Elimination of Human Papillomavirus and Cervical Pathological Microbiota with Photodynamic Therapy in Women from Mexico City with Cervical Intraepithelial Neoplasia I.

Cervical carcinoma (CC) is the second cause of cancer death in Mexican women. It starts with premalignant lesions known as Intraepithelial Cervical Neoplasia (CIN) that can develop due to infection by Human Papillomavirus (HPV) and other microorganisms. Current CIN therapy involves invasive methods that affect cervix integrity and fertility; we propose the use of photodynamic therapy (PDT) as a strategy with few side effects. In this work, the effectiveness of PDT for CIN I, HPV and pathogenic vaginal microbiota elimination in 29 women of Mexico City with CIN I, CIN I + HPV and HPV diagnosis was determined. After 6 months of PDT application, HPV infection was eliminated in 100% of the patients (P < 0.01), CIN I + HPV in 64.3% (P < 0.01) and CIN I in 57.2% (P > 0.05). PDT also eliminated pathogenic microorganisms: Chlamydia trachomatis in 81% of the women (P < 0.001) and Candida albicans in 80% (P < 0.05), without affecting normal microbiota since Lactobacillus iners was eliminated only in 5.8% of patients and the opportunistic Gardnerella vaginalis in 20%. These results show that PDT was highly effective in eradicating HPV and pathogenic microorganisms, suggesting that PDT is a promising therapy for cervical infections.

Aloe vera and Fermented Extracts Exhibit an Anti-Inflammatory Effect on Human Glioblastoma/Astrocytoma U373 MG Cells.

The anti-inflammatory effects of Aloe vera (AV), polysaccharide extract from AV, and extracts from the digestion and colonic fermentation of AV were evaluated using an immortal astrocyte cell line (U373 MG) that develops a neuro-inflammatory profile. Cell viability and inflammatory markers were assessed after stimulation with neuropeptide substance P (SP) that activates the pro-inflammatory MAPK (mitogen-activated protein kinase) pathway. Cell viability after SP treatment was over 50% at 10 mg/mL AV, polysaccharide extract from AV, extracts from the digestion: non-digestible fraction of AV non-digestible fraction of polysaccharide extract from AV and extracts from the colonic fermentation of AV, at 4 and 24 h. Moreover, cells exposed to SP and treated with these extracts showed lower protein-activated ERK1/ERK2 (extracellular signal-regulated kinases 1 and 2), p38 (MAPK protein p38), and NFκB (nuclear factor κB) levels with respect to the SP-stimulated control. Inflammation inhibition by extracts of polysaccharide extract from AV and extracts from the colonic fermentation of AV, at 24 h in the study of p38 was not as statistically significant in ERK1/ERK2 and NFκB. Nevertheless, there was a significant decrease (p < 0.05) in pro-inflammatory cytokine IL-6 levels in cells exposed to all samples. Samples with extracts from the colonic fermentation of AV, at 4 or 24 h showed the highest inhibitory effect on IL-6 production.

Immunogenic analysis of epitope-based vaccine candidate induced by photodynamic therapy in MDA-MB-231 triple-negative breast cancer cells.

BACKGROUND: Photodynamic therapy (PDT) is used to treat tumors through selective cytotoxic effects. PDT induces damage-associated molecular patterns (DAMPs) expression, which can cause an immunogenic death cell (IDC). In this study we identified potential immunogenic epitopes generated by PDT on triple-negative breast cancer cell line (MDA-MB-231). METHODS: MDA-MB-231 cells were exposed to PDT using ALA (160 µg/mL)/630 nm at 8 J/cm2. Membrane proteins were extracted and separated by 2D PAGE. Proteins overexpressed were identified by LC-MS/MS and analyzed in silico through a peptide-HLA docking in order to identify the epitopes with more immunogenicity and antigenicity properties, as well as lower allergenicity and toxicity activity. The selected peptides were evaluated in response to macrophage activation and cytokine release by flow cytometry. RESULTS: Differential proteins were overexpressed in the cells treated with PDT. A group of 16 peptides were identified from them, established in a rigorous selection by measuring antigenicity, immunogenicity, allergenicity, and toxicity in silico. The final selection was based on molecular dynamics, where 2 peptides showed the highest stability regarding to the RMSD value. These peptides were obtained from the proteins calreticulin and HSP90. The cytokine analysis evidenced macrophage activation by the releasing of TNF. CONCLUSION: Two peptides were identified from calreticulin and HSP90; proteins induced by PDT in MDA-MB-231 cells. Both epitopes showed immunogenic potential as a peptide-based vaccine for triple-negative breast cancer.

