Multiple Na,K-ATPase Subunits Colocalize in the Brush Border of Mouse Choroid Plexus Epithelial Cells

(1) Background: The unusual accumulation of Na,K-ATPase complexes in the brush border membrane of choroid plexus epithelial cells have intrigued researchers for decades. However, the full range of the expressed Na,K-ATPase subunits and their relation to the microvillus cytoskeleton remains unknown. (2) Methods: RT-PCR analysis, co-immunoprecipitation, native PAGE, mass spectrometry, and differential centrifugation were combined with high-resolution immunofluorescence histochemistry, proximity ligase assays, and stimulated emission depletion (STED) microscopy on mouse choroid plexus cells or tissues in order to resolve these issues. (3) Results: The choroid plexus epithelium expresses Na,K-ATPase subunits α1, α2, β1, β2, β3, and phospholemman. The α1, α2, β1, and β2, subunits are all localized to the brush border membrane, where they appear to form a complex. The ATPase complexes may stabilize in the brush border membrane via anchoring to microvillar actin indirectly through ankyrin-3 or directly via other co-precipitated proteins. Aquaporin 1 (AQP1) may form part of the proposed multi-protein complexes in contrast to another membrane protein, the Na-K-2Cl cotransporter 1 (NKCC1). NKCC1 expression seems necessary for full brush border membrane accumulation of the Na,K-ATPase in the choroid plexus. (4) Conclusion: A multitude of Na,K-ATPase subunits form molecular complexes in the choroid plexus brush border, which may bind to the cytoskeleton by various alternative actin binding proteins.     

Interactions between immunoglobulin-like and catalytic modules in Clostridium thermocellum cellulosomal cellobiohydrolase CbhA

Cellobiohydrolase CbhA from Clostridium thermocellum cellulosome is a multi-modular protein composed starting from the N-terminus of a carbohydrate-binding module (CBM) of family 4, an immunoglobulin(Ig)-like module, a catalytic module of family 9 glycoside hydrolases (GH9), X1(1) and X1(2) modules, a CBM of family 3 and a dockerin module. Deletion of the Ig-like module from the Ig-GH9 construct results in complete inactivation of the GH9 module. The crystal structure of the Ig-GH9 module pair reveals the existence of an extensive module interface composed of over 40 amino acid residues of both modules and maintained through a large number of hydrophilic and hydrophobic interactions. To investigate the importance of these interactions between the two modules, we compared the secondary and tertiary structures and thermostabilities of the individual Ig-like and GH9 modules and the Ig-GH9 module pair using both circular dichroism (CD) spectroscopy and differential scanning calorimetry (DSC). Thr230, Asp262 and Asp264 of the Ig-like module are located in the module interface of the Ig-GH9 module pair and are suggested to be important in 'communication' between the modules. These residues were mutated to alanyl residues. The structure, stability and catalytic properties of the native Ig-GH9 and its D264A and T230A/D262A mutants were compared. The results indicate that despite being able to fold relatively independently, the Ig-like and GH9 modules interact and these interactions affect the final fold and stability of each module. Mutations of one or two amino acid residues lead to destabilization and change of the mechanism of thermal unfolding of the polypeptides. The enzymatic properties of native Ig-GH9, D264A and T230A/D262A mutants are similar. The results indicate that inactivation of the GH9 module occurs as a result of multiple structural disturbances finally affecting the topology of the catalytic center.     

Designing smarter touch-based interfaces for educational contexts

In next-generation classrooms and educational environments, interactive technologies such as surface computing, natural gesture interfaces, and mobile devices will enable new means of motivating and engaging students in active learning. Our foundational studies provide a corpus of over 10,000 touch interactions and nearly 7,000 gestures collected from nearly 70 adults and children aged 7 years and above, which can help us understand the characteristics of children’s interactions in these modalities and how they differ from adults. Based on these data, we identify key design and implementation challenges of supporting children’s touch and gesture interactions, and we suggest ways to address them. For example, we find children have more trouble successfully acquiring onscreen targets and having their gestures recognized than do adults, especially the youngest age group (7–10 years old). The contributions of this work provide a foundation that will enable touch-based interactive educational apps that increase student success.     

