Medizin
Refine
Document Type
- Conference Proceeding (2) (remove)
Language
- English (2) (remove)
Has Fulltext
- yes (2)
Is part of the Bibliography
- no (2)
Institute
- Medizin (2)
- Georg-Speyer-Haus (1)
- Pharmazie (1)
Cytoprotective functions of amyloid precursor protein family members in stress signaling and aging
(2013)
Poster presentation: Molecular Neurodegeneration: Basic biology and disease pathways Cannes, France. 10-12 September 2013.
Background: The amyloid precursor protein (APP) is processed via two different metabolic pathways: the amyloidogenic and the non-amyloidogenic pathway, the latter of which leading to generation of the secreted N-terminal APP fragment sAPPα [1]. Previous studies from our group suggest that sAPPα exerts potent neuroprotective effects and inhibits stress-triggered cell death via modulation of gene expression, as well as by antagonizing different types of neurotoxic stress [2]. It was also observed that the biochemical processing of APP is downregulated during aging which in turn reduced the secretion of sAPPα [3]. Based on these observations, we have studied the potential physiological function of sAPPα/APP and APLPs (APP like proteins) on the regulation of age-associated, stress induced signaling pathways, apoptosis and senescence.
Materials and methods: SH-SY5Y, PC12, IMR90 cells were used as cellular models. Depletion of APP, APLP1 (APP like protein 1) and APLP2 (APP like protein 2) in SH-SY5Y cells was achieved by stable lentiviral knockdown. To analyze the protective function of sAPPα, we have used conditioned supernatants of wild type APP overexpressing HEK cells and recombinant His-tagged sAPPα purified from yeast. The cells were treated with sAPPα prior to the addition of different stress stimuli (MG132, epoxomicin, UV, H2O2) after which cell death, gene expression and senescence were analyzed by MTT assays, caspase activity assays, Western blots and X-Gal staining respectively.
Results: Our data show that sAPPα can antagonize premature senescence induced by repetitive short term induction of proteasomal stress in IMR-90 cells and apoptosis triggered by prolonged proteasomal stress and other death stimuli in PC12, SH-SY5Y and IMR90 cells which was accompanied by a sAPPα-dependent inhibition of the JNK stress signaling pathway. In contrast, no significant changes in cell viability and apoptosis were observed when APP knockdown cells were pretreated with sAPPα.
Conclusions: Our observations suggest that sAPPα can antagonize both apoptosis and cellular senescence and requires expression of holo-APP to mediate its cytoprotective effects. They also support the notion that the physiological function of APP is linked to modulation of neuronal and brain aging.
Objective: Establishment of an immunocompetent mouse model representing the typical progressive stages observed in malignant human gliomas for the in vivo evaluation of novel target-specific regimens.
Methods: Isolated clones from tumours that arose spontaneously in GFAP-v-src transgenic mice were used to develop a transplantable brain tumour model in syngeneic B6C3F1 mice. STAT3 protein was knocked down by infection of tumour cells with replication-defective lentivirus encoding STAT3-siRNA. Apoptosis is designed to be induced by soluble recombinant TRAIL + chemical Bcl-2/Bcl-xL inhibitors.
Results: Striatal implantation of 105 mouse tumour cells resulted in the robust development of microscopically (2 – 3 mm) infiltrating malignant gliomas. Immunohistochemically, the gliomas displayed the astroglial marker GFAP and the oncogenic form of STAT3 (Tyr-705-phosphorylated) which is found in many malignancies including gliomas. Phosphorylated STAT3 was particularly prominent in the nucleus but was also found at the plasma membrane of peripherally infiltrating glioma cells. To evaluate the role of STAT3 in tumour progression, we stably expressed siRNA against STAT3 in several murine glioma cell lines. The effect of STAT3 depletion on proliferation, invasion and survival will be first assessed in vitro and subsequently after transplantation in vivo. Upstream and downstream components of the STAT3 signalling pathway as well as possible non-specific side effects of STAT3-siRNA expression after lentiviral infection will be examined, too.
Conclusions: Its high rate of engraftment, its similarity to the malignant glioma of origin, and its rapid locally invasive growth should make this murine model useful in testing novel therapies for malignant gliomas.