In silico analysis of the association of hsa-miR-16 expression and cell survival in MDA-MB-231 breast cancer cells subjected to photodynamic therapy.

BACKGROUND: Breast cancer is the most common malignancy effecting women, and the triple-negative breast cancer (TNBC) subtype is particularly aggressive. This study aimed to evaluate the differential expression pattern of microRNAs (miRNAs) between untreated MDA-MB-231 cells (TNBC cell model) and those that survived photodynamic therapy (PDT) to gain insights into cell survival mechanisms. METHODS: Two PDT cycles were applied to MDA-MB-231 cells, using δ-aminolevulinic acid (ALA) followed by laser light at 635 nm. RNA was obtained from cells surviving PDT and untreated cells. The miRNAs expression profile was analyzed to detect the differences between the two groups. The potential target network of hsa-miR-16 was examined in silico with the integrative database Ingenuity® Pathway Analysis software. RESULTS: After the first and second PDT cycles, 17.8% and 49.6% of the MDA-MB-231 cells were viable. Microarray profiling of miRNAs showed decreased hsa-miR-16 expression (p < 0.05) in MDA-MB-231 cells surviving PDT when compared to the control cells. The predicted downstream targets of hsa-miR-16 were: 1) tumor suppressor protein 53; 2) molecules related to the cell cycle, such as cyclin D1, D3, and E1, and checkpoint kinase 1; 3) cell proliferation molecules, including fibroblast growth factor 1, 2 and 7 and fibroblast growth factor receptor 1; and 4) apoptosis-related molecules, consisting of BCL-2, B-cell leukemia/lymphoma 2, caspase 3, and cytochrome c. CONCLUSIONS: The differential expression of hsa-miR-16 between untreated MDA-MB-231 cells and those surviving PDT has not been previously reported. There was a lower expression of hsa-miR-16 in treated cells, which probably altered its downstream target network. In silico analysis predicted, a network related to the cell cycle, proliferation and apoptosis. These results are congruent with previous descriptions of hsa-miR-16 as a tumor suppressor and suggest that the treated population has increased their capacity to survive.

Glycosylated Nanoparticles for Cancer-Targeted Drug Delivery.

Nanoparticles (NPs) are novel platforms that can carry both cancer-targeting molecules and drugs to avoid severe side effects due to nonspecific drug delivery in standard chemotherapy treatments. Cancer cells are characterized by abnormal membranes, metabolic changes, the presence of lectin receptors, glucose transporters (GLUT) overexpression, and glycosylation of immune receptors of programmed death on cell surfaces. These characteristics have led to the development of several strategies for cancer therapy, including a large number of carbohydrate-modified NPs, which have become desirable for use in cell-selective drug delivery systems because they increase nanoparticle-cell interactions and uptake of carried drugs. Currently, the potential of NP glycosylation to enhance the safety and efficacy of carried therapeutic antitumor agents has been widely acknowledged, and much information is accumulating in this field. This review seeks to highlight recent advances in NP stabilization, toxicity reduction, and pharmacokinetic improvement and the promising potential of NP glycosylation from the perspective of molecular mechanisms described for drug delivery systems for cancer therapy. From preclinical proof-of-concept to demonstration of therapeutic value in the clinic, the challenges and opportunities presented by glycosylated NPs, with a focus on their applicability in the development of nanodrugs, are discussed in this review.

Use of AKR1C1 and TKTL1 in the Diagnosis of Low-grade Squamous Intraepithelial Lesions from Mexican Women.