Structural study of titanium oxide films synthesized by ion beam-assisted deposition

The application of titanium dioxide (TiO(2)) films as surgical implant coatings for antibiotic attachment depends crucially on their available surface area and thus their surface morphology and crystallinity. Here, we report our fabrication of high Wenzel ratio TiO(2) films targeted to increase the film surface area using the ion beam-assisted deposition (IBAD) technique at high-deposition temperatures (approximately 610 degrees C). The modulation of the films' surface morphology was accomplished by varying the chemical identity of the concurrent ion beams bombarded on the films during the e-beam evaporation process. Atomic force microscopy (AFM) and scanning electron microscopy (SEM) were employed to investigate the surface morphology of the as-deposited films. X-ray diffractometry (XRD) revealed that these nanocrystalline films primarily consist of anatase phase TiO(2). Wenzel ratio, the ratio of the actual surface area to the projected area, of IBAD films prepared with argon, oxygen, and nitrogen ion beams was measured to be 1.52, 1.31 and 1.49, respectively. The effect of the differences in chemical reactivity and ion size of these three type ion beams are discussed to explain the present results.     

Evolution of the cranial computed tomography scan in child abuse

Computed tomography (CT) scans obtained at the time of clinical presentation have occasionally been reported to be normal in children with history and findings of significant abusive head injury. We have retrospectively observed abnormalities in "normal" scans of some similar children. We have also seen abnormalities develop on serial scanning. To determine how frequently these situations occur, we reviewed charts of 34 children with a final diagnosis of child abuse who also had cranial CT scans performed. Their CT scans were retrospectively reviewed by a pediatric radiologist. Eleven (11/34) CT scans had initially been interpreted as normal. Four (4/11) of these had been reinterpreted during the hospitalization as abnormal, affecting medical (1) and legal (3) outcome. Repeat scanning in three of the remaining seven resulted in surgical drainage of a subdural effusion (1) and affected legal outcome (2). Four of the seven initial scans felt normal throughout the hospitalizations were judged abnormal on retrospective review. This evaluation was confirmed in the two rescanned. Initial CT interpretation most often failed to appreciate changes in parenchymal density and small amounts of falcine or cortical subdural blood. Subsequent scans also showed evolving effusions and infarcts. Changes were noted in 1 1/2 to 5 days. The CT scan frequently shows subtle changes in the immediate posttrauma period. If the child does not recover promptly, subsequent scans frequently result in significant changes in clinical and legal management.     

Human immunodeficiency virus infection, Part II

The acceptance of highly active antiretroviral therapy (HAART) among patients and health care providers has had a dramatic impact on the epidemiology and clinical characteristics of many opportunistic infections associated with human immunodeficiency virus (HIV). Previously intractable opportunistic infections and syndromes are now far less common. In addition, effective antibiotic prophylactic therapies have had a profound impact on the risk of patients developing particular infections and on the incidence of these infections overall. Most notable among these are Pneumocystis carinii, disseminated Mycobacterium avium complex, tuberculosis, and toxoplasmosis. Nevertheless, infections continue to cause significant morbidity and mortality among patients who are infected with HIV. The role of HAART in many clinical situations is unquestioned. Compelling data from clinical trials support the use of these therapies during pregnancy to prevent perinatal transmission of HIV. HAART is also recommended for health care workers who have had a "significant" exposure to the blood of an HIV-infected patient. Both of these situations are discussed in detail in this article. In addition, although more controversial, increasing evidence supports the use of HAART during the acute HIV seroconversion syndrome. An "immune reconstitution syndrome" has been newly described for patients in the early phases of treatment with HAART who develop tuberculosis, M avium complex, and cytomegalovirus disease. Accumulating data support the use of hydroxyurea, an agent with a long history in the field of myeloproliferative disorders, for the treatment of HIV. Newer agents, particularly abacavir and adefovir dipivoxil, are available through expanded access protocols, and their roles are being defined and clarified.     

