Mesenchymal stem cells differentiate into keratinocytes and express epidermal kallikreins: Towards an in vitro model of human epidermis

Epidermal differentiation is a complex process in which keratinocytes go through morphological and biochemical changes in approximately 15 to 30 days. Abnormal keratinocyte differentiation is involved in the pathophysiology of several skin diseases. In this scenario, mesenchymal stem cells (MSCs) emerge as a promising approach to study skin biology in both normal and pathological conditions. Herein, we have studied the differentiation of MSC from umbilical cord into keratinocytes. MSC were cultured in Dulbecco's modified Eagle's medium (DMEM) (proliferation medium) and, after characterization, differentiation was induced by culturing cells in a defined keratinocyte serum-free medium (KSFM) supplemented with epidermal growth factor (EGF) and calcium chloride ions. Cells cultivated in DMEM were used as control. Cultures were evaluated from day 1 to 23, based on the cell morphology, the expression of p63, involucrin and cytokeratins (KRTs) KRT5, KRT10 and KRT14, by quantitative polymerase chain reaction, Western blot analysis or immunofluorescence, and by the detection of epidermal kallikreins activity. In cells grown in keratinocyte serum-free medium with EGF and 1.8 mM calcium, KRT5 and KRT14 expression was shown at the first day, followed by the expression of p63 at the seventh day. KRT10 expression was detected from day seventh while involucrin was observed after this period. Data showed higher kallikrein (KLK) activity in KSFM-cultured cells from day 11th in comparison to control. These data indicate that MSC differentiated into keratinocytes similarly to that occurs in the human epidermis. KLK activity detection appears to be a good methodology for the monitoring the differentiation of MSC into the keratinocyte lineage, providing useful tools for the better understanding of the skin biology.     

How many copies of GSTM1 and GSTT1 are associated with head and neck cancer risk?

Purpose: GSTM1 and GSTT1 present a polymorphism that can drive complete gene deletions. The current methodology can powerfully determine GSTM1 and GSTT1 copy number variations (CNVs), which may clarify the real contribution of each gene copies to the cellular detoxification process and tumour risk. However, only analysing the presence/absence of these genes yielded controversial results for several disorders, including cancer. Because head and neck cancer (HNC) is becoming a serious global health problem, this study determined the CNVs of GSTM1 and GSTT1 in an HNC case-control population and investigated the possible association between gene copy numbers and tumour risk.

Methods: CNV was evaluated by (Ct) 2^?ΔΔCt qPCR methodology in 619 HNC patients and 448 patients with no tumour diagnosis.

Results: The genes copy number range was 0–3. The CNV of GSTM1 and GSTT1 frequencies were similar between the cases and control. Thus, none copy of GSTM1 and GSTT1 were associated with HNC risk. Notwithstanding, one copy of both genes had higher frequencies among individuals who carried GSTM1 and GSTT1.

Conclusions: One copy number of GSTM1 and GSTT1 presented a higher frequency among carrier genes, but the CNV of GSTM1 and GSTT1 was not associated with HNC risk.     

Flaxseed and dried tomato waste used together in laying hens diet

The aim of this study was to investigate the effects of simultaneous supplementation of laying hens with dietary sources of n-3 polyunsaturated fatty acids (PUFA) and carotenoids on egg quality, fatty acids and carotenoid profile of the egg yolk and on feed and yolk lipid peroxidation. A 6-week experiment was carried out with 53-week old laying hens (96 Tetra SL) assigned to a control and three treatment groups supplemented with 5% flaxseeds and different levels of dried tomato waste (DTW, 2.5%, 5.0% and 10.0%). Hens from the groups supplemented with 5% and 7.5% DTW had a significantly lower average daily feed intake and laying percentage as compared to the control. Increased doses of dietary DTW enhanced yolk Roche colour score in direct correlation with the enrichment of egg yolk in carotenoids but decreased their transfer efficiency from feed to egg. After 4 weeks, egg yolk from hens fed with 5% flaxseeds and 7.5% DTW had increased lutein and zeaxanthin levels (by 29% and 24%, respectively) and the colour score was 3.5 fold higher compared to the control group. As a result of the dietary supplementation with flaxseed, the n-3 fatty acid content was 3.1–3.7-fold higher in egg yolk compared with the control and the n-6/n-3 ratio decreased from 18.3 (control) to 4.1–5.4 in supplemented diets. Dietary supplementation with 5% DTW effectively prevented lipid oxidation of eggs enriched with n-3 PUFA, but the increase in DTW content depressed the absorption and deposition of n-3 PUFA in egg yolk.     

