Orally delivered microencapsulated probiotic formulation favorably impacts polyp formation in APC (Min/+) model of intestinal carcinogenesis.

The development of intestinal polyps in an orthotopic colorectal mouse model, receiving a probiotic yogurt formulation containing microencapsulated live Lactobacillus acidophilus cells was investigated. The expression of various immunohistochemical markers namely CD8, Mac-1, Ki-67, and cleaved caspase-3, was evaluated. Results suggest that the probiotic formulation decreases overall intestinal inflammation. Mice receiving the probiotic formulation were found to develop almost two-fold fewer tumors in the small intestines. In the large intestine, however, there was no significant difference observed among polyp numbers. The formulation appears to have potential application in the prevention of various GI pathological conditions.

Oral probiotic microcapsule formulation ameliorates non-alcoholic fatty liver disease in Bio F1B Golden Syrian hamsters.

The beneficial effect of a microencapsulated feruloyl esterase producing Lactobacillus fermentum ATCC 11976 formulation for use in non-alcoholic fatty liver disease (NAFLD) was investigated. For which Bio F1B Golden Syrian hamsters were fed a methionine deficient/choline devoid diet to induce non-alcoholic fatty liver disease. Results, for the first time, show significant clinical benefits in experimental animals. Examination of lipids show that concentrations of hepatic free cholesterol, esterified cholesterol, triglycerides and phospholipids were significantly lowered in treated animals. In addition, serum total cholesterol, triglycerides, uric acid and insulin resistance were found to decrease in treated animals. Liver histology evaluations showed reduced fat deposits. Western blot analysis shows significant differences in expression levels of key liver enzymes in treated animals. In conclusion, these findings suggest the excellent potential of using an oral probiotic formulation to ameliorate NAFLD.

Microencapsulated Saccharomyces cerevisiae column bioreactor for potential use in renal failure uremia.

A novel bioreactor containing viable APA microencapsulated yeast cells was designed. Rat plasma was used for perfusion. Yeast cell loading and perfusion flow rate were studied to maximize urea removal. An increase in column loading from 25% to 100%, increased urea removal from 5.67 ± 1.34% to 30.45 ± 0.48%. An increase in flow rate from low to high, increased urea removal from 30.46% to 40.4%. At 100% column loading and high flow rate, the creatinine and phosphate concentrations decreased by 22% and 10%, respectively, while ammonia concentrations increased by 58.9% (p < 0.05). Our in-vitro perfusion study demonstrates that microencapsulated yeast cells can remove urea efficiently.

Diet-induced metabolic hamster model of nonalcoholic fatty liver disease.

BACKGROUND: Obesity, hypercholesterolemia, elevated triglycerides, and type 2 diabetes are major risk factors for metabolic syndrome. Hamsters, unlike rats or mice, respond well to diet-induced obesity, increase body mass and adiposity on group housing, and increase food intake due to social confrontation-induced stress. They have a cardiovascular and hepatic system similar to that of humans, and can thus be a useful model for human pathophysiology. METHODS: Experiments were planned to develop a diet-induced Bio F(1)B Golden Syrian hamster model of dyslipidemia and associated nonalcoholic fatty liver disease in the metabolic syndrome. Hamsters were fed a normal control diet, a high-fat/high-cholesterol diet, a high-fat/high-cholesterol/methionine-deficient/choline-devoid diet, and a high-fat/high-cholesterol/choline-deficient diet. Serum total cholesterol, high-density lipoprotein cholesterol, low-density lipoprotein cholesterol, triglycerides, glucose, atherogenic index, and body weight were quantified biweekly. Fat deposition in the liver was observed and assessed following lipid staining with hematoxylin and eosin and with oil red O. RESULTS: In this study, we established a diet-induced Bio F(1)B Golden Syrian hamster model for studying dyslipidemia and associated nonalcoholic fatty liver disease in the metabolic syndrome. Hyperlipidemia and elevated serum glucose concentrations were induced using this diet. Atherogenic index was elevated, increasing the risk for a cardiovascular event. Histological analysis of liver specimens at the end of four weeks showed increased fat deposition in the liver of animals fed with a high-fat/high cholesterol diet, as compared to animals fed with the control diet. CONCLUSION: Our study established that hamsters fed with a high-fat/high-cholesterol diet developed fatty liver and mild diabetes. Bio F(1)B hamsters fed with a high-fat/high-cholesterol diet may thus be a good animal model for research on the treatment of diet-induced metabolic syndrome complicated by nonalcoholic fatty liver disease.

