Diverse antiepileptic drugs increase the ratio of background synaptic inhibition to excitation and decrease neuronal excitability in neurones of the rat entorhinal cortex in vitro

Although most anti-epileptic drugs are considered to have a primary molecular target, it is clear that their actions are unlikely to be limited to effects on a single aspect of inhibitory synaptic transmission, excitatory transmission or voltage-gated ion channels. Systemically administered drugs can obviously simultaneously access all possible targets, so we have attempted to determine the overall effect of diverse agents on the balance between GABAergic inhibition, glutamatergic excitation and cellular excitability in neurones of the rat entorhinal cortex in vitro. We used an approach developed for estimating global background synaptic excitation and inhibition from fluctuations in membrane potential obtained by intracellular recordings. We have previously validated this approach in entorhinal cortical neurones [. Greenhill and Jones (2007a) Neuroscience 147:884-892]. Using this approach, we found that, despite their differing pharmacology, the drugs tested (phenytoin, lamotrigine, valproate, gabapentin, felbamate, tiagabine) were unified in their ability to increase the ratio of background GABAergic inhibition to glutamatergic excitation. This could occur as a result of decreased excitation concurrent with increased inhibition (phenytoin, lamotrigine, valproate), a decrease in excitation alone (gabapentin, felbamate), or even with a differential increase in both (tiagabine). Additionally, we found that the effects on global synaptic conductances agreed well with whole cell patch recordings of spontaneous glutamate and GABA release (our previous studies and further data presented here). The consistency with which the synaptic inhibition:excitation ratio was increased by the antiepileptic drugs tested was matched by an ability of all drugs to concurrently reduce intrinsic neuronal excitability. Thus, it seems possible that specific molecular targets among antiepileptic drugs are less important than the ability to increase the inhibition:excitation ratio and reduce overall neuronal and network excitability. © 2010 IBRO.

Blood-brain barrier in vitro model: A tissue engineering approach and validation

This dissertation evaluated the feasibility of using commercially available immortalized cell lines in building a tissue engineered in vitro blood-brain barrier (BBB) co-culture model for preliminary drug development studies. Mouse endothelial cell line and rat astrocyte cell lines purchased from American Type Culture Collections (ATCC) were the building blocks of the co-culture model. An astrocyte derived acellular extracellular matrix (aECM) was introduced in the co-culture model to provide a novel in vitro biomimetic basement membrane for the endothelial cells to form endothelial tight junctions. Trans-endothelial electrical resistance (TEER) and solute mass transport studies were engaged to quantitatively evaluate the tight junction formation on the in-vitro BBB models. Immuno-fluorescence microscopy and Western Blot analysis were used to qualitatively verify the in vitro expression of occludin, one of the earliest discovered tight junction proteins. Experimental data from a total of 12 experiments conclusively showed that the novel BBB in vitro co-culture model with the astrocyte derived aECM (CO+aECM) was promising in terms of establishing tight junction formation represented by TEER values, transport profiles and tight junction protein expression when compared with traditional co-culture (CO) model setups and endothelial cells cultured alone. Experimental data were also found to be comparable with several existing in vitro BBB models built from various methods. In vitro colorimetric sulforhodamine B (SRB) assay revealed that the co-cultured samples with aECM resulted in less cell loss on the basal sides of the insert membranes than that from traditional co-culture samples. The novel tissue engineering approach using immortalized cell lines with the addition of aECM was proven to be a relevant alternative to the traditional BBB in vitro modeling.

Multifunctional nanoparticles in cancer: In vitro characterization, in vivo distribution, and cellular response after laser-NIR dye-induced heating

