PMC

A172 modulates transcription of the genes coding for pro-inflammatory cytokines (TNF-α, INF-γ and IL-1β) and induces transcription of the genes coding for anti-inflammatory cytokines (Il-4 and Il-6).

Engagement of platelet-derived growth factor receptor (PDGF-R) is critical for PDGF-BB induced monocyte chemoattractant protein 1 (MCP-1) expression in astrocytes. (A) Human primary as well as A172 primary astrocytes express both PDGF-α and PDGF-β receptors. Untreated human primary and A172 astrocytes were subjected to flow cytometric analyses using antibodies against human PDGF-αR (CD140α) and PDGF-βR (CD140β). (B,C) PDGF-BB-mediated induction of MCP-1 mRNA and protein expression in human primary and A172 astrocytes. (B) Human primary and A172 astrocytes were exposed to PDGF-BB for 6 h. Total RNA isolated from human A172 astrocytes was subjected to real-time reverse-transcriptase polymerase chain reaction (RT-PCR) analysis using primers for human MCP-1. (C) Supernatant fluid from astrocytes treated with PDGF-BB for 24 h were assessed for release of chemokine MCP-1 using the human MCP-1 enzyme-linked immunosorbent assay (ELISA) kit. (D) Total RNA from A172 cells transfected with either nonsense (Non) or PDGF-Rβ short interfering (si)RNAs was subjected to RT-PCR analysis using primers specific for PDGF-Rβ. (E) siRNA, but not Non siRNA inhibited PDGF-BB-mediated induction of MCP-1 RNA. (F) Supernatants from A172 cells transfected with either Non or siRNAs were subjected to MCP-1 ELISA assay. PDGF-Rβ siRNA, but not Non siRNA inhibited PDGF-BB-mediated induction of MCP-1 protein. **P <0.01, ***P <0.001 versus control group.

The strain A172 grows in larger clumps (A) compared to the strain A170 (B). Bacteria were grown in liquid culture and collected for microscopic examination at the early stationary phase. (C) The A172 strains displays also a slower growth rate, compared to A170.

(A) Logarithmically growing A172, and T98G cells were incubated with varying concentrations of AG490 (μM) with or without 200 nM UCN-01 for 24 h. The cells were lysed, and proteins were separated by SDS-PAGE and probed with the indicated antibodies. Phosphorylation of BAD has been noted to inhibit apoptotic signaling. BAD phosphorylation at Ser¹¹², critical for BAD function, was unaffected by either UCN-01 or AG490 in the T98G cell line. Opposing effects were observed in A172. There is no phosphorylation of BAD at Ser¹³⁶ either in A172 or T98G cells. (B). Logarithmically growing T98G and A172 cells were pretreated with either LY294002 (25 μM - a PI3K/Akt specific inhibitor) or with U0126 (25 μM- a MEK-specific inhibitor) 2 h prior to AG490 (25 μM) treatment for 24 h. Equal amounts of proteins were separated by SDS-PAGE and probed with p-BAD (Ser¹¹²) antibody. The blots were subsequently stripped and reprobed with antibodies against total BAD. U0126 potently blocked AG490-induced Ser112 phosphorylation of BAD in A172 cells, no effect was observed with administration of LY294002. This observation suggests that AG490 may trigger a MAPK cascade to phosphorylate and inactivate BAD in p53 wild type but not in mutants.