The sound of silence: Quantification of typical absence seizures by sonifying EEG signals from a custom-built wearable device.

OBJECTIVE: To develop and validate a method for long-term (24-h) objective quantification of absence seizures in the EEG of patients with childhood absence epilepsy (CAE) in their real home environment using a wearable device (waEEG), comparing automatic detection methods with auditory recognition after seizure sonification. METHODS: The waEEG recording was acquired with two scalp electrodes. Automatic analysis was performed using previously validated software (Persyst® 14) and then fully reviewed by an experienced clinical neurophysiologist. The EEG data were converted into an audio file in waveform format with a 60-fold time compression factor. The sonified EEG was listened to by three inexperienced observers and the number of seizures and the processing time required for each data set were recorded blind to other data. Quantification of seizures from the patient diary was also assessed. RESULTS: Eleven waEEG recordings from seven CAE patients with an average age of 8.18 ± 1.60 years were included. No differences in the number of seizures were found between the recordings using automated methods and expert audio assessment, with significant correlations between methods (ρ > .89, p < .001) and between observers (ρ > .96, p < .001). For the entire data set, the audio assessment yielded a sensitivity of .830 and a precision of .841, resulting in an F1 score of .835. SIGNIFICANCE: Auditory waEEG seizure detection by lay medical personnel provided similar accuracy to post-processed automatic detection by an experienced clinical neurophysiologist, but in a less time-consuming procedure and without the need for specialized resources. Sonification of long-term EEG recordings in CAE provides a user-friendly and cost-effective clinical workflow for quantifying seizures in clinical practice, minimizing human and technical constraints.

Plastic rearrangement of basal forebrain parvalbumin-immunoreactive neurons in the kainite model of epilepsy.

Temporal lobe epilepsy (TLE) is the most prevalent form of epilepsy, through the neuronal mechanisms of this syndrome remain elusive. In addition to the temporal lobe structures, it was found that the basal forebrain cholinergic cells are also involved in epileptogenesis. However, little is known about the involvement of the basal forebrain GABAergic neurons in epilepsy; despite this, they largely project to the temporal lobe and are crucial for the regulation of the hippocampal circuitry. In this study, we assessed epilepsy-induced changes in parvalbumin (PARV) immunoreactive neurons of the medial septum (MS) and of the magnocellular preoptic nucleus (MCPO) using the kainic acid (KA) model in rats. In addition, we estimated the respective changes in the cholinergic varicosities in the MS, where we observed a significant reduction in the PARV cell number (12849 ± 2715 vs. 9372 ± 1336, p = .029) and density (16.2 ± 2.62 vs. 10.5 ± 1.00 per .001 mm3, p =.001), and an increase in the density of cholinergic varicosities (47.9 ± 11.1 vs. 69.4 ± 17.8 per 30,000 µm2, p =.036) in KA-treated animals. In the MCPO, these animals showed a significant increase in somatic volume (827.9 ± 235.2 µm3 vs. 469.9 ± 79.6 µm3, p = .012) and total cell number (2268.6 ± 707.1 vs. 1362.4 ± 262.0, p =.028). These results show that the basal forebrain GABAergic cell populations undergo numerical and morphological changes in epileptic animals, which may contribute to an increased vulnerability of brain circuits to epilepsy and epilepsy-related functional impairments.

GuttaFlow® Bioseal Cytotoxicity Assessment: In Vitro Study.