Microencapsulation to reduce mechanical loss of microspheres: implications in myocardial cell therapy.

OBJECTIVE: Previous regenerative studies have demonstrated massive cell losses after intramyocardial cellular delivery. Therefore, efforts at reducing mechanical losses may prove more successful in optimising cellular therapy. In this study, we hypothesized that escalating mesenchymal stem cells (MSCs) dose will not produce corresponding improvement in cardiac function due to washout of the small cells in microcirculation. Using microspheres similar in size to MSCs, that are encapsulated in alginate-poly-l-lysine-alginate (APA), we tested the hypothesis that size is an important factor in early losses. METHODS: In experiment I, five groups of rats (n=9 each) underwent coronary ligation; group I had no treatment; the other groups received escalating 0.5 × 10(6), 1.5 × 10(6), 3 × 10(6) and 5 × 10(6) of MSCs each. Echocardiogram was performed at baseline, 2 days and 7 weeks after surgery. In experiment II, cell-sized microspheres (10 µm) were encapsulated in APA microcapsules. In group I (n=16), rats received bare microspheres, group II (n=16) microspheres within 200 µm microcapsules and in group III (n=16), microspheres within 400 µm microcapsules. After 20 min, hearts were quantified for the amount retained. RESULTS: Myocardial function did not improve further with escalating cell doses beyond an initial response at 1.5 × 10(6) cells. Encapsulated microspheres in 200 µm and 400 µm microcapsules demonstrated a fourfold increase in retention rate compared with 10 µm microspheres. CONCLUSION: We concluded that suboptimal functional improvement in this animal model starts at 1.5 × 10(6) cells and does not respond to escalating cell doses. Improving mechanical retention is possible by increasing the size of the injectate. Microencapsulation could be used to encapsulate donor cells and facilitate functional improvement in cellular heart failure therapy.

Suppression of tumorigenesis: modulation of inflammatory cytokines by oral administration of microencapsulated probiotic yogurt formulation.

The objective of this study was to examine the ability of a novel microencapsulated probiotic yogurt formulation to suppress the intestinal inflammation. We assessed its anticancer activity by screening interleukin-1, 6, and 12 (IL-1, 6, 12), secretory levels of tumor necrosis factor-alpha (TNF-α), interferon-gamma (IFN-γ), prostaglandin E(2) (PGE(2)), and thromboxane B2 in the digesta obtained from the duodenum, jejunum, proximal, and distal segments of the ileum of C57BL/6J-Apc(Min)/J mice. Formulation-receiving animals showed consistently lower proinflammatory cytokines' levels when compared to control group animals receiving empty alginate-poly-L-lysine-alginate (APA) microcapsules suspended in saline. The concentrations of IL-12 found in serum in control and treatment group animals were significant: 46.58 ± 16.96 pg/mL and 158.58 ± 28.56 pg/mL for control and treatment animals, respectively. We determined a significant change in plasma C-reactive protein: 81.04 ± 23.73 ng/mL in control group and 64.21 ± 16.64 ng/mL in treatment group. Western blots showed a 71% downregulation of cyclooxygenase-2 (COX-2) protein in treatment group animals compared to control. These results point to the possibility of using this yogurt formulation in anticancer therapies, in addition to chronic gut diseases such as Crohn's disease, irritable bowel syndrome (IBS), and inflammatory bowel disease (IBD) thanks to its inflammation lowering properties.

Oral microencapsulated live Saccharomyces cerevisiae cells for use in renal failure uremia: preparation and in vivo analysis.

Orally administrable alginate-poly-L-lysine-alginate (APA) microcapsules containing live yeast cells was investigated for use in renal failure. At all times, yeast cells remain inside the microcapsules, which are then excreted in the stool. During their gastrointestinal passage, small molecules, like urea, diffuse into the yeast microcapsules where they are hydrolyzed. Orally administrating these microcapsules to uremic rats was found to decrease urea concentrations from 7.29 +/- 0.89 mmol/L to 6.12 +/- 0.90 mmol/L over a treatment period of eight weeks. After stopping the treatment, the urea concentrations increased back to uremic levels of 7.64 +/- 0.77 mmol/L. The analysis of creatinine concentrations averaged 39.19 +/- 4.33 micromol/L, 50.83 +/- 5.55 micromol/L, and 50.28 +/- 7.10 micromol/L for the normal-control, uremic-control and uremic-treatment groups, respectively. While creatinine concentrations for both uremic-control and uremic-treatment groups did not differ among each other (P > .05), they were, however, significantly higher than those of the normal control group (P < .05). Uric acid concentrations averaged 80.08 +/- 26.49 micromol/L, 99.92 +/- 26.55 micromol/L, and 86.49 +/- 28.42 micromol/L for the normal-control, uremic-control and uremic-treatment groups, respectively. There were no significant differences in both calcium and phosphate concentrations among all three groups (P > .05). The microbial populations of five tested types of bacteria were not substantially altered by the presence of the yeast APA encapsulated yeast (P > .05).

