Toxicological and pharmacokinetic properties of sucralose-6-acetate and its parent sucralose: in vitro screening assays.

The purpose of this study was to determine the toxicological and pharmacokinetic properties of sucralose-6-acetate, a structural analog of the artificial sweetener sucralose. Sucralose-6-acetate is an intermediate and impurity in the manufacture of sucralose, and recent commercial sucralose samples were found to contain up to 0.67% sucralose-6-acetate. Studies in a rodent model found that sucralose-6-acetate is also present in fecal samples with levels up to 10% relative to sucralose which suggest that sucralose is also acetylated in the intestines. A MultiFlow® assay, a high-throughput genotoxicity screening tool, and a micronucleus (MN) test that detects cytogenetic damage both indicated that sucralose-6-acetate is genotoxic. The mechanism of action was classified as clastogenic (produces DNA strand breaks) using the MultiFlow® assay. The amount of sucralose-6-acetate in a single daily sucralose-sweetened drink might far exceed the threshold of toxicological concern for genotoxicity (TTCgenotox) of 0.15 µg/person/day. The RepliGut® System was employed to expose human intestinal epithelium to sucralose-6-acetate and sucralose, and an RNA-seq analysis was performed to determine gene expression induced by these exposures. Sucralose-6-acetate significantly increased the expression of genes associated with inflammation, oxidative stress, and cancer with greatest expression for the metallothionein 1 G gene (MT1G). Measurements of transepithelial electrical resistance (TEER) and permeability in human transverse colon epithelium indicated that sucralose-6-acetate and sucralose both impaired intestinal barrier integrity. Sucralose-6-acetate also inhibited two members of the cytochrome P450 family (CYP1A2 and CYP2C19). Overall, the toxicological and pharmacokinetic findings for sucralose-6-acetate raise significant health concerns regarding the safety and regulatory status of sucralose itself.

Sex-specific effects of neonatal oral sucrose treatment on growth and liver choline and glucocorticoid metabolism in adulthood.

Hospitalized preterm infants experience painful medical procedures. Oral sucrose is the nonpharmacological standard of care for minor procedural pain relief. Infants are treated with numerous doses of sucrose, raising concerns about potential long-term effects. The objective of this study was to determine the long-term effects of neonatal oral sucrose treatment on growth and liver metabolism in a mouse model. Neonatal female and male mice were randomly assigned to one of two oral treatments (n = 7-10 mice/group/sex): sterile water or sucrose. Pups were treated 10 times/day for the first 6 days of life with 0.2 mg/g body wt of respective treatments (24% solution; 1-4 µL/dose) to mimic what is given to preterm infants. Mice were weaned at age 3 wk onto a control diet and fed until age 16 wk. Sucrose-treated female and male mice gained less weight during the treatment period and were smaller at weaning than water-treated mice (P ≤ 0.05); no effect of sucrose treatment on body weight was observed at adulthood. However, adult sucrose-treated female mice had smaller tibias and lower serum insulin-like growth factor-1 than adult water-treated female mice (P ≤ 0.05); these effects were not observed in males. Lower liver S-adenosylmethionine, phosphocholine, and glycerophosphocholine were observed in adult sucrose-treated compared with water-treated female and male mice (P ≤ 0.05). Sucrose-treated female, but not male, mice had lower liver free choline and higher liver betaine compared with water-treated female mice (P < 0.01). Our findings suggest that repeated neonatal sucrose treatment has long-term sex-specific effects on growth and liver methionine and choline metabolism.

Quantification and cytotoxicity of degradation products (chloropropanols) in sucralose containing e-liquids with propylene glycol and glycerol as base.

