OBJECTIVES:Exploring associations between the gut microbiota and colonic inflammation and assessing sequential changes during exclusive enteral nutrition (EEN) may offer clues into the microbial origins of Crohn's disease (CD).METHODS:Fecal samples (n=117) were collected from 23 CD and 21 healthy children. From CD children fecal samples were collected before, during EEN, and when patients returned to their habitual diets. Microbiota composition and functional capacity were characterized using sequencing of the 16S rRNA gene and shotgun metagenomics.RESULTS:Microbial diversity was lower in CD than controls before EEN (P=0.006); differences were observed in 36 genera, 141 operational taxonomic units (OTUs), and 44 oligotypes. During EEN, the microbial diversity of CD children further decreased, and the community structure became even more dissimilar than that of controls. Every 10 days on EEN, 0.6 genus diversity equivalents were lost; 34 genera decreased and one increased during EEN. Fecal calprotectin correlated with 35 OTUs, 14 of which accounted for 78% of its variation. OTUs that correlated positively or negatively with calprotectin decreased during EEN. The microbiota of CD patients had a broader functional capacity than healthy controls, but diversity decreased with EEN. Genes involved in membrane transport, sulfur reduction, and nutrient biosynthesis differed between patients and controls. The abundance of genes involved in biotin (P=0.005) and thiamine biosynthesis decreased (P=0.017), whereas those involved in spermidine/putrescine biosynthesis (P=0.031), or the shikimate pathway (P=0.058), increased during EEN.CONCLUSIONS:Disease improvement following treatment with EEN is associated with extensive modulation of the gut microbiome.

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A study of the plasma pharmacokinetics, tumor localization, and safety of a single dose of doxorubicin encapsulated in liposomes containing surface-bound polyethylene glycol (PEG-liposomal doxorubicin) was conducted in patients with Kaposi's sarcoma (KS) as a manifestation of acquired immune deficiency syndrome (AIDS). Eighteen patients with AIDS-KS diagnosed by examination of biopsy specimens were randomly assigned to receive either standard doxorubicin or PEG-liposomal doxorubicin. Consecutive participants were entered at three dose levels (10, 20, and 40 mg/m2) in ascending fashion. Clearance of PEG-liposomal doxorubicin was 0.034 L/h/m2 to 0.108 L/h/m2, volume of distribution (Vd) was 2.2 L/m2 to 4.4 L/m2, and half-lives (t1/2) of the initial decline in the plasma concentration-time curve and of the terminal decline were 3.77 hours and 41.3 hours, respectively. Seventy-two hours after administration, doxorubicin levels observed in lesions of patients receiving PEG-liposomal doxorubicin were 5.2 to 11.4 times greater than those found in patients given comparable doses of standard doxorubicin. PEG-liposomal doxorubicin and standard doxorubicin were roughly equipotent in producing toxicity. Encapsulation in liposomes containing surface-bound PEG significantly limits the distribution and elimination of doxorubicin, results in greater accumulation of the drug in KS lesions 72 hours after dosing than does standard doxorubicin, and may improve drug efficacy and therapeutic index in the treatment of AIDS-KS.

A cell suspension culture of tobacco (Nicotiana tabacum L.) was used as a model to study injury to cells during biolistic transformation. Lawns of cells were bombarded with tungsten particles that were coated with a plasmid containing the jl-glucuronidase and the neomycin phosphotransferase 11 genes. When a gunpowder-driven biolistic device was used, numerous transiently expressing cells were focused around the epicenter of the blast which was manifested by a hole blown in the filter paper supporting the cells. However, transformed cells nearest the blast epicenter were injured and could not be recovered as stable transformants. The injury was primarily caused by physical trauma to the cells from gas blast and acoustic shock generated by the device. Postlaunch baffles or meshes placed in the gunpowder device reduced cell injury and increased the recovery of kanamycin-resistant colonies 3.5-and 2.5-fold, respectively. A newly developed helium-driven device was more gentle to the cells and also increased the number of transformants. Cell injury could be further moderated by using a mesh and a prelaunch baffle in the helium device. Toxicity of the tungsten microprojectiles also contributed to cell injury. Gold microprojectiles were not toxic and resulted in fourfold more kanamycin-resistant colonies than when similar quantities of similarly sized tungsten particles were used. gene delivery into cultured animal cells and live animals (for refs. see ref. 18).Although biolistics is rapidly being adopted to new applications, improvements in the technology are still needed to make it more efficient. For plant species, transient gene expression is relatively easy to achieve, but usually only a few percent of the transiently expressing cells can be recovered as stable transformants. One factor that limits the recovery of stable transformants is injury to the cells. With the commercially available GP2-driven biolistic device (PDS-1000, DuPont), a portion of the cells/tissues are commonly dislodged and/or killed at the epicenter of the blast, creating a central zone without transformation (1, 9, 10). It is likely that a portion of the cells outside of this zone are also injured and impaired from subsequent division and growth.In this study, we used a tobacco cell suspension culture as a model to investigate the causes of cell injury during biolistic transformation. Our results suggest that the primary cause of cell injury is the gas blast and acoustic shock generated by the device. Additionally, the tungsten microprojectiles can themselves be toxic to cells. A new helium-driven device was found to be more gentle to the cells. Shock-attenuating mechanisms placed within the sample chamber also moderated cell injury. Gold particles could be substituted for tungsten and were not toxic.Biolistics (biological ballistics) is a process by which DNA or other biological materials are delivered into cells in association with high-velocity microprojectiles (17). Since its introduction in 1987, there have been numerous reports ...