Transmission electron microscopy (TEM) of hygroscopic, permeable, and electron-absorbing biological cells has been an important challenge due to the volumetric shrinkage, electrostatic charging, and structural degradation of cells under high vacuum and fixed electron beam.(1-3) Here we show that bacterial cells can be encased within a graphenic chamber to preserve their dimensional and topological characteristics under high vacuum (10(-5) Torr) and beam current (150 A/cm(2)). The strongly repelling π clouds in the interstitial sites of graphene's lattice(4) reduces the graphene-encased-cell's permeability(5) from 7.6-20 nm/s to 0 nm/s. The C-C bond flexibility(5,6) enables conformal encasement of cells. Additionally, graphene's high Young's modulus(6,7) retains cell's structural integrity under TEM conditions, while its high electrical(8) and thermal conductivity(9) significantly abates electrostatic charging. We envision that the graphenic encasement approach will facilitate real-time TEM imaging of fluidic samples and potentially biochemical activity.

Adiponectin (Acrp30) is an insulin-sensitizing hormone produced and secreted exclusively by adipose tissue. Confocal fluorescent microscopy demonstrated the colocalization of adiponectin with the Golgi membrane markers p115, ␤-COP, and the transGolgi network marker, syntaxin 6. Treatment of cells with brefeldin A redistributed adiponectin to the endoplasmic reticulum where it colocalized with the chaperone protein BIP and inhibited secretion of adiponectin demonstrating a requirement for a functional Golgi apparatus for adiponectin release. Confocal fluorescent microscopy also demonstrated a colocalization of endogenous adiponectin with that of expressed GGA1myc (Golgi-localizing ␥-adaptin ear homology ARF-binding protein) but with no significant overlap between adiponectin and the GGA2myc or GGA3myc isoforms. Consistent with confocal fluorescent microscopy, transmission electron microscopy demonstrated the colocalization of GGA1 with adiponectin. Although GGA1 did not directly interact with the adiponectin protein, the adiponectin enriched membrane compartments of adipocyte were precipitated by a GST-GGA1 cargo binding domain (VHS) fusion protein but not with a GST-GGA2 VHS or GST-GGA3 VHS fusion proteins. Moreover, co-expression of adiponectin with a GGA1 dominant-interfering mutant (GGA1-VHS GAT domain) resulted in a marked inhibition of adiponectin secretion in both 3T3L1 adipocytes and HEK293 cells, whereas no inhibition was detected with the truncated mutants GGA2-VHSGAT or GGA3-VHSGAT. Moreover, co-expression of wild type GGA1 with adiponectin enhanced secretion of adiponectin. Interestingly, leptin secretion was unaffected by neither the wild type form or GGA1 mutant. Taken together these data demonstrate that the trafficking of adiponectin through its secretory pathway is dependent on GGAcoated vesicles.Over the past several years many studies have documented that in addition to being a fat storage depot, adipocytes are a bona fide endocrine tissue that secrete several hormones that control insulin sensitivity and energy balance (1-3). In particular, adiponectin, also called adipocyte complement-related protein of 30 kDa (Acrp30), adipoQ, GBP28, and apM1, was originally isolated as a highly induced gene following adipocyte differentiation (4). This hormone is secreted exclusively by adipocytes (4 -7) and functions in vivo as an insulin sensitizer (8 -10), reducing glucose production by the liver (11) and enhancing fatty acid oxidation in skeletal muscle (10) through the activation of two distinct receptor isoforms (12). These receptors mediate increased AMP-dependent kinase activation (13-15) and peroxisome proliferating activated receptor-␣ ligand activity (16).Adiponectin serum levels inversely correlate with insulin resistance in both in animals and humans (17-21) in contrast to that observed for other adipokines such as tumor necrosis factor-␣ and resistin (22-25). Type 2 diabetic patients also display reduced levels of adiponectin (26 -29). Moreover, injection of purified adiponectin decreases gluc...

C. difficile infects thousands of hospitalized patients every year, causing significant morbidity and mortality. C. difficile spores play a pivotal role in the transmission of the pathogen in the hospital environment. During infection, the spores germinate, and the vegetative bacterial cells produce toxins that damage host tissue. Thus, sporulation and toxin production are two important traits of C. difficile. In this study, we showed that a mutation in tcdR, the toxin gene regulator, affects both toxin production and sporulation in epidemic-type C. difficile strain R20291.

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Background. Clinical placements are an integral component of physiotherapy education as they give physiotherapy students the opportunity to apply their academic knowledge and skills and at the same time universities aim to provide an education that responds to the demands of practice settings.