Electronic cigarettes (e-cigarettes) have gained increasing popularity in recent years, mostly because they are supposed to be less harmful than regular cigarettes. Therefore, it is highly imperative to investigate possible noxious effects to protect the consumers. E-liquids consist of propylene glycol, glycerol, aroma compounds and sweeteners. One of these sweeteners is a chlorinated version of sucrose, namely sucralose. The aim of this work was to investigate degradation products of sucralose in the presence of propylene glycol and glycerol at different temperatures of commercially available e-cigarettes. Chemical analysis and biological tests were simultaneously performed on e-liquid aerosol condensates. The results of the chemical analysis, which was executed by employing GC-MS/GC-FID, demonstrated high amounts of various chloropropanols. The most abundant one is extremely toxic, namely 3-chloropropane-1,2-diol, which can be detected at concentrations ranging up to 10,000 mg/kg. Furthermore, a cytotoxicity investigation of the condensates was performed on HUVEC/Tert2 cells in which metabolic activity was determined by means of resazurin assay. The cellular metabolic activity significantly decreased by treatment with e-liquid aerosol condensate. Due to the results of this study, we advise against the use of sucralose as sweetener in e-liquids.

Cd36 gene expression in adipose and hepatic tissue mediates the lipids accumulation in liver of obese rats with sucrose-induced hepatic steatosis.

Obesity is considered a global epidemic and is mainly associated with the development of diabetes, cardiovascular diseases and Non-Alcoholic Fatty Liver (NAFLD). The pathogenesis between obesity and hepatic steatosis is partially known, but could involve differentiated or tissue-specific participation of the expression of Cd36 mRNA that codes for a receptor which is a transporter of free fatty acids (FFA) in different tissues, favoring the lipids storage. This relative expression was evaluated in adipose and liver tissue in rats with steatosis after consumption of sucrose for 30 and 40 weeks. Ten Wistar rats were divided into two experimental groups (St-30 and St-40), which received a standard diet plus 30 % sucrose in their water intake. These rats showed a significant increase in abdominal fat, serum biochemical determinations, HOMA-IR; as well as, changes in adipocytes size and mild portal hepatitis and grade 2 hepatic steatosis. The relative expression of Cd36 mRNA increased in liver tissue after 30 (4.5-fold) and 40 (8.5-fold) weeks of sucrose ingestión but no in adipose tissue; with respect to control group (P < 0.05). This expression was associated with a significant increase in the levles of sCD36 in serum, which is indicator of the presence of the FFA transporter in the hepatocyte membrane causing lipids accumulation. The above shows the link between the adipose and hepatic tissue for the accumulation of steatotic fat in the liver through time, mediated by the relative expression of cd36 mRNA that encodes for the FFA transporter.

Artificial sweeteners impair endothelial vascular reactivity: Preliminary results in rodents.

BACKGROUND AND AIMS: Prospective epidemiological studies highlighted recently the link between artificial sweeteners (AS) consumption and the risk of developing cardiometabolic diseases. However, underlying mechanisms remain unknown. Thus, the aim of this preliminary study was to characterize, in a healthy rat population, the effect of chronic AS consumption on body composition and vascular function, an early marker for cardiovascular disease. METHODS AND RESULTS: Healthy Wistar rats followed a 10-week standard diet including the consumption of water sweetened or not with a sucralose/acesulfame potassium solution at different concentrations: for moderate consumption at 1 and 2 mg.kg-1.day-1, respectively or high intake at 15 and 15 mg.kg-1.day-1 for both molecules (acceptable daily intake). Body fat composition has been evaluated and ex vivo aortic vasomotor function has been investigated with a pharmacological approach. CONCLUSION: Both groups of AS-treated rats showed a significant increase in subcutaneous and perirenal adipose tissue mass storage, without changes in total body mass. However, rats that have consumed AS at Acceptable Daily Intake (ADI) concentration revealed a significant vascular endothelial dysfunction compared to other groups. These results are interesting because they will help to better explain the observed increase in cardiometabolic risk.

Evaluation of long-term effects of artificial sweeteners on rat brain: a biochemical, behavioral, and histological study.