The sealers used for root canal treatment should be biocompatible for the peri-radicular tissues, to evaluate the cytotoxic effects of GuttaFlow® bioseal sealer and to compare them with AH26® epoxy resin. Culture media were conditioned with the GuttaFlow® bioseal and AH26® pellets. MDPC-23 odontoblast cell cultures were treated with conditioned medium and serial dilutions. To evaluate the metabolic activity and cellular viability, the MTT and SRB assays were performed. To determine the production of reactive oxygen species, the DHE and DCF-DA probes were used. Cell cycle and cell-death types were assessed by cytometry, and to evaluate the mineralization capacity, the Alizarin Red S coloration was used. Statistical analysis was performed using analysis of variance (ANOVA) when normality was found and Kruskal-Wallis on the opposite case. For the comparison with normality values, the Student t-test was used. Cells exposed to the GuttaFlow® bioseal conditioned medium maintained high metabolic activities, except at higher concentrations. Likewise, viability was maintained, but a significant decrease was observed after exposure to the highest concentration (p < 0.001), associated with cell death by late apoptosis and necrosis. When cell cultures were exposed to AH26®, metabolic activity was highly compromised, resulting in cell death. An imbalance in the production of peroxides and superoxide anion was observed. GuttaFlow® bioseal showed higher biocompatibility than AH26®.

Platinum(II) ring-fused chlorins as efficient theranostic agents: Dyes for tumor-imaging and photodynamic therapy of cancer.

The discovery of Pt-chlorin-type theranostic agents is described. Luminescent Pt(II) 4,5,6,7-tetrahydropyrazolo[1,5-a]pyridine-fused chlorins, with different degrees of hydrophilicity, have been synthesized and their in vitro photocytotoxicity against human melanoma, oesophageal and bladder carcinomas was studied. A di(hydroxymethyl)-substituted chlorin was identified as a privileged molecule to explore imaging-guided photodynamic therapy. In addition to the high activity as PDT agent and absence of cytotoxicity per se, this molecule showed the ideal photophysical and photochemical properties. In vivo studies using a A375 melanoma mouse model, proved the extraordinary properties of this chlorin as a luminescent probe and the ability to impair tumor growth, making image guided treatment and follow up a possibility.

Calcium Phosphate Ceramics Can Prevent Bisphosphonate-Related Osteonecrosis of the Jaw.

Bisphosphonate-associated osteonecrosis of the jaw (BRONJ), a post-surgical non-healing wound condition, is one of the most common side effects in patients treated with nitrogen-containing bisphosphonates. Its physiopathology has been related with suppression of bone turnover, of soft tissue healing and infection. Biphasic calcium phosphates (BCP) are used as a drug delivery vehicle and as a bone substitute in surgical wounds. Due to their capacity to adsorb zoledronate, it was hypothesized these compounds might have a protective effect on the soft tissues in BRONJ wounds. To address this hypothesis, a reproducible in vivo model of BRONJ in Wistar rats was used. This model directly relates chronic bisphosphonate administration with the development of osteonecrosis of the jaw after tooth extraction. BCP granules were placed in the alveolus immediately after tooth extraction in the test group. The animals were evaluated through nuclear medicine, radiology, macroscopic observation, and histologic analysis. Encouragingly, calcium phosphate ceramics were able to limit zoledronate toxicity in vivo and to favor healing, which was evidenced by medical imaging (nuclear medicine and radiology), macroscopically, and through histology. The studied therapeutic option presented itself as a potential solution to prevent the development of maxillary osteonecrosis.

Cytotoxic Effects of Zoom® Whitening Product in Human Fibroblasts.

Tooth whitening procedures are increasing; however, side effects can occur, such as damage to pulp cells, by the whitening products. This study aims to assess the cellular effects promoted by a whitening product, namely, the oxidative stress fostered by the active agent hydrogen peroxide, with and without photoactivation. Additionally, if cellular recovery occurred, we intended to determine the time point where cells recover from the tooth whitening induced damage. Human fibroblasts were exposed to hydrogen peroxide, Zoom®, Zoom® + irradiation, and irradiation alone. The following analysis was performed: metabolic activity evaluation by the MTT assay; cell viability, mitochondrial membrane potential, peroxides production, superoxide radical production, and reduced glutathione expression by flow cytometry. We determined the IC50 value for all groups, and a dose-dependent cytotoxic effect was verified. At the times analyzed, hydrogen peroxide groups showed no metabolic activity recovery while a cell recovery was observed after 24 h (Zoom®) and 48 h (Zoom® + irradiation). Cell death was seen in hydrogen peroxide and Zoom® + irradiation groups, mainly by apoptosis, and the irradiation had a cytotoxic effect per se. This in vitro study supports that whitening products with moderate hydrogen peroxide (HP) concentration have a temporary effect on cells, allowing a cellular recovery.