Ultrafine chitosan nanoparticles as an efficient nucleic acid delivery system targeting neuronal cells.

BACKGROUND: Cell transfection with nanoscaled cationic polymeric particles using Chitosan has been extensively explored. Because of its properties such as cationic charges, biocompatibility, biodegradability, and low toxicity, it has been used as a potential gene, siRNA, protein (including antibodies), and drug carrier system. METHOD: This work describes the development of chitosan nanoparticles of a 20-nm diameter for a potential siRNA delivery application. The particles were prepared using an ionic gelation method, using sodium tripolyphosphate as a cross-linker. The effect of variation in pH was investigated on particle size and surface charge. Gene loading efficiency by chitosan nanoparticles was performed by varying weight ratios of chitosan: siRNA. Transfection efficiency was evaluated on Neuro2a cells. RESULTS: It was observed that 20-nm-sized nanoscale complexes induced significant transfection in neuronal cells. CONCLUSION: These particles have potential in the delivery of siRNA to neural tissues.

Orally delivered microencapsulated live probiotic formulation lowers serum lipids in hypercholesterolemic hamsters.

Elevated serum cholesterol is a major risk factor for coronary artery disease. Nutritional therapies such as probiotics have been suggested to manage elevated cholesterol. This study investigates the cholesterol and triglyceride lowering potential of a microencapsulated feruloyl esterase-producing Lactobacillus fermentum 11976 (LF11976) probiotic formulation. Male Bio F(1)B hamsters were assigned to two groups to receive either the microcapsule probiotic formulation (containing LF11976 cells at 12.51 log colony-forming units/mL) or placebo formulation (empty) microcapsules, twice daily, by oral gavage for 18 weeks. For the duration of the study, animals were fed a hypercholesterolemic diet. Serum total cholesterol, low-density lipoprotein (LDL) cholesterol, and the atherogenic index were 21.36%, 31.43%, and 32.59% lower in the group gavaged with the microencapsulated probiotic formulation than in the placebo control group after 18 weeks (P < .05). Histology studies showed reduced progression of atherosclerotic lesions in animals treated with microencapsulated LF11976 as compared to control animals. Treatment with microencapsulated LF11976 formulation produces significant reductions in serum total cholesterol, LDL cholesterol, and serum triglyceride levels in diet-induced hypercholesterolemic hamsters. Findings suggest the potential of the oral microencapsulated probiotic cell formulation as a functional nutritional alternative for managing excessive serum cholesterol and triglyceride levels.

Microcapsule carbon nanotube devices for therapeutic applications.

Carbon nanotubes are a new class of nanomaterials that have immense potential in the field of biomedicine. Their ability to carry large quantities of therapeutic molecules makes them prime candidates for providing targeted delivery of therapeutics for use in various diseases. However, their utility is limited due to the problems faced during their delivery to target sites. This article for the first time describes the design of a novel microcapsule carbon nanotube targeted delivery device. This device has potential in the targeted delivery of carbon nanotubes in suitable membranes along with their cargo, safely and effectively to the target loci.

Estimation of the potential antitumor activity of microencapsulated Lactobacillus acidophilus yogurt formulation in the attenuation of tumorigenesis in Apc(Min/+) mice.

There is a strong correlation between orally administered probiotics and suppression of the low-grade inflammation that can lead to restoration of normal local immune functions. We studied the potential immunomodulatory and antitumorigenic properties of microencapsulated probiotic bacterial cells in a yogurt formulation in Min mice carrying a germline APC mutation. Daily oral administration of microencapsulated Lactobacillus acidophilus bacterial cells in the yogurt formulation mice resulted in significant suppression of colon tumor incidence, tumor multiplicity, and reduced tumor size. Results show that oral administration of microencapsulated L. acidophilus contributed to the stabilization of animal body weight and decreased the release of bile acids. Histopathological analyses revealed fewer adenomas in treated versus untreated animals. Furthermore, treated animals exhibited fewer gastrointestinal intra-epithelial neoplasias with a lower grade of dysplasia in detected tumors. Results suggest that oral administration of microencapsulated probiotic L. acidophilus exerts anti-tumorous activity, which consequently leads to reduced tumor outcome.