The aim of the present study was to compare the effects of artificial sweeteners (aspartame, saccharin, and sucralose) on rat brain. Twenty-four adult male Sprague-Dawley rats were included in the study. The control group (n = 6) received regular tap water, whereas other groups received aspartame (3 mg/kg/day, n = 6,) or saccharin (3 mg/kg/day, n = 6) or sucralose (1.5 mg/kg/day, n = 6) in the drinking water. Following 6 weeks, the passive avoidance learning (PAL) test was performed to evaluate the neurobehavioral effects of sweeteners. The brains were assessed for lipid peroxides, neuron count, and Glial fibrillary acidic protein (GFAP) immunohistochemistry. Our results demonstrated that chronic intake of sweeteners significantly impaired PAL performance in all groups. Hippocampal CA1-CA3 areas revealed significantly lower neuronal count in aspartame and increased GFAP expression in all groups. Brain lipid peroxides were significantly higher in all groups. Our findings suggest that long-term consumption of artificial sweeteners may have harmful effects on cognition and hippocampal integrity in rats.

A histological assessment of effects of sucralose on liver of albino rats.

OBJECTIVE OF THE STUDY: Sucralose is an artificial sweetener freely available under different brand names over the counter. This study is aimed at evaluating the histopathological changes in the liver after administration of sub lethal dose of pure sucralose. MATERIALS AND METHODS: Inbred adult Wistar albino rats weighing about 150-200g of either sex, were divided into 6 control rats and 6 experimental rats. Experimental rats were given sucralose orally by gavage in the dose of 3g/kg/day dissolved in distilled water for 30 days whereas Control rats received equal quantity of distilled water by the same route. The animals were anesthetized with anesthetic ether and then perfused with 10% formal saline. Livers were dissected out. Pieces each having thickness 5mm were taken for paraffin sectioning. 5 micron thick sections were cut using a rotary microtome. Hundred slides were made from each liver and stained with hematoxylin and eosin, periodic acid-schiff (PAS) and Masson's trichrome stain. Slides were evaluated for histomorphological changes. RESULTS: Experimental rats showed features of patchy degeneration of hepatocytes along with Kupffer cells hyperplasia, lymphocytic infiltration, sinusoidal dilatation and fibrosis indicating a definite hepatic damage on regular ingestion of sucralose. Sinusoidal width was also found to be increased in experimental animals as compared to controls. CONCLUSION: It is suggestive that sucralose should be taken with caution to avoid hepatic damage. Effects of ingestion of sucralose on human subjects should be investigated further.

Maternal obesity characterized by gestational diabetes increases the susceptibility of rat offspring to hepatic steatosis via a disrupted liver metabolome.

Maternal obesity is associated with a high risk for gestational diabetes mellitus (GDM), which is a common complication of pregnancy. The influence of maternal obesity and GDM on the metabolic health of the offspring is poorly understood. We hypothesize that GDM associated with maternal obesity will cause obesity, insulin resistance and hepatic steatosis in the offspring. Female Sprague-Dawley rats were fed a high-fat (45%) and sucrose (HFS) diet to cause maternal obesity and GDM. Lean control pregnant rats received low-fat (LF; 10%) diets. To investigate the interaction between the prenatal environment and postnatal diets, rat offspring were assigned to LF or HFS diets for 12 weeks, and insulin sensitivity and hepatic steatosis were evaluated. Pregnant GDM dams exhibited excessive gestational weight gain, hyperinsulinaemia and hyperglycaemia. Offspring of GDM dams gained more weight than the offspring of lean dams due to excess adiposity. The offspring of GDM dams also developed hepatic steatosis and insulin resistance. The postnatal consumption of a LF diet did not protect offspring of GDM dams against these metabolic disorders. Analysis of the hepatic metabolome revealed increased diacylglycerol and reduced phosphatidylethanolamine in the offspring of GDM dams compared to offspring of lean dams. Consistent with altered lipid metabolism, the expression of CTP:phosphoethanolamine cytidylyltransferase, and peroxisomal proliferator activated receptor-α mRNA was reduced in the livers of GDM offspring. GDM exposure programs gene expression and hepatic metabolite levels and drives the development of hepatic steatosis and insulin resistance in young adult rat offspring.

