SummaryThe synthesis of alpha-2-macroglobulin (a 2 M) is low in adult rat liver and elevated in fetal liver. During the acute-phase (AP) response it becomes significantly increased in both adult and fetal liver. In this work, the cross talk of STAT3 and NF-k B transcription factors during a 2 M gene expression was analysed. Using immunoblotting, their cellular compartmentalization was examined by comparing the cytoplasmic levels of STAT3 and NF-k B with their active equivalents, the 86 and 91 kDa isoforms and p65-subunit, respectively, in the nuclear extract and nuclear matrix. Different partitioning dynamics of the transcription factors were observed. At the level of protein-DNA interactions, studied by a 2 M promoter affinity chromatography, it was established that different ratios of promoter-binding STAT3 isoforms participated in elevated hepatic transcription in the basal state fetus and the AP-adult, but only the 91 kDa isoform in the AP-fetus. Unchanged levels of DNA-bound p65 in the control and AP-fetus suggest that it participated in constitutive transcription. The promoter-binding of p65 observed in the AP-adult suggests that it was involved in transcriptional stimulation of a 2 M expression. The selective enrichment of the AP-adult nuclear matrix with promoterbinding STAT3 disclosed the importance of this association in the induction of transcription. Protein-protein interactions were examined by co-immunoprecipitation. Interactions between the 86 kDa STAT3 isoform and p65 that were observed in the control and AP-fetus and of both the 86 and 91 kDa STAT3 isoforms with p65 in the AP-adult, suggest that protein-protein interactions were functionally connected to increased transcription. We concluded that a 2 M gene expression is driven by developmental-and AP-related mechanisms that rely on STAT3/NF-k B interplay.
We previously reported that the NO-modified form of HIV protease inhibitor Saquinavir (Saq) is a potent antitumoral agent efficient against numerous tumor cell lines in vitro and in vivo. In acute toxicity studies, doses of Saq-NO equivalent to DL100 of the parental drug were completely nontoxic. Beside direct effect on malignant cell growth, Saq-NO sensitizes certain type of cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-mediated cell death. In this study, we evaluated the effects of Saq-NO on androgen-dependent prostate cancer LNCaP. Saq-NO inhibited both the growth of LNCaP cells in vitro and in xenograft models. Suppression of tumor growth was accompanied with cell cycle arrest in G 0/G 1 phase and established a persistent inhibition of proliferation. Furthermore, Saq-NO reverted sensitivity of LNCaP cells to TRAIL but not to TNF. Treatment of cells with Saq-NO induced transient upregulation of Akt and ERK1/2. This, however, did not represent the primary mode of action of Saq-NO, as elimination with specific inhibitors did not compromise the chemotherapic efficacy of the drug. However, permanent abrogation of phosphorylation of the S6 protein, which is the downstream target of both signaling pathways, was observed. Diminished S6 phosphorylation was associated with re-established sensitivity to TRAIL and reduction of X-linked inhibitor of apoptosis protein (XIAP). In summary, NO modification of Saq led to a new chemical entity with stronger and more pleiotropic antitumor activity than the parental drug.
The biggest drawback of a current diabetes therapy is the treatment of the consequences not the cause of the disease. Regardless of the diabetes type, preservation and recovery of functional pancreatic beta cells stands as the biggest challenge in the treatment of diabetes. Free radicals and oxidative stress are among the major mediators of autoimmune destruction of beta cells in type 1 diabetes (T1D) or beta cell malfunction and death provoked by glucotoxicity and insulin resistance in type 2 diabetes (T2D). Additionally, oxidative stress reduces functionality of beta cells in T2D by stimulating their de-/trans-differentiation through the loss of transcription factors critical for beta cell development, maturity and regeneration. This review summarizes up to date clarified redox-related mechanisms involved in regulating beta cell identity and death, underlining similarities and differences between T1D and T2D. The protective effects of natural antioxidants on the oxidative stress-induced beta cell failure were also discussed. Considering that oxidative stress affects epigenetic regulatory mechanisms involved in the regulation of pancreatic beta cell survival and insulin secretion, this review highlighted huge potential of epigenetic therapy. Special attention was paid on application of the state-of-the-art CRISPR/Cas9 technology, based on targeted epigenome editing with the purpose of changing the differentiation state of different cell types, making them insulin-producing with ability to attenuate diabetes. Clarification of the above-mentioned mechanisms could provide better insight into diabetes etiology and pathogenesis, which would allow development of novel, potentially more efficient therapeutic strategies for the prevention or reversion of beta cell loss.
Background Poly(ADP-ribosyl)ation (PARylation), a posttranslational modification introduced by PARP-1 and PARP-2, has first been implicated in DNA demethylation due to its role in base excision repair. Recent evidence indicates a direct influence of PARP-dependent PARylation on TET enzymes which catalyse hydroxymethylation of DNA—the first step in DNA demethylation. However, the exact nature of influence that PARylation exerts on TET activity is still ambiguous. In our recent study, we have observed a negative influence of PARP-1 on local TET-mediated DNA demethylation of a single gene and in this study, we further explore PARP–TET interplay. Results Expanding on our previous work, we show that both TET1 and TET2 can be in vitro PARylated by PARP-1 and PARP-2 enzymes and that TET1 PARylation negatively affects the TET1 catalytic activity in vitro. Furthermore, we show that PARylation inhibits TET-mediated DNA demethylation at the global genome level in cellulo. Conclusions According to our findings, PARP inhibition can positively influence TET activity and therefore affect global levels of DNA methylation and hydroxymethylation. This gives a strong rationale for future examination of PARP inhibitors' potential use in the therapy of cancers characterised by loss of 5-hydroxymethylcytosine.
The expression of CYP1A, a biomarker for the presence of xenobiotic compounds, was examined in three fish species from the wider vicinity of Bar harbor in winter and spring. Induction of CYP1A was observed in winter and increased further in spring. Several PCBs were found in seawater in winter. They decreased below the limit of detection in spring, when the PAH fluorene was detected. It is concluded that the constant presence of CYP1A expression is probably due to pollutants in the environment, whereas increased expression of CYP1A in spring results from exposure of the fish to fluorene.
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