Multiscale Biodistribution Analysis of Lipophilic, Poorly Soluble Drugs.

Clofazimine is a poorly soluble drug that accumulates as solid deposits in the body during prolonged oral administration. At the outset, we hypothesized that clofazimine accumulated intracellularly by a passive and spontaneous crystallization, and in various levels of experimental set-ups, from a tissue culture to mouse models. We found that clofazimine readily formed amorphous inclusions in complexes with intracellular membranes in MDCK cells, while different types of inclusions were found in the tissue macrophages of clofazimine-diet fed mice. Most of the inclusions in vivo appeared as vibrant red, birefringent, 10 – 20 µm length crystal-like structures; however, their physicochemical and morphological characteristics were inconsistent with those of pure clofazimine crystals. Most remarkably, among the inclusions from macrophages, we discovered a new cytoplasmic structure delimited by double membranes with internal supramolecular organizations resembling stacks of lipidic lamellae. Upon prolonged dosing, the intact clofazimine was redistributed from adipose tissues to the lymphatic organs paralleled by anti-inflammatory responses such as splenomegaly, liver microgranulomas, and an expansion of macrophage populations. In conclusion, instead of passive intracellular crystallization hypothesis, I propose that clofazimine accumulates in vivo by active sequestration in the immune system. By constructing intracellular crystal- and organelle-like “polyhedrosomes”, the macrophages can impact the clofazimine’s systemic pharmacokinetics and biodistribution, from micro to macro scale.Ph.D.Pharmaceutical SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/91404/1/jsbaik_1.pd

Melatonin receptor 1 B polymorphisms associated with the risk of gestational diabetes mellitus

<p>Abstract</p> <p>Backgrounds</p> <p>Two SNPs in <it>melatonin receptor 1B </it>gene, <it>rs10830963 </it>and <it>rs1387153 </it>showed significant associations with fasting plasma glucose levels and the risk of Type 2 Diabetes Mellitus (T2DM) in previous studies. Since T2DM and gestational diabetes mellitus (GDM) share similar characteristics, we suspected that the two genetic polymorphisms in <it>MTNR1B </it>may be associated with GDM, and conducted association studies between the polymorphisms and the disease. Furthermore, we also examined genetic effects of the two polymorphisms with various diabetes-related phenotypes.</p> <p>Methods</p> <p>A total of 1,918 subjects (928 GDM patients and 990 controls) were used for the study. Two <it>MTNR1B </it>polymorphisms were genotyped using TaqMan assay. The allele distributions of SNPs were evaluated by <it>x</it>²models calculating odds ratios (ORs), 95% confidence intervals (CIs), and corresponding <it>P </it>values. Multiple regressions were used for association analyses of GDM-related traits. Finally, conditional analyses were also performed.</p> <p>Results</p> <p>We found significant associations between the two genetic variants and GDM, <it>rs10830963</it>, with a corrected <it>P </it>value of 0.0001, and <it>rs1387153</it>, with the corrected <it>P </it>value of 0.0008. In addition, we also found that the two SNPs were associated with various phenotypes such as homeostasis model assessment of beta-cell function and fasting glucose levels. Further conditional analyses results suggested that <it>rs10830963 </it>might be more likely functional in case/control analysis, although not clear in GDM-related phenotype analyses.</p> <p>Conclusion</p> <p>There have been studies that found associations between genetic variants of other genes and GDM, this is the first study that found significant associations between SNPs of <it>MTNR1B </it>and GDM. The genetic effects of two SNPs identified in this study would be helpful in understanding the insight of GDM and other diabetes-related disorders.</p

Deep-etch freeze-fracture microscopy of the intracellular CLDI from 6 wk treated mouse liver.

<p>Note the outer membrane of the CLDI is studded with globular protein-like features (white triangle). The CLDI was broken during freeze-fracture, revealing a lamellar internal structure at the top.</p

Outer membrane and internal organization of a CLDI from 6 wk treated mouse liver.