Nitrosative stress and apoptosis in non-anemic healthy rats induced by intravenous iron sucrose similars versus iron sucrose originator.

Iron can both induce and inhibit nitrosative stress. Intracellular iron levels play an important role in nitric oxide (NO(•)) signaling mechanisms. Depending on various factors, such as the cell's redox state and transition metal levels, NO(•) generation may lead to lipid peroxidation and DNA damage as well as both anti- and pro-apoptotic effects. Administration of intravenous iron sucrose originator (IS(ORIG)) has been shown not to cause significant tyrosine nitration or significantly increased caspase 3 levels in non-anemic rats. In this study, the potential of several marketed iron sucrose similars (ISSs) to induce tyrosine nitration and caspase 3 expression in non-anemic rats was assessed. Although the physico-chemical properties of most of the analyzed ISSs complied with the United States Pharmacopeia for iron sucrose injection, all ISSs resulted in higher levels of tyrosine nitration and increased the expression of caspase 3 versus IS(ORIG). Moreover, significant differences were detected in tissue iron distribution between IS(ORIG)- and ISS-treated animals. In general, ISORIG resulted in higher levels of ferritin deposits versus ISSs whereas ISSs showed higher Prussian blue-stainable iron(III) deposits than IS(ORIG). This result suggests that some iron from ISSs bypassed the tightly regulated pathway through resident macrophages of the liver, spleen and bone marrow thus, ending up in the cellular compartment that favors oxidative and or nitrosative stress as well as apoptosis. The results also confirm that polynuclear iron(III)-oxyhydroxide carbohydrates, such as iron sucrose, cannot be fully characterized by physico-chemical methods alone.

The effect of five artificial sweeteners on Caco-2, HT-29 and HEK-293 cells.

CONTEXT: Artificial sweeteners (AS) have been associated with tumor development (including colon cancer) in both animals and humans although evidence has been conflicting. OBJECTIVES: Additional research was thus conducted by studying the effects of 5 AS on the morphology, cell proliferation and DNA in cells by utilizing Caco-2, HT-29 (colon) and HEK-293 (kidney) cell lines. MATERIALS AND METHODS: Cells were exposed to sodium cyclamate, sodium saccharin, sucralose and acesulfame-K (0-50 mM) and aspartame (0-35 mM) over 24, 48 and 72 hours. Morphological changes were presented photographically and % cell viability was determined by using the MTT cell viability assay. Possible DNA damage (comet assay) induced by the AS (0.1, 1 and 10 mM, treated for 24, 48 and 72 hours) was studied. The appearance of "comets" was scored from no damage to severe damage (0-4). RESULTS: Cells became flatter and less well defined at higher AS concentrations (>10 mM). At concentrations >10 mM, decreased cell viability was noted with both increasing concentration and increasing incubation time for all cell lines tested. In general, HEK-293 cells seemed to be less affected then the colon cancer cells. Sucralose and sodium saccharin seemed to elicit the greatest degree of DNA fragmentation of all the sweeteners tested in all the cell lines used. DISCUSSION: Morphological cell alterations, cell viability and DNA fragmentation seemed to be more in the colon cancer cells. CONCLUSIONS: Further studies have to be performed to clarify mechanisms involved causing these alterations in mammalian cells.

Acute effects of feeding fructose, glucose and sucrose on blood lipid levels and systemic inflammation.

