of Mettl3 abolished the protective effects of DAPA on CFs under HG condition. Mechanistically, Mettl3 interacted with Marcks in CFs and induced Marcks mRNA m6A modification. HG induced high expression of Marcks in CFs. The overexpression of Marcks could counteract DAPA or Mettl3 knockdown-evoked inhibitory effects on CF proliferation, oxidative stress, and fibrosis under HG condition. Dapagliflozin suppressed HG-induced proliferation, oxidative stress, and fibrosis by reducing Mettl3-induced m6A modification in Marcks mRNA.

NAP-22 and MARCKS mRNA expression levels in spontaneously hypertensive rat kidneys: the effect of drinking water with different calcium contents

Objective. The aim of study was to assess the effect of the level of calcium intake with drinking water on NAP-22 and MARCKS mRNA expression in cortical and medullar kidney layers of spontaneously hypertensive rats.Design and methods. The study involved 90-day-old SHR (n = 8) and WKY (n = 8) strain rats of both sexes. We assessed tissue samples from cortical and medullar kidney layers. NAP-22 and MARCKS mRNA expression levels were determined by RT-PCR.Results. Sufficient drinking water calcium intake was associated with similar the expression of NAP-22 and MARCKS mRNA in kidneys of spontaneously hypertensive and normotensive rats. Consumption of drinking water with insufficient calcium content it decreases in both rat strains, being more evident in spontaneously hypertensive rats, especially in the medullar layer.Conclusions. Our results show that genetically determined impairments of calcium metabolism in cells of spontaneously-hypertensive rats (SHR line) and their effect on intracellular signaling processes are more evident with the reduced intake of exogenous calcium.

NAP-22 and MARCKS mRNA expression levels in spontaneously hypertensive rat kidneys: the effect of drinking water with different calcium contents

Objective. The aim of study was to assess the effect of the level of calcium intake with drinking water on NAP-22 and MARCKS mRNA expression in cortical and medullar kidney layers of spontaneously hypertensive rats.Design and methods. The study involved 90-day-old SHR (n = 8) and WKY (n = 8) strain rats of both sexes. We assessed tissue samples from cortical and medullar kidney layers. NAP-22 and MARCKS mRNA expression levels were determined by RT-PCR.Results. Sufficient drinking water calcium intake was associated with similar the expression of NAP-22 and MARCKS mRNA in kidneys of spontaneously hypertensive and normotensive rats. Consumption of drinking water with insufficient calcium content it decreases in both rat strains, being more evident in spontaneously hypertensive rats, especially in the medullar layer.Conclusions. Our results show that genetically determined impairments of calcium metabolism in cells of spontaneously-hypertensive rats (SHR line) and their effect on intracellular signaling processes are more evident with the reduced intake of exogenous calcium.

MARcKS and nAP-22 proteins mRnA expression in renal cortex and renal medulla of rats with spontaneous hypertension

Objective. To study the changes in mRNA expression level of two main protein kinase C substrates — MARCKS and NAP-22 — in rats with spontaneous hypertension (SHR rats) and in normotensive control rats (WKY rats) in renal cortex, renal medulla and total kidney. We also aimed at the identification of possible interstrain differences between the mRNA expression levels.Design and methods.We assessed the level of MARCKS and NAP-22 mRNA by real-time polymerase chain reaction in male SHR and WKY (as a normotensive control) rats.Results. In SHR rats, MARCKS mRNA expression level in renal cortex was 1,5 times higher than in renal medulla (p = 0,0001) and also higher than in total kidney (p = 0,001), in renal medulla it was lower than in total kidney (p = 0,002). In WKY rats, MARCKS mRNA expression level in renal cortex was higher than in renal medulla (p = 0,0005). There was no differences neither between renal cortex and total kidney (p = 0,011), nor between renal medulla and total kidney (p = 0,716). In SHR rats, NAP-22 mRNA expression level in renal cortex was twofold higher than in renal medulla (p = 0,001), in renal medulla it was lower than in total kidney (p = 0,005), the differences between renal cortex and total kidney were less significant (p = 0,011). In WKY rats, NAP-22 mRNA expression level in renal cortex was 1,5 times higher than in renal medulla (p = 0,001), while in renal medulla it was lower than in total kidney (p = 0,002). There was no significant difference in NAP-22 mRNA expression level between renal medulla and total kidney (p = 0,011). There were no significant interstrain differences in the animal groups either in the levels of MARCKS mRNA expression in renal cortex (p = 0,872), in renal medulla (p = 0,024) or in total kidney (p = 0,520). Neither there were differences in the levels of NAP-22 mRNA expression in cortex (p = 0,028), in medulla (p = 0,028) and in total kidney (p = 0,978).Conclusions. In both SHR and WKY rat strains, the level of MARCKS and NAP-22 mRNA expression in cortical and medullary kidney layers is different, in WKY rats these differences are less pronounced. At the same time, interstrain differences in NAP-22 and MARCKS mRNA expression levels in cortical, medullary layers and in total kidney of SHR and WKY rats were not found.

