Semaphorin3A signalling is mediated via sequential Cdk5 and GSK3beta phosphorylation of CRMP2: implication of common phosphorylating mechanism underlying axon guidance and Alzheimer's disease

Collapsin response mediating protein-2 (CRMP2) has been identified as an intracellular protein mediating Semaphorin3A (Sema3A), a repulsive guidance molecule. In this study, we demonstrate that cyclin-dependent kinase 5 (Cdk5) and glycogen synthase kinase 3beta (GSK3beta) plays a critical role in Sema3A signalling. In In vitro kinase assay, Cdk5 phosphorylated CRMP2 at Ser522, while GSK3beta did not induce any phosphorylation of CRMP2. Phosphorylation by GSK3beta was exclusively observed in Cdk5-phosphorylated CRMP2, but barely in CRMP2T509A. These results indicate that Cdk5 primarily phosphorylates CRMP2 at Ser522 and GSK3beta secondarily phosphorylates at Thr509. The dual-phosphorylated CRMP2, but not non-phosphorylated or single-phosphorylated CRMP2, is recognized with the antibody 3F4, which is highly reactive with the neurofibrillary tangles of Alzheimer's disease. 3F4 recognized the CRMP2 in the wild-type but not cdk5-/- mouse embryonic brain lysates. The phosphorylation of CRMP2 at Ser522 caused reduction of its affinity to tubulin. In dorsal root ganglion neurones, Sema3A stimulation enhanced the levels of the phosphorylated form of CRMP2 detected by 3F4. Over-expression of CRMP2 mutant substituting either Ser522 or Thr509 to Ala attenuates Sema3A-induced growth cone collapse response. These results suggest that the sequential phosphorylation of CRMP is an important process of Sema3A signalling and the same mechanism may have some relevance to the pathological aggregation of the microtubule-associated proteins.     

Antiphospholipid antibodies

The concept of antiphospholipid syndrome(APS) has been widely accepted. Antiphospholipid antibodies originally included anticardiolipin antibodies and lupus anticoagulants as serological marker of APS. However, recent advances have shown that most pathogenic antiphospholipid antibodies are directed to phospholipid binding proteins such as beta 2-glycoprotein I and prothrombin as well as phospholipids. The preliminary classification criteria for definite APS have been advocated as the "Sapporo criteria". Further prospective investigations are required to re-evaluate the clinical significance of so-called antiphospholipid antibodies.     

Blepharophimosis sequence and de novo balanced autosomal translocation [46, XY,t(3;4)(q23;p15.2)]: Possible assignment of the trait to 3q23

We report on a boy with the blepharophimosis sequence and de novo, apparently balanced reciprocal translocation between 3q23 and 4p15.2 [46, XY,t(3;4)(q23;p15.2)de novo]. Possible assignment of this autosomal dominant disorder is discussed. A 3q23 band is a more preferable gene locus of the blepharophi mosis sequence, based on the comparison of clinical manifestations between 4p- and 3q-syndromes.     

Concurrent visual task effects on evoked and emitted auditory p300 in adolescents

Using an oddball stimulus presentation paradigm, the effects of divided attention on auditory P300s were studied. Auditory attention was either divided or focused, depending on the demands placed on subjects during the performance of a concomitantly presented visual task. Two types of auditory tasks were performed under each of the two auditory attention conditions. In one, subjects responded to infrequently presented high pitched tones (oddball stimuli). In the other they responded to the occasional omission of a stimulus in an otherwise rhythmically presented chain of stimuli. P300s and reaction times were recorded to both the rare tones and the omissions. The Sternberg visual memory task was used to manipulate the subject's auditory attention state. Subjects actively performed the Sternberg task during the divided auditory attention condition, whereas during the focused attention condition they were not required to respond to the visual stimuli. During focused auditory attention, evoked auditory P300s were both larger and faster than their emitted counterparts. During divided attention, auditory P300s were reduced in amplitude but latency was unaffected. Evoked auditory P300s showed evidence of containing P300a as well as P300b components, particularly when attention was shared with the visual task.     

