Engineered Bacillus subtilis as oral probiotics to target circulating lactic acid.

Lactic acid or lactate, a key byproduct of anaerobic glycolysis, plays pivotal roles in routine metabolism. An increase in lactic acid is observed in various pathological conditions such as cancer, diabetes, genetic mitochondrial disease, and aging. While several groups have proposed small molecule inhibitors to reduce circulating lactic acid, there are few clinically relevant ways to manage acute or chronic elevations in lactic acid in patients. In addition, recent evidence suggests that lactic acid exchanges between the gut, blood, and peripheral tissues, and professional marathon runners harbor specific gut microbial species that more efficiently metabolize lactic acid. Inspired by these findings, this work sought to engineer probiotic B. subtilis strains to express lactate oxidase that could increase circulating lactic acid catabolism after delivery to the gut. After optimization, oral administration of engineered B. subtilis to the gut of mice reduced the elevation in blood lactic acid levels after exogenous lactic acid challenge without affecting normal gut microbiota composition, inflammation or liver enzymes. Taken together, through the oral delivery of engineered probiotics to the gastrointestinal tract, our proof-of-concept study offers a new opportunity to therapeutically target diseases where blood lactic acid is elevated, and provides a new approach to "knocking down" metabolites to help understand the roles of metabolites in host physiological and pathological processes.

Inhibition of inflammatory microglia by dietary fiber and short-chain fatty acids.

Microglia play a vital role maintaining brain homeostasis but can also cause persistent neuroinflammation. Short-chain fatty acids (SCFAs) produced by the intestinal microbiota have been suggested to regulate microglia inflammation indirectly by signaling through the gut-brain axis or directly by reaching the brain. The present work evaluated the anti-inflammatory effects of SCFAs on lipopolysaccharide (LPS)-stimulated microglia from mice fed inulin, a soluble fiber that is fermented by intestinal microbiota to produce SCFAs in vivo, and SCFAs applied to primary microglia in vitro. Feeding mice inulin increased SCFAs in the cecum and in plasma collected from the hepatic portal vein. Microglia isolated from mice fed inulin and stimulated with LPS in vitro secreted less tumor necrosis factor α (TNF-α) compared to microglia from mice not given inulin. Additionally, when mice were fed inulin and injected i.p with LPS, the ex vivo secretion of TNF-α by isolated microglia was lower than that secreted by microglia from mice not fed inulin and injected with LPS. Similarly, in vitro treatment of primary microglia with acetate and butyrate either alone or in combination downregulated microglia cytokine production with the effects being additive. SCFAs reduced histone deacetylase activity and nuclear factor-κB nuclear translocation after LPS treatment in vitro. Whereas microglia expression of SCFA receptors Ffar2 or Ffar3 was not detected by single-cell RNA sequencing analysis, the SCFA transporters Mct1 and Mct4 were. Nevertheless, inhibiting monocarboxylate transporters on primary microglia did not interfere with the anti-inflammatory effects of SCFAs, suggesting that if SCFAs produced in the gut regulate microglia directly it is likely through an epigenetic mechanism following diffusion.

Dietary Fiber as a Counterbalance to Age-Related Microglial Cell Dysfunction.

With increasing age, microglia shift toward a pro-inflammatory phenotype that may predispose individuals to neurodegenerative disease. Because fiber fermentation in the colon produces bioactive short-chain fatty acids (SCFAs; e.g., acetate, butyrate, and propionate) that signal through the gut-brain axis, increasing dietary fiber may prevent or reverse age-related dysregulation of microglia. Adult (3-4 months old) and aged (23-24 months old) male and female mice were given ad libitum access to a modified AIN-93M diet with 1% cellulose or the same diet with 2.5 or 5.0% inulin for 8 weeks. Several adult and aged male mice fed 0 or 5% inulin were randomly selected for whole brain single-cell RNA sequencing (scRNA-seq) and differential gene expression analysis to classify brain microglia according to gene expression profile; and identify additional genetic markers of aging as possible targets for dietary interventions. Microglia were isolated from remaining mice and expression of selected aging-, inflammatory-, and sensome-related genes was assessed by Fluidigm as was the ex vivo secretion of tumor necrosis factor-alpha (TNF-α). SCFAs were measured in samples collected from the cecum. Microglia from adult and aged mice segregated into distinct phenotypes according to their gene expression profile. In aged mice, a considerably greater proportion of the population of microglia was identified being "activated" and a considerably smaller proportion was identified being "quiescent." These findings using whole brain scRNA-seq were largely corroborated using highly purified microglia and Fluidigm analysis to assess a selected panel of genes. Aged mice compared to adults had lower levels of SCFA's in cecum. Dietary inulin increased SCFAs in cecum and mostly restored microglial cell gene expression and TNF-α secretion to that seen in adults. Sex differences were observed with females having lower levels of SCFAs in cecum and increased neuroinflammation. Overall, these data support the use of fiber supplementation as a strategy to counterbalance the age-related microglial dysregulation.

Green Synthesis of Gold Nanoparticles Using Upland Cress and Their Biochemical Characterization and Assessment.

This research focuses on the plant-mediated green synthesis process to produce gold nanoparticles (Au NPs) using upland cress (Barbarea verna), as various biomolecules within the upland cress act as both reducing and capping agents. The synthesized gold nanoparticles were thoroughly characterized using UV-vis spectroscopy, surface charge (zeta potential) analysis, scanning electron microscopy-energy-dispersive X-ray spectroscopy (SEM-EDX), atomic force microscopy (AFM), attenuated total reflection Fourier transform infrared spectroscopy (ATR-FTIR), and X-ray diffraction (XRD). The results indicated the synthesized Au NPs are spherical and well-dispersed with an average diameter ~11 nm and a characteristic absorbance peak at ~529 nm. EDX results showed an 11.13% gold content. Colloidal Au NP stability was confirmed with a zeta potential (ζ) value of -36.8 mV. X-ray diffraction analysis verified the production of crystalline face-centered cubic gold. Moreover, the antimicrobial activity of the Au NPs was evaluated using Gram-negative Escherichiacoli and Gram-positive Bacillus megaterium. Results demonstrated concentration-dependent antimicrobial properties. Lastly, applications of the Au NPs in catalysis and biomedicine were evaluated. The catalytic activity of Au NPs was demonstrated through the conversion of 4-nitrophenol to 4-aminophenol which followed first-order kinetics. Cellular uptake and cytotoxicity were evaluated using both BMSCs (stem) and HeLa (cancer) cells and the results were cell type dependent. The synthesized Au NPs show great potential for various applications such as catalysis, pharmaceutics, and biomedicine.

The COVID-19 pandemic and physical activity.

The SARS-CoV-2-caused COVID-19 pandemic has resulted in a devastating threat to human society in terms of health, economy, and lifestyle. Although the virus usually first invades and infects the lung and respiratory track tissue, in extreme cases, almost all major organs in the body are now known to be negatively impacted often leading to severe systemic failure in some people. Unfortunately, there is currently no effective treatment for this disease. Pre-existing pathological conditions or comorbidities such as age are a major reason for premature death and increased morbidity and mortality. The immobilization due to hospitalization and bed rest and the physical inactivity due to sustained quarantine and social distancing can downregulate the ability of organs systems to resist to viral infection and increase the risk of damage to the immune, respiratory, cardiovascular, musculoskeletal systems and the brain. The cellular mechanisms and danger of this "second wave" effect of COVID-19 to the human body, along with the effects of aging, proper nutrition, and regular physical activity, are reviewed in this article.