Laetitia Rodes

Gut microbiota: next frontier in understanding human health and development of biotherapeutics.

The gut microbiota is a remarkable asset for human health. As a key element in the development and prevention of specific diseases, its study has yielded a new field of promising biotherapeutics. This review provides comprehensive and updated knowledge of the human gut microbiota, its implications in health and disease, and the potentials and limitations of its modification by currently available biotherapeutics to treat, prevent and/or restore human health, and future directions. Homeostasis of the gut microbiota maintains various functions which are vital to the maintenance of human health. Disruption of the intestinal ecosystem equilibrium (gut dysbiosis) is associated with a plethora of human diseases, including autoimmune and allergic diseases, colorectal cancer, metabolic diseases, and bacterial infections. Relevant underlying mechanisms by which specific intestinal bacteria populations might trigger the development of disease in susceptible hosts are being explored across the globe. Beneficial modulation of the gut microbiota using biotherapeutics, such as prebiotics, probiotics, and antibiotics, may favor health-promoting populations of bacteria and can be exploited in development of biotherapeutics. Other technologies, such as development of human gut models, bacterial screening, and delivery formulations eg, microencapsulated probiotics, may contribute significantly in the near future. Therefore, the human gut microbiota is a legitimate therapeutic target to treat and/or prevent various diseases. Development of a clear understanding of the technologies needed to exploit the gut microbiota is urgently required.

Bioengineered baculoviruses as new class of therapeutics using micro and nanotechnologies: Principles, prospects and challenges☆

Designing a safe and efficient gene delivery system is required for success of gene therapy trials. Although a wide variety of viral, non-viral and polymeric nanoparticle based careers have been widely studied, the current gene delivery vehicles are limited by their suboptimal, non-specific therapeutic efficacy and acute immunological reactions, leading to unwanted side effects. Recently, there has been a growing interest in insect-cell-originated baculoviruses as gene delivery vehicles for diverse biomedical applications. Specifically, the emergence of diverse types of surface functionalized and bioengineered baculoviruses is posed to edge over currently available gene delivery vehicles. This is primarily because baculoviruses are comparatively non-pathogenic and non-toxic as they cannot replicate in mammalian cells and do not invoke any cytopathic effect. Moreover, emerging advanced studies in this direction have demonstrated that hybridizing the baculovirus surface with different kinds of bioactive therapeutic molecules, cell-specific targeting moieties, protective polymeric grafts and nanomaterials can significantly improve the preclinical efficacy of baculoviruses. This review presents a comprehensive overview of the recent advancements in the field of bioengineering and biotherapeutics to engineer baculovirus hybrids for tailored gene therapy, and articulates in detail the potential and challenges of these strategies for clinical realization. In addition, the article illustrates the rapid evolvement of microfluidic devices as a high throughput platform for optimizing baculovirus production and treatment conditions.

Biotransformation of polyphenols in a dynamic multistage gastrointestinal model.

A multi-reactor gastrointestinal model was used to digest a mixture of pure polyphenol compounds, including non-flavonoid phenolic acids (chlorogenic acid, caffeic acid, ferulic acid) and a flavonoid (rutin) to identify phenolic metabolites and short chain fatty acids (SCFAs) and compare relative antioxidant capacities following a 24h digestion. Biotransformation of these polyphenols occurred in the colonic compartments generating phenylpropionic, benzoic, phenylacetic and cinnamic acids. Total SCFAs increased in all colonic vessels with a rise in the proportion of propionic to acetic acid. Antioxidant capacity increased significantly in all compartments, but first in the stomach, small intestine and ascending colon. After 24h, the colonic vessels without parent polyphenols, but containing new metabolites, had antioxidant capacities similar to the stomach and small intestine, containing parent compounds. Biotransformation of pure polyphenols resulted in different phenolic metabolite and SCFAs profiles in each colonic segment, with important health implications for these colonic compartments.

Investigation of probiotic bacteria as dental caries and periodontal disease biotherapeutics.

Oral diseases, specifically dental caries and periodontal disease, are characterised by increases in pathogenic microorganisms, increased demineralisation and increased inflammation and levels of inflammatory markers. Despite the therapeutic strategies, oral diseases have elevated prevalence rates. Recent work has demonstrated that probiotic bio-therapeutics can decrease oral pathogen counts, including caries-causing Streptococcus mutans and oral inflammation. The aim of this work was to investigate putative probiotic bacteria, selected for S. mutans inhibition and for their oral health-promoting characteristics. The probiotic bacteria were screened for S. mutans inhibition, probiotic bacteriocin activity, salivary pH modulation, probiotic nutrient (sucrose) competition, probiotic co-aggregation with S. mutans, bacterial attachment to oral epithelial keratinocytes, bacterial nitric oxide production and bacterial antioxidant activity. The results indicate that Lactobacillus reuteri strains NCIMB 701359, NCIMB 701089, NCIMB 702655 and NCIMB 702656 inhibited S. mutans to non-detectable levels (<10 cfu/ml). L. reuteri strains also demonstrated the highest antioxidant capacity of the tested strains (7.73-13.99 µM Trolox equivalents), suggesting their use as both caries and periodontal disease therapeutics. Although Lactobacillus fermentum NCIMB 5221 inhibited S. mutans at lower levels, it significantly buffered the pH (4.18) of saliva containing S. mutans, co-aggregated with S. mutans (10.09%), demonstrated high levels of sucrose consumption (138.11 mM) and successfully attached to gingival epithelial cells (11%). This study identified four L. reuteri strains and one L. fermentum strain to be further investigated as oral disease biotherapeutics.

