Metformin inhibits proliferation and migration of glioblastoma cells independently of TGF-β2

To this day, glioblastoma (GBM) remains an incurable brain tumor. Previous research has shown that metformin, an oral anti-diabetic drug, may decrease GBM cell proliferation and migration especially in brain tumor initiating cells (BTICs). As transforming growth factor β 2 (TGF-β₂) has been reported to promote high-grade glioma and is inhibited by metformin in other tumors, we explored whether metformin directly interferes with TGF-β₂-signaling. Functional investigation of proliferation and migration of primary BTICs after treatment with metformin+/?TGF-β₂ revealed that metformin doses as low as 0.01 mM metformin thrice a day were able to inhibit proliferation of susceptible cell lines, whereas migration was impacted only at higher doses. Known cellular mechanisms of metformin, such as increased lactate secretion, reduced oxygen consumption and activated AMPK-signaling, could be confirmed. However, TGF-β₂ and metformin did not act as functional antagonists, but both rather inhibited proliferation and/or migration, if significant effects were present. We did not observe a relevant influence of metformin on TGF-β₂ mRNA expression (qRT-PCR), TGF-β₂ protein expression (ELISA) or SMAD-signaling (Western blot). Therefore, it seems that metformin does not exert its inhibitory effects on GBM BTIC proliferation and migration by altering TGF-β₂-signaling. Nonetheless, as low doses of metformin are able to reduce proliferation of certain GBM cells, further exploration of predictors of BTICs' susceptibility to metformin appears justified.     

Enzymes of phenylpropanoid metabolism in the important medicinal plant Melissa officinalis L.

Lemon balm (Melissa officinalis, Lamiaceae) is a well-known medicinal plant. Amongst the biologically active ingredients are a number of phenolic compounds, the most prominent of which is rosmarinic acid. To obtain better knowledge of the biosynthesis of these phenolic compounds, two enzymes of the general phenylpropanoid pathway, phenylalanine ammonia-lyase (PAL) and 4-coumarate:coenzyme A-ligase (4CL), were investigated in suspension cultures of lemon balm. MoPAL1 and Mo4CL1 cDNAs were cloned and heterologously expressed in Escherichia coli and the enzymes characterised. Expression analysis of both genes showed a correlation with the enzyme activities and rosmarinic acid content during a cultivation period of the suspension culture. Southern-blot analysis suggested the presence of most probably two gene copies in the M. officinalis genome of both PAL and 4CL. The genomic DNA sequences of MoPAL1 and Mo4CL1 were amplified and sequenced. MoPAL1 contains one phase 2 intron of 836 bp at a conserved site, whilst Mo4CL1 was devoid of introns.     

Combined RNAi-Mediated Suppression of Rictor and EGFR Resulted in Complete Tumor Regression in an Orthotopic Glioblastoma Tumor Model

The PI3K/AKT/mTOR pathway is commonly over activated in glioblastoma (GBM), and Rictor was shown to be an important regulator downstream of this pathway. EGFR overexpression is also frequently found in GBM tumors, and both EGFR and Rictor are associated with increased proliferation, invasion, metastasis and poor prognosis. This research evaluated in vitro and in vivo whether the combined silencing of EGFR and Rictor would result in therapeutic benefits. The therapeutic potential of targeting these proteins in combination with conventional agents with proven activity in GBM patients was also assessed. In vitro validation studies were carried out using siRNA-based gene silencing methods in a panel of three commercially available human GBM cell lines, including two PTEN mutant lines (U251MG and U118MG) and one PTEN-wild type line (LN229). The impact of EGFR and/or Rictor silencing on cell migration and sensitivity to chemotherapeutic drugs in vitro was determined. In vivo validation of these studies was focused on EGFR and/or Rictor silencing achieved using doxycycline-inducible shRNA-expressing U251MG cells implanted orthotopically in Rag2M mice brains. Target silencing, tumor size and tumor cell proliferation were assessed by quantification of immunohistofluorescence-stained markers. siRNA-mediated silencing of EGFR and Rictor reduced U251MG cell migration and increased sensitivity of the cells to irinotecan, temozolomide and vincristine. In LN229, co-silencing of EGFR and Rictor resulted in reduced cell migration, and increased sensitivity to vincristine and temozolomide. In U118MG, silencing of Rictor alone was sufficient to increase this line’s sensitivity to vincristine and temozolomide. In vivo, while the silencing of EGFR or Rictor alone had no significant effect on U251MG tumor growth, silencing of EGFR and Rictor together resulted in a complete eradication of tumors. These data suggest that the combined silencing of EGFR and Rictor should be an effective means of treating GBM.     

