SGIP1alpha is an endocytic protein that directly interacts with phospholipids and Eps15

SGIP1 has been shown to be an endophilin-interacting protein that regulates energy balance, but its function is not fully understood. Here, we identified its splicing variant of SGIP1 and named it SGIP1alpha. SGIP1alpha bound to phosphatidylserine and phosphoinositides and deformed the plasma membrane and liposomes into narrow tubules, suggesting the involvement in vesicle formation during endocytosis. SGIP1alpha furthermore bound to Eps15, an important adaptor protein of clathrin-mediated endocytic machinery. SGIP1alpha was colocalized with Eps15 and the AP-2 complex. Upon epidermal growth factor (EGF) stimulation, SGIP1alpha was colocalized with EGF at the plasma membrane, indicating the localization of SGIP1alpha at clathrin-coated pits/vesicles. SGIP1alpha overexpression reduced transferrin and EGF endocytosis. SGIP1alpha knockdown reduced transferrin endocytosis but not EGF endocytosis; this difference may be due to the presence of redundant pathways in EGF endocytosis. These results suggest that SGIP1alpha plays an essential role in clathrin-mediated endocytosis by interacting with phospholipids and Eps15.     

Accumulation of multiacetylated forms of histones by trichostatin A and its developmental consequences in early starfish embryos

Summary External application of 10 rig/ml (R)-trichostatin A (TSA), a potent and specific inhibitor of mammalian histone deacetylase, to the embryo of the starfish Asterina pectinifera inhibited development during the early gastrula stage before formation of mesenchyme cells. The TSA-sensitive period was limited to the mid-blastula stage before hatching. The pulse-chase experiment clearly demonstrated that TSA induced an accumulation of acetylated histone species in blastulae through inhibition of historic deacetylation. Similar blockage of development at the early gastrula stage was observed with n-butyrate, which has been known as a weak inhibitor of historic deacetylase. These results suggest an intimate role for historic acetylation-deacetylation equilibria in starfish development. Correspondence to: S. Ikegami     

Mice lacking alpha1,3-fucosyltransferase IX demonstrate disappearance of Lewis x structure in brain and increased anxiety-like behaviors

The 3-fucosyl-N-acetyllactosamine [Lewis x (Le(x)), CD15, SSEA-1] carbohydrate structure is expressed on several glycolipids, glycoproteins, and proteoglycans of the nervous system and has been implicated in cell-cell recognition, neurite outgrowth, and neuronal migration during development. To characterize the functional role of Le(x) carbohydrate structure in vivo, we have generated mutant mice that lack alpha1,3-fucosyltransferase IX (Fut9(-/-)). Fut9(-/-) mice were unable to synthesize the Le(x) structure carried on glycoproteins and glycolipids in embryonic and adult brain. However, no obvious pathological differences between wild-type and Fut9(-/-) mice were found in brain. In behavioral tests, Fut9(-/-) mice exhibited increased anxiety-like responses in dark-light preference and in elevated plus maze tests. Immunohistochemical analysis showed that the number of calbindin-positive neurons was decreased in the basolateral amygdala in Fut9(-/-) mice. These observations indicated that the carbohydrates synthesized by Fut9 play critical roles in functional regulations of interneurons in the amygdalar subdivisions and suggested a role for the Le(x) structure in some aspects of emotional behavior in mice.     

The formation of Zn-Chl a in Chlorella heterotrophically grown in the dark with an excessive amount of Zn2+

Chlorella, when heterotrophically cultivated in the dark, is able to grow with Zn2+ at 10-40 mM, which is 10 times the concentration lethal to autotrophically grown cells. However, the lag phase is prolonged with increasing concentrations of Zn2+; for example, in this study, 1 d of the control lag phase was prolonged to about 16 d with Zn2+ at 16.7 mM (x2,000 of the control). Once the cells started to grow, the log phase was finished within 4-6 d regardless of Zn concentration, which was almost the same as that of the control. The photosysystem I reaction center chlorophyll, P700, and the far-red fluorescence were detected only after the late log phase of the growth curve, suggesting that chlorophyll-protein complexes can be organized after cell division has ceased. Interestingly, at more than 16.7 mM of Zn2+, Zn-chlorophyll a was accumulated and finally accounted for about 25% of the total chlorophyll a in the late stationary phase. We found that the Zn-chlorophyll a was present in the thylakoid membranes and not in the soluble fractions of the cells. The rather low fluorescence yield at around 680 nm in the stationary phase suggests that Zn-chlorophyll a can transfer its excitation energy to other chlorophylls. Before accumulation of Zn-chlorophyll a, a marked amount of pheophytin a was temporally accumulated, suggesting that Zn-chlorophyll a could be chemically synthesized via pheophytin a.     

