Nitensidine B affects proteins of the glycolytic pathway and induces apoptosis in cervical carcinoma cells immortalized by HPV16.

BACKGROUND: Cervical cancer, the fourth most common type of cancer among women worldwide, accounts for approximately 12% of all types of malignancies that affect women. Natural products have contributed significantly to the development of modern therapies; approximately 70% of the drugs available for chemotherapy are naturally based products. PURPOSE: The purpose of this study was to examine the biological activities of nitensidine B (NTB), a guanidinic alkaloid isolated from the leaves of Pterogyne nitens Tul. (Fabaceae) in a cervical cancer cell line. METHODS: In vitro experiments were performed using cervical carcinoma cells immortalized by human papillomavirus type 16 (HPV16, SiHa cells), since epidemiological and molecular studies have demonstrated robust associations between the etiologies of cervical cancer and HPV infection. Cytotoxicity as well as the effect of NTB treatment on intracellular signals of apoptosis, fragmentation of internucleosomal DNA via terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL), and levels of apoptosis effectors (Caspase 3/7) were evaluated. In addition, differential proteomic analysis (iTRAQ) and protein validation using western blot were performed. RESULTS: The cytotoxicity of NTB treatment in the SiHa cell line was concentration-dependent, with the minimum inhibitory concentration of 50% of the cells of 40.98 µM. In the TUNEL assay, SiHa cell apoptosis with 3/7 caspase activation was reported at 12 h following treatment. Differential proteomic analysis by iTRAQ demonstrated that proteins of the glycolytic pathway, aldolase A, alpha-enolase, pyruvate kinase, and glyceraldehyde 3-phosphate dehydrogenase were underexpressed. CONCLUSION: These results indicated that NTB could play a role in decreasing glycolysis . Since tumor cells prefer the glycolytic pathway to generate energy, these findings suggest that NTB may be a reliable model for the study of human cervical cancer cell lines immortalized by HPV16, however more experiments can be performed.

Emodin, Physcion, and Crude Extract of Rhamnus sphaerosperma var. pubescens Induce Mixed Cell Death, Increase in Oxidative Stress, DNA Damage, and Inhibition of AKT in Cervical and Oral Squamous Carcinoma Cell Lines.

There have been few studies on the pharmacological properties of Rhamnus sphaerosperma var. pubescens, a native Brazilian species popularly known as "fruto-de-pombo." The aim of this study was to investigate the scavenging capacity of emodin, physcion, and the ethanolic crude extract of Rhamnus sphaerosperma var. pubescens against reactive oxygen and nitrogen species, as well as their role and plausible mechanisms in prompting cell death and changes in AKT phosphorylation after cervical (SiHa and C33A) and oral (HSC-3) squamous cell carcinoma treatments. Emodin was shown to be the best scavenger of NO• and O2•-, while all samples were equally effective in HOCl/OCl- capture. Emodin, physcion, and the ethanolic extract all exhibited cytotoxic effects on SiHa, C33A, HSC-3, and HaCaT (immortalized human keratinocytes, nontumorigenic cell line), involving mixed cell death (apoptosis and necrosis) independent of the caspase activation pathway. Emodin, physcion, and the ethanolic extract increased intracellular oxidative stress and DNA damage. Emodin decreased the activation of AKT in all tumor cells, physcion in HSC-3 and HaCaT cells, and the ethanolic extract in C33A and HaCaT cells, respectively. The induction of cancer cell death by emodin, physcion, and the ethanolic crude extract of Rhamnus sphaerosperma var. pubescens was related to an increase in intracellular oxidative stress and DNA damage and a decrease in AKT activation. These molecules are therefore emerging as interesting candidates for further study as novel options to treat cervical and oral carcinomas.

Photodynamic therapy combined to cisplatin potentiates cell death responses of cervical cancer cells.

BACKGROUND: Photodynamic therapy (PDT) has proven to be a promising alternative to current cancer treatments, especially if combined with conventional approaches. The technique is based on the administration of a non-toxic photosensitizing agent to the patient with subsequent localized exposure to a light source of a specific wavelength, resulting in a cytotoxic response to oxidative damage. The present study intended to evaluate in vitro the type of induced death and the genotoxic and mutagenic effects of PDT alone and associated with cisplatin. METHODS: We used the cell lines SiHa (ATCC® HTB35™), C-33 A (ATCC® HTB31™) and HaCaT cells, all available at Dr. Christiane Soares' Lab. Photosensitizers were Photogem (PGPDT) and methylene blue (MBPDT), alone or combined with cisplatin. Cell death was accessed through Hoechst and Propidium iodide staining and caspase-3 activity. Genotoxicity and mutagenicity were accessed via flow cytometry with anti-gama-H2AX and micronuclei assay, respectively. Data were analyzed by one-way ANOVA with Tukey's posthoc test. RESULTS: Both MBPDT and PGPDT induced caspase-independent death, but MBPDT induced the morphology of typical necrosis, while PGPDT induced morphological alterations most similar to apoptosis. Cisplatin predominantly induced apoptosis, and the combined therapy induced variable rates of apoptosis- or necrosis-like phenotypes according to the cell line, but the percentage of dead cells was always higher than with monotherapies. MBPDT, either as monotherapy or in combination with cisplatin, was the unique therapy to induce significant damage to DNA (double strand breaks) in the three cell lines evaluated. However, there was no mutagenic potential observed for the damage induced by MBPDT, since the few cells that survived the treatment have lost their clonogenic capacity. CONCLUSIONS: Our results elicit the potential of combined therapy in diminishing the toxicity of antineoplastic drugs. Ultimately, photodynamic therapy mediated by either methylene blue or Photogem as monotherapy or in combination with cisplatin has low mutagenic potential, which supports its safe use in clinical practice for the treatment of cervical cancer.

Piperine, a natural bioenhancer, nullifies the antidiabetic and antioxidant activities of curcumin in streptozotocin-diabetic rats.

Knowing that curcumin has low bioavailability when administered orally, and that piperine has bioenhancer activity by inhibition of hepatic and intestinal biotransformation processes, the aim of this study was to investigate the antidiabetic and antioxidant activities of curcumin (90 mg/kg) and piperine (20 or 40 mg/kg), alone or co-administered, incorporated in yoghurt, in streptozotocin (STZ)-diabetic rats. The treatment for 45 days of STZ-diabetic rats with curcumin-enriched yoghurt improved all parameters altered in this experimental model of diabetes: the body weight was increased in association with the weight of skeletal muscles and white adipose tissues; the progressive increase in the glycemia levels was avoided, as well as in the glycosuria, urinary urea, dyslipidemia, and markers of liver (alanine and aspartate aminotransferases and alkaline phosphatase) and kidney (urinary protein) dysfunction; the hepatic oxidative stress was decreased, since the activities of the antioxidant enzymes superoxide dismutase, catalase and glutathione peroxidase were increased, and the levels of malondialdehyde and protein carbonyl groups were reduced. The dose of 20 mg/kg piperine also showed antidiabetic and antioxidant activities. The treatment of STZ-diabetic rats with both curcumin and 20 mg/kg piperine in yoghurt did not change the antidiabetic and antioxidant activities of curcumin; notably, the treatment with both curcumin and 40 mg/kg piperine abrogated the beneficial effects of curcumin. In addition, the alanine aminotransferase levels were further increased in diabetic rats treated with curcumin and 40 mg/kg piperine in comparison with untreated diabetic rats. These findings support that the co-administration of curcumin with a bioenhancer did not bring any advantage to the curcumin effects, at least about the antidiabetic and antioxidant activities, which could be related to changes on its biotransformation.