Insulin resistance and decreased spexin in Indian Patients with Type 2 Diabetes Mellitus

Spexin is novel biomarker, which plays a potential role in glucose and lipid metabolisms. However, there was paucity of serum spexin levels in obesity and diabetes mellitus subjects. Hence the current study was aimed to find the relationship between the serum spexin levels in type 2 Diabetes mellitus (type 2 DM) with extrapolation of cardiovascular disease (CVD) risk. A cross-sectional study included 330 participants, subdivided as control (n=110), type 2 DM (n=110) and type 2 DM with CVD groups (n=110). HbA1c, insulin, lipid profile, spexin & leptin including blood pressure and body mass index were analyzed from all the participants. The serum spexin levels (ng/ml) were significantly decreased in type 2 DM (mean ± sd: 0.65 ± 0.03) and type 2 DM with CVD (0.48 ± 0.02) groups compared to the control (0.79 ± 0.03) group (p<0.001). The decreased spexin levels were observed in type 2 DM, and further more decreased in type 2 DM with CVD patients compared to controls indicating that spexin levels could be served as an early prediction of obesity-induced T2DM with CVD risk.     

Real-time imaging of the dynamics of secretory granules in growth cones

Secretory granules containing a hybrid protein consisting of the regulated secretory protein tissue plasminogen activator and an enhanced form of green fluorescent protein were tracked at high spatial resolution in growth cones of differentiated PC12 cells. Tracking shows that granules, unlike synaptic vesicles, generally are mobile in growth cones. Quantitative analysis of trajectories generated by granules revealed two dominant modes of motion: diffusive and directed. Diffusive motion was observed primarily in central and peripheral parts of growth cones, where most granules diffused two to four orders of magnitude more slowly than comparably sized spheres in dilute solution. Directed motion was observed primarily in proximal parts of growth cones, where a subset of granules underwent rapid, directed motion at average speeds comparable to those observed for granules in neurites. This high-resolution view of the dynamics of secretory granules in growth cones provides insight into granule organization and release at nerve terminals. In particular, the mobility of granules suggests that granules, unlike synaptic vesicles, are not tethered stably to cytoskeletal structures in nerve terminals. Moreover, the slow diffusive nature of this mobility suggests that secretory responses involving centrally distributed granules in growth cones will occur slowly, on a time scale of minutes or longer.     

Mucociliary transport determined by in vivo microdialysis in the airways of normal and CF mice

We report a novel method to measure mucociliary transport (MCT) in both the upper and lower airways of normal and CF mice. The in vivo microdialysis technique involves placing a small quantity of dye on the airway surface and a microdialysis probe a defined distance from the site of dye deposition. The dye is transported toward the probe by ciliary transport and, upon reaching the microdialysis probe, diffuses across the dialysis membrane and is collected in the dialysate leaving the probe. The rate of MCT is calculated from the length of time from dye deposition to recovery. The rate of tracheal MCT in normal mice was 2.2 +/- 0.45 (SE) mm/min (n = 6), a value similar to that in reports using other techniques. MCT in CF mice was not different (2.3 +/- 0.29, n = 6), consistent with previous observations suggesting that tracheal ion transport properties are not different between CF and normal mice. The rate of MCT in the nasal cavity of normal mice was slower than in the trachea (1.3 +/- 0.26, n = 4). MCT in the CF mouse nasal cavity (1.4 +/- 0.31, n = 8), a region in which the CF mouse exhibits bioelectric properties similar to the human CF patient, was, again, not different from the normal mouse, perhaps reflecting copious gland secretion offsetting Na(+) and liquid hyperabsorption. In conclusion, we have developed a versatile, simple in vivo method to measure MCT in both upper and lower airways of mice and larger animals.     

Conformational control of Bax localization and apoptotic activity by Pro168

In healthy cells, Bax resides inactive in the cytosol because its COOH-terminal transmembrane region (TMB) is tucked into a hydrophobic pocket. During apoptosis, Bax undergoes a conformational change involving NH2-terminal exposure and translocates to mitochondria to release apoptogenic factors. How this process is regulated remains unknown. We show that the TMB of Bax is both necessary and sufficient for mitochondrial targeting. However, its availability for targeting depends on Pro168 located within the preceding loop region. Pro168 mutants of Bax lack apoptotic activity, cannot rescue the apoptosis-resistant phenotype of Bax/Bak double knockout cells, and are retained in the cytosol even in response to apoptotic stimuli. Moreover, the mutants have their NH2 termini exposed. We propose that Pro168 links the NH2 and the COOH terminus of Bax and is required for COOH-terminal release and mitochondrial targeting once this link is broken.     

The influence of early experience on the development of sensory systems

Once sensory stimuli become able to alter firing patterns in the developing brain, they can influence the maturation of neuronal circuits. Recent experimental studies add to our understanding of precisely which developmental events are affected by early experience. In particular, it appears that experience of the external environment can affect the brain earlier in development and at earlier stages of sensory processing than previously thought. These studies emphasise the developmental importance of the patterning of neuronal firing produced either by sensory stimuli or by spontaneous activity. The timing of action potentials is also an important aspect of several exciting studies describing the mechanisms - anatomical, synaptic, and molecular - by which early experience brings about alterations in the maturation of sensory circuitry. Importantly, this kind of approach can lead to predictions concerning the nature of sensory stimulation that is most effective in instructing brain development.     