Rye bread and synthetic bread odorants – effective trap bait and lure for German cockroaches

Bread‐in‐beer and bread‐in‐water are prevalent home recipe trap baits for attracting German cockroaches (GCRs), Blattella germanica (L.) (Dictyoptera: Blattellidae), which are significant urban pests. Our objectives were to (1) test the attractiveness of these baits, (2) study the underlying factors of GCR attraction, and (3) determine whether a blend of synthetic bread odorants could replace bread in a trap lure. In large‐arena laboratory experiments with laboratory‐reared GCR males, traps baited with rye bread not only captured eightfold more males than unbaited control traps but also most males released into bioassay arenas. Neither beer nor water enhanced the attractiveness of bread. Bread crust as a bait was more effective than bread crumbs. As Porapak Q headspace volatile extracts of rye bread attracted GCRs, all rye bread odorants in extracts were identified by gas chromatography‐mass spectrometry. Synthetic rye bread odorants and other known bread odorants were then assembled into a master blend. This master blend, and even partial blends lacking certain groups of organic volatiles such as aldehydes and ketones, proved very attractive to GCRs. We conclude that rye bread could be used as an effective bait in retainer traps or, laced with insecticide, as a food source in bait stations. A lure of synthetic bread odorants may eventually replace bread as bait, but the minimum number of essential odorants for that lure has yet to be determined.     

Do western boxelder bugs sunbathe for sanitation? Inferences from in vitro experiments

When western boxelder bugs, Boisea rubrolineata (Barber) (Hemiptera: Rhopalidae), form aggregations in warm sunlight, they release from their posterior dorsal abdominal gland an odorous blend of monoterpenes with heretofore unknown biological function. In laboratory analyses and experiments, we show that bugs in warm sunlight, but not in shade, exude and spread copious amounts of monoterpenes onto their cuticle. These monoterpenes do not serve as a pheromone, but rather as a means of sanitation. They inhibit germination of conidia of the fungal pathogen Beauveria bassiana (Bals.‐Criv.) Vuill. (Hypocreales) as well as halt the growth of germinated spores. This prophylactic defense against pathogens appears adaptive for phytophagous insects, like B. rubrolineata, that are prone to infections by microbes thriving on leaf surfaces and in the insects' overwintering microhabitat.     

The EJC factor eIF4AIII modulates synaptic strength and neuronal protein expression

Proper neuronal function and several forms of synaptic plasticity are highly dependent on precise control of mRNA translation, particularly in dendrites. We find that eIF4AIII, a core exon junction complex (EJC) component loaded onto mRNAs by pre-mRNA splicing, is associated with neuronal mRNA granules and dendritic mRNAs. eIF4AIII knockdown markedly increases both synaptic strength and GLUR1 AMPA receptor abundance at synapses. eIF4AIII depletion also increases ARC, a protein required for maintenance of long-term potentiation; arc mRNA, one of the most abundant in dendrites, is a natural target for nonsense-mediated decay (NMD). Numerous new NMD candidates, some with potential to affect synaptic activity, were also identified computationally. Two models are presented for how translation-dependent decay pathways such as NMD might advantageously function as critical brakes for protein synthesis in cells such as neurons that are highly dependent on spatially and temporally restricted protein expression.