BACKGROUND: Recent studies have demonstrated a relationship between fructose consumption and risk of developing metabolic syndrome. Mechanisms by which dietary fructose mediates metabolic changes are poorly understood. This study compared the effects of fructose, glucose and sucrose consumption on post-postprandial lipemia and low grade inflammation measured as hs-CRP. METHODS: This was a randomized, single blinded, cross-over trial involving healthy subjects (n=14). After an overnight fast, participants were given one of 3 different isocaloric drinks, containing 50 g of either fructose or glucose or sucrose dissolved in water. Blood samples were collected at baseline, 30, 60 and 120 minutes post intervention for the analysis of blood lipids, glucose, insulin and high sensitivity C-reactive protein (hs-CRP). RESULTS: Glucose and sucrose supplementation initially resulted in a significant increase in glucose and insulin levels compared to fructose supplementation and returned to near baseline values within 2 hours. Change in plasma cholesterol, LDL and HDL-cholesterol (measured as area under curve, AUC) was significantly higher when participants consumed fructose compared with glucose or sucrose (P<0.05). AUC for plasma triglyceride levels however remained unchanged regardless of the dietary intervention. Change in AUC for hs-CRP was also significantly higher in subjects consuming fructose compared with those consuming glucose (P<0.05), but not sucrose (P=0.07). CONCLUSION: This study demonstrates that fructose as a sole source of energy modulates plasma lipids and hsCRP levels in healthy individuals. The significance of increase in HDL-cholesterol with a concurrent increase in LDL-cholesterol and elevated hs-CRP levels remains to be delineated when considering health effects of feeding fructose-rich diets. REGISTRATION NUMBER FOR CLINICAL TRIALS: ACTRN12614000431628.

Early life stage (ELS) toxicity of sucralose to fathead minnows, Pimephales promelas.

Sucralose, an intense artificial sweetener, has been detected in wastewater and surface waters at concentrations ranging from ng/L to low µg/L. Although over a hundred studies have been conducted to evaluate the safety of sucralose for human consumption, few studies have focused on the chronic ecotoxicological effects of this compound in fish. As a remedy to this data gap, an early-life stage toxicity test was conducted to assess the effects of sucralose on hatching, survival, and growth of fathead minnows (Pimephales promelas). Hatching, survival, and growth were unaffected by 98 mg/L of sucralose. The Lowest-Observed-Effect Concentration (LOEC) and the No-Observed-Effect Concentration (NOEC) for fathead minnows determined by this study are >98 and 98 mg/L, respectively. The results from this study suggest that the concentrations of sucralose detected in the environment are well below those required to cause adverse effects to developing aquatic organisms.

Pathologists' perspective on the study design, analysis, and interpretation of proliferative lesions in a lifetime rodent carcinogenicity bioassay of sucralose.

Sucralose, a sugar substitute first approved for use in 1991, is a non-caloric sweetener regulated globally as a food additive. Based on numerous experimental animal studies (dating to the 1980s) and human epidemiology studies, international health agencies have determined that sucralose is safe when consumed as intended. A single lifetime rodent carcinogenicity bioassay conducted by the Ramazzini Institute (RI) reported that mice fed diets containing sucralose develop hematopoietic neoplasia, but controversy continues regarding the validity and relevance of these data for predicting health effects in humans. The present paper addresses the controversy by providing the perspective of experienced pathologists on sucralose-related animal toxicity and carcinogenicity data generally, and the RI carcinogenicity bioassay findings specifically, using results from publicly available papers and international regulatory authority decisions. In the authors' view, flaws in the design, methodology, data evaluation, and reporting of the RI carcinogenicity bioassay for sucralose diminish the value of the data as evidence that this agent represents a carcinogenic hazard to humans. This limitation will remain until the RI bioassay is repeated under Good Laboratory Practices and the design, data, and accuracy of the pathology diagnoses and interpretations are reviewed by qualified pathologists with experience in evaluating potential chemically-induced carcinogenic hazards.

The Neonicotinoid Imidacloprid Impairs Learning, Locomotor Activity Levels, and Sucrose Solution Consumption in Bumblebees (Bombus terrestris).