Practical removal of radioactivity from sediment mud in a swimming pool in Fukushima, Japan by immobilized photosynthetic bacteria

About 90% of the radioactive Cs in the sediment mud of a school's swimming pool in Fukushima, Japan was removed by treatment for 3 d using the alginate immobilized photosynthetic bacterium Rhodobcater sphaeroides SSI. Even though batch treatment was carried out 3 times repeatedly, the activity of immobilized cells in removing Cs was maintained at levels of about 84% (second batch) and 78% (third batch). Cs was strongly attached to the sediment mud because, even with HNO(3) treatment at pH of 2.00-1.60 for 24 h, it was not eluted into the water. Furthermore, more than 75% of the Cs could be removed without solubilization with HNO(3). This suggests that the Cs attached to the sediment mud was transformed into immobilized cells via the Cs(+) ion by the negative charge of the immobilized cell surface and/or the potassium transport system of the photosynthetic bacterium.     

Inhibitors of inflammation and endogenous surfactant pool size as modulators of lung injury with initiation of ventilation in preterm sheep

Background

Increased pro-inflammatory cytokines in tracheal aspirates correlate with the development of BPD in preterm infants. Ventilation of preterm lambs increases pro-inflammatory cytokines and causes lung inflammation.

Objective

We tested the hypothesis that selective inhibitors of pro-inflammatory signaling would decrease lung inflammation induced by ventilation in preterm newborn lambs. We also examined if the variability in injury response was explained by variations in the endogenous surfactant pool size.

Methods

Date-mated preterm lambs (n = 28) were operatively delivered and mechanically ventilated to cause lung injury (tidal volume escalation to 15 mL/kg by 15 min at age). The lambs then were ventilated with 8 mL/kg tidal volume for 1 h 45 min. Groups of animals randomly received specific inhibitors for IL-8, IL-1, or NF-κB. Unventilated lambs (n = 7) were the controls. Bronchoalveolar lavage fluid (BALF) and lung samples were used to quantify inflammation. Saturated phosphatidylcholine (Sat PC) was measured in BALF fluid and the data were stratified based on a level of 5 μmol/kg (~8 mg/kg surfactant).

Results

The inhibitors did not decrease the cytokine levels or inflammatory response. The inflammation increased as Sat PC pool size in BALF decreased. Ventilated lambs with a Sat PC level > 5 μmol/kg had significantly decreased markers of injury and lung inflammation compared with those lambs with < 5 μmol/kg.

Conclusion

Lung injury caused by high tidal volumes at birth were decreased when endogenous surfactant pool sizes were larger. Attempts to decrease inflammation by blocking IL-8, IL-1 or NF-κB were unsuccessful.     

Lymphatic ontogeny and effect of hypoplasia in developing lung

The pulmonary lymphatic vasculature plays a vital role in maintaining fluid homeostasis required for efficient gas exchange at capillary alveolar barriers and contributes to lung fluid clearance at birth. To further understanding of pulmonary lymphatic function at birth, lineage-tracing analysis of mouse lung was used. Lineage analysis confirmed that lymphatic endothelial cells (LEC) bud from extrapulmonary lymphatics and demonstrated that LEC migrate into developing lung along precise pathways. LEC cluster first in the primary bronchovascular region then along the secondary broncho-arterial regions and along veins. Small lymphatic vessels in distal lung develop from LEC that have migrated into lung mesenchyme from the extrapulmonary lymphatics. Finally, proximal and distal lymphatics remodel to form vessels with lumens in stereotypical locations. Loss of function analysis with lung-specific expression of a secreted form of the extracellular domain of vascular endothelial growth factor receptor-3 (dnR3) caused significant embryonic pulmonary lymphatic hypoplasia with fourfold reduction in distal LEC. Lung-specific expression of dnR3 did not affect blood vascular development, overall lung organogenesis or lymphatic development in other organs. Neonatal mice with pulmonary lymphatic hypoplasia developed respiratory distress with significantly increased mortality. During the transition to air breathing, lymphatic hypoplasia adversely affected fetal lung fluid clearance as determined by wet/dry weight analysis and morphometric analysis of bronchovascular cuffing and mesenchymal thickening. Surfactant synthesis was unaffected. Together, these data demonstrate that lung lymphatics develop autonomously and that pulmonary lymphatic hypoplasia is detrimental to survival of the neonate due to impaired lung fluid clearance.