MicroRNA 429 Regulates Mucin Gene Expression and Secretion in Murine Model of Colitis.

miRNAs are non-coding RNAs that play important roles in the pathogenesis of human diseases by regulating target gene expression in specific cells or tissues. We aimed to detect miRNAs related to ulcerative colitis [UC], identify their target molecules, and analyse the correlation between the miRNAs and their target genes in colorectal cells and dextran sulphate sodium [DSS]-induced mouse colitis. UC-associated miRNAs were identified by miRNA microarray analysis using DSS-induced colitis and normal colon tissues. The results were validated by quantitative real-time polymerase chain reaction [RT-PCR]. We identified target genes of MIR429, a colitis-associated miRNA, from our screen by comparing the mRNA microarray analysis in MIR429-overexpressed cells with predicted candidate target genes. We constructed luciferase reporter plasmids to confirm the effect of MIR429 on target gene expression. The protein expression of the target genes was measured by western blot,enzyme-linked immunosorbent assay [ELISA] analysis, or immunohistochemistry. We identified 37 DSS-induced colitis associated miRNAs. We investigated MIR429 that is down-regulated in DSS-induced colitis, and identified 41 target genes of MIR429. We show that the myristoylated alanine-rich protein kinase C substrate [MARCKS] is a direct target of MIR429. MARCKS mRNA and protein expression levels are down-regulated by MIR429, and MIR429 regulates the expression of MARCKS and MARCKS-mediated mucin secretion in colorectal cells and DSS-induced colitis. In addition, anti-MIR429 up-regulates MARCKS expression in colorectal cell lines. Our findings suggest that MIR429 modulates mucin secretion in human colorectal cells and mouse colitis tissues by up-regulating of MARCKS expression, thereby making MIR429 a candidate for anti-colitis therapy in human UC.

MARCKS and protein F1/GAP-43 mRNA in chick brain: effects of imprinting

The phosphorylation of MARCKS, but not protein F1/GAP-43, is increased in the intermediate and medial portion of the hyperstriatum ventrale (IMHV) after chick imprinting. Here we investigated if MARCKS, but not F1/GAP-43, gene expression would also be altered after imprinting. We first investigated the constitutive mRNA distribution of MARCKS and F1/GAP-43 in chick brain. MARCKS mRNA was expressed in most cells and exhibited a relatively homogeneous distribution. In contrast, F1/GAP-43 mRNA levels were elevated in discrete brain regions, as we had observed in mammals. The highest F1/GAP-43 mRNA levels in the chick brain were in sensory and associational structures such as the hyperstriatal complex and neostriatum, and lower levels were in structures involved in motor control, such as paleostriatum. These results in chick are consistent with the previously drawn generalization that F1/GAP-43 mRNA is expressed in those brain regions which exhibit synaptic plasticity. After imprinting, MARCKS mRNA levels in IMHV were higher in good learners than poor learners. In contrast, analysis of F1/GAP-43 mRNA levels revealed no differences related to training in any brain region sampled. These selective results for MARCKS but not F1/GAP-43 parallel the prior findings on their phosphorylation, and are consistent with our hypothesis that the very same proteins that are post-translationally modified in association with learning and memory also undergo alterations in their gene expression.

Decreased expression of the myristoylated alanine-rich C kinase substrate in transformed [formula omitted] 3T3 mouse fibroblasts

Expression of the myristoylated alanine-rich C kinase substrate was studied in the untransformed A31 and the transformed BPA31, DA31 and KA31 BALB c 3T3 mouse fibroblast cell lines. The MARCKS transcript was markedly reduced in the three transformed cell lines. Southern analysis revealed similar gene copy number and no apparent DNA rearrangement. In the untransformed A31 cells, the MARCKS mRNA was constitutive in the cell cycle.