Evaluation of dentinogenesis inducer biomaterials: an in vivo study.

When exposure of the pulp to external environment occurs, reparative dentinogenesis can be induced by direct pulp capping to maintain pulp tissue vitality and function. These clinical situations require the use of materials that induce dentin repair and, subsequently, formation of a mineralized tissue. OBJECTIVE: This work aims to assess the effect of tricalcium silicate cements and mineral trioxide aggregate cements, including repairing dentin formation and inflammatory reactions over time after pulp exposure in Wistar rats. METHODOLOGY: These two biomaterials were compared with positive control groups (open cavity with pulp tissue exposure) and negative control groups (no intervention). The evaluations were performed in three stages; three, seven and twenty-one days, and consisted of an imaging (nuclear medicine) and histological evaluation (H&E staining, immunohistochemistry and Alizarin Red S). RESULTS: The therapeutic effect of these biomaterials was confirmed. Nuclear medicine evaluation demonstrated that the uptake of 99mTc-Hydroxymethylene diphosphonate (HMDP) showed no significant differences between the different experimental groups and the control, revealing the non-occurrence of differences in the phosphocalcium metabolism. The histological study demonstrated that in mineral trioxide aggregate therapies, the presence of moderate inflammatory infiltration was found after three days, decreasing during follow-ups. The formation of mineralized tissue was only verified at 21 days of follow-up. The tricalcium silicate therapies demonstrated the presence of a slight inflammatory infiltration on the third day, increasing throughout the follow-up. The formation of mineralized tissue was observed in the seventh follow-up day, increasing over time. CONCLUSIONS: The mineral trioxide aggregate (WhiteProRoot®MTA) and tricalcium silicate (Biodentine™) present slight and reversible inflammatory signs in the pulp tissue, with the formation of mineralized tissue. However, the exacerbated induction of mineralized tissue formation with the tricalcium silicate biomaterial may lead to the formation of pulp calcifications.

Evaluation of dentinogenesis inducer biomaterials: an in vivo study

Abstract When exposure of the pulp to external environment occurs, reparative dentinogenesis can be induced by direct pulp capping to maintain pulp tissue vitality and function. These clinical situations require the use of materials that induce dentin repair and, subsequently, formation of a mineralized tissue. Objective: This work aims to assess the effect of tricalcium silicate cements and mineral trioxide aggregate cements, including repairing dentin formation and inflammatory reactions over time after pulp exposure in Wistar rats. Methodology: These two biomaterials were compared with positive control groups (open cavity with pulp tissue exposure) and negative control groups (no intervention). The evaluations were performed in three stages; three, seven and twenty-one days, and consisted of an imaging (nuclear medicine) and histological evaluation (H&E staining, immunohistochemistry and Alizarin Red S). Results: The therapeutic effect of these biomaterials was confirmed. Nuclear medicine evaluation demonstrated that the uptake of 99mTc-Hydroxymethylene diphosphonate (HMDP) showed no significant differences between the different experimental groups and the control, revealing the non-occurrence of differences in the phosphocalcium metabolism. The histological study demonstrated that in mineral trioxide aggregate therapies, the presence of moderate inflammatory infiltration was found after three days, decreasing during follow-ups. The formation of mineralized tissue was only verified at 21 days of follow-up. The tricalcium silicate therapies demonstrated the presence of a slight inflammatory infiltration on the third day, increasing throughout the follow-up. The formation of mineralized tissue was observed in the seventh follow-up day, increasing over time. Conclusions: The mineral trioxide aggregate (WhiteProRoot®MTA) and tricalcium silicate (Biodentine™) present slight and reversible inflammatory signs in the pulp tissue, with the formation of mineralized tissue. However, the exacerbated induction of mineralized tissue formation with the tricalcium silicate biomaterial may lead to the formation of pulp calcifications

A cross-sectional analysis of patient characteristics, health conditions and patient experience at a Portuguese medical acupuncture teaching appointment.

