Quantification of cAMP and cGMP analogs in intact cells: pitfalls in enzyme immunoassays for cyclic nucleotides.

Naunyn Schmiedebergs Arch Pharmacol

Institute for Clinical Biochemistry and Pathobiochemistry, University of Wuerzburg, Grombuehlstrasse 12, Wuerzburg, Germany.

Published: August 2011

Immunoassays are routinely used as research tools to measure intracellular cAMP and cGMP concentrations. Ideally, this application requires antibodies with high sensitivity and specificity. The present work evaluates the cross-reactivity of commercially available cyclic nucleotide analogs with two non-radioactive and one radioactive cAMP and cGMP immunoassay. Most of the tested cyclic nucleotide analogs showed low degree competition with the antibodies; however, with Rp-cAMPS, 8-Br-cGMP and 8-pCPT-cGMP, a strong cross-reactivity with the corresponding cAMP and cGMP, respectively, immunoassays was observed. The determined EIA-binding constants enabled the measurement of the intracellular cyclic nucleotide concentrations and revealed a time- and lipophilicity-dependent cell membrane permeability of the compounds in the range of 10-30% of the extracellular applied concentration, thus allowing a more accurate prediction of the intracellular analog levels in a given experiment.

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3145891PMC
http://dx.doi.org/10.1007/s00210-011-0662-6DOI Listing

Publication Analysis

Top Keywords

camp cgmp
16
cyclic nucleotide
12
nucleotide analogs
8
quantification camp
4
cgmp
4
cgmp analogs
4
analogs intact
4
intact cells
4
cells pitfalls
4
pitfalls enzyme
4

Similar Publications

Article Synopsis
  • Strychni Semen has shown anti-tumor properties against non-small cell lung cancer (NSCLC), but its molecular mechanisms require further investigation.
  • The study utilized network pharmacology and molecular docking to identify 21 active components and 67 targets related to NSCLC, with key findings showing strong interactions between the components and core targets such as PTGS2 and NR3C1.
  • Pathway analysis indicated that Strychni Semen's therapeutic effects on NSCLC primarily involve the Calcium, Estrogen, and cGMP-PKG/cAMP pathways, helping to elucidate its underlying mechanisms of action.
View Article and Find Full Text PDF

Phosphodiesterase 2 A (PDE2A) function is stimulated by cGMP to catabolize cAMP. However, neurological and neurochemical effects of PDE2A deficiency are poorly understood. To address this gap, we studied behavioral characteristics and cerebral morpho-chemical changes of adult male heterozygous C57BL/6-PDE2A+/- (HET), and wild type C57BL/6-PDE2A+/+ (WT) mice.

View Article and Find Full Text PDF

Objectives: To investigate which phosphodiesterase (PDE) isoforms are expressed in fibroblasts isolated from the tunica albuginea (TA) of patients with Peyronie's disease (PD), and to measure the potency of PDE inhibitors in preventing transformation of these fibroblasts to profibrotic myofibroblasts.

Materials And Methods: Fibroblasts isolated from the TA of men undergoing surgery for correction of PD curvature were transformed to myofibroblasts using transforming growth factor beta-1. The expression of 21 PDE isoforms was investigated using quantitative reverse transcriptase-polymerase chain reaction and protein analysis, as were the effects of various PDE inhibitors on prevention of myofibroblast transformation.

View Article and Find Full Text PDF

[Molecular mechanism of Xiangsha Liujunzi Decoction in treating chronic atrophic gastritis based on transcriptome sequencing technology].

Zhongguo Zhong Yao Za Zhi

September 2024

Key Laboratory of Traditional Chinese Medicine for Prevention and Control of Regional High Incidence Diseases in Ningxia,Ministry of Education, Ningxia Medical University Yinchuan 750004, China.

Based on transcriptomics technology, this study investigated the molecular mechanisms of Xiangsha Liujunzi Decoction in treating chronic atrophic gastritis(CAG), which were confirmed through experimental validation. The CAG rat model was built by the MNNG composite multi-factor method, followed by a 90-day administration of Xiangsha Liujunzi Decoction. The study measured the rat body mass and 3-hour food intake in each group and observed the pathological changes in gastric tissue using HE staining.

View Article and Find Full Text PDF

cAMP signalling is critical for memory consolidation and certain forms of long-term potentiation (LTP). Phosphodiesterases (PDEs), enzymes that degrade the second messengers cAMP and cGMP, are highly conserved during evolution and represent a unique set of drug targets, given the involvement of these enzymes in several pathophysiological states including brain disorders. The PDE4 family of cAMP-selective PDEs exert regulatory roles in memory and synaptic plasticity, but the specific roles of distinct PDE4 isoforms in these processes are poorly understood.

View Article and Find Full Text PDF

Want AI Summaries of new PubMed Abstracts delivered to your In-box?

Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!