Evaluation of time-resolved fluorometric immunoassay of CA 50

Serum CA 50 was determined by a time resolved fluorometric immunoassay (TR-FIA) with CANAG CA-50 DELFIA kit. Evaluation of the assay system gave satisfactory results in its sensitivity, accuracy, reproducibility, dynamic range and easy handling. No prozone phenomenon was observed up to 347,000 U/ml. From a histogram of 134 normal sera, the cut off point was determined at 34 U/ml. CA 50 in 202 patients' sera was determined with this assay. Nineteen of 20 patients pancreatic cancer, 6 of 21 gastric cancer, 14 of 25 hepatoma gave positive values. In comparison with CA 19-9, higher values and higher rates of positive CA 50 were observed in benign and malignant liver diseases, suggesting its non-cancerous origin in the liver. A high correlation was observed between the level of CA 50 and CA 19-9 of 157 patients' sera. Serum CA 50 was completely correlated with CA 19-9 in the clinical course of patients with pancreatic cancer, but not in patients with hepatoma. Thus we conclude that the CANAG CA-50 DELFIA System is useful for the diagnosis and monitoring cancer patients but must be used with care because of its elevation in benign liver diseases.     

Tumor necrosis factor-alpha in the placenta is not elevated in pre-eclamptic patients despite its elevation in peripheral blood

PROBLEM: Tumor necrosis factor-alpha (TNF-alpha) is present in human placental and uterine cells at the early and late stages of gestation and promotes the regulation of trophoblast growth and invasion. We evaluated whether TNF-alpha levels in the placenta and blood of pre-eclamptic women differed from those with normal pregnancies. METHOD OF STUDY: The subjects were 39 pregnant women carrying single fetuses (21 normal-pregnant and 18 pre-eclamptic patients). Their average gestational age at entry was 38-39 weeks. Peripheral blood was collected before the onset of labor and separated serum was stored at -20 degrees C. A tissue segment of the placenta was cut and frozen in liquid nitrogen immediately after delivery at -80 degrees C. The frozen placental tissue was added to phosphate-buffered saline. The tissue was fully homogenized and centrifuged. Separated supernatant was stored at -80 degrees C. TNF-alpha levels in separated serum and TNF-alpha and total protein (TP) levels in separated supernatant were measured. The presence of TNF-alpha in the placenta was evaluated by immunohistochemistry in five pre-eclamptic and five normal-pregnant patients. RESULTS: Serum TNF-alpha levels were higher in pre-eclampsia than in normal pregnancies. However, TNF-alpha/TP levels in the placenta did not differ significantly between the two groups. As for TNF-alpha immunostaining of trophoblastic cells in the placenta, it was weak in three and moderate in two of the normal pregnancies, while it was absent in two, weak in one, and moderate in two in the pre-eclampsia group. CONCLUSIONS: We demonstrated no significant increase in TNF-alpha/TP levels in the placenta in pre-eclampsia despite a significant increase in serum TNF-alpha levels. There was no strong immunostaining for TNF-alpha detected by immunohistochemistry in the pre-eclampsia group. These findings suggest that TNF-alpha in the placenta is not a key cytokine to interfere with normal trophoblast invasion into the myometrium in pre-eclampsia, and that sources other than the placenta may contribute to the elevated levels of TNF-alpha found in the circulation of pre-eclamptic patients.     

Synthesis and characterization of poly(1-benzoylthiosemicarbazide) and poly(1,3,4-thiadiazole amine) from 2-(p-aminophenyl)-1,3,4-oxadiazoline-5-thione via ring-opening process