Microencapsulated bile salt hydrolase producing Lactobacillus reuteri for oral targeted delivery in the gastrointestinal tract.

This is the first study of its kind to screen probiotic lactic acid bacteria for the purpose of microencapsulating a highly bile salt hydrolase (BSH)-active strain. A Lactobacillus reuteri strain and a Bifidobacterium longum strain were isolated as the highest BSH producers among the candidates. Microcapsules were prepared with a diameter of 619 +/- 31 mum and a cell load of 5 x 10(9) cfu/ml. Post de Man, Rogosa, and Sharpe broth-acid challenge, L. reuteri microcapsules metabolized glyco- and tauro-conjugated bile salts at rates of 10.16 +/- 0.46 and 1.85 +/- 0.33 micromol/g microcapsule per hour, respectively, over the first 2 h. Microencapsulated B. longum had minimal BSH activity and were significantly (P < 0.05) more susceptible to acid challenge. Further testing of L. reuteri microcapsules in a simulated human gastrointestinal (GI) model showed an improved rate, with 49.4 +/- 6.21% of glyco-conjugates depleted after 60 min and complete deconjugation after 4 h. Microcapsules protected the encased cells in the simulated stomach maintaining L. reuteri viability above 10(9), 10(8), and 10(6) cfu/ml after 2 h at pH 3.0, 2.5, and 2.0, respectively. Results show excellent potential for this highly BSH-active microencapsulation system in vitro, highlighted by improved viability and substrate utilization in simulated GI transit.

Preparation and in vitro analysis of microencapsulated live Lactobacillus fermentum 11976 for augmentation of feruloyl esterase in the gastrointestinal tract.

FA (ferulic acid) is a well-known phenolic phytochemical present in plant cell walls. Various studies have indicated that FA has many physiological functions in the prevention of chronic disease. It has been shown to play an important chemoprotective role in degenerative diseases. FA also shows strong antioxidant and nitrite-scavenging potential and anticarcinogenic and antiinflammatory properties. The in vivo physiological importance of FA depends on its availability for absorption. Dietary fibre-bound FA is partially released by gut micro-organisms; however, the concentration of the released FA is too low to act as a chemopreventive agent. Therefore it is important to augment the bioavailability of FA to appreciate more fully its real physiological effect. This paper evaluates the suitability of the alginate-poly(L-lysine)-alginate microcapsules for oral delivery of live feruloyl esterase-producing Lactobacillus fermentum 11976 cells, in vitro, by using a dynamic simulated human GI (gastrointestinal) model. The present study shows that microencapsulated L. fermentum 11976 cells can efficiently break down a FA-containing substrate, and establishes the biotechnological basis for their use in supplementing the bioavailability of dietary FA in the intestine.

Live encapsulated Lactobacillus acidophilus cells in yogurt for therapeutic oral delivery: preparation and in vitro analysis of alginate-chitosan microcapsules.

Targeted delivery of live microencapsulated bacterial cells has strong potential for application in treating various diseases, including diarrhea, kidney failure, liver failure, and high cholesterol, among others. This study investigates the potential of microcapsules composed of two natural polymers, alginate and chitosan (AC), and the use of these artificial cells in yogurt for delivery of probiotic Lactobacillus acidophilus bacterial live cells. Results show that the integrity of AC microcapsules was preserved after 76 h of mechanical shaking in MRS broth and after 12 h and 24 h in simulated gastric and intestinal fluids. Using an in vitro computer-controlled simulated human gastrointestinal (GI) model, we found 8.37 log CFU/mL of viable bacterial cells were present after 120 min of gastric exposure and 7.96 log CFU/mL after 360 min of intestinal exposure. In addition, AC microcapsules composed of chitosan 10 and 100 at various concentrations were subjected to 4-week storage in 2% milk fat yogurt or 0.85% physiological solution. It was found that 9.37 log CFU/mL of cells encapsulated with chitosan 10 and 8.24 log CFU/mL of cells encapsulated with chitosan 100 were alive after 4 weeks. The AC capsule composed of 0.5% chitosan 10 provided the highest bacterial survival of 9.11 log CFU/mL after 4 weeks. Finally, an investigation of bacterial viability over 72 h in different pH buffers yielded highest survival of 6.34 log CFU/mL and 10.34 log CFU/mL at pH 8 for free and AC-encapsulated cells, respectively. We conclude from these findings that encapsulation allows delivery of a higher number of bacteria to desired targets in the GI tract and that microcapsules containing bacterial cells are good candidates for oral artificial cells for bacterial cell therapy.