A novel biocompatible and biodegradable polymer, termed poly(Glycerol malate co-dodecanedioate) (PGMD), was prepared by thermal condensation method and used for fabrication of nanoparticles (NPs). PGMD NPs were prepared using the single oil emulsion technique and loaded with an imaging/hyperthermia agent (IR820) and a chemotherapeutic agent (doxorubicin, DOX). The size of the void PGMD NPs, IR820-PGMD NPs and DOX-IR820-PGMD NPs were approximately 90 nm, 110 nm, and 125 nm respectively. An acidic environment (pH=5.0) induced higher DOX and IR820 release compared to pH=7.4. DOX release was also enhanced by exposure to laser, which increased the temperature to 42°C. Cytotoxicity of DOX-IR820-PGMD NPs was comparable in MES-SA but was higher in Dx5 cells compared to free DOX plus IR820 (p<0.05). The combination of hyperthermia (HT) and chemotherapy improved cytotoxicity in both cell lines. We also explored the cellular response after rapid, short-term and low thermal dose (laser/Dye/NP) induced-heating, and compared it to slow, long-term and high thermal dose cell incubator heating by investigating the reactive oxygen species (ROS) level, hypoxia-inducible factor-1&agr; (HIF-1&agr;) and vascular endothelial growth factor (VEGF) expression. The cytotoxicity of IR820-PGMD NPs after laser/Dye/NP HT resulted in higher cancer cell killing compared to incubator HT. ROS level, HIF-1&agr; and VEGF expression were elevated under incubator HT, while maintained at the baseline level under the laser/Dye/NP HT. In vivo mouse studies showed that NP formulation significantly improved the plasma half-life of IR820 after tail vein injection. Significant lower IR820 content was observed in kidney in DOX-IR820-PGMD NP treatment as compared to free IR820 treatment in our biodistribution studies (p<0.05). In conclusion, both IR820-PGMD NPs and DOX-IR820-PGMD NPs were successfully developed and used for both imaging and therapeutic purposes. Rapid and short-term laser/Dye/NP HT, with a low thermal dose, did not up-regulate HIF-1&agr; and VEGF expression, whereas slow and long-term incubator HT, with a high thermal dose, can enhance expression of both HIF-1&agr; and VEGF.^

Targeted Brain Derived Neurotropic Factors (BDNF) Delivery across the Blood-Brain Barrier for Neuro-Protection Using Magnetic Nano Carriers: An In-Vitro Study

Parenteral use of drugs; such as opiates exert immunomodulatory effects and serve as a cofactor in the progression of HIV-1 infection, thereby potentiating HIV related neurotoxicity ultimately leading to progression of NeuroAIDS. Morphine exposure is known to induce apoptosis, down regulate cAMP response element-binding (CREB) expression and decrease in dendritic branching and spine density in cultured cells. Use of neuroprotective agent; brain derived neurotropic factor (BDNF), which protects neurons against these effects, could be of therapeutic benefit in the treatment of opiate addiction. Previous studies have shown that BDNF was not transported through the blood brain barrier (BBB) in-vivo.; and hence it is not effectivein-vivo. Therefore development of a drug delivery system that can cross BBB may have significant therapeutic advantage. In the present study, we hypothesized that magnetically guided nanocarrier may provide a viable approach for targeting BDNF across the BBB. We developed a magnetic nanoparticle (MNP) based carrier bound to BDNF and evaluated its efficacy and ability to transmigrate across the BBB using an in-vitro BBB model. The end point determinations of BDNF that crossed BBB were apoptosis, CREB expression and dendritic spine density measurement. We found that transmigrated BDNF was effective in suppressing the morphine induced apoptosis, inducing CREB expression and restoring the spine density. Our results suggest that the developed nanocarrier will provide a potential therapeutic approach to treat opiate addiction, protect neurotoxicity and synaptic density degeneration.

Development of a Lab-on-a-Chip Device for Rapid Nanotoxicity Assessment In Vitro

Increasing useof nanomaterials in consumer products and biomedical applications creates the possibilities of intentional/unintentional exposure to humans and the environment. Beyond the physiological limit, the nanomaterialexposure to humans can induce toxicity. It is difficult to define toxicity of nanoparticles on humans as it varies by nanomaterialcomposition, size, surface properties and the target organ/cell line. Traditional tests for nanomaterialtoxicity assessment are mostly based on bulk-colorimetric assays. In many studies, nanomaterials have found to interfere with assay-dye to produce false results and usually require several hours or days to collect results. Therefore, there is a clear need for alternative tools that can provide accurate, rapid, and sensitive measure of initial nanomaterialscreening. Recent advancement in single cell studies has suggested discovering cell properties not found earlier in traditional bulk assays. A complex phenomenon, like nanotoxicity, may become clearer when studied at the single cell level, including with small colonies of cells. Advances in lab-on-a-chip techniques have played a significant role in drug discoveries and biosensor applications, however, rarely explored for nanomaterialtoxicity assessment. We presented such cell-integrated chip-based approach that provided quantitative and rapid response of cellhealth, through electrochemical measurements. Moreover, the novel design of the device presented in this study was capable of capturing and analyzing the cells at a single cell and small cell-population level. We examined the change in exocytosis (i.e. neurotransmitterrelease) properties of a single PC12 cell, when exposed to CuOand TiO2 nanoparticles. We found both nanomaterials to interfere with the cell exocytosis function. We also studied the whole-cell response of a single-cell and a small cell-population simultaneously in real-time for the first time. The presented study can be a reference to the future research in the direction of nanotoxicity assessment to develop miniature, simple, and cost-effective tool for fast, quantitative measurements at high throughput level. The designed lab-on-a-chip device and measurement techniques utilized in the present work can be applied for the assessment of othernanoparticles' toxicity, as well.