BACKGROUND: Acupuncture is one of the most popular and most frequently used complementary medicines worldwide, with benefits for several health conditions when integrated into Western medical practice. OBJECTIVE: To perform a retrospective analysis of patient characteristics, health conditions and patient experience in a teaching medical acupuncture appointment at the Faculty of Medicine, University of Coimbra/Coimbra Hospital and University Centre. METHODS: 500 medical records between January 2010 and December 2015 were accessed and 324 were included. The characteristics examined include gender, age, health conditions treated, number of acupuncture needles used in each treatment, professional who referred the patient, type of stimulation, number of treatment sessions and patient experience regarding the degree of improvement at the end of treatment. RESULTS: Patients range from 13 to 92 years old, with most between 40 and 59 yo (40.7%). In total, 71.3% were female and 28.7% male. The most commonly treated health conditions were musculoskeletal symptoms (60.4%), nervous and headaches (18.2%) and orofacial (11.3%). The median number of appointments was 6 and the median needles used per treatment was 12. In 52.2% of consultations, electrostimulation was performed. The majority of patients were referred by physical and rehabilitative medicine and dentists. 85.3% of patients reported improvement at the end of the treatment, with those who performed more sessions presenting a higher improvement. CONCLUSIONS: This pioneering study in Portugal presents similar results to other countries regarding patient characterization. The high success rate (85.3%) regarding patient improvement recommends the use of acupuncture as an effective complementary therapy.

Direct Pulp Capping: Which is the Most Effective Biomaterial? A Retrospective Clinical Study.

(1) Background: Recently, tricalcium silicate cements, such as Biodentine™, have emerged. This biomaterial has a calcium hydroxide base and characteristics like mineral aggregate trioxide cements, but has tightening times that are substantially more suitable for their application and other clinical advantages. (2) Methods: A retrospective clinical study was conducted with 20 patients, which included a clinical evaluation of the presence or absence of pulp inflammation compatible symptoms, radiographic evaluation of the periapical tissues, and structural alterations of the coronary restoration that supports pulp capping therapies with Biodentine™ and WhiteProRoot®MTA. (3) Results: This clinical study revealed similar success rates between mineral trioxide cement and tricalcium silicates cements at 6 months, with 100% and 95% success rates, respectively. There were no statistically significant differences between both biomaterials and between these and the various clinical circumstances, namely the absolute isolation of the operating field, exposure size, the aetiology of exposure, and even the type of restorative material used. (4) Conclusions: Biodentine™ demonstrated a therapeutic effect on the formation of a dentin bridge accompanied by slight inflammatory signs, with a high clinical success rate, indicating the possibility of its effective and safe use in dental pulp direct capping in humans, similar to the gold standard material.

Synthetic Calcium Phosphate Ceramics as a Potential Treatment for Bisphosphonate-Related Osteonecrosis of the Jaw.