<p>(<b>A</b>) Zoomed out image, revealing the exposed, inner multilamellar core with the outer double membrane layers peeled back towards the cytoplasm of the cell. The CLDI was broken open during the sample preparation process. Regions of interest (corresponding to panels B to F) are marked with letters. (<b>B</b>) Zoomed image of the inner lamellar surface. (<b>C</b>) Zoomed image of the multilamellar core region, showing variable spacings in the order of ranging from 6 to 14 nanometers. (<b>D</b>) Zoomed image of the innermost face of the inner bounding double membrane without the globular, protein-like features. (<b>E</b>) Zoomed image of the innermost face of the outer bounding double membrane. Protein-sized features are observed on the inner surface of the outer membrane (arrowheads). (<b>F</b>) Zoomed image of the outer double membrane. Note the large space between the double membranes, with “pillars” bridging the membranes (arrows).</p

Molecular Imaging of Intracellular Drug–Membrane Aggregate Formation

Clofazimine is a lipophilic antibiotic with an extremely long pharmacokinetic half-life associated with the appearance of crystal-like drug inclusions, in vivo. Here, we studied how clofazimine accumulates inside cells in the presence of supersaturating, extracellular concentrations of the drug (in the range of physiological drug concentrations). Based on a combination of molecular imaging, biochemical analysis and electron microscopy techniques, clofazimine mass increased inside cells in vitro, over a period of several days, with discrete clofazimine inclusions forming in the cytoplasm. These inclusions grew in size, number and density, as long as the drug-containing medium was replenished. With Raman confocal microscopy, clofazimine’s spectral signature in these inclusions resembled that of amorphous clofazimine precipitates and was unlike that of clofazimine crystals. Additional experiments revealed that clofazimine first accumulated in mitochondria, with ensuing changes in mitochondrial structure and function. In turn, the degenerating organelles coalesced, fused with each other and condensed to form prominent drug–membrane aggregates (dubbed autophagosome-like drug inclusions or “aldis”). Like clofazimine, it is possible that intracellular drug–membrane aggregate formation is a common phenomenon underlying the reported phenotypic effects of many other small molecule drugs

Social Media Activism and Convergence in Tweet Topics After the Initial #MeToo Movement for Two Distinct Groups of Twitter Users

Online social media movements are now common and support cultural discussions on difficult health and social topics. The #MeToo movement, focusing on the pervasiveness of sexual assault and harassment, has been one of the largest and most influential online movements. Our study examines topics of conversation on Twitter by supporters of the #MeToo movement and by Twitter users who were uninvolved in the movement to explore the extent to which tweet topics for these two groups converge over time. We identify and collect one year’s worth of tweets for supporters of the #MeToo movement ( N = 168 users; N = 105,538 tweets) and users not involved in the movement ( N = 147 users; N = 112,301 tweets referred to as the Neutral Sample). We conduct topic frequency analysis and implement an unsupervised machine learning topic modeling algorithm, latent Dirichlet allocation, to explore topics of discussion on Twitter for these two groups of users before and after the initial #MeToo movement. Our results suggest that supporters of #MeToo discussed different topics compared to the Neutral Sample of Twitter users before #MeToo with some overlap on politics. The supporters were already discussing sexual assault and harassment issues six months before #MeToo, and discussion on this topic increased 13.7-fold in the six months after. For the Neutral Sample, sexual assault and harassment was not a key topic of discussion on Twitter before #MeToo, but there was some limited increase afterward. Results of bigram frequency analysis and topic modeling showed a clear increase in topic related to gender for the supporters of #MeToo but gave mixed results for the Neutral Sample comparison group. Our results suggest limited shifts in the conversation on Twitter for the Neutral Sample. Our methods and results have implications for measuring the extent to which online social media movements, like #MeToo, reach a broad audience. </jats:p