Spatial Analysis of Slowly Oscillating Electric Activity in the Gut of Mice Using Low Impedance Arrayed Microelectrodes

Smooth and elaborate gut motility is based on cellular cooperation, including smooth muscle, enteric neurons and special interstitial cells acting as pacemaker cells. Therefore, spatial characterization of electric activity in tissues containing these electric excitable cells is required for a precise understanding of gut motility. Furthermore, tools to evaluate spatial electric activity in a small area would be useful for the investigation of model animals. We thus employed a microelectrode array (MEA) system to simultaneously measure a set of 8×8 field potentials in a square area of ∼1 mm². The size of each recording electrode was 50×50 µm², however the surface area was increased by fixing platinum black particles. The impedance of microelectrode was sufficiently low to apply a high-pass filter of 0.1 Hz. Mapping of spectral power, and auto-correlation and cross-correlation parameters characterized the spatial properties of spontaneous electric activity in the ileum of wild-type (WT) and W/W^v mice, the latter serving as a model of impaired network of pacemaking interstitial cells. Namely, electric activities measured varied in both size and cooperativity in W/W^v mice, despite the small area. In the ileum of WT mice, procedures suppressing the excitability of smooth muscle and neurons altered the propagation of spontaneous electric activity, but had little change in the period of oscillations. In conclusion, MEA with low impedance electrodes enables to measure slowly oscillating electric activity, and is useful to evaluate both histological and functional changes in the spatio-temporal property of gut electric activity.     

cDNA clones coding for the structural subunit of a chicken brain nicotinic acetylcholine receptor

Nicotinic acetylcholine receptors (AChRs) immunoaffinity-purified from brains are composed of only two kinds of subunits rather than the four kinds present in muscle-type AChRs. Here we report the N-terminal protein sequences of the structural subunits of AChRs from rat and chicken brains and the cloning of full-length cDNAs for the chicken brain AChR structural subunit. Previously, the N-terminal amino acid sequence of the ACh-binding subunit of AChR immunoaffinity-purified from rat brain was shown to correspond to the cDNA alpha 4. Thus, cDNA sequences are now known for both of the subunits that form one AChR subtype in vivo.     

Effects of surfactant proteins SP-B and SP-C on dynamic and static mechanics of immature lungs

Kobayashi, Tsutomu, Katsumi Tashiro, Ken Yamamoto, ShunichiNitta, Shigeo Ohmura, and Yasuhiro Suzuki. Effects of surfactant proteins SP-B and SP-C on dynamic and static mechanics of immature lungs. J. Appl. Physiol. 83(6):1849-1856, 1997.To investigate the effects of surfactantproteins B (SP-B) and C (SP-C) on lung mechanics, we compared tidal andstatic lung volumes of immature rabbits anesthetized with pentobarbitalsodium and given reconstituted test surfactants (RTS).With a series of RTS having various SP-B concentrations (0-0.7%)but a fixed SP-C concentration (1.4%), both the tidal volume with25-cmH₂O insufflation pressure and the static volume deflated to5-cmH₂O airway pressure increased, significantly correlating with the SP-B concentration: the former increased from 6.5 to 26.0 ml/kg (mean), and the latter increased from6.4 to 31.8 ml/kg. With another series of RTS having afixed SP-B concentration (0.7%) but various SP-C concentrations(0-1.4%), the tidal volume increased from 5.1 to 24.8 ml/kg,significantly correlating with the SP-C concentration, whereas thestatic volume increased from 3.4 to 32.0 ml/kg, the ceiling value, inthe presence of a minimal concentration of SP-C (0.18%). Inconclusion, certain doses of SP-B and SP-C were indispensable foroptimizing dynamic lung mechanics; the static mechanics, however,required significantly less SP-C.

     

Epitope mapping of the human immunodeficiency virus type 1 gp120 with monoclonal antibodies.

A soluble form of recombinant gp120 of human immunodeficiency virus type 1 was used as an immunogen for production of murine monoclonal antibodies. These monoclonal antibodies were characterized for their ability to block the interaction between gp120 and the acquired immunodeficiency syndrome virus receptor, CD4. Three of the monoclonal antibodies were found to inhibit this interaction, whereas the other antibodies were found to be ineffective at blocking binding. The gp120 epitopes which are recognized by these monoclonal antibodies were mapped by using a combination of Western blot (immunoblot) analysis of gp120 proteolytic fragments, immunoaffinity purification of fragments of gp120, and antibody screening of a random gp120 gene fragment expression library produced in the lambda gt11 expression system. Two monoclonal antibodies which blocked gp120-CD4 interaction were found to map to adjacent sites in the carboxy-terminal region of the glycoprotein, suggesting that this area is important in the interaction between gp120 and CD4. One nonblocking antibody was found to map to a position that was C terminal to this CD4 blocking region. Interestingly, the other nonblocking monoclonal antibodies were found to map either to a highly conserved region in the central part of the gp120 polypeptide or to a highly conserved region near the N terminus of the glycoprotein. N-terminal deletion mutants of the soluble envelope glycoprotein which lack these highly conserved domains but maintain the C-terminal CD4 interaction sites were unable to bind tightly to the CD4 receptor. These results suggest that although the N-terminal and central conserved domains of intact gp120 do not appear to be directly required for CD4 binding, they may contain information that allows other parts of the molecule to form the appropriate structure for CD4 interaction.     