Microencapsulated bacterial cells can be used to produce the enzyme feruloyl esterase: preparation and in-vitro analysis.

Biotechnological production of ferulic acid, a precursor of vanillin, is an attractive alternative for various industries due to the high price and demand for natural ferulic acid. Feruloyl esterase has been identified as a key enzyme involved in microbial transformations of ferulic acid to vanillin. Several fungal feruloyl esterases have been purified and characterized for their use in the production of ferulic acid. This paper, for the first time, discusses the use of lactic acid bacteria for the production of ferulic acid. Specifically, we have used Lactobacillus cells and microencapsulation so that ferulic acid can be produced continuously using various types of fermentation systems. Bacteria were encapsulated in alginate-poly-L-lysine-alginate (APA) microcapsules, and the production of ferulic acid by lactobacilli was detected using a real-time high-performance liquid chromatography (HPLC)-based assay. Results show that ferulic acid can be produced using microencapsulated Lactobacillus fermentum (ATCC 11976) with significant levels of biological feruloyl esterase activity.

Investigation of microencapsulated BSH active lactobacillus in the simulated human GI tract.

This study investigated the use of microencapsulated bile salt hydrolase (BSH) overproducing Lactobacillus plantarum 80 cells for oral delivery applications using a dynamic computer-controlled model simulating the human gastrointestinal (GI) tract. Bile salt deconjugation rates for microencapsulated BSH overproducing cells were 4.87 +/- 0.28 mumol/g microcapsule/h towards glycoconjugates and 0.79 +/- 0.15 mumol/g microcapsule/h towards tauroconjugates in the simulated intestine, a significant (P< .05) increase over microencapsulated wild-type cells. Microcapsules protected the encased cells in the simulated stomach prior to intestinal release, maintaining cell viability above 109 cfu/mL at pH 2.5 and 3.0 and above 106 cfu/mL at pH 2.0 after 2-hour residence times. In the simulated intestine, encased cell viability was maintained above 1010 cfu/mL after 3, 6, and 12-hour residence times in bile concentrations up to 1.0%. Results show that microencapsulation has potential in the oral delivery of live BSH active bacterial cells. However, in vivo testing is required.

Effects of nitrogen and phosphorus limitation on the activated sludge biomass in a kraft mill biotreatment system.

Unlike wastewater, pulp and paper mill effluents are generally severely deficient in bioavailable nitrogen and phosphorus. The influence of nitrogen and phosphorus limitations on steady-state or typical pulp and paper mill activated sludge floc properties and performance was studied using a bioreactor-fed synthetic raw mill effluent and seeded with mill activated sludge. Limitation of either nitrogen or phosphorus decreased growth, five-day biochemical oxygen demand, and suspended solids removal. Nitrogen limitation greatly enhanced activated sludge floc poly-beta3-hydroxybutyrate (PHB), but not carbohydrate or extracellular polymeric substances (EPS). In contrast, phosphorus limitation increased total floc carbohydrate and EPS, but not PHB. The flocs showed little ability to store either nitrogen or phosphorus. Nitrogen limitation, but not phosphorus limitation, produced much more negative net floc surface charge, increasing fines, while phosphorus limitation, but not nitrogen limitation, increased the floc bound water content and surface hydrophobicity and decreased fines.

Live bacterial cells as orally delivered therapeutics.

Despite the swift escalation in research regarding the use of live bacterial cells for therapeutic purposes, the prophylactic and curative use of probiotic microorganisms still remains a wide and controversial field. In addition, the acknowledgement that live bacterial cells can be genetically engineered to synthesise products that have therapeutic potential has generated substantial interest among clinicians and health professionals. Clinical trials have increasingly provided an insightful scientific derivation for the use of live bacterial cells in medicinal practice in diseases such as diarrhoea, cancer, Crohn's disease, enhancement of the host's immune response, and numerous other diseases. A key constraint in the use of live bacterial cells, however, is the complexity of delivering them to the correct target sites. Oral delivery of free live cells, lyophilised cells and immobilised cells has been attempted, but with restricted success, chiefly because bacterial cells are unable to survive passage through the gastrointestinal tract in sufficient dosage. On many occasions, when given orally, these cells have been found to provoke immunogenic responses that are undesirable. Recent studies show that these problems can be overcome by delivering live bacterial cells using artificial cell microcapsules. This review abridges recent developments in the therapeutic use of live bacterial cells, addresses the potential and restrictions for their application in therapy, and provides insights into the future course of this emerging therapy.