Avaliação da atividade anti-Helicobacter pylori e citotóxica in vitro de extratos orgânicos obtidos das folhas de Encholirium spectabile e Syzygium cumini

Helicobacter pylori is a spiral, Gram negative, mobile, and microaerophilic bacteria recognized as a major cause of gastritis, ulcer, gastric cancer, and gastric low grade, B cell, mucosa – associated lymphoid tissue (MALT) lymphoma, constituting an important microorganism in medical microbiology. Its importance comes from the difficulty of treatment because the requirement of multiple drugs use, besides the increasing emergence of resistant and multiresistant strains to antibiotics used in th e clinic. In order to expand safe and effective therapeutic options , chemical studies on medicinal plants by obtaining extracts, fractions, isolated compounds or essential oils with some biological activity has been intensified . Given the above, the objective was to evaluate the inhi bitory activity of organic extracts derived from Syzygium cumini and Encholirium spectabile, with antiulcer history, and the essential oil, obtained from S. cumini, against H. pylori (ATCC 43504) by the disk diffusion method, for qualitative evaluation, an d determination of minimum inhibitory concentration (MIC) using the broth microdilution method, for quantitative analysis. Also was evaluated the extracts in vitro toxicity by a hemolytic assay using sheep red blood cells, and VERO and HeLa cells using the MTT assay to analyze cell viability. The extracts of both plant used in antimicrobial assays did not inhibit bacterial growth, however the essential oil of S. cumini (SCFO) proved effective, showing MIC value of 205 μg/mL (0.024 % dilution of the original oil). In the hemolytic assay, the same oil shows moderate toxicity, by promote 25% hemolysis at 1000 μg/mL. Regarding the cytotoxicity in cell culture, the SCFO, at 260 μg/mL, affected the cell viability around 80% of HeLa and 50% of VERO cells. So the oi l obtained from S. cumini leaves has antimicrobial activity against H. pylori and cytotoxicity potential, suggesting a source of new molecule drug candidates, since new stages of toxicity in vitro and in vivo, as well, chemical characterization be evaluate d. Moreover, the development of a prospective drug delivery system can result in a prototype to be used in preclinical tests.

Avaliação dos efeitos tóxicos in vitro e in vivo do extrato hidroetanólico dos frutos de Genipa americana L. (Rubiaceae) em camundongos Swiss

The therapeutic use of medicinal plants has contributed since antiquity in a beneficial way for health. However, many species lacks of scientific evidence which provide basis for their use in therapeutic practice. In this context is the Genipa americana L. species (Rubiaceae), popularly known as jenipapo and used to treat syfilis, ulcer and hemorrhagic disturbs. It's also used against bruising, as tonic and as aphrodisiac. Due this species lacks toxicological studies, the aim of this study was to evaluate the toxicity in vivo (acute and sub-chronic toxicity) and in vitro (cytotoxicity) of the hydroethanolic extract from G. americana fruits. The hydroethanolic extract of G. americana fruits was prepared by maceration. A preliminary phytochemical analysis was performed to assess the presence of secondary metabolites in the extract. The cytotoxicity study of the extract (0.1, 1.0, 10, 100 and 1000 mg / 100 ul) were performed against normal cells (3T3) and tumor (786-0, HepG2 and B16), analyzed by the MTT assay. To evaluate the acute (single dose of 2000 mg / Kg) and subchronic (100, 500 and 1000 mg / kg for 30 days) toxicity Swiss mice of both sexes were used. At the end of the experiment, blood samples and organs were collected for analysis. Data between groups were compared by t test or ANOVA with Dunnett's post-test with 5% significance level. The phytochemical study of the extracts mainly indicated the presence of iridoids. Results for cytotoxicity tests showed up to 70% inhibition of B16 cell line at a dose of 1000 mg / 100 ul, and up to 29% inhibition of 786-0 at a dose of 10 ug / 100 ul. The extract did not cause death in 3T3 and HepG2 cells. During the in vivo assays, there were no animal deaths. Analysis of blood samples revealed that the animals submitted to the evaluation of acute toxicity had changes in AST and ALT, and that the animals evaluated for subchronic toxicity showed changes in the relative wet weight of the kidney and plasma urea concentration. No differences were observed between groups on histopathological evaluation of the collected organs. Despite the changes found in the in vivo toxicity tests, using the criteria described by the OECD Guidelines, it is suggested that the hydroethanolic extract of the fruits of the G. americana is classified as low toxicity. The cytotoxicity of the extract suggests that they have potential against melanoma cell lines (B16).