(1) Background: Bisphosphonate-related osteonecrosis of the jaw (BRONJ) is one of the most often seen side effects in patients treated with nitrogen-containing bisphosphonates (BPs), a post-surgical non-healing wound condition. Since calcium phosphate (CP) compounds are able to adsorb zoledronate (ZOL) when used as a drug delivery vehicle, we aimed to verify if these ceramics might have a potential protective effect for soft tissues surrounding surgical osseous wounds. (2) Methods: The chemical reaction between ZOL and CP compounds was evaluated through ultraviolet-visible spectroscopy and elemental analysis. A primary culture of human gingival fibroblasts (HGF) was established as a model to evaluate the cytotoxicity of the association of ZOL (5-500 µM) and of ZOL/biphasic calcium phosphates (BCP). Metabolic activity, cell viability, types of cell death, the cell cycle through, and the migration ability of human gingival fibroblasts were evaluated. (3) Results: ZOL was adsorbed by biphasic calcium phosphate compounds in an aqueous solution. The HGF were sensitive to ZOL toxicity; nevertheless, ZOL/BCP showed a significant protective effect regarding metabolic activity, cell viability, and cell migration. (4) Conclusions: BCP interaction with ZOL reduces or abolishes its toxicity in HGF. This finding represents a potential solution for BRONJ in the case of patients undergoing therapy with ZOL.

Biodentine™ Boosts, WhiteProRoot®MTA Increases and Life® Suppresses Odontoblast Activity.

(1) Background: When pulp exposure occurs, reparative dentinogenesis can be induced by direct pulp capping to maintain the vitality and function of the tissue. The aim of this work was to assess the cytotoxicity and bioactivity of three different direct pulp capping materials, calcium hydroxide (Life®), mineral trioxide aggregate (WhiteProRoot®MTA) and calcium silicate (Biodentine™), in an odontoblast-like mouse cell line (MDPC-23). (2) Methods: Metabolic activity was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide test (MTT)assay, viability by the sulforhodamine B (SRB) assay, and the type of death and cell cycle analysis by flow cytometry. Alkaline phosphatase was evaluated by polymerase chain reaction (PCR), and dentin sialoprotein expression was assessed by immunocytochemistry. Mineralization was determined by the Alizarin Red S colorimetric assay and quantified by spectrophotometry. (3) Results: Life® induced a decrease in metabolic activity and viability, which is associated with an increase cell death. WhiteProRoot®MTA and Biodentine™ induced similar effects in cytotoxicity assays, with an increase in the expression of dentin sialoprotein (DSP) and formation of mineralized deposits, especially with Biodentine™. (4) Conclusions: The results of WhiteProRoot®MTA confirm its indication for these therapies, justifying its recognition as the "gold standard". Biodentine™ may be an alternative, since they promote the same cellular response that mineral trioxide aggregate (MTA) does.

Prevalence of vitamin D deficiency amongst soccer athletes and effects of 8 weeks supplementation.

BACKGROUND: High prevalence of vitamin D deficiency is well known around the world in risk populations. Although less is known about the athletic population, some studies report vitamin D deficiency amongst athletic population and adequate vitamin D levels are crucial for athletic population as they can prevent injuries such as stress fractures and might even have ergogenic effects for example on muscle function. The main objectives were to evaluate the basal serum levels of 25(OH)D and calcium in professional soccer athletes on the latitude 40°N, to evaluate the effects in 25(OH)D and calcium serum levels following supplementation of 1667 IU/day of cholecalciferol during a period of 8 weeks and evaluate eventual toxicity arising from it. METHODS: Twenty-eight professional athletes were evaluated according to the skin type. Basal serum levels of 25(OH)D and calcium were evaluated during winter months. Athletes were then supplemented with cholecalciferol 25.000 IU every two weeks. Serum levels of 25(OH)D and calcium were evaluated after supplementation. RESULTS: 25(OH)D initially ranged between 9.9 ng/mL and 32.9 ng/mL with a median of 19.2 IQR 7.24 ng/mL. A statistically significant inverse correlation exists between vitamin D deficiency and the Fitzpatrick Scale (ρ=-0.555 P=0.003). After 8 weeks, 25(OH)D ranged between 10.6 ng/mL and 43.4 ng/mL with a median of 33.2 ng/mL IQR 6.1 ng/mL. We verified a statistically significant increase of serum 25(OH) D levels (11.74±5.988; CI 95% [9,02; 14,47]; P<0.001. In addition, there was a statistically significant reduction of calcium: -0.36±0.457; CI 95% [- 0.57; -0.15]; P=0.002. CONCLUSIONS: Professional athletes have a high prevalence of vitamin D deficiency. Supplementation with cholecalciferol in winter months during 8 weeks is safe and effective in raising 25(OH)D serum levels. However, it may not be sufficient for athletes to reach adequate vitamin D levels.

