Research highlights

The unbalance between intracellular reactive oxygen species (ROS) and the cells’ antioxidant defense systems is generally called oxidative stress and has been clearly associated with several pathologies from cardiac ischemia, neurodegenerative diseases, diabetes or cancer.

One of the main objectives of the group is the elucidation of targets for new synthetic compounds that can be useful in the overcoming of oxidative stress situations. The compounds are being synthesized in the Department of Chemistry and have shown promising properties as RONS scavengers protecting, by that way, cells from oxidative stress events and/or inducing cells’ natural defenses. We use different biochemical approaches and different mammalian cell lines (examples bellow) to screen and elucidate molecular mechanisms of action of the new compounds in development.

The biochemical studies include analysis of damage to biomolecules (lipid peroxidation, protein oxidation or DNA damage) and elucidation of signaling pathways involved in cell death. Identification of other action targets, namely study of the interaction with specific receptors is also in the scope of our studies.

Another field of research deals with phytochemicals that are worldwide accepted to be beneficial to human health and help prevent several diseases such as cancer, diabetes, neurodegenerative and cardiovascular diseases. It is also known that there is a negative correlation between the consumption of natural antioxidants and aging. However, the biochemical and molecular mechanisms of action these phytochemicals are poorly understood. So another aim of the group is to investigate the health improving properties of phytochemicals elucidate their mechanisms of action and to estimate their potential for use in functional foods and as new leads for the pharmaceutical industry. As a result of our work in the past years the mechanisms of hepatoprotection conferred by phenolic compounds have been elucidated; the antidiabetic metformin-like effect of extracts of plants of the genus Salvia has been identified, and the impact on glucose transporters characterised. In addition, we have identified anticarcinogenic properties at the levels of antigenotoxic protection, pro-apoptotic and induction of DNA repair of some natural compounds and plant extracts.

Ongoing and future Work

Oxidative stress protection by newly synthesized compounds

The work already performed, with two different classes of compounds (nitrogen structures and diarylamines) suggests a good potential for their development as drugs with pharmacological interest in diseases where oxidative stress is involved. The nitrogen compounds studied revealed to be potent ROS/RNS scavengers able to act against different deleterious stimuli. The results obtained with the catecholaminergic cell model of PC12 cells, allows some correlation with which occurs in neurodegenerative diseases. The cardiomiocyte cell model (H9c2) is useful to correlate with some cardiomiomathies. Some of the new synthetic nitrogen compounds seem to interfere in intracellular cascade of events conducting to cell death, namely at caspases mediated apoptosis. Another group of compounds, the di(hetero)arylamines, proved to be potent antioxidants effective at the nanomolar range, in cell lines and in isolated mitochondria. They also proved to protect from the collapse of mitochondrial membrane potential and interfere with other bioenergetic parameters (RCI or ADP/O ratio). This work is being supported by an FCT grant (PTDC/QUI/68382/2006).
Induction of apoptosis by estrogen receptor-mediated mechanisms. Some of the new diarylamines in study have in their structure a benzo-b-thiophene ring, like raloxifen, a drug used in the treatment of breast cancer. They proved to inhibit cell proliferation and induce apoptosis in an estrogen positive (ER+) cell line. Their potential to bind to estrogen-receptors is currently under investigation. We intend to show that these new synthetic drugs, “raloxifen-like”, might have potential to be further developed for breast cancer treatment.

Antioxidant and anti-aging effects of natural compounds

We intend to investigate the role of phenolic compounds in slowing down the aging processes by mechanisms involving improvement of the cell defensive mechanisms against damaged proteins, in particular by modulating the heat shock response and protein turnover pathways. Antidiabetic effects. We will be pursuing the investigation of antidiabetic effects of plant extracts and isolated compounds at the levels of the liver, gastrointestinal and pancreatic function. We will be focusing on the modulation of the expression of the sodium dependent glucose transporter 1 in enterocytes at transcriptional and posttranscriptional levels. Pilot studies with humans showed benefits on lipid profiles which will also be further investigated.
Anticarcinogenic effects. Work on antigenotoxic, pro-apoptotic and inhibitor of malignant progression will be continued to elucidate at the molecular level anticancer mechanisms of plant food constituents. Nutritional chemoprevention of colorectal cancer will be evaluated in AOM treated animals where aberrant crypt foci formation can be prevented as a result of protection of initiation or progression steps. In freshly isolated colonocytes and lymphocytes we will evaluate the usefulness of lymphocyte DNA damage as a predictor of in vivo protection. The role of effects on autophagy by natural compounds as inducer of cancer cell death are being examined in vitro and in vivo in a xenograft model.

