Potent and selective activity of a new carbocyclic nucleoside analog (carbovir: NSC 614846) against human immunodeficiency virus in vitro

Carbocyclic 2',3'-didehydro-2',3'-dideoxyguanosine (Carbovir: NSC 614846), a novel nucleoside analog, emerged as a potent and selective anti-HIV agent from a large screening program conducted by the National Cancer Institute and its contractors. Its hydrolytic stability and its ability to inhibit the infectivity and replication of HIV in T-cells at concentrations of approximately 200- to 400-fold below toxic concentrations make carbovir a top-priority candidate for development as a potential antiretroviral agent in the treatment of AIDS patients.     

Effect of nifedipine on the contractile function of the rat diaphragm in vitro

The isometric contractile response of the directly-stimulated rat diaphragm was studied before and following addition of the calcium channel blocker, nifedipine. Nifedipine (10 micrograms/ml and 30 micrograms/ml bath concentrations) significantly increased isometric force output during twitch and unfused tetanic stimulation. Force potentiation during unfused tetanic stimulation was equivalent during either high or low voltage stimulation. Nifedipine had no effect on the time to peak force, half relaxation time, or relaxation time during twitch stimulation; thus, both activation and relaxation rates were increased. The force potentiating actions of nifedipine persisted in a calcium-free bathing solution and were enhanced by d-tubocurarine. In contrast to the force enhancing effects found with twitch and unfused tetanic stimulation, nifedipine caused a small but significant reduction in isometric force during maximal fused tetanic stimulation. It is concluded that the force potentiating effects of nifedipine on rat diaphragm are not due to fiber recruitment, enhancement of neuromuscular excitation, or altered inward trans-sarcolemmal calcium flux, but may result from a direct effect of the drug on the rate of activation of the contractile apparatus.     

Reduced IRF4 expression promotes lytic phenotype in Type 2 EBV-infected B cells

Humans are infected with two types of EBV (Type 1 (T1) and Type 2 (T2)) that differ substantially in their EBNA2 and EBNA 3A/B/C latency proteins and have different phenotypes in B cells. T1 EBV transforms B cells more efficiently than T2 EBV in vitro, and T2 EBV-infected B cells are more lytic. We previously showed that both increased NFATc1/c2 activity, and an NFAT-binding motif within the BZLF1 immediate-early promoter variant (Zp-V3) contained in all T2 strains, contribute to lytic infection in T2 EBV-infected B cells. Here we compare cellular and viral gene expression in early-passage lymphoblastoid cell lines (LCLs) infected with either T1 or T2 EBV strains. Using bulk RNA-seq, we show that T2 LCLs are readily distinguishable from T1 LCLs, with approximately 600 differentially expressed cellular genes. Gene Set Enrichment Analysis (GSEA) suggests that T2 LCLs have increased B-cell receptor (BCR) signaling, NFAT activation, and enhanced expression of epithelial-mesenchymal-transition-associated genes. T2 LCLs also have decreased RNA and protein expression of a cellular gene required for survival of T1 LCLs, IRF4. In addition to its essential role in plasma cell differentiation, IRF4 decreases BCR signaling. Knock-down of IRF4 in a T1 LCL (infected with the Zp-V3-containing Akata strain) induced lytic reactivation whereas over-expression of IRF4 in Burkitt lymphoma cells inhibited both NFATc1 and NFATc2 expression and lytic EBV reactivation. Single-cell RNA-seq confirmed that T2 LCLs have many more lytic cells compared to T1 LCLs and showed that lytically infected cells have both increased NFATc1, and decreased IRF4, compared to latently infected cells. These studies reveal numerous differences in cellular gene expression in B cells infected with T1 versus T2 EBV and suggest that decreased IRF4 contributes to both the latent and lytic phenotypes in cells with T2 EBV.     

Phospholamban decreases the energetic efficiency of the sarcoplasmic reticulum Ca pump

We tested the hypothesis that increased Sarcoplasmic reticulum (SR) Ca content ([Ca](SRT)) in phospholamban knockout mice (PLB-KO) is because of increased SR Ca pump efficiency defined by the steady-state SR [Ca] gradient. The time course of thapsigargin-sensitive ATP-dependent (45)Ca influx into and efflux out of cardiac SR vesicles from PLB-KO and wild-type (WT) mice was measured at 100 nm free [Ca]. We found that PLB decreased the initial SR Ca uptake rate (0.13 versus 0.31 nmol/mg/s) and decreased steady-state (45)Ca content (0.9 versus 4.1 nmol/mg protein). Furthermore, at similar total SR [Ca], the pump-mediated Ca efflux rate was higher in WT (0.065 versus 0.037 nmol/mg/s). The pump-independent leak rate constant (k(leak)) was also measured at 100 nm free [Ca]. The results indicate that k(leak) was < 1% of pump-mediated backflux and was not different among nonpentameric mutant PLB (PLB-C41F), WT pentameric PLB (same expression level), and PLB-KO. Therefore differences in passive SR Ca leak cannot be the cause of the higher thapsigargin-sensitive Ca efflux from the WT membranes. We conclude that the decreased total SR [Ca] in WT mice is caused by decreased SR Ca influx rate, an increased Ca-pump backflux, and unaltered leak. Based upon both thermodynamic and kinetic analysis, we conclude that PLB decreases the energetic efficiency of the SR Ca pump.     

Potential role of muscarinic receptors in schizophrenia

The role of muscarinic receptors in schizophrenia was investigated using the muscarinic agonist PTAC. PTAC was highly selective for muscarinic receptors, was a partial agonist at muscarinic M2/M4 receptors and an antagonist at M1, M3 and M5 receptors. PTAC was highly active in animal models predictive of antipsychotic behavior including inhibition of conditioned avoidance responding in rats and blockade of apomorphine-induced climbing behavior in mice. d-Amphetamine-induced Fos expression in rat nucleus accumbens was inhibited by PTAC, thus directly demonstrating the ability of PTAC to modulate DA activity. In electrophysiological studies in rats, PTAC acutely inhibited the firing of A10 DA cells and after chronic administration decreased the number of spontaneously firing DA cells in the A10 brain area. However, PTAC did not appreciably alter the firing of A9 DA cells. Thus, PTAC appears to have novel antipsychotic-like activity and these data suggest that muscarinic compounds such as PTAC may represent a new class of antipsychotic agents.