BACKGROUND/AIM: To determine the differential protein profiles of cervical cancer cell lines in order to find potential targets that can be used as biomarkers in low-grade squamous intraepithelial lesions (LSIL) diagnosis. MATERIALS AND METHODS: Proteomic analysis was performed on cervical cancer cell lines by 2D electrophoresis and liquid chromatography-mass spectrometry. Biomarker validation was performed in histological samples by immunofluorescence. RESULTS: Aldo-keto reductase C1 (AKR1C1) and transketolase-like 1 (TKTL1) proteins were selected as biomarkers and their expression was increased in samples with LSIL diagnosis. TKTL1 in combination with AKR1C1 increased sensitivity and specificity to 75% and 66%, respectively, with an area under curve of 0.76 in receiver operating characteristics curve analysis. CONCLUSION: AKR1C1 and TKTL1 showed potential as biomarkers for diagnosis of LSIL in Mexican women, with similar sensitivity and specificity to the biomarkers used in clinical trials for diagnosis of LSIL.

CRISPR-dCas9-Based Artificial Transcription Factors to Improve Efficacy of Cancer Treatment With Drug Repurposing: Proposal for Future Research.

Due to the high resistance that cancer has shown to conventional therapies, it is difficult to treat this disease, particularly in advanced stages. In recent decades, treatments have been improved, being more specific according to the characteristics of the tumor, becoming more effective, less toxic, and invasive. Cancer can be treated by the combination of surgery, radiation therapy, and/or drug administration, but therapies based on anticancer drugs are the main cancer treatment. Cancer drug development requires long-time preclinical and clinical studies and is not cost-effective. Drug repurposing is an alternative for cancer therapies development since it is faster, safer, easier, cheaper, and repurposed drugs do not have serious side effects. However, cancer is a complex, heterogeneous, and highly dynamic disease with multiple evolving molecular constituents. This tumor heterogeneity causes several resistance mechanisms in cancer therapies, mainly the target mutation. The CRISPR-dCas9-based artificial transcription factors (ATFs) could be used in cancer therapy due to their possibility to manipulate DNA to modify target genes, activate tumor suppressor genes, silence oncogenes, and tumor resistance mechanisms for targeted therapy. In addition, drug repurposing combined with the use of CRISPR-dCas9-based ATFs could be an alternative cancer treatment to reduce cancer mortality. The aim of this review is to describe the potential of the repurposed drugs combined with CRISPR-dCas9-based ATFs to improve the efficacy of cancer treatment, discussing the possible advantages and disadvantages.

Gold nanoparticle uptake is enhanced by estradiol in MCF-7 breast cancer cells.

Purpose: In the present study, we investigated the effects of 17ß-estradiol (E2) on membrane roughness and gold nanoparticle (AuNP) uptake in MCF-7 breast cancer cells. Methods: Estrogen receptor (ER)-positive breast cancer cells (MCF-7) were exposed to bare 20 nm AuNPs in the presence and absence of 1×10-9 M E2 for different time intervals for up to 24 hrs. The effects of AuNP incorporation and E2 incubation on the MCF-7 cell surface roughness were measured using atomic force microscopy (AFM). Endocytic vesicle formation was studied using confocal laser scanning microscopy (CLSM). Finally, the results were confirmed by hyperspectral optical microscopy. Results: High-resolution AFM images of the surfaces of MCF-7 membranes (up to 250 nm2) were obtained. The incubation of cells for 12 hrs with AuNP and E2 increased the cell membrane roughness by 95% and 30% compared with the groups treated with vehicle (ethanol) or AuNPs only, respectively. This effect was blocked by an ER antagonist (7α,17ß-[9-[(4,4,5,5,5-Pentafluoropentyl)sulfinyl]nonyl]estra-1,3,5(10)-triene-3,17-diol [ICI] 182,780). Higher amounts of AuNPs were localized inside MCF-7 cells around the nucleus, even after 6 hrs of E2 incubation, compared with vehicle-treated cells. Endolysosome formation was induced by E2, which may be associated with an increase in AuNP-uptake. Conclusions: E2 enhances AuNP incorporation in MCF-7 cells by modulating of plasma membrane roughness and inducing lysosomal endocytosis. These findings provide new insights into combined nanotherapies and hormone therapies for breast cancer.