Bumblebees carry out the complex task of foraging to provide for their colonies. They also conduct pollination, an ecosystem service of high importance to both wild plants and entomophilous crops. Insecticides can alter different aspects of bumblebee foraging behavior, including the motivation to leave the hive, finding the right flowers, handling flowers, and the ability to return to the colony. In the present study, we assessed how the neonicotinoid imidacloprid affects bumblebees' foraging behavior after exposure to four different treatment levels, including field-realistic concentrations (0 [control], 1, 10, and 100 µg/L), through sucrose solution over 9 days. We observed the behavior of several free-flying bumblebees simultaneously foraging on artificial flowers in a flight arena to register the bees' complex behavior postexposure. To conduct a detailed assessment of how insecticides affect bumblebee locomotor behavior, we used video cameras and analyzed the recordings using computer vision. We found that imidacloprid impaired learning and locomotor activity level when the bumblebees foraged on artificial flowers. We also found that imidacloprid exposure reduced sucrose solution intake and storage. By using automated analyses of video recordings of bumblebee behavior, we identified sublethal effects of imidacloprid exposure at field-realistic doses. Specifically, we observed negative impacts on consumption of sucrose solution as well as on learning and locomotor activity level. Our results highlight the need for more multimodal approaches when assessing the sublethal effects of insecticides and plant protection products in general. Environ Toxicol Chem 2023;42:1337-1345. © 2023 SETAC.

Phytochemical­rich herbal formula ATG­125 protects against sucrose­induced gastrocnemius muscle atrophy by rescuing Akt signaling and improving mitochondrial dysfunction in young adult mice.

The antioxidant capability of herbal remedies has attracted widespread attention, but their molecular mechanisms in a muscle atrophy model have not been explored. The aim of the present study was to compare the bioactivity of sucrose challenged mice following treatment with ATG­125. Here, through a combination of transcriptomic and biomedical analysis, herbal formula ATG­125, a phytochemical­rich formula, was identified as a protective factor against muscle atrophy in sucrose challenged mice. Gene ontology (GO) identified differentially expressed genes that were primarily enriched in the 'negative regulation of proteolysis', 'cellular amino acid metabolic process', 'lipoprotein particle' and 'cell cycle', all of which were associated with the ATG­125­mediated prevention of muscle atrophy, particularly with regard to mitochondrial biogenesis. In skeletal muscle, a set of mitochondrial­related genes, including angiopoietin­like 4, nicotinamide riboside kinase 2 (Nmrk2), pyruvate dehydrogenase lipoamide kinase isozyme 4, Asc­type amino acid transporter 1 and mitochondrial uncoupling protein 3 (Ucp3) were markedly upregulated following ATG­125 intervention. An increase in Nmrk2 and Ucp3 expression were noted after ATG­125 treatment, in parallel with upregulation of the 'nicotinate and nicotinamide metabolism' pathway, as determined using the Kyoto Encyclopedia of Genes and Genomes (KEGG). Furthermore, KEGG pathway analysis revealed the downregulation of 'complement and coagulation cascades', 'cholesterol metabolism', 'biosynthesis of amino acids' and 'PPAR signaling pathway', which were associated with the downregulation of serine (or cysteine) peptidase inhibitor clade A member (Serpina)3, Serpina1b, Serpina1d, Serpina1e, apolipoprotein (Apo)a1 and Apoa2, all of which were cardiovascular and diabetes­associated risk factors and were regulated by ATG­125. In addition, ATG­125 treatment resulted in downregulated mRNA expression levels of ATPase sarcoplasmic/endoplasmic reticulum Ca2+ transporting 2, troponin­I1, troponin­C1 and troponin­T1 in young adult gastrocnemius muscle compared with the sucrose group. Nuclear factor­κB­hypoxia inducible factor­1α­TGFß receptor type­II­vascular endothelial growth factor staining indicated that ATG­125 decreased sucrose­induced chronic inflammation. ATG­125 was sufficient to prevent muscle atrophy, and this protective effect may be mediated through upregulation of AKT phosphorylation, upregulating the insulin growth factor­1R­insulin receptor substrate­PI3K­AKT pathway, which in turn resulted in a forkhead box O­dependent decrease in protein degradation pathways, including regulation of atrogin1 and E3 ubiquitin­protein ligase TRIM63. Peroxisome­proliferator activated receptor γ coactivator 1α (PGC1α) was decreased in young adult mice challenged with sucrose. ATG­125 treatment significantly increased PGC1α and significantly increased UCP­1,2,3 expression levels, which suggested ATG­125 poised the mitochondria for uncoupling of respiration. This effect is consistent with the increased SIRT1 levels and may explain an increase in mitochondria biogenesis. Taken together, the present study showed that ATG­125, as an integrator of protein synthesis and degradative pathways, prevented muscle wasting.