Endometrial Cancer Spheres Show Cancer Stem Cells Phenotype and Preference for Oxidative Metabolism.

This study aimed to characterize endometrial cancer regarding cancer stem cells (CSC) markers, regulatory and differentiation pathways, tumorigenicity and glucose metabolism. Endometrial cancer cell line ECC1 was submitted to sphere forming protocols. The first spheres generation (ES1) was cultured in adherent conditions (G1). This procedure was repeated and was obtained generations of spheres (ES1, ES2 and ES3) and spheres-derived cells in adherent conditions (G1, G2 and G3). Populations were characterized regarding CD133, CD24, CD44, aldehyde dehydrogenase (ALDH), hormonal receptors, HER2, P53 and ß-catenin, fluorine-18 fluorodeoxyglucose ([18F]FDG) uptake and metabolism by NMR spectroscopy. An heterotopic model evaluated differential tumor growth. The spheres self-renewal was higher in ES3. The putative CSC markers CD133, CD44 and ALDH expression were higher in spheres. The expression of estrogen receptor (ER)α and P53 decreased in spheres, ERß and progesterone receptor had no significant changes and ß-catenin showed a tendency to increase. There was a higher 18F-FDG uptake in spheres, which also showed a lower lactate production and an oxidative cytosol status. The tumorigenesis in vivo showed an earlier growth of tumours derived from ES3. Endometrial spheres presented self-renewal and differentiation capacity, expressed CSC markers and an undifferentiated phenotype, showing preference for oxidative metabolism.

Direct Pulp Capping: What is the Most Effective Therapy?-Systematic Review and Meta-Analysis.

INTRODUCTION: Direct pulp capping therapies use biomaterials to protect exposed tissues, inducing repair through the production of a mineralized barrier. The purpose of this study was to compare the effectiveness of biomaterials and techniques by means of a systematic review and meta-analysis. METHODS: The PubMed, Cochrane, and Embase databases were used to search the literature published from January 1, 1980 until August 31, 2017. Studies that met inclusion criteria were screened by 2 authors individually. The meta-analysis was performed on mineral trioxide aggregate (MTA) cement vs calcium hydroxide cement, tricalcium silicate cement vs MTA cement, and adhesive systems vs CaOH cement and evaluated the success rate, inflammatory response, and dentin bridge formation. RESULTS: Forty-six studies were included in the systematic review, while 22 studies were included in the meta-analysis. There was no significant heterogeneity between the studies. MTA cements showed a significantly higher success rate, in all parameters, compared with calcium hydroxide cements (odds ratio = 2.72; 95% confidence interval [CI] = 1.90-3.90; P = 0.000). However, when compared with the tricalcium silicate cements, there were no statistically significant differences (odds ratio = 1.18; 95% CI = 0.53-2.65; P = 0.672). Adhesive systems showed a significantly lower success rate, in all parameters, compared with calcium hydroxide cements (odds ratio = 0.062; 95% CI = 0.024-0.157; P = 0.000). CONCLUSIONS: MTA cements have a higher success rate, with a lower inflammatory response and a more predictable hard dentin barrier formation than calcium hydroxide cements. However, there were no differences, in these parameters, when MTA cement was compared with tricalcium silicate cements. Dental adhesives systems showed the lowest success rates.

Ascorbic Acid Chemosensitizes Colorectal Cancer Cells and Synergistically Inhibits Tumor Growth.