Vaginosis-associated bacteria and its association with HPV infection. / Bacterias relacionadas con vaginosis bacteriana y su asociación a la infección por virus del papiloma humano.

BACKGROUND AND OBJECTIVE: Cervical cancer is an important health problem in our country. It is known that there are several risk factors for this neoplasm, and it has been suggested that cervical microbiome alterations could play a role in the development and progress of cancer. Bacterial vaginosis associated bacteria such as Atopobium vaginae and Gardnerella vaginalis has been suggested as potential risk factor for cervical lesions and cervical cancer. MATERIAL AND METHODS: DNA from 177 cervical scraping samples was studied: 104 belonged to women without cytological or colposcopic alterations and 73 samples from precursor lesions with previous human papillomavirus (HPV) infection history. All samples were screened for Atopobium vaginae, Gardnerella vaginalis and HPV by PCR. RESULTS: High HPV prevalence was found in precursor samples, and 30% of samples without lesions were positive for HPV. Virtually all samples contained sequences of both bacteria, and interestingly, there was not HPV association observed; these results could suggest that these microorganisms could be part of the cervical microbiome in Mexican population. CONCLUSIONS: The results obtained indicate that the bacteria analysed could be part of normal biome in Mexican women, suggesting a potential reconsideration of the pathogen role of these microorganisms.

Theranostic-PDT with the antibody anti isoform 4 SOD mitocondrial labeled with PpIX in the lung cancer cell line A-549.

BACKGROUND: In this work, a drug product composed of an IgM antibody derived from a hybridoma subclone 4C1F6D5G7B8 was prepared and further labeled with PpIX to be used in cell lines A-549 and MRC-5. The aim of this work was to evaluate the potential theranostic activity of the obtained product together with photodynamic therapy (PDT). METHODS: The IgM antibody labeled with PpIX was used in different concentrations to perform theranostics with PDT in cell lines A-549 and MRC-5 in order to identify the specificity of IgM antibody in lung cancer cells by means of a LED-irradiation system set at 630 nm. The location of the conjugate was further determined by confocal microscopy. RESULTS: The theranostic with conjugate Ab-PpIX in the A-549 cell lines showed fluorescence by confocal microscopy, whereas the MRC-5 cell line showed no reactivity. The PDT with the conjugate in the cell line A-549 decreased its viability 70% compared to the control. On the contrary, with the MRC-5 cell line no viability diference was shown. The confocal microscopy applied to the cell line A-549 showed that the Ab-PpIX was majorly located at the cytoplasm. CONCLUSION: Ab-PpIX showed therapeutical potential in lung cancer cells A-549 and had no activity in non-cancerous lung cells (MCR-5).

Effectiveness of Photodynamic Therapy in Elimination of HPV-16 and HPV-18 Associated with CIN I in Mexican Women.

This study aimed to determine the effectiveness of photodynamic therapy (PDT), using δ-aminolevulinic acid (5-ALA), in the elimination of premalignant cervical lesions in Mexican patients with human papillomavirus (HPV) infection and/or cervical intraepithelial neoplasia (CIN). Thirty women diagnosed with CIN I and/or positive for HPV participated in the study. Topical 6% 5-ALA in gel form was applied to the uterine cervix; after 4 h, the lesion area was irradiated with a light dose of 200 J cm-2 at 635 nm. This procedure was performed three times at 48-h intervals. Clinical follow-up was performed at 3, 6, and 12 months after the initial PDT administration, by colposcopy, cervical cytology, histopathological analysis, polymerase chain reaction, and hybrid capture. Of HPV-infected patients without evidence of CIN I, 80% cleared the infection, while HPV associated with CIN I was eliminated in 83% of patients (P < 0.05). At 12 months, CIN I had regressed in 57% of patients, although this response was not statistically significant. PDT using 6% 5-ALA is concluded to be effective in eliminating HPV infection associated or not with CIN I.

[Use of nanoparticles (NP) in photodynamic therapy (PDT) against cancer]. / Uso de nanopartículas (NP) en la terapia fotodinámica (photodynamic therapy [PDT]) contra el cáncer.