Colorimetric detection of oxalate in aqueous solution by a pyrogallol red‐based Cu2+ complex

The pyrogallol red (PR)‐based Cu²⁺ complex was proven to be an effective and selective colorimetric chemosensing ensemble for recognition of oxalate over other anions in a perfect aqueous solution. The addition of oxalate to the PR–Cu²⁺ complex resulted in a colour change from purple to orange colour due to the regeneration of PR by the chelation of oxalate with Cu²⁺, while other anions did not induce any significant colour change. Moreover, it was revealed that no obvious interference was observed during the titrations with oxalate into each other anion. Therefore, the PR–Cu²⁺ complex can be used as a simple and practical colorimetric chemosensor for detecting oxalate.     

Camellidins,Antifungal Saponins Isolated from Camellia japonica

Two triterpenoid saponins were isolated from an aqueous or a methanolic extract of camellia (Camellia japonica) leaf. They had an antifungal activity characterized by abnormal germination of conidia. These saponins were composed of 3βhydroxy-18β-acetoxy-28-norolean-12-en-16-one or 3β, 8β-dihydroxy-28-norolean-12-en-16-one as aglycon, and d-glucuronic acid, dglucose and two moles of dgalactose as the sugar moiety. The authors have named these new saponins “Camellidin,” which might have value for studies in the fields of phytopathology and biochemistry.     

Studies on Molecular Properties of αs1-κ-Casein Complex by the Hydrodynamical Methods

Some molecular properties of α_s1-κ-casein complex, α_s1- and κ-casein polymers were examined by gel filtration, ultracentrifugation, and viscometry at pH 7.1. The Stoke’s radii of α_s1-κ-casein complex, α_s1- and κ-casein polymers were 99, 44 and 108 Å, respectively. The molecular weight of the above proteins were approximately 45 × 10⁴, 10 × 10⁴ and 80 × 10⁴, respectively. The stokes radius of α_s1-κ casein complex reduced compared with that of κ-casein polymer and the molecular weight of the complex was about half that of κ-casein polymer. These results suggest that κ-casein polymer dissociates into 4 smaller particles when α_s1-κ-casein complex is formed. The frictional coefficient and Scheraga-Mandelkern constants for each protein suggest that the molecular shape of α_s1-casein polymer is globular and that of α_s1-κ-casein complex and κ-casein polymer is rod-like.     

Aberrant Glycogen Synthase Kinase 3β Is Involved in Pancreatic Cancer Cell Invasion and Resistance to Therapy

Background and Purpose

The major obstacles to treatment of pancreatic cancer are the highly invasive capacity and resistance to chemo- and radiotherapy. Glycogen synthase kinase 3β (GSK3β) regulates multiple cellular pathways and is implicated in various diseases including cancer. Here we investigate a pathological role for GSK3β in the invasive and treatment resistant phenotype of pancreatic cancer.

Methods

Pancreatic cancer cells were examined for GSK3β expression, phosphorylation and activity using Western blotting and in vitro kinase assay. The effects of GSK3β inhibition on cancer cell survival, proliferation, invasive ability and susceptibility to gemcitabine and radiation were examined following treatment with a pharmacological inhibitor or by RNA interference. Effects of GSK3β inhibition on cancer cell xenografts were also examined.

Results

Pancreatic cancer cells showed higher expression and activity of GSK3β than non-neoplastic cells, which were associated with changes in its differential phosphorylation. Inhibition of GSK3β significantly reduced the proliferation and survival of cancer cells, sensitized them to gemcitabine and ionizing radiation, and attenuated their migration and invasion. These effects were associated with decreases in cyclin D1 expression and Rb phosphorylation. Inhibition of GSK3β also altered the subcellular localization of Rac1 and F-actin and the cellular microarchitecture, including lamellipodia. Coincident with these changes were the reduced secretion of matrix metalloproteinase-2 (MMP-2) and decreased phosphorylation of focal adhesion kinase (FAK). The effects of GSK3β inhibition on tumor invasion, susceptibility to gemcitabine, MMP-2 expression and FAK phosphorylation were observed in tumor xenografts.

Conclusion

The targeting of GSK3β represents an effective strategy to overcome the dual challenges of invasiveness and treatment resistance in pancreatic cancer.