A novel thermally stable and semiconducting polyheterocycle, poly(1,3,4-thiadiazole amine), was synthesized from 2-(p-aminophenyl)-1,3,4-oxadiazoline-5-thione via ring-opening. The polymer is a new class of ordered alternating copoly(aniline) containing 1,3,4-thiadiazole heterocyclic units. An investigation of the reaction of 2-phenyl-1,3,4-oxadiazoline-5-thione with aniline was conducted as a model reaction for the polymerization, and poly(phosphoric acid) (PPA) and phosphorus pentoxide/methanesulfonic acid (PPMA) were found to be favorable both as condensing agent and solvent for the formation of 2-anilino-5-phenyl-1,3,4-thiadiazole as a model compound. The polymerization was carried out both by two-step procedure that included ring-opening self-polyaddition giving poly(1-benzoylthiosemicarbazide), followed by cyclodehydration to poly(1,3,4-thiadiazole amine), and by a one-step procedure including cyclodehydration in situ. The poly(1-benzoylthiosemicarbazide) which was formed in the first step in m-cresol had reduced viscosities up to 0,42 dL·g^?1, and it was converted to poly(1,3,4-thiadiazole amine) by treating in PPA or PPMA. Poly(1,3,4-thiadiazole amine) having reduced viscosities up to 0,25 dL·g^?1 was also synthesized by the direct one-step polymerization in PPA or PPMA. The polymer is highly thermally stable and exhibited no weight loss up to 350°C under nitrogen. Its electric conductivity was less than 10^?10 S·cm^?1 at ambient temperature, but markedly increased to 2,9·10^?7 S·cm^?1 upon doping with iodine.     

DNA analysis of two patients with a non-fluorescent y chromosome

Summary Results of DNA study on two patients of gonadal dysgenesis with a 45,X/46,X,Ynf (non-fluorescent Y chromosome) karyotype are described. In one patient who developed gonadoblastoma, all 12 loci on the non-fluorescent part of Yq were detected. Another patient did not have gonadoblastoma at 20 years, and only the proximal 6 loci out of 12 were detected.     

Specific tension of elbow flexor and extensor muscles based on magnetic resonance imaging

Series cross-section images of the upper extremity were obtained for four men by magnetic resonance imaging (MRI) and anatomical cross-sectional areas (ACSA) of elbow flexor muscles [biceps brachii (BIC), brachialis (BRA), brachioradialis (BRD)] and extensor muscles [triceps brachii (TRI)] were measured. Physiological cross-sectional area (PCSA) was calculated from the muscle volume and muscle fibre length, the former from the series ACSA and the latter from the muscle length multiplied by previously reported fibre/muscle length ratios. Elbow flexion/extension torque was measured using an isokinetic dynamometer and the force at the tendons was calculated from the torque and moment arms of muscles measured by MRI. Maximal ACSA of TRI was comparable to that of total flexors, while PCSA of TRI was greater by 1.9 times. Within flexors, BRA had the greatest contribution to torque (47%), followed by BIC (34%) and BRD (19%). Specific tension related to the estimated velocity of muscle fibres were similar for elbow flexors and extensors, suggesting that the capacity of tension development is analogous between two muscle groups.     

Orientational behavior of side-chain liquid-crystalline polysiloxanes deduced from measurements of second harmonic generation

Side-chain functionalized polysiloxanes were prepared via polymer-analogous esterification of poly[(3-chloroformylpropyl)methylsiloxane] with 4-(4-hydroxyphenylazo)nitrobenzene ( P1 ), 4-[4-(ω-hydroxyalkyloxy)phenylazo]nitrobenzene ( P2 – P4 ), 4-{4-[N-(2-hydroxyethyl)-N-methyl]anilinoazo}nitrobenzene ( P5 ), 4-(4-hydroxypiperidino)nitrobenzene ( P6 ), or 4-[4-(2-hydroxyethyl)piperidino]nitrobenzene ( P7 )., P1 , P3 , P4 and P5 exhibit liquid crystallinity, as deduced from differential scanning calorimetry, polarized microscopic observations and X-ray diffraction measurements. The liquid-crystalline phase of P1 and P5 is a nematic phase, and that of P3 and P4 is a smectic one. The second harmonic generation (SHG) measurement of a spin-coated film of P1 was carried out by the Maker fringe method using a Q-switched Nd:YAG laser (1064 nm). The SHG profile after the heat treatment of a spin-coated film suggests a perpendicular orientation of the mesogenic molecules to the glass substrate. The SH light intensity of a corona-poled film was 20-fold higher than that of a film which was only heated, though no differences were observed in their UV-vis absorption spectra. These findings suggest that the mesogenic-molecular dipole moments are aligned to the same direction in the crystalline or liquid-crystalline phase by a poling treatment.