Nanotechnology is a promising interdisciplinary field for developing improved methods of diagnosis and treatment of different diseases, including cancer. Give their optical, magnetic, and structural property, the nanoparticles have been proposed to be use in the development of unconventional treatments for cancer such as photodynamic therapy (PDT). In PDT, a photosensitizing agent is used that accumulates in tumor cells, generating reactive oxygen species that causes the death of malignant cells after irradiation with light at a particular wavelength. However, the use of PDT presents different problems in its application due to the characteristics of hydrophobicity of the photosensitizers, which hinder the efficiency of administration and treatment. It is here where the use of nanoparticles is proposed as a delivery vehicle to optimize treatment application. In this review we describe the use of nanoparticles coupled to PDT in the treatment of cancer and its molecular mechanism of action.

Markers of Pluripotency in Human Amniotic Epithelial Cells and Their Differentiation to Progenitor of Cortical Neurons.

Human pluripotent stem cells (hPSC) have promise for regenerative medicine due to their auto-renovation and differentiation capacities. Nevertheless, there are several ethical and methodological issues about these cells that have not been resolved. Human amniotic epithelial cells (hAEC) have been proposed as source of pluripotent stem cells. Several groups have studied hAEC but have reported inconsistencies about their pluripotency properties. The aim of the present study was the in vitro characterization of hAEC collected from a Mexican population in order to identify transcription factors involved in the pluripotency circuitry and to determine their epigenetic state. Finally, we evaluated if these cells differentiate to cortical progenitors. We analyzed qualitatively and quantitatively the expression of the transcription factors of pluripotency (OCT4, SOX2, NANOG, KLF4 and REX1) by RT-PCR and RT-qPCR in hAEC. Also, we determined the presence of OCT4, SOX2, NANOG, SSEA3, SSEA4, TRA-1-60, E-cadherin, KLF4, TFE3 as well as the proliferation and epigenetic state by immunocytochemistry of the cells. Finally, hAEC were differentiated towards cortical progenitors using a protocol of two stages. Here we show that hAEC, obtained from a Mexican population and cultured in vitro (P0-P3), maintained the expression of several markers strongly involved in pluripotency maintenance (OCT4, SOX2, NANOG, TFE3, KLF4, SSEA3, SSEA4, TRA-1-60 and E-cadherin). Finally, when hAEC were treated with growth factors and small molecules, they expressed markers characteristic of cortical progenitors (TBR2, OTX2, NeuN and ß-III-tubulin). Our results demonstrated that hAEC express naïve pluripotent markers (KLF4, REX1 and TFE3) as well as the cortical neuron phenotype after differentiation. This highlights the need for further investigation of hAEC as a possible source of hPSC.

Effectiveness of Toki's criteria and determination of variables for identification of HPV L1 protein in oral lesions.

OBJECTIVE: To evaluate the effectiveness of Toki's criteria in identifying the HPV L1 protein in oral lesions with the use of immunohistochemistry (IHC) and to determine which criteria optimize such identification. STUDY DESIGN: Retrospective study of 277 cases diagnosed as HPV lesions at 22 years. Tests of sensitivity, specificity, positive predictive value (PPV), negative predictive value (NPV), kappa coefficients, and chi2 values, as well as two logistic regression analyses (p≤0.05), were conducted. RESULTS: Of the lesions studied, 96.4% (267 of 277) were positive for HPV using Toki's criteria and 28.5% (79 of 277) were positive for L1 by IHC. Toki's criteria showed sensitivity=93.67%, specificity=2.53%, PPV=6.99%, and NPV=46.55%. Neither concordance nor statistically significant associations were observed between both tests. The logistic regression of Toki's criteria was useful in the diagnosis of L1, correctly classified 71.8% of the lesions positive for L1, and showed a Hosmer-Lemeshow adjustment of p=0.614 and a Nagelkerke's coefficient of determination of 6.8%. The explanatory variables statistically significant at p≤0.05 were dyskeratosis (p=0.01) and papillomatosis (p=0.04). Forty-nine independent variables (clinical and histopathologic) were involved in the second regression analysis. The model correctly classified 85.2% of the lesions and showed a Hosmer-Lemeshow adjustment of p=0.696 and a Nagelkerke's coefficient of determination of 60.2%. The explanatory variables statistically significant at p≤0.05 were: age younger than 35 years (p=0.001), multiple lesions (p=0.031), hyperorthokeratosis (p=0.019), focal intracellular edema (p=0.002), and the presence of 1 to more than 5 cells with degenerative changes in their nucleus (p=0.048). CONCLUSIONS: Toki's criteria are not adequate to make a diagnosis of lesions by HPV in the mouth, but the logistic regression analysis showed clinical and histopathologic variables which optimize the identification of lesions through the L1 protein. However, a PCR study is advisable when the presence of high-risk HPV is suspected.

