Detecting Dna Synthesis Of Neointimal Formation After Catheter Balloon Injury In Gk And In Wistar Rats: Using 5-ethynyl-2 '-deoxyuridine
alkynes; arterial injury; balloon injury; Cardiovascular System & Cardiology; carotid-artery; Catheter; cell-cycle progression; click chemistry; diabetes-mellitus; diabetes-mellitus; DNA synthesis; EdU; eluting stents; Endocrinology & Metabolism; in-vitro; mammalian target; Neointimal formation; PCNA; proliferation; terminal
Background: Neointimal formation plays an important role in the pathogenesis of coronary restenosis after percutaneous coronary intervention (PCI), especially in patients with diabetes mellitus. Recently, some studies have shown that 5-ethynyl-2'-deoxyuridine (EdU) incorporation can serve as a novel alternative to the 5-bromo-2'-deoxyuridine (BrdU) antibody detection method for detection of DNA synthesis in regenerating avian cochlea, chick embryo and the adult nervous system. However, few studies have been performed to assess the suitability of EdU for detecting DNA synthesis in vascular neointima. Methods: The carotid artery balloon injury model was established in Goto-Kakizaki (GK) and Wistar rats. A Cell-Light (TM) EdU Kit was used to detect EdU-labeled cell nuclei of common carotid arteries at day 7 after catheter balloon injury. Different methods of injecting EdU were tested. The protein levels of proliferating cell nuclear antigen (PCNA) and p-Akt (Ser473), as well as the mRNA levels of PCNA were evaluated by Western blotting and quantitative real-time PCR (qRT-PCR), respectively. Immunohistochemical staining was also employed to visualize PCNA-positive cells. Results: At day 7 after catheter balloon injury, far more EdU-positive and PCNA-positive cells were observed in GK rats. When comparing groups that received different EdU doses, it was found that the percentage of EdU-positive cells at a dose of 100 mg/kg body weight was than at doses of 25 mg/kg and 50 mg/kg. The number of positive cells was significantly higher in the repeated injection group compared to the single injection group. Further, after balloon injury DNA synthesis in GK rats was more notable than in Wistar rats. Neointimal formation in GK rats was more obvious than in Wistar rats. The protein levels of PCNA and p-Akt (Ser473) and the mRNA levels of PCNA were increased in injured rats as compared to uninjured rats, and were significantly higher in GK rats than in Wistar rats. Conclusion: By intraperitoneal injections of EdU at a dose of 100 mg/kg three times, EdU incorporation can detect carotid arterial DNA synthesis caused by neointimal formation in GK rats and Wistar rats at day 7 after balloon injury by the EdU click reaction quickly and effectively. Moreover, more obvious DNA synthesis in the vascular neointima could be observed in GK rats than in Wistar rats.
Guo J S; Li D Y; Bai S R; Xu T D; Zhou Z M; Zhang Y B
Cardiovascular Diabetology
2012
2012-12
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1186/1475-2840-11-150" target="_blank" rel="noreferrer noopener">10.1186/1475-2840-11-150</a>
Sex and temporally-dependent effects of methamphetamine toxicity on dopamine markers and signaling pathways
dopamine; Akt; Neurosciences & Neurology; Pharmacology & Pharmacy; gene-expression; estrous-cycle; striatum; protein-phosphorylation; markers; glycogen-synthase kinase-3-beta; Methamphetamine; Sex difference; evoked striatal dopamine; induced neurotoxicity; monoamine; neuronal apoptosis; neurotoxicity; physiological functions; terminal; transporters
Methamphetamine induces a greater neurodegenerative effect in male versus female mice. In order to investigate this sex difference we studied the involvement of Akt and extracellular signal-regulated kinase (ERK1/2) in methamphetamine toxicity as a function of time post-treatment (30 min, 1 and 3 days). Methamphetamine-induced decreases in dopamine concentrations and dopamine transporter (DAT) specific binding in the medial striatum were similar in female and male mice when evaluated 1 day post-methamphetamine (40 mg/kg). At 3 days post-methamphetamine, striatal dopamine concentration and DAT specific binding continued to decline in males, whereas females showed a recovery with increases in dopamine content and DAT specific binding in medial striatum at day 3 versus day 1 post-methamphetamine. The reduction in striatal vesicular monoamine transporter 2 specific binding observed at 1 and 3 days post-methamphetamine showed neither a sex- nor temporal-dependant effect. Under the present experimental conditions, methamphetamine treatments had modest effects on dopamine markers measured in the substantia nigra. Proteins assessed by Western blots showed similar reductions in both female and male mice for DAT proteins at 1 and 3 days post-methamphetamine. An increase in the phosphorylation of striatal Akt (after 1 day), glycogen synthase kinase 3 beta (at 1 and 3 days) and ERK1/2 (30 min post-methamphetamine) was only observed in females. Striatal glial fibrillary acidic protein levels were augmented in both females and males at 3 days post-methamphetamine. These results reveal some of the sex- and temporally-dependent effects of methamphetamine toxicity on dopaminergic markers and suggest some of the signaling pathways associated with these responses. (C) 2012 Elsevier Ltd. All rights reserved.
Bourque M; Dluzen D E; Di Paolo T
Neuropharmacology
2012
2012-06
Journal Article or Conference Abstract Publication
<a href="http://doi.org/10.1016/j.neuropharm.2012.02.009" target="_blank" rel="noreferrer noopener">10.1016/j.neuropharm.2012.02.009</a>