Microencapsulated Bifidobacterium longum subsp. infantis ATCC 15697 Favorably Modulates Gut Microbiota and Reduces Circulating Endotoxins in F344 Rats

The gut microbiota is a bacterial bioreactor whose composition is an asset for human health. However, circulating gut microbiota derived endotoxins cause metabolic endotoxemia, promoting metabolic and liver diseases. This study investigates the potential of orally delivered microencapsulated Bifidobacterium infantis ATCC 15697 to modulate the gut microbiota and reduce endotoxemia in F344 rats. The rats were gavaged daily with saline or microencapsulated B. infantis ATCC 15697. Following 38 days of supplementation, the treated rats showed a significant (P < 0.05) increase in fecal Bifidobacteria (4.34 ± 0.46 versus 2.45 ± 0.25% of total) and B. infantis (0.28 ± 0.21 versus 0.52 ± 0.12 % of total) and a significant (P < 0.05) decrease in fecal Enterobacteriaceae (0.80 ± 0.45 versus 2.83 ± 0.63% of total) compared to the saline control. In addition, supplementation with the probiotic formulation reduced fecal (10.52 ± 0.18 versus 11.29 ± 0.16 EU/mg; P = 0.01) and serum (0.33 ± 0.015 versus 0.30 ± 0.015 EU/mL; P = 0.25) endotoxins. Thus, microencapsulated B. infantis ATCC 15697 modulates the gut microbiota and reduces colonic and serum endotoxins. Future preclinical studies should investigate the potential of the novel probiotic formulation in metabolic and liver diseases.

Transit Time Affects the Community Stability of Lactobacillus and Bifidobacterium Species in an In Vitro Model of Human Colonic Microbiotia

Abstract: Retention time, which is analogous to transit time, is an index for bacterial stability in the intestine. Its consideration is of particular importance to optimize the delivery of probiotic bacteria in order to improve treatment efficacy. This study aims to investigate the effect of retention time on Lactobacilli and Bifidobacteria stability using an established in vitro human colon model. Three retention times were used: 72, 96, and 144 h. The effect of retention time on cell viability of different bacterial populations was analyzed with bacterial plate counts and PCR. The proportions of intestinal Bifidobacteria, Lactobacilli, Enterococci, Staphylococci and Clostridia populations, analyzed by plate counts, were found to be the same as that in human colonic microbiota. Retention time in the human colon affected the stability of Lactobacilli and Bifidobacteria communities, with maximum stability observed at 144 h. Therefore, retention time is an important parameter that influences bacterial stability in the colonic microbiota. Future clinical studies on probiotic bacteria formulations should take into consideration gastrointestinal transit parameters to improve treatment efficacy.

Lactobacillus fermentum NCIMB 5221 and NCIMB 2797 as cholesterol-lowering probiotic biotherapeutics: in vitro analysis

Cardiovascular and coronary artery disease risk are correlated with cholesterol levels and are significant health concerns. Current cholesterol-lowering approaches includes lifestyle and diet modifications, as well as statins which presents numerous shortcomings. The probiotic bacteria, Lactobacillus fermentum NCIMB 5221 and NCIMB 2797, have demonstrated cholesterol-lowering potential in animal studies. However, there is a lack in understanding the mechanism(s) behind these observed effects. The goal of this work is to investigate, in vitro, the cholesterol-lowering mechanisms of these two strains. To determine the cholesterol-lowering mechanisms, probiotic cholesterol assimilation, colon epithelial adhesion and inhibition of cholesterol uptake by colon epithelial (Caco-2) cells were investigated. L. fermentum NCIMB 2797 (P=0.012) and NCIMB 5221 (P=0.003) assimilated cholesterol and their cell surface hydrophobicity was 70.30±8.85% and 55.60±2.59%, respectively. Both L. fermentum strains showed no significant impact (P>0.05) on Caco-2 cell viability. Of most interest, Caco-2 pre-exposure to L. fermentum NCIMB 5221 significantly decreased (P=0.015) cholesterol uptake, with 85.98±2.07% uptake compared to the untreated cells. Similarly, L. fermentum NCIMB 2797 probiotic cells significantly decreased (P=0.019) cholesterol uptake by Caco-2 cells, with 86.45±1.71% uptake observed compared to the control cells. The results demonstrate that L. fermentum NCIMB 5221 and L. fermentum NCIMB 2797 have the potential via various modes of action to lower cholesterol. Additional studies are required to understand the mechanism(s) of action behind probiotic cholesterol assimilation and behind the cholesterol uptake inhibition by colon epithelial cells.