Effect of germinated soy protein on the growth of HeLa cervical cancer cells in female athymic mice.

Previous studies showed that germination could improve the antiproliferative effect of soy protein on cervical cancer cells and that a peptide fraction (MAPF) from germinated soybeans decreases the expression of PTTG1 and TOP2A (2 genes considered as therapeutic targets) causing apoptosis of cancer cells. The aim of this work was to study the effect of feeding germinated soybean protein diets on the tumor growth in nude mice inoculated with cervical cancer cells and identify the bioactive component. Mice were randomly assigned to 1 of the 6 dietary groups based in AIN-93G formulation with 6 protein sources: casein, ungerminated soy protein (SP), and SP from 2 and 6 days of germination, with and without ethanol-soluble phytochemicals (ESPC). Compared with casein-fed controls, the tumor volumes after 5 wk were reduced by 44.6% by ungerminated SP, 98.9% by 2-day-germinated SP, 97.7% by 2-day-germinated SP without ESPC, 94.7% by 6-day-germinated SP, and 92.7% by 6-day-germinated SP without ESPC (P < 0.05). Liquid chromatography coupled with tandem mass spectrometry analysis of MAPF showed that the bioactive peptide might be the leginsulin, a peptide involved in signal transduction of soybean cells. Germination is a simple procedure that could help to increase the anticancer activity of soy protein probably through generation of biologically active peptides.

Sodium butyrate increases the effect of the photodynamic therapy: a mechanism that involves modulation of gene expression and differentiation in astrocytoma cells.

OBJECTIVES: In order to evaluate the improvement of the photodynamic therapy (PDT) due to sodium butyrate (NaBu), its effectiveness in U373-MG and D54-MG astrocytoma cell lines was evaluated. METHODS: Cells were exposed to delta-aminolevulinic acid (δ-ALA) as a precursor to endogenous photosensitizer protoporphyrin IX (PpIX). In both astrocytoma cells, an important increase by ALA was observed in uroporphyrinogen synthetase gene expression: 1.8- and 52-fold for D54-MG and U373-MG cells, respectively. After irradiation, they showed 16.67 and 28.9% of mortality in U373-MG and D54-MG, respectively. These mortalities increased to 70.62 and 96.7% when U373-MG and D54-MG cells, respectively, were exposed 24 h to 8 mM NaBu, before to PpIX induction. NaBu induced expression of caspase-3, caspase-9, and Bcl-2 and increased Bax in U373-MG cells. ALA-induced morphological changes are compatible to differentiation. CONCLUSIONS: Genes and differentiation induced mainly by NaBu improve cell death performed by PDT in astrocytoma cells. These facts prove the synergistic effect of NaBu on cytotoxic damage induced by PDT.

A peptide fraction from germinated soybean protein down-regulates PTTG1 and TOP2A mRNA expression, inducing apoptosis in cervical cancer cells.

The aim of this study was to evaluate the effect of a peptide fraction, obtained from a germinated soybean protein hydrolysate, on the viability, apoptosis and cancer related gene expression in HeLa cells. Soybean was germinated for 0-6 days and proteins were isolated from the seeds. Protein isolates, without ethanol-soluble phytochemicals, were hydrolyzed with digestive enzymes and their effect on growth in HeLa cells was evaluated. The most active hydrolysate was separated by ultrafiltration into five peptide fractions. A >10 kDa fraction was the most active against cancer cells. This fraction down-regulated PTTG1 and TOP2A mRNA expression (two genes considered as therapeutic targets) and induced apoptosis in cancer cells activating the caspase cascade and causing DNA fragmentation. Germinated soy protein isolates could be a bioactive ingredient of functional food.