Avaliação dos efeitos tóxicos in vitro e in vivo do extrato hidroetanólico dos frutos de Genipa americana L. (Rubiaceae) em camundongos Swiss

The therapeutic use of medicinal plants has contributed since antiquity in a beneficial way for health. However, many species lacks of scientific evidence which provide basis for their use in therapeutic practice. In this context is the Genipa americana L. species (Rubiaceae), popularly known as jenipapo and used to treat syfilis, ulcer and hemorrhagic disturbs. It's also used against bruising, as tonic and as aphrodisiac. Due this species lacks toxicological studies, the aim of this study was to evaluate the toxicity in vivo (acute and sub-chronic toxicity) and in vitro (cytotoxicity) of the hydroethanolic extract from G. americana fruits. The hydroethanolic extract of G. americana fruits was prepared by maceration. A preliminary phytochemical analysis was performed to assess the presence of secondary metabolites in the extract. The cytotoxicity study of the extract (0.1, 1.0, 10, 100 and 1000 mg / 100 ul) were performed against normal cells (3T3) and tumor (786-0, HepG2 and B16), analyzed by the MTT assay. To evaluate the acute (single dose of 2000 mg / Kg) and subchronic (100, 500 and 1000 mg / kg for 30 days) toxicity Swiss mice of both sexes were used. At the end of the experiment, blood samples and organs were collected for analysis. Data between groups were compared by t test or ANOVA with Dunnett's post-test with 5% significance level. The phytochemical study of the extracts mainly indicated the presence of iridoids. Results for cytotoxicity tests showed up to 70% inhibition of B16 cell line at a dose of 1000 mg / 100 ul, and up to 29% inhibition of 786-0 at a dose of 10 ug / 100 ul. The extract did not cause death in 3T3 and HepG2 cells. During the in vivo assays, there were no animal deaths. Analysis of blood samples revealed that the animals submitted to the evaluation of acute toxicity had changes in AST and ALT, and that the animals evaluated for subchronic toxicity showed changes in the relative wet weight of the kidney and plasma urea concentration. No differences were observed between groups on histopathological evaluation of the collected organs. Despite the changes found in the in vivo toxicity tests, using the criteria described by the OECD Guidelines, it is suggested that the hydroethanolic extract of the fruits of the G. americana is classified as low toxicity. The cytotoxicity of the extract suggests that they have potential against melanoma cell lines (B16).

Reprogramming of distinct astroglial populations into specific neuronal subtypes in vitro and in vivo

Recently, the field of cellular reprogramming has been revolutionized by works showing the potential to directly lineage-reprogram somatic cells into neurons upon overexpression of specific transcription factors. This technique offers a promising strategy to study the molecular mechanisms of neuronal specification, identify potential therapeutic targets for neurological diseases and eventually repair the central nervous system damaged by neurological conditions. Notably, studies with cortical astroglia revealed the high potential of these cells to reprogram into neurons using a single neuronal transcription factor. However, it remains unknown whether astroglia isolated from different regions of the central nervous system have the same neurogenic potential and generate induced neurons (iN) with similar phenotypes. Similarly, little is known about the fate that iNs could adopt after transplantation in the brain of host animals. In this study we compare the potential to reprogram astroglial cells isolated from the postnatal cerebral cortex and cerebellum into iNs both in vitro and in vivo using the proneural transcription factors Neurogenin-2 (Neurog2) and Achaete scute homolog-1 (Ascl1). Our results indicate cerebellar astroglia can be reprogrammed into induced neurons (iNs) with similar efficiencies to cerebral cortex astroglia. Notably however, while iNs in vitro adopt fates reminiscent of cortical or cerebellar neurons depending on the astroglial population used for reprogramming, in situ, after transplantation in the postnatal and adult mouse brain, iNs adopt fates compatible with the region of integration. Thus, our data suggest that the origin of the astroglial population used for lineage-reprogramming affects the fate of iNs in vitro, but this imprinting can be overridden by environmental cues after grafting.

