Induction of IgG antibodies against GD3 ganglioside in rabbits by an anti-idiotypic monoclonal antibody.

Anti-idiotypic MAb were raised in syngeneic mice against a mouse MAb recognizing GD3 ganglioside (MAb R24). Two anti-idiotypic MAb, designated BEC2 and BEC3, recognized distinct determinants on MAb R24 that mapped near or within the GD3-binding site. New Zealand white rabbits, which express GD3 on normal tissues, were immunized with either BEC2, BEC3, or control MAb FLOPC-21. All rabbits developed high and equivalent titers of antibodies against mouse immunoglobulins. Immunization with BEC2 and BEC3 induced rabbit antibodies expressing R24 idiotype as demonstrated by their ability to inhibit BEC2 binding to R24. Antibodies (IgG and IgM) reacting with GD3 developed in five of eight rabbits immunized with BEC2 but not in rabbits immunized with BEC3 or with control MAb. Serum antibodies against GD3 did not cross-react with other gangliosides. These results show that MAb BEC2 can mimic GD3 ganglioside and can induce antibodies against GD3 ganglioside despite expression of GD3 on normal rabbit tissue.     

Metabolic clearance and secretion rates of subunits of human thyrotropin.

Metabolic clearance rates (MCR) of the alpha and beta subunits of human thyrotropin (hTSH-alpha and hTSH-beta) were determined by a constant infusion to equilibrium method. In 15 normal individuals (six men, six premenopausal women, and three post-menopausal women), the mean MCR of hTSH-alpha (68 ml/min per m2) was significantly faster than that of hTSH-beta (48 ml/min per m2) was significantly faster than that of hTSH-beta (48 ml/min per m2); both were two to three times more rapid than the previously determined MCR of hTSH. In patients with primary hypothyroidism, MCR were significantly slower with a mean value of 55 ml/min per m2 for hTSH-alpha and 37 ml/min per m2 for hTSH-beta. However, MCR of subunits were not significantly faster than normal in hyperthyroid patients. Serum concentrations of alpha subunits and hTSH-beta were measured by radioimmunoassay, and secretion rates of alpha and hTSH-beta from the pituitary were calculated using hTSH-alpha and hTSH-beta MCR, respectively. In the normal individuals, alpha secretion rates averaged 91 mug/day per m2, greater than those previously determined for hTSH and human follicle-stimulating hormone. Alpha secretion rates were significantly elevated in the normal postmenopausal women (211 mug/day per m2) and in the premenopausal hypothyroid women (202 mug/day per m2); they were also elevated in the postmenopausal hypothyroid women (277 mug/day per m2). Alpha secretion rates were significantly decreased in the premenopausal hyperthyroid women (66 mug/day per m2). Usually, the secretion rates of hTSH-beta could not be calculated in normal individuals, and the rates in hyperthyroid patients could never be calculated because serum hTSH-beta was not detected. Six normals had detectable hTSH-beta secretion rates (17 mug/day per m2); hTSH-beta secretion rates were significantly increased in patients with primary hypothyroidism (28 mug/day per m2). Although we had previously demonstrated a 50-fold increase in hTSH secretion rates in primary hypothyroidism, there was only a 2-fold increase in alpha and hTSH-beta secretion rates. Thus, increased subunit synthesis appears to be utilized predominantly for production of complete hTSH.     

Nonlinear (amplified) relationship between nuclear occupancy by triiodothyronine and the appearance rate of hepatic alpha-glycerophosphate dehydrogenase and malic enzyme in the rat.

Three separate approaches were applied to examine the general relationship between R, the rate of induction of specific enzymes (mitochondrial alpha-glycero-phosphate dehydrogenase and cytosolic malic enzyme) and q, the fractional nuclear occupancy by triiodothyronine in male Sprague-Dawley rats. Daily 200-microgram injections of triiodothyronine per 10u g body wt for 7 days resulted in saturation of the hepatic nuclear sites and the achievement of an apparent new steady state of enzyme levels. The increase achieved over base-line hypothyroid levels was then compared with the increment over hypothyroid base line characteristic of intact euthyroid animals with 47% of nuclear sites occupied. The maximal theoretical reate of steady-state enzyme induction could be protected on the basis of the observed maximal increase in enzyme activity observed 1 day after the injection of graded doses of hormone and lambda, the known fractional rate of enzyme dissipation. The 24-h dose-response studies were used to generate R as a continuous function of q, both in hypothyroid as well as in euthyroid animals. This approach involved the numerical solution of an ordinary differential equation describing the rate of change of enzyme as a function of R, which was assumed to be uniquely related to q. Results of these analyses indicated that the ratio of the maximal rate of induction of enzyme at full occupancy to the rate of induction under euthyroid conditions assumes a value between 9.0 and 19.5, depending on the precise analytic and experimental approach applied. This value is far in excess of the theoretical ratio 2.13 which on would anticipate if R were linearly related to q and 47% of the nuclear sites occupied under physiological conditions. Thus, the signal for enzyme induction appears to undergo progressjive amplification with increasing nuclear occupancy. Moreover, the curve describing the relationship between R and q appears highly nonlinear throughout (concave upwards). Although the molecular mechanism responsible for amplification is unknown, recognition of this phenomenon may be helpful in understanding tissue effects of thyroid hormone excess. Moreover, the analytic technique for determining R as a function of q may be of general applicability in studying hormonal response systems under nonsteady-state conditions.     

Evaluation of dot-ELISA for serological diagnosis of amebiasis

 We improved the dot enzyme-linked immunosorbent assay (dot-ELISA) reported by Itoh and Sato, and assessed the usefulness of this test for the diagnosis of amebiasis