Key References

Silva, J.P., Machado, V., Calhelha, R., Queiroz, M.J.R.P. and Coutinho, O.P. 2010. Di(heteroarylamines in the benzo(b)thiophene series as novel antioxidants. Drug Discoveries & Therapeutics, 4(4): 246-256.

Lima, C.F., Pereira-Wilson, C., Rattan, S.I.S. 2010. Curcumin induces heme oxygenase-1 in normal human skin fibroblasts through redox signaling: relevance for anti-aging intervention. Molecular Nutrition and Food Research (in press)

Ramos, A.A., Pereira-Wilson, C., Collins, A.R. 2010. Protective effects of Ursolic acid and Luteolin against oxidative DNA damage include enhancement of DNA repair in Caco-2 cells. Mutat Res. (in press).

Ramos, A.A., Azqueta, A., Pereira-Wilson, C., Collins, A.R. 2010. Polyphenolic compounds from Salvia species protect cellular DNA from oxidation and stimulate DNA repair in cultured human cells. J Agric Food Chem. 58(12): 7465-71.

Azevedo, M.F., Camsari, C., Sá, C.M., Lima, C.F., Fernandes-Ferreira, M. and Pereira-Wilson, C. 2010. Ursolic acid and luteolin-7-glucoside improve lipid profiles and increase liver glycogen content through glycogen synthase kinase-3. Phytotherapy Research, 24(S2): S220-S224.

Pinto-Basto, D., Silva, J.P., Queiroz, M.J., Moreno, A.J., Coutinho, O.P. 2009. Antioxidant activity of synthetic diarylamines: a mitochondrial and cellular approach. Mitochondrion, 9(1): 17-26.

Silva, J.P., Gomes, A.C., Proença, M.F. and Coutinho, O.P. 2009. Novel nitrogen compounds enhance protection and repair of oxidative DNA damage in a neuronal cell model: comparison with quercetin. Chemico-Biological Interactions, 181: 328-337.

Sá, C.M., Ramos, A.A., Azevedo, M.F., Lima, C.F., Fernandes-Ferreira, M. and Pereira-Wilson, C. 2009. Sage tea drinking improves lipid profile and antioxidant defences in humans. International Journal of Molecular Sciences, 10(9): 3937-3950.

Xavier, C.P.R., Lima, C.F., Preto, A., Seruca, R., Fernandes-Ferreira, M. and Pereira-Wilson, C. 2009. Luteolin, quercetin and ursolic acid are potent inhibitors of proliferation and inducers of apoptosis in both KRAS and BRAF mutated human colorectal cancer cells. Cancer Letters, 281(2): 162-170.

Xavier, C.P.R., Lima, C.F., Fernandes-Ferreira, M. and Pereira-Wilson, C. 2009. Salvia fruticosa, Salvia officinalis and rosmarinic acid induce apoptosis and inhibit proliferation of human colorectal cell lines: the role in MAPK/ERK pathway. Nutrition and Cancer, 61(4): 564-571.

Silva, J.P., Proença, M.F., Coutinho, O.P. 2008. Protective role of new nitrogen compounds on ROS/RNS-mediated damage to PC12 cells. Free Radical Research 42 (1): 57-69.

Ramos, A.A., Lima, C.F., Pereira, M.L., Fernandes-Ferreira, M. and Pereira-Wilson, C. 2008. Antigenotoxic effects of quercetin, rutin and ursolic acid on HepG2 cells: Evaluation by the comet assay. Toxicology Letters, 177(1): 66-73.