Computer assisted synthesis and in-vitro cytotoxic evaluation of new pyrazole-fused isoquinolinoquinones derivatives as PI3K receptor antagonist with promising antitumoral activity

The importance of pyrazole and isoquinoline-5,8-dione scaffolds in medical chemistry is underlined by the high number of drugs currently on trading that contains these active ingredients. Due to their cytotoxic capability, the interest of medicinal chemists in these heterocyclic rings has grown exponentially especially, for cancer therapy. In this project, the first synthesis of pyrazole-fused isoquinoline-5,8-diones has been developed. 1,3-Dipolar cycloaddition followed by oxidative aromatization, established by our research group, has been employed. Screening of reaction conditions and characterization studies about the regioselectivity have been successfully performed. A remote control of regioselectivity, to achieve the two possible regioisomers has been accomplished. Through Molecular Docking studies, Structure-Activity relationship of differently substituted scaffolds containing our central core proved that a family of PI3K inhibitors have been discovered. Finally, in order to verify the promising antitumor activity, a first test of cell viability in vitro on T98G cell line of a solid brain tumor, the Glioblastoma Multiforme, showed cytotoxic inhibition comparable to currently trade anticancer drugs.

An in vitro investigation on the cytotoxic and nuclear receptor transcriptional activity of the mycotoxins fumonisin B1 and beauvericin

Fumonisin B1 (FB1) and beauvericin (BEA) are secondary metabolites of filamentous fungi, which under appropriate temperature and humidity conditions may develop on various foods and feeds. To date few studies have been performed to evaluate the toxicological and endocrine disrupting effects of FB1 and BEA. The present study makes use of various in vitro bioassays including; oestrogen, androgen, progestagen and glucocorticoid reporter gene assays (RGAs) for the study of nuclear receptor transcriptional activity, the thiazolyl blue tetrazolium bromide (MTT) assay to monitor cytotoxicity and high content analysis (HCA) for the detection of pre-lethal toxicity in the RGA and Caco-2 human colon adenocarcinoma cells. At the receptor level, 0.001-10μM BEA or FB1 did not induce any agonist responses in the RGAs. However at non-cytotoxic concentrations, an antagonistic effect was exhibited by FB1 on the androgen nuclear receptor transcriptional activity at 10μM and BEA on the progestagen and glucocorticoid receptors at 1μM. MTT analysis showed no decrease in cell viability at any concentration of FB1, whereas BEA showed a significant decrease in viability at 10μM. HCA analysis confirmed that the reduction in the progestagen receptor transcriptional activity at 1μM BEA was not due to pre-lethal toxicity. In addition, BEA (10μM) induced significant toxicity in both the TM-Luc (progestagen responsive) and Caco-2 cells.

PEG-based solid dispersions enhanced the solubility and dissolution of eprosartan mesylate in-vitro

Abstract - This study investigates the effect of solid dispersions prepared from of polyethylene glycol (PEG) 3350 and 6000 Da alone or combined with the non-ionic surfactant Tween 80 on the solubility and dissolution rate of a poorly soluble drug eprosartan mesylate (ESM) in attempt to improve its bioavailability following its oral administration.

INTRODUCTION

ESM is a potent anti-hypertension [1]. It has low water solubility and is classified as a Class II drug as per the Biopharmaceutical Classification Systems (BCS) leading to low and variable oral bioavailability (approximately 13%). [2]. Thus, improving ESM solubility and/or dissolution rate would eventually improve the drug bioavailability. Solid dispersion is widely used technique to improve the water solubility of poorly water-soluble drugs employing various biocompatible polymers. In this study, we aimed to enhance the solubility and dissolution of EMS employing solid dispersion (SD) formulated from two grades of poly ethylene glycol (PEG) polymers (i.e. PEG 3350 & PEG 6000 Da) either individually or in combination with Tween 80.