Fate of sucralose through environmental and water treatment processes and impact on plant indicator species.

The degradation and partitioning of sucralose during exposure to a variety of environmental and advanced treatment processes (ATP) and the effect of sucralose on indicator plant species were systematically assessed. Bench scale experiments were used to reproduce conditions from environmental processes (microbial degradation, hydrolysis, soil sorption) and ATPs (chlorination, ozonation, sorption to activated carbon, and UV radiation). Degradation only occurred to a limited extent during hydrolysis, ozonation, and microbial processes indicating that breakdown of sucralose will likely be slow and incomplete leading to accumulation in surface waters. Further, the persistence of sucralose was compared to suggested human tracer compounds, caffeine and acesulfame-K. In comparison sucralose exhibits similar or enhanced characteristics pertaining to persistence, prevalence, and facile detection and can therefore be considered an ideal tracer for anthropogenic activity. Ecological effects of sucralose were assessed by measuring sucrose uptake inhibition in plant cotelydons and aquatic plant growth impairment. Sucralose did not inhibit plant cotelydon sucrose uptake, nor did it effect frond number, wet weight, or growth rate in aquatic plant, Lemna gibba. Though sucralose does not appear toxic to plant growth, the peristent qualities of sucralose may lead to chronic low-dose exposure with largely unknown consequences for human and environmental health.

Cytotoxicity effects of cryoprotectants as single-component and cocktail vitrification solutions.

Cryoprotectant (CPA) cytotoxicity constitutes a challenge in developing cryopreservation protocols, specifically in vitrification where high CPA concentrations are necessary to achieve the ice-free, vitreous state. Few cytotoxicity studies have investigated vitrification-relevant concentrations of CPAs, and the benefits and disadvantages of cocktail solutions and of incorporating non-permeating solutes have not been fully evaluated. In this study, we address these issues by determining the cytotoxicity kinetics for dimethylsulfoxide (Me(2)SO) and 1,2-propanediol (PD) on alginate-encapsulated ßTC-tet mouse insulinomas for a range of concentrations and temperatures. Cytotoxicity kinetics were also determined for two cocktails, DPS (3M Me(2)SO+3M PD+0.5M sucrose) and PEG400 (1M Me(2)SO+5M PD+0.34M poly(ethylene)glycol with M.W. of 400). PD was found to be more cytotoxic than Me(2)SO at higher concentrations and temperatures. This was reflected in PEG400 being more cytotoxic at room temperature than PEG400 at 4°C or DPS at either temperature. Addition of non-permeating solutes increased the cytotoxicity of cocktails. Furthermore, results indicate that CPA cytotoxicity may not be additive and that combining CPAs may increase cytotoxicity synergistically. Finally, when comparing cytotoxic effects towards encapsulated HepG2 and ßTC-tet cells, and towards ßTC-tet cells in capsules and in monolayers, CPAs appear more cytotoxic towards cells with higher metabolic activity. The incorporation of these results in the rational design of CPA addition/removal processes in vitrification is discussed.

Impact of controlled high-sucrose and high-fat diets on eosinophil recruitment and cytokine content in allergen-challenged mice.