NIDA technical review: nitrite inhalants

Nitrite inhalants are commonly abused substances in the US and Europe. "Nitrite inhalants and AIDS" was a popular topic in the early 1980s when the cause of AIDS was not known. With the discovery of HIV, concern about nitrite use wained. However, nitrite inhalant use is associated with behavioral relapse and HIV transmission among gay men, with decreased lymphocyte counts and natural killer cell activity in laboratory studies, and remains a candidate "cofactor" in the pathogenesis of AIDS-related Karposi sarcoma. Discouraging nitrite use continues to be a worthwhile public health goal. Participants recommend specific research efforts.     

Reduction of disulfide bonds in an amyloidogenic Bence Jones protein leads to formation of "amyloid-like" fibrils in vitro

Motivated by the finding that the amino acid sequence of the Bence Jones protein BJP-DIA was identical to that of the main protein component of the amyloid fibrils obtained from the same patient with AL-amyloidosis, (Klafki, H.-W., Kratzin, H.-D., Pick, A.-I., Eckart, K., Karas, M. & Hilschmann, N. (1992) Biochemistry 31, 3265-3272.), we attempted to create "amyloid-like" fibrils from the Bence Jones protein in vitro, without addition of proteolytic enzymes. Reduction of BJP-DIA, solubilized in PBS, pH 7.4, overnight at 37 degrees C resulted in the formation of a precipitate which had affinity for the dye Congo red. Electron microscopy of negatively stained samples of the reduced protein revealed aggregates of linear unbranched fibrils. SDS-polyacrylamide gel electrophoresis demonstrated that the precipitate consisted almost exclusively of intact light chain molecules. This result makes it possible to deduce a molecular model of these amyloid fibrils generated in vitro.     

Definition of inseparably fused ventricular myocardium in thoracopagus: fetal echocardiographic utility and pathologic refinement

Correlative echocardiographic and pathological findings in a thoracopagus with conjoined hearts are reported. One twin had tricuspid atresia with discordant atrioventricular connections and concordant ventriculoarterial connections. The morphologic right ventricle was hypoplastic and there was a large muscular ventricular septal defect. The other twin had hypoplasia of the mitral valve anulus and left ventricle with double-outlet right ventricle and pulmonary valve atresia. The tricuspid valve was severely insufficient in part because of a large orifice and redundant, elongated leaflets with abnormal chordal attachments. The left ventricles of these two twins shared a perforated common "free wall" with at least two large defects allowing mixing of the circulations at that level. Not all anatomic details were established conclusively by fetal echocardiography; however, sufficient diagnostic information was obtained to support a decision not to aggressively resuscitate these twins after elective cesarean delivery at 31 weeks' gestation.     

MVA-vaccine in primovaccination against smallpox after the age of three (author's transl)

These are the results of 417 primary vaccinations against smallpox with MVA, an attenuated vaccine. All of the patients were over three years old. In Europe, this group has an increased risk of postvaccinal encephalitis. The intracutaneous inoculation produced a light local infiltration and only 11.7% showed a light generalized reaction. In order to attain a reliable immunity, a second inoculation was performed one week later, epicutaneously with Elstree-vaccine. The local reaction was in general that of a normal revaccination, and 79.4% of our patients had neither fever nor general symptoms. Among the others no one became seriously ill. We did not observe any complications specific for a smallpox vaccination. In comparison to the other vaccination methods currently used, the MVA "Two-step"-vaccination is probably the safest. Its risk of a postvaccinal encephalitis is supposed to be lower according to animal experiments and theoretical considerations. This hypothesis has been confirmed meanwhile by the results of several thousand vaccinations in the already mentioned age-group.