MATERIALS AND METHODS

ESM SDs were prepared by solvent evaporation method using either PEG 3350 or PEG 6000 at various (drug: polymer, w/w) ratios 1:1, 1:2, 1:3, 1:4, 1:5 alone or combined with Tween 80 added at fixed percentage of 0.1 of drug by weight?. Physical mixtures (PMs) of drug and carriers were also prepared at same ratios. Drug solid dispersions and physical mixtures were characterized in terms of drug content, drug dissolution using dissolution apparatus USP II and assayed using HPLC method. Drug dissolution enhancement ratio (ER %) from SD in comparison to the plain drug was calculated. Drug-polymer interactions were evaluated using Differential Scanning Calorimetry (DSC) and FT-IR.

RESULTS AND DISCUSSION

The in vitro solubility and dissolution studies showed SDs prepared using both polymers produced a remarkable improvement (p<0.05) in comparison to the plain drug which reached around 32% (Fig. 1). The dissolution enhancement ratio was polymer type and concentration-dependent. Adding Tween 80 to the SD did not show further dissolution enhancement but reduced the required amount of the polymer to get the same dissolution enhancement. The DSC and FT-IR studies indicated that using SD resulted in transformation of drug from crystalline to amorphous form.

CONCLUSIONS

This study indicated that SDs prepared by using both polymers i.e. PEG 3350 and PEG 6000 improved the in-vitro solubility and dissolution of ESM remarkably which may result in improving the drug bioavailability in vivo.

Acknowledgments

This work is a part of MSc thesis of O.M. Ali at the Faculty of Pharmacy, Aleppo University, Syria.

REFERENCES

[1] Ruilope L, Jager B: Eprosartan for the treatment of hypertension. Expert Opin Pharmacother 2003; 4(1):107-14

[2] Tenero D, Martin D, Wilson B, Jushchyshyn J, Boike S, Lundberg, D, et al. Pharmacokinetics of intravenously and orally administered Eprosartan in healthy males: absolute bioavailability and effect of food. Biopharm Drug Dispos 1998; 19(6): 351- 6.

In vitro efficacy of selected medicinal plants from Cholistan desert, Pakistan, against gastrointestinal helminths of sheep and goats

Gastrointestinal helminths are a major constraint to small ruminants in extensive husbandry systems of tropical regions. Yet, unavailability, high prices, side effects, and development of parasite resistance often limit the use of synthetic anthelmintics. Traditional medicinal plants might be an effective low-cost alternative. Therefore the in vitro anthelmintic activity of leaf extracts of the ligneous plants Capparis decidua, Salsola foetida, Suaeda fruticosa, Haloxylon salicornicum, and Haloxylon recurvum from Cholistan, Pakistan, was investigated against adult worms of Haemonchus contortus, Trichuris ovis, and Paramphistomum cervi. Various concentrations (from 7.8 to 500 mg dry matter ml^(−1)) of three extracts (aqueous, methanol, and aqueous-methanol) of each plant were tested at different time intervals for their anthelmintic activity via adult motility assay. Plant species (p<=0.01), extract type (p<=0.001), parasite species (p<=0.001), extract concentration (p<=0.001), time of exposure (p<=0.001) and their interactions (p<=0.001) affected the number of immobile or dead helminths. The 50% lethal concentration (LC_(50)) values indicated that the methanol and aqueous-methanol extracts of C. decidua, H. recurvum, and H. salicornicum as well as the methanol extract of S. fruticosa have the potential to be developed into plant-based remedies against the studied helminths. Further studies are needed to investigate the in vivo anthelmintic activity of these extracts, in order to develop effective, cheap and locally available anthelmintics for pastoralists in Cholistan and neighbouring desert regions.