Despite an ongoing focus on the role of diet in health and disease, we have only a limited understanding of these concepts at the cellular and molecular levels. While obesity has been clearly recognized as contributing to metabolic syndrome and the pathogenesis of adult asthma, recent evidence has linked high sugar intake alone to an increased risk of developing asthma in childhood. In this study, we examined the impact of diet in a mouse model of allergic airways inflammation with a specific focus on eosinophils. As anticipated, male C57BL/6 mice gained weight on a high-calorie, high-fat diet. However, mice also gained weight on an isocaloric high-sucrose diet. Elevated levels of leptin were detected in the serum and airways of mice maintained on the high-fat, but not the high-sucrose diets. We found that diet alone had no impact on eosinophil numbers in the airways at baseline or their recruitment in response to allergen (Alternaria alternata) challenge in either wild-type or leptin-deficient ob/ob mice. However, both bronchoalveolar lavage fluid and eosinophils isolated from lung tissue of allergen-challenged mice exhibited profound diet-dependent differences in cytokine content. Similarly, while all wild-type mice responded to allergen challenge with significant increases in methacholine-dependent total airway resistance (Rrs), airway resistance in mice maintained on the isocaloric high-sucrose (but not the high-calorie/high-fat) diet significantly exceeded that of mice maintained on the basic diet. In summary, our findings revealed that mice maintained on an isocaloric high-sucrose diet responded to allergen challenge with significant changes in both BAL and eosinophil cytokine content together with significant increases in Rrs. These results provide a model for further exploration of the unique risks associated with a high-sugar diet and its impact on allergen-associated respiratory dysfunction.

Effect of different intravenous iron preparations on lymphocyte intracellular reactive oxygen species generation and subpopulation survival.

BACKGROUND: Infections in hemodialysis (HD) patients lead to high morbidity and mortality rates and are associated with early cardiovascular mortality, possibly related to chronic inflammation. Intravenous (IV) iron is widely administered to HD patients and has been associated with increased oxidative stress and dysfunctional cellular immunity. The purpose of this study was to examine the effect of three commercially available IV iron preparations on intracellular reactive oxygen species generation and lymphocyte subpopulation survival. METHODS: Peripheral blood mononuclear cells (PBMC) were isolated from healthy donor buffy coat. PBMC were cultured and incubated with 100 microg/mL of sodium ferric gluconate (SFG), iron sucrose (IS) or iron dextran (ID) for 24 hours. Cells were then probed for reactive oxygen species (ROS) with dichlorofluorescein-diacetate. In separate studies, isolated PBMCs were incubated with the 25, 50 or 100 microg/mL iron concentrations for 72 hours and then stained with fluorescein conjugated monoclonal antibodies for lymphocyte subpopulation identification. Untreated PBMCs at 24 hours and 72 hours served as controls for each experiment. RESULTS: All three IV iron preparations induced time dependent increases in intracellular ROS with SFG and IS having a greater maximal effect than ID. The CD4+ lymphocytes were most affected by IV iron exposure, with statistically significant reduction in survival after incubation with all three doses (10, 25 and 100 microg/mL) of SFG, IS and ID. CONCLUSION: These data indicate IV iron products induce differential deleterious effects on CD4+ and CD16+ human lymphocytes cell populations that may be mediated by intracellular reactive oxygen species generation. Further studies are warranted to determine the potential clinical relevance of these findings.

Heating of food containing sucralose might result in the generation of potentially toxic chlorinated compounds.

Sucralose is widely used as non-caloric intense artificial sweetener. It was previously considered to be thermally stable and safe. This was based on studies performed in the early 1990s. However, significant concerns have been raised more recently regarding the physicochemical stability of sucralose at high temperatures in the context of food processing. Over the last decades different independently performed studies indicated that sucralose is decomposed at high temperatures, e.g. through cooking. This - in turn - was considered to be associated with the formation of chlorinated potentially toxic compounds, such as chloropropanols and dioxins. In this review, the literature on thermal stability of sucralose and the generation of potentially toxic compounds was assessed and comparatively discussed. Considering the validity of published data, we conclude that sucralose can be degraded at high temperatures, e.g. during cooking or baking of sucralose-containing foods. As a consequence potentially toxic chlorinated compounds might be generated.