Colorectal cancer (CRC) is continuously classified as one of the most incidental and mortal types of cancer worldwide. The positive outcomes of the conventional chemotherapy are frequently associated with high toxicity, which often leads to the suspension of the treatment. Growing evidences consider the use of pharmacological concentrations of ascorbic acid (AA), better known as vitamin C, in the treatment of cancer. The use of AA in a clinical context is essentially related to the adoption of new therapeutic strategies based on combination regimens, where AA plays a chemosensitizing role. The reduced sensitivity of some tumors to chemotherapy and the highly associated adverse effects continue to be some of the major obstacles in the effective treatment of CRC. So, this paper aimed to study the potential of a new therapeutic approach against this neoplasia with diminished side effects for the patient. This approach was based on the study of the combination of high concentrations of AA with reduced concentrations of drugs conventionally used in CRC patients and eligible for first and second line chemotherapeutic regimens, namely 5-fluorouracilo (5-FU), oxaliplatin (Oxa) or irinotecan (Iri). The evaluation of the potential synergy between the compounds was first assessed in vitro in three CRC cell lines with different genetic background and later in vivo using one xenograft animal model of CRC. AA and 5-FU act synergistically in vitro just for longer incubation times, however, in vivo showed no benefit compared to 5-FU alone. In contrast to the lack of synergy seen in in vitro studies with the combination of AA with irinotecan, the animal model revealed the therapeutic potential of this combination. AA also potentiated the effect of Oxa, since a synergistic effect was demonstrated, in almost all conditions and in the three cell lines. Moreover, this combined therapy (CT) caused a stagnation of the tumor growth rate, being the most promising tested combination. Pharmacological concentrations of AA increased the efficacy of Iri and Oxa against CRC, with promising results in cell lines with more aggressive phenotypes, namely, tumors with mutant or null P53 expression and tumors resistant to chemotherapy.

Revisiting colorectal cancer animal model - An improved metastatic model for distal rectosigmoid colon carcinoma.

Colorectal cancer (CRC) is the second most frequent and fatal cancer in Western countries. Understanding its biology with different incidence along the colon and rectum, genetic profile and how these factors contribute to local/distant progression, has been hampered by the lack of a suitable CRC model. We report a reproducible model, using human CRC cell lines (CL) (WiDr, LS1034, C2BBe1) injected (1 × 107 cells/animal) in RNU rats (n = 55) which underwent cecostomy and descending colostomy with mucosal-cutaneous fistula of the sigmoid colon. CL were characterized by immunohistochemistry: CK20, CDX2, P53, vimentin, Ki67, CD44, CD133, E-cadherin, ß-catenin and CEA; cancer stem cells-immune system interaction was studied and tumor progression was assessed with nuclear medicine imaging (99mTc-MIBI). Animals developed locally invasive tumors and with WiDr neural invasion was registered. Cancer stem cells were detected in WiDr (CD44 positive). All the cell lines stimulated the immune system, being WiDr the most aggressive. Imaging studies demonstrated tumor uptake. With this CRC model we can study the microenvironment role and tumor-stroma interactions. All CL developed primary disease, but only the WiDR established neural invasion which may represent a metastatic pathway. This model can help unveiling the underlying metastatic mechanisms, and ultimately test better therapeutic approaches for CRC.

Butyrate, a dietary fiber derivative that improves irinotecan effect in colon cancer cells.