Effect of ditaxin and heteranthin and inhibitory effect of Ditaxis heterantha extract on L5178Y tumor development in mice.

Ditaxis heterantha seeds are used as spices for flavoring and coloring food. Two new apocarotenoids derived from the seeds, heteranthin and ditaxin, were evaluated for their in vitro cytotoxic effects in murine lymphoma cells lines. Bioabsorption in mice and preventive and antitumor effects of the apocarotenoids were determined. Ditaxin and heteranthin showed cytotoxic effects in vitro against murine malignant cells and normal splenocyte cells. The 50% inhibitory concentration (IC(50)) for ditaxin in splenocytes was 0.1825 mM; in L5178Y, the IC(50) was 0.1923 mM. The heteranthin IC(50) in splenocytes was 0.1325 mM; in L5178Y, the value was 0.3889 mM. The maximum ditaxin plasma concentration was found after 2 hours of administration (mean±standard deviation, 7.5±2.05 µg/mL). Oral administration of the D. heterantha extract (100 mg/kg per day) for 14 days after the L5178Y lymphoma cell implantation showed no significant effect compared with groups that were not pretreated. However, tumor inhibition in groups treated intraperitoneally before inoculation with the L5178Y cells showed a significant difference (P<.001) compared with the groups not pretreated.

Spirulina maxima and its protein extract protect against hydroxyurea-teratogenic insult in mice.

Congenital malformations are one of the major causes of child mortality all over the world. In order to prevent them it is necessary to find substances that act as anti-teratogenic agents. In this study hydroxyurea (HU), an antineoplastic and teratogenic drug, was administered to pregnant mice because one of its major mechanisms of teratogenesis is the production of reactive oxygen species (ROS). The aim of this work was to determine if Spirulina maxima (SP) and its aqueous protein extract could protect against HU-teratogenic insult in mouse embryos. SP has been used for a long time because of its nutritional and pharmacological properties. The antioxidant activity, one of the most important, is related to the protein extract due to its content of phycobiliproteins. It was observed that neither SP nor its extract provoked teratogenic effects at any dose tested and even increased vitelline yolk sac circulation. Dams exposed to HU (30 mg/kg, i.p.) presented embryos with multiple alterations in their development. Groups treated with SP or its extract, before and after HU exposure, showed a protector effect in a dose-dependent manner. TBARS test confirmed that the protection effect was related to the antioxidant activity of both SP and its extract.

Effect of protein hydrolysates from germinated soybean on cancerous cells of the human cervix: an in vitro study.

Consumption of soybeans can reduce the risk of different types of cancer. Little is known about the effect of germination on the anticancer properties of soya. This study was done to determine if germination improves the anticancer properties of soybean protein through generation of amino acids or bioactive peptides. Soybean was germinated for 0, 2, 3, 4, 5, and 6 days and proteins were isolated from the seeds. Isolates with and without ethanol-soluble phytochemicals were hydrolyzed with digestive enzymes and their effect on growth in HeLa and C-33 (epidermoid cervical carcinoma) and HaCaT (non-cancerous human keratinocytes) cells were evaluated with the Alamar Blue method. Germination induced degradation of the alpha and alpha' fractions of beta-conglycinin and acid fraction of glycinin, generating low molecular weight peptides. Degrees of hydrolysis ranged from 73-77%. Hydrolysates inhibited the growth of HeLa cells and C-33 at concentrations exceeding 1.25 mg/ml. Major inhibition was observed with the hydrolysate germinated for 2 days and containing ethanolsoluble phytochemicals (IC(50) 2.15 and 2.27 mg/ml for HeLa and C-33, respectively). Interestingly, hydrolysate cytoxicity for normal cells was minimal in comparison to cancer cells.