Silencing the PTEN gene is protective against neuronal death induced by human immunodeficiency virus type 1 Tat.

Neurons are targets of toxicity induced by the human immunodeficiency virus (HIV)-1 protein Tat (transactivator of transcription). Exposure to Tat increases [Ca(2+)](i) in striatal neurons and activates multiple cell death pathways. In earlier studies the authors showed that Tat activated both caspase-3 and endonuclease-G, a caspase-independent effector of apoptosis, and that Tat-induced neurotoxicity was not attenuated by a caspase-3 inhibitor. Because Tat activates multiple, parallel death pathways, the authors attempted to reduce Tat-induced neurotoxicity by manipulating signaling pathways upstream of mitochondrial apoptotic events. PTEN (phosphatase and tensin homolog deleted on chromosome 10), a negative regulator of Akt/PKB (protein kinase B) phosphorylation, was chosen as a target for silencing. Akt/PKB activity directs multiple downstream pathways mediated by GSK3beta, BAD, forkhead transcription factors, nuclear factor kappa B (NFkappaB), and others, in a manner that promotes proliferation and survival. Striatal neurons were nucleofected with short interfering RNA (siRNA) vectors targeting PTEN, or a negative-control siRNA. Although Tat(1-86) significantly increased the death of neurons transfected with control construct by 72 h, PTEN-silenced neurons were completely protected. These findings indicate that Akt is a critical intermediary in the direct neurotoxicity induced by HIV-1 Tat, and identify Akt regulation as a possible therapeutic strategy for Tat-induced neurotoxicity in HIV encephalitis (HIVE).

Epidermal growth factor promotes oligodendrocyte process formation and regrowth after injury.

Oligodendrocytes (OLs) form myelin within the central nervous system and are targets in numerous demyelinating diseases and injuries. OLs grown in culture maintain the developmental timetable which occurs in vivo and mature into cells with a relatively normal phenotype. In this study, cultured cells are used to test whether EGF can modulate process formation in OLs both before and after transection injury. EGF had no effect on the formation of new processes by OLs at any stage of development. To test the effect of EGF on process outgrowth after injury, mature OLs were selected and injured by laser transection of a single process, then imaged at 24-h intervals for 120 h. EGF promoted the recovery and regrowth of injured processes and also significantly increased outgrowth in uninjured processes. As well, it increased the number of new sprouts formed by OLs after injury. Results suggest that the effects of EGF on process outgrowth are a consequence of EGF interaction with a signaling pathway that is specifically activated within injured OLs. The potent effect of EGF on OL process formation after an injury suggests that modulation of the signaling pathways involved might provide a mechanism to promote remyelination.

Oxidant-induced hypertrophy of A549 cells is accompanied by alterations in eukaryotic translation initiation factor 4E and 4E-binding protein-1.

Control of protein synthesis resides at the level of eukaryotic translation initiation (eIF) complex formation. Complex formation is regulated by the mRNA cap-binding protein, eIF4E, whose activity is influenced by phosphorylation and binding to 4E-binding protein 1 (4E-BP1). To provide a link between alterations in protein synthesis and the pathogenesis of oxidant-mediated lung disease, we investigated the effect of hydrogen peroxide (H2O2) on actively growing A549 cells. Cells were exposed to 200 or 400 microM H2O2 for 4 h and then assessed for changes in proliferation, protein synthesis, and eIF4E and 4E-BP1 status over 72 h. We found that both concentrations of H2O2 inhibited [3H]thymidine incorporation and cell division while inducing a G2/M-predominant growth arrest within 24 h. In addition, H2O2 increased cell size, [3H]leucine incorporation/cell, and total cell protein. Although time had little effect on eIF4E and 4E-BP1 expression and phosphorylation state of control cells, H2O2 induced a 2- to 3-fold increase in eIF4E and 4E-BP1 expression, a 5-fold increase in eIF4E phosphorylation, and a shift in the distribution of 4E-BP1 phosphorylation favoring lesser phosphorylated forms. These findings suggest that oxidant-mediated alterations in protein synthesis and cell morphology occur in concert with changes in factors known to regulate translation kinetics.