Calcium ion promotes yeast Dmc1 activity via formation of long and fine helical filaments with single-stranded DNA

Dmc1 is specifically required for homologous recombination during meiosis. Here we report that the calcium ion enabled Dmc1 from budding yeast to form regular helical filaments on single-stranded DNA (ssDNA) and activate its strand assimilation activity. Relative to magnesium, calcium increased the affinity of Dmc1 for ATP and but reduces its DNA-dependent ATPase activity. These effects, together with previous studies of other RecA-like recombinases, support the view that ATP binding to Dmc1 protomers is required for functional filament structure. The helical pitch of the Saccharomyces cerevisiae Dmc1-ssDNA helical filament was estimated to be 13.4 +/- 2.5 nm. Analysis of apparently "complete" Dmc1-ssDNA filaments indicated a stoichiometry of 24 +/- 2 nucleotides per turn of the Dmc1 helix. This finding suggests that the number or protomers per helical turn and/or the number of nucleotides bound per Dmc1 protomer differs from that reported previously for Rad51 and RecA filaments. Our data support the view that the active form of Dmc1 protein is a helical filament rather than a ring. We speculate that Ca(2+) plays a significant role in regulating meiotic recombination.     

Reduced three-dimensional motility in dehydrated airway mucus prevents neutrophil capture and killing bacteria on airway epithelial surfaces

Cystic fibrosis (CF) lung disease is characterized by persistent lung infection. Thickened (concentrated) mucus in the CF lung impairs airway mucus clearance, which initiates bacterial infection. However, airways have other mechanisms to prevent bacterial infection, including neutrophil-mediated killing. Therefore, we examined whether neutrophil motility and bacterial capture and killing functions are impaired in thickened mucus. Mucus of three concentrations, representative of the range of normal (1.5 and 2.5% dry weight) and CF-like thickened (6.5%) mucus, was obtained from well-differentiated human bronchial epithelial cultures and prepared for three-dimensional studies of neutrophil migration. Neutrophil chemotaxis in the direction of gravity was optimal in 1.5% mucus, whereas 2.5% mucus best supported neutrophil chemotaxis against gravity. Lateral chemokinetic movement was fastest on airway epithelial surfaces covered with 1.5% mucus. In contrast, neutrophils exhibited little motility in any direction in thickened (6.5%) mucus. In in vivo models of airway mucus plugs, neutrophil migration was inhibited by thickened mucus (CF model) but not by normal concentrations of mucus ("normal" model). Paralleling the decreased neutrophil motility in thickened mucus, bacterial capture and killing capacity were decreased in CF-like thickened mucus. Similar results with each mucus concentration were obtained with mucus from CF cultures, indicating that inhibition of neutrophil functions was mucus concentration dependent not CF source dependent. We conclude that concentrated ("thick") mucus inhibits neutrophil migration and killing and is a key component in the failure of defense against chronic airways infection in CF.     

Human cystatin C, an amyloidogenic protein, dimerizes through three-dimensional domain swapping

The crystal structure of human cystatin C, a protein with amyloidogenic properties and a potent inhibitor of cysteine proteases, reveals how the protein refolds to produce very tight two-fold symmetric dimers while retaining the secondary structure of the monomeric form. The dimerization occurs through three-dimensional domain swapping, a mechanism for forming oligomeric proteins. The reconstituted monomer-like domains are similar to chicken cystatin except for one inhibitory loop that unfolds to form the 'open interface' of the dimer. The structure explains the tendency of human cystatin C to dimerize and suggests a mechanism for its aggregation in the brain arteries of elderly people with amyloid angiopathy. A more severe 'conformational disease' is associated with the L68Q mutant of human cystatin C, which causes massive amyloidosis, cerebral hemorrhage and death in young adults. The structure of the three-dimensional domain-swapped dimers shows how the L68Q mutation destabilizes the monomers and makes the partially unfolded intermediate less unstable. Higher aggregates may arise through the three-dimensional domain-swapping mechanism occurring in an open-ended fashion in which partially unfolded molecules are linked into infinite chains.     

The CF salt controversy: in vivo observations and therapeutic approaches

There is controversy over whether abnormalities in the salt concentration or volume of airway surface liquid (ASL) initiate cystic fibrosis (CF) airway disease. In vivo studies of CF mouse nasal epithelia revealed an increase in goblet cell number that was associated with decreased ASL volume rather than abnormal [Cl(-)]. Aerosolization of osmolytes in vivo failed to raise ASL volume. In vitro studies revealed that osmolytes and pharmacological agents were effective in producing isotonic volume responses in human airway epithelia but were typically short acting and less effective in CF cultures with prolonged volume hyperabsorption and mucus accumulation. These data show that (1) therapies can be designed to normalize ASL volume, without producing deleterious compositional changes in ASL, and (2) therapeutic efficacy will likely depend on development of long-acting pharmacologic agents and/or an increased efficiency of osmolyte delivery.