Cyclic AMP phosphodiesterases in human lymphocytes

Abstract

Summary. The function of lymphocytes, like platelets, has been shown to be inhibited by agents which increase intracellular cyclic AMP. Two high-affinity cAMP phosphodiesterases (PDEs), the cyclic GMP-inhibited cAMP phosphodiesterase, PDE3, and the cAMP-specific phosphodiesterase PDE4, are known to regulate cAMP concentration in haemopoietic cells by degrading cAMP to AMP. We characterized the relative contribution of the two PDEs to total lymphocyte PDE activity. We then determined which of the different gene products, PDE3A, typical of myocardium and platelets, or PDE3B, typical of adipocytes, were present in lymphocytes. The PDE3-specific inhibitor, milrinone, and the PDE4 inhibitor, rolipram, suppressed hydrolysis by 70% and 30% respectively, which indicated that both PDE4 and PDE3were present, and that PDE3 was predominant. RT-PCR yields the expected size fragment for the primer pair PDE3B and not for PDE3A. The DNA sequence obtained had > 95% identity with PDE3B. PDE3B appears to be the major cAMP PDE in lymphocytes. In contrast to human platelets, human lymphocytes appear to contain the PDE3B subtype. Since PDE3B in adipocytes is subject to hormonal regulation, lymphocytes may be similarly modulated. Understanding the role of cAMP regulation and the involvement of cAMP in lymphocyte function may have important implications in drug development.Keywords: phosphodiesterase, lymphocytes, cyclic AMP, milrinone, rolipram.Cyclic AMP, an important second messenger in cells, is maintained at steady state levels as a result of formation and destruction by adenylate cyclase and cAMP phosphodiesterases (PDEs) respectively. Currently, the group of PDEs isolated from mammalian tissues is divided into at least seven distinct classes, based on a variety of biochemical characteristics including their substrate specificity, response to selective inhibitors, mode of regulation and kinetic characteristics (Beavo et al, 1994). These classes appear to be coded for by related but distinct genes yet they share a conserved region of about 250 amino acids at the C terminal end (Charbonneau, 1990). This region demonstrates a homology of about 28-40% between classes and contains the catalytic domain (Charbonneau, 1990). The N terminal region of each class appears to be distinct and may contain the regulatory domain. Each class has at least two isotypes coded for by different genes or the product of differential mRNA splicing. Lymphocytes contain two high-affinity cAMP PDEs, the cyclic GMP-inhibited cAMP phosphodiesterase or PDE3 and the cAMP-specific phosphodiesterase or PDE 4 (Robicsek et al, 1991). In monocytes and neutrophils PDE4 predominates (Wright et al, 1990) whereas in platelets most of PDE activity is PDE3A (Degerman et al, 1994;Cheung et al, 1996). Two rat and two human enzymes have been cloned from heart (PDE3A) (Meacci et al, 1992) and adipocyte (PDE3B) (Taira et al, 1993), and their results show that the PDE3 class appears to have a conserved region of 44 amino acids at the C terminal end distinct from the catalytic re...

Inhibition of PDE4 phosphodiesterase activity induces growth suppression, apoptosis, glucocorticoid sensitivity, p53, and p21WAF1/CIP1 proteins in human acute lymphoblastic leukemia cells

“…In addition to PDE4, lymphocytes express PDE3 activity. 39 However, the PDE3 inhibitor trequinsin caused little effect on CEM cell growth, even at concentrations well above the inhibitory concentration of 50% (IC 50 ) for PDE3 activity in vitro of 0.3 nM ( Figure 1G). Similarly, the PDE1-specific inhibitor vinpocetine induced no growth suppression of CEM cells until the drug rose to concentrations at which PDE subclass selectivity is lost (IC 50 for PDE1 activity in vitro ϭ 20 M; Figure 1H).…”

“…[30][31][32] Normal lymphocytes and several lymphoid leukemia cell lines express at least 2 high-affinity cAMP PDEs, cyclic guanosine monophosphate (cGMP)-inhibited cAMP PDE (PDE3) and the cAMP-specific PDE (PDE4), whereas PDE4 is dominant in monocytes and neutrophils and PDE3 in platelets. [33][34][35][36][37][38][39][40] Vinpocetine, an inhibitor of PDE1, induces apoptosis in ALL cells. 41 Rolipram, an inhibitor of PDE4 enzymes, selectively induces apoptosis in CLL patient cells in vitro.…”

Among circulating hematopoietic cells, B-CLL uniquely expresses functional EPAC1, but EPAC1-mediated Rap1 activation does not account for PDE4 inhibitor-induced apoptosis

“…At a minimum, lymphoid cells express PDE1B, PDE3B, PDE4A, B, and D, and PDE7A; all enzymes that can catabolize 3Ј:5Ј cyclic adenosine monophosphate (cAMP). [1][2][3][4][5] In studies of the mature B-cell malignancy B-cell chronic lymphocytic leukemia (B-CLL), we and others have found that both nonspecific PDE inhibitors such as theophylline and the PDE4-specific inhibitor rolipram induce apoptosis in leukemic cells over a 48-to 72-hour period. [6][7][8] While rolipram also induces apoptosis in peripheral B cells, albeit superimposed on a high normal basal apoptotic rate, the same agent has little or no apoptotic effect on peripheral T cells.…”

Effects of Forskolin on Kupffer Cell Production of Interleukin-10 and Tumor Necrosis Factor Alpha Differ from Those of Endogenous Adenylyl Cyclase Activators: Possible Role for Adenylyl Cyclase 9

“…Following stimulation, intracellular cAMP levels are rapidly down-regulated by phosphodiesterases that transform cAMP into inactive AMP (29). A range of downstream effects of the cAMP signal have been described in different immune cells, regulated by strict localization of the signal (reviewed in reference 36).…”