Études in vivo et in vitro sur le potentiel néphroprotecteur de plantes médicinales anti-diabétiques de la pharmacopée traditionnelle des Cris de la Baie-James

Notre équipe a identifié le thé Labrador [Rhododendron groenlandicum L. (Ericaceae)] comme une plante potentiellement antidiabétique de la pharmacopée traditionnelle des Cris de la Baie James orientale. Dans la présente étude, nous avons évalué les effets néphroprotecteurs potentiels de la plante. De la microalbuminurie et de la fibrose rénale ont été développées chez des souris alimentées avec une diète grasse (DG). Le R. groenlandicum améliore d’une façon non-significative la microalbuminurie, avec des valeurs de l’aire sous la courbe (ACR) diminuant de 0.69 à 0.53. La valeur de la fibrose rénale qui était, à l’origine, de 4.85 unités arbitraires (UA) dans des souris alimentées à la DG, a chuté à 3.27 UA après avoir reçu un traitement de R. groenlandicum. Le R. groenlandicum a réduit la stéatose rénale de presque la moitié alors que l’expression du facteur de modification Bcl-2 (Bmf) a chuté de 13.96 UA à 9.43 UA. Dans leur ensemble les résultats suggèrent que le traitement avec R. groenlandicum peut améliorer la fonction rénale altérée par DG. Dans l’étude subséquente, notre équipe a identifié 17 espèces de la forêt boréale, de la pharmacopée traditionnelle des Cris de la Baie James orientale, qui ont présenté des activités biologiques prometteuses in vitro et in vivo dans le contexte du DT2. Nous avons maintenant examiné ces 17 extraits afin d’identifier lesquels possèdent un potentiel cytoprotecteur rénale en utilisant des cellules Madin Darby Canine Kidney (MDCK) mises à l’épreuve dans un médium hypertonique. Nous concluons que plusieurs plantes antidiabétiques Cris exercent une activité de protection rénale qui pourrait être pertinente dans le contexte de la néphropathie diabétique (ND) qui affecte une proportion importante des Cris. La G. hispidula et la A. balsamea sont parmi les plantes les plus puissantes dans ce contexte et elles semblent protectrices principalement en inhibant la caspase 9 dans la voie de signalisation apoptotique mitochondriale. Finalement, nous avons utilisé une approche de fractionnement guidée par un test biologique pour identifier les fractions actives et les composés de A. balsamea avec un potentiel de protection rénale in vitro dans des cellules MDCK mises au défi avec un médium hypertonique. La fraction d’hexane (Hex) possède le potentiel le plus élevé parmi toutes les fractions de solvant contre les dommages cellulaires induits par le stress hypertonique. Dans des études précédentes, trois composés purs ont été identifiés à partir de la fraction Hex, à savoir, l’acide abiétique, l’acide déhydroabiétique et le squalène. L’acide abiétique se distinguait par son effet puissant dans le maintien de la viabilité des cellules MDCK (AnnV-/PI-) à un niveau relativement élevé (augmentation de 25.48% relative au stress hypertonique, P<0.0001), ainsi qu’une réduction significative (diminution de 20.20% par rapport au stress hypertonique, P<0.0001) de l’apoptose de stade précoce (AnnV+/PI-). L’acide abiétique peut donc servir à normaliser les préparations traditionnelles d’A. balsamea et à trouver des applications potentielles dans le traitement de la néphropathie diabétique. Les trois études ont été intrinsèquement liées les unes aux autres, par conséquent, nous avons réussi à identifier R. groenlandicum ainsi que A. balsamea comme nouvelles plantes prometteuses contre la néphropathie diabétique. Nous croyons que ces résultats profiteront à la communauté crie pour la gestion des complications diabétiques, en particulier la néphropathie diabétique. En parallèle, nos données pourraient faire avancer l'essai clinique de certaines plantes médicinales de la pharmacopée traditionnelle des Cris de la Baie James orientale du Canada.

Assessment of genotoxicity of aflatoxin M1 and B1 contaminated milks after in vitro human digestion

Introduction - Milk is considered a complete food from the nutritional point of view. Milk can be exposed to various types of contamination, such as mycotoxins. These metabolites are naturally occurring toxic compounds produced by fungi. Several studies on milk samples have reported the presence of aflatoxin B1 (AFB1) and M1 (AFM1), due to the high incidence in samples intended for human consumption, carcinogenicity proven AFB1 and resistance of the contaminants to the process of digestion, making those available for intestinal absorption. Considering these aspects, the objective of this study was to evaluate the genotoxicity of milk samples contaminated by AFB1 and AFM1 before and after the action of lactic acid bacteria using Caco-2 intestinal human cells.