An in vitro intestinal model captures immunomodulatory properties of the microbiota in inflammation.

Considerable effort has been put forth to understand mechanisms by which the microbiota modulates and responds to inflammation. Here, we explored whether oxidation metabolites produced by the host during inflammation, sodium nitrate and trimethylamine oxide, impact the composition of a human stool bacterial population in a gut simulator. We then assessed whether an immune-competent in vitro intestinal model responded differently to spent medium from bacteria exposed to these cues compared to spent medium from a control bacterial population. The host-derived oxidation products were found to decrease levels of Bacteroidaceae and overall microbiota metabolic potential, while increasing levels of proinflammatory Enterobacteriaceae and lipopolysaccharide in bacterial cultures, reflecting shifts that occur in vivo in inflammation. Spent microbiota media induced elevated intracellular mucin levels and reduced intestinal monolayer integrity as reflected in transepithelial electrical resistance relative to fresh medium controls. However, multiplexed cytokine analysis revealed markedly different cytokine signatures from intestinal cultures exposed to spent medium with added oxidation products relative to spent control medium, while cytokine signatures of cultures exposed to fresh media were similar regardless of addition of host-derived cues. Further, the presence of immune cells in the intestinal model was required for this differentiation of cytokine signatures. This study indicates that simple in vitro immune-competent intestinal models can capture bacterial-mammalian cross-talk in response to host-derived oxidation products and supports utility of these systems for mechanistic studies of interactions between the gut microbiome and host in inflammation.

Tissue distribution of ethanol after intraprostatic injection using a porous needle.

Purpose: To develop a safe and precise method for intraprostatic injection, and to establish correlation between the volume of ethanol injectate and the volume of subsequent infiltrated prostate tissue. Materials and methods: We performed intraprostatic injection of 96% ethanol using a needle which has a segment of its wall made of capillary membrane with hundreds of pores in an acute and chronic canine experiment, in heart-beating cadaveric organ donors, and in a xenograft model of human prostate cancer. Whole mount tissue sections were used for three-dimensional reconstruction of the necrotic lesions and calculation of their volumes. Results: The ethanol injection resulted in oval shaped lesions of well-delineated coagulative necrosis. In both healthy human and canine prostates, the prostatic pseudocapsule and neurovascular bundle remained intact without evidence of disruption. There was a linear correlation between administered volume of ethanol and the volume of necrotic lesion. Regression analysis showed strong correlation in the acute canine experiments and in experiments performed on xenografts of human prostate cancer. A formula was calculated for each experiment to estimate the relationship between the injected volume and the volume of infiltrated prostate tissue area. Conclusions: Intraprostatic injection using a porous needle allows for effective and predictable tissue distribution of the injectate in the prostate. Through varying the volume of the agent injected and use of needles with a different length of the porous segment, the volume of infiltrated tissue could be adjusted allowing for targeted focal treatment.

5-year retrospective analysis of patients with phenylketonuria (PKU) and hyperphenylalaninemia treated at two specialized clinics.

BACKGROUND: Phenylketonuria (PKU) is an autosomal recessive disease caused by mutations in the PAH gene, resulting in deficiency of phenylalanine hydroxylase (PAH), an enzyme that converts phenylalanine (Phe) to tyrosine (Tyr). The purpose of this study was to capture real-world data associated with managing PKU under current standard of care and to characterize a representative population for a planned gene therapy trial. METHODS: A retrospective chart review was conducted at two U.S. clinics for individuals 10-40 years old diagnosed with PKU-related hyperphenylalaninemia (HPA). Demographics, medical history, treatments and blood Phe data were collected from electronic medical records spanning a five-year period ending in November 2017. RESULTS: 152 patients were enrolled (65.8% had classical PKU). Although >95% of patients were prescribed a Phe-restricted diet, blood Phe concentrations remained substantially elevated, particularly in patients diagnosed with classical PKU. As the Phe threshold was lowered (Phe < 600, 360, 120 or 30 µmol/L), the number of patients with consecutive lab values below the threshold decreased, suggesting that many patients' Phe levels are inadequately controlled. 62.5% of patients were reported as having a history of at least one neuropsychiatric comorbidity, and adults were more likely than adolescents (69.5% vs. 54.3%). 92 of 98 PAH genotypes collected were distinct mutations; the 6 null-null genotypes were associated with classical PKU. Overall the demographics and clinical data were consistent across both sites. CONCLUSION: Despite dietary restrictions, mean Phe concentrations were > 360 µmol/L (a level considered well-controlled based on current U.S. treatment guidelines) for mild, moderate, and classical PKU patients. There remains an unmet need for therapies to control Phe concentrations.