A diet rich in fiber is associated with a low risk of developing colorectal cancer. Dietary fiber fermentation by intestinal microflora results in the production of butyrate, which has been reported as a chemopreventive agent and a histone deacetylase inhibitor (HDACi). Irinotecan is used as second-line treatment and induces adverse effects with serious life-threatening toxicities in at least 36% of patients. Our study intends to find a synergy that could improve the efficacy and decrease the toxicity of chemotherapy. Results demonstrate that milimolar concentrations of butyrate has an anti-proliferative effect in all three colon cancer cell lines under study, leading to a decrease on cell viability, expression of P21, P53 and ß-catenin, being able to modulate P-glycoprotein activity and to induce apoptosis by modulation of BAX/BCL-2 ratio. Combined therapy has a cytotoxic potential, resulting in a synergistic effect, and allows a reduction in irinotecan concentration needed to reduce IC50. This potential was verified in terms of cell viability and death, cell cycle and expression of P21 and P53. Butyrate and irinotecan act synergistically in the three cancer cell lines, despite the different genetic background and location, and inhibited tumor growth in a xenograft model. Butyrate is able to influence the mechanism of LS1034 cell line chemoresistance. Butyrate in combination with chemotherapeutic agents has an important role for the treatment of colorectal cancer. Such understanding can guide decisions about which patients with colorectal cancer may benefit from therapy with butyrate demonstrating the important role of diet in colorectal cancer treatment.

Advances on photodynamic therapy of melanoma through novel ring-fused 5,15-diphenylchlorins.

The synthesis, photophysical behaviour and photosensitization ability of novel 4,5,6,7-tetrahydropyrazolo[1,5-a]pyridine-fused 5,15-diphenylchlorins against melanoma cells are described. All studied chlorins were found to be extremely active against melanoma cell lines A375 showing IC50 values below 20 nM. Furthermore, a dihydroxymethyl diphenylchlorin was identified as an excellent candidate to allow modulating of different types of cell death, apoptosis vs. necrosis, by varying its concentration. This can be explored as a tool to improve the effectiveness of PDT since inflammatory response resulting from necrotic cell death after PDT can activate the antitumor immune response with implications also regarding the vascular damage. This feature combined with very low cytotoxicity against human melanoma cells in the absence of light activation and against human fibroblast HFF-1 cells makes this chlorin a candidate of choice as a photosensitizer for PDT. A comprehensive photophysical investigation including the determination of quantum yields for fluorescence, singlet oxygen sensitization and internal conversion, lifetimes and rate constants of all the excited state deactivation processes has been undertaken.

Mammospheres of hormonal receptor positive breast cancer diverge to triple-negative phenotype.

OBJECTIVES: This study aimed to characterize mammospheres from hormonal receptor (HR) positive and triple-negative breast cancer (TNBC), hypothesizing a differential profile of CSC and differentiation markers, and a stemness enrichment when successive sphere forming-protocols are performed. METHODS: Breast cancer cells MCF-7 and HCC1806 were submitted to sphere-forming protocols. The first sphere generation (MS1) was cultured in adherent conditions (G1). This procedure was repeated and generations of mammospheres (MS1, MS2, and MS3) and sphere-derived cells in adherent conditions (G1, G2, and G3) were obtained. The mammosphere forming capacity, self-renewal, area and doubling time were evaluated. Flow cytometry regarding CD133, CD24, and CD44 and western-blot regarding aldehyde dehydrogenase (ALDH), hormonal receptors and P53 expression was performed. RESULTS: Breast cancer cell lines harboured the capacity to form spheres, which originated derived adherent populations. The sphere-forming capacity was enhanced in HCC1806-MS3 compared to MS1. Self-renewal was higher in MCF-7 mammospheres, which also had an increased area. The putative CSC markers CD133 showed tendency to be enhanced in mammospheres but the CD44+/CD24-/low phenotype was not identified. The expression of ALDH was greater in mammospheres from MCF-7 and HCC1806 than in the respectively derived adherent cells. The expression of oestrogen receptor (ER)-α, progesterone receptor (PR) and P53 decreased in MCF-7 spheres. ER-ß expression was lower in mammospheres from both cell lines compared with parental and derived adherent populations. CONCLUSIONS: Loss of HR and P53 expression in HR-positive mammospheres evidences the minor population of CSC which shares characteristics with the TNBC phenotype.