Nebulization as a tool for photosensitizer delivery to the respiratory tract

To this day, any photosensitizers for the photodynamic treatment of pulmonary illnesses have been administered intravenously. There is, however, an intrinsic difficulty in reaching the target cells or bacteria in the respiratory system. Nebulization could overcome distribution problems and alleviate side effects by delivering the photosensitizers directly to the lungs. In this study, we evaluated the viability of three photosensitizers (indocyanine green, the chlorine Photodithazine, and the porphyrin Photogem) was evaluated comparatively in a jet nebulizer. Quantitative analysis was performed by looking at the droplet size, extent of nebulization, output over time and stability of the solutions. All of the tested photosensitizers were found to be adequately nebulized. We also demonstrated the delivery of indocyanine green to the pulmonary tract and its activation with infrared light in a murine model using extracorporeal detection of fluorescence. This was an important step toward clinical implementation of the extracorporeally illuminated photodynamic inactivation of pneumonia, recently demonstrated in vivo by this research group.      

Characterization of Core-Shell Alginate Capsules

Food Biophysics - A new droplets millifluidic/inverse gelation based process was used to produce core-shell alginate milli-capsules. Water-in-oil (W/O) emulsion dispersed phase containing Ca2+ ions...     

3,4‐Methylenedioxypyrovalerone prevents while methylone enhances methamphetamine‐induced damage to dopamine nerve endings: β‐ketoamphetamine modulation of neurotoxicity by the dopamine transporter

Methylone, 3,4‐methylenedioxypyrovalerone (MDPV), and mephedrone are psychoactive ingredients of ‘bath salts’ and their abuse represents a growing public health care concern. These drugs are cathinone derivatives and are classified chemically as β‐ketoamphetamines. Because of their close structural similarity to the amphetamines, methylone, MDPV, and mephedrone share most of their pharmacological, neurochemical, and behavioral properties. One point of divergence in their actions is the ability to cause damage to the CNS. Unlike methamphetamine, the β‐ketoamphetamines do not damage dopamine (DA) nerve endings. However, mephedrone has been shown to significantly accentuate methamphetamine neurotoxicity. Bath salt formulations contain numerous different psychoactive ingredients, and individuals who abuse bath salts also coabuse other illicit drugs. Therefore, we have evaluated the effects of methylone, MDPV, mephedrone, and methamphetamine on DA nerve endings. The β‐ketoamphetamines alone or in all possible two‐drug combinations do not result in damage to DA nerve endings but do cause hyperthermia. MDPV completely protects against the neurotoxic effects of methamphetamine while methylone accentuates it. Neither MDPV nor methylone attenuates the hyperthermic effects of methamphetamine. The potent neuroprotective effects of MDPV extend to amphetamine‐, 3,4‐methylenedioxymethamphetamine‐, and MPTP‐induced neurotoxicity. These results indicate that β‐ketoamphetamine drugs that are non‐substrate blockers of the DA transporter (i.e., MDPV) protect against methamphetamine neurotoxicity, whereas those that are substrates for uptake by the DA transporter and which cause DA release (i.e., methylone, mephedrone) accentuate neurotoxicity.

     

Study of the antimalarial properties of hydroxyethylamine derivatives using green fluorescent protein transformed Plasmodium berghei

A rapid decrease in parasitaemia remains the major goal for new antimalarial drugs and thus, in vivo models must provide precise results concerning parasitaemia modulation. Hydroxyethylamine comprise an important group of alkanolamine compounds that exhibit pharmacological properties as proteases inhibitors that has already been proposed as a new class of antimalarial drugs. Herein, it was tested the antimalarial property of new nine different hydroxyethylamine derivatives using the green fluorescent protein (GFP)-expressing Plasmodium berghei strain. By comparing flow cytometry and microscopic analysis to evaluate parasitaemia recrudescence, it was observed that flow cytometry was a more sensitive methodology. The nine hydroxyethylamine derivatives were obtained by inserting one of the following radical in the para position: H, 4Cl, 4-Br, 4-F, 4-CH3, 4-OCH3, 4-NO2, 4-NH2 and 3-Br. The antimalarial test showed that the compound that received the methyl group (4-CH3) inhibited 70% of parasite growth. Our results suggest that GFP-transfected P. berghei is a useful tool to study the recrudescence of novel antimalarial drugs through parasitaemia examination by flow cytometry. Furthermore, it was demonstrated that the insertion of a methyl group at the para position of the sulfonamide ring appears to be critical for the antimalarial activity of this class of compounds.