Design of a novel gut bacterial adhesion model for probiotic applications

Abstract A new gut bacterial adhesion model has been developed. For this, a continuous-flow bioreactor packed with bacteria-coated beads was designed to simulate the gut lining and other features. In vitro model efficacy shows successful bacterial cell gut adhesions: bacterial adhesion was higher with mucin–alginate compared to controls. In feasibility study, adhesion of Lactobacillus fermentum NCIMB 5221 and Lactobacillus reuteri NCIMB 701359 was investigated for their metabolic activities for bile salt. Bile salt hydrolase (BSH)-active Lactobacillus reuteri exerted higher activity than non-BSH-active L. fermentum. This model has potential use in gut health, probiotic, bacterial cell gut adhesion and other delivery applications.

Identification of Lactobacillus Fermentum Strains with Potential against Colorectal Cancer by Characterizing Short Chain Fatty Acids Production, Anti-Proliferative Activity and Survival in an Intestinal Fluid: In Vitro Analysis

The use of probiotics as preventive agents in colorectal cancer (CRC), as widely suggested in many clinical and pre-clinical studies, was often linked to the potency of short chain fatty acids (SCFAs) in the gut. However, there remains an incomplete understanding of the fatty-acid-producing activity of certain probiotics and their cancer preventive potential. In the current study, L. fermentum strains were investigated for their potential use with CRC treatments. Using cell-free extracts, L. fermentum NCIMB -5221, - 2797, and -8829 were first compared based on their SCFAs production and anti-proliferative activity against Caco-2 colon cancer cells. The corresponding SCFAs synthetic formulations, similar to the ones produced by the bacteria, were prepared and compared with the latter to determine the role and efficacy of naturally produced SCFAs in inhibiting the proliferation of colon cancer cells. Subsequently, the bioactivity and stability of L. fermentum bacterial strains in a simulated intestinal fluid (SIF) was determined. Results showed that L. fermentum NCIMB -5221 and -8829 were the most potent in producing SCFAs, in particular, acetic (192.3 ± 4 mg/L minimum), propionic (69.2 ± 1.6 mg/L minimum), and butyric (35.4 ± 2.9 mg/L minimum) acids. They were also found to inhibit the growth of Caco-2 cells (53.4 ± 1.6%, 72 h, p = 0.021) in comparison with L. acidophilus ATCC 314. Additionally, they showed resistance to SIF (16.3 ± 1.9% minimum, 72 h, p = 0.006) and produced SCFAs in SIF at concentrations high enough to significantly inhibit Caco-2 proliferation (74.73 ± 2.1%, 72 h). Based on characteristics related to bacterial cell survival, SCFA production, and anti-proliferative activity, L. fermentum NCIMB -5221 and - 2797 could potentially be considered as biotherapeutic agents against CRC.

Enrichment of Bifidobacterium longum subsp. infantis ATCC 15697 within the human gut microbiota using alginate-poly-l-lysine-alginate microencapsulation oral delivery system: an in vitro analysis using a computer-controlled dynamic human gastrointestinal model

Abstract This study evaluates alginate-poly-l-lysine-alginate Bifidobacterium longum subsp. infantis ATCC 15697-loaded microcapsules to enrich the human gut microbiota. The cell survival of alginate-poly-l-lysine-alginate microencapsulated B. infantis ATCC 15697 in gastric acid, bile, and through human gastrointestinal transit was investigated, as well as the formulation’s effect on the gut microbiota. Results show that microencapsulation increases B. infantis ATCC 15697 cell survival at pH1.0 (33.54 ± 2.80% versus <1.00 ± 0.00%), pH1.5 (41.15 ± 2.06% versus <1.00 ± 0.00%), pH2.0 (60.88 ± 1.73% versus 36.01 ± 2.63%), pH3.0 (75.43 ± 1.23% versus 46.30 ± 1.43%), pH4.0 (71.40 ± 2.02% versus 47.75 ± 3.12%) and pH5.0 (73.88 ± 3.79% versus 58.93 ± 2.26%) (p < 0.05). In addition, microencapsulation increases cell survival at 0.5% (76.85 ± 0.80% versus 70.77 ± 0.64%), 1.0% (59.99 ± 0.97% versus 53.47 ± 0.58%) and 2.0% (53.10 ± 1.87% versus 44.59 ± 1.52%) (p < 0.05) (w/v) bile. Finally, daily administration of alginate-poly-l-lysine-alginate microencapsulated B. infantis ATCC 15697 in a human gastrointestinal model induces a significant enrichment of B. infantis within the ascending (184.51 ± 17.30% versus 53.83 ± 17.82%; p < 0.05), transverse (174.79 ± 25.32% versus 73.17 ± 15.30%; p < 0.05) and descending (94.90 ± 25.22% versus 46.37 ± 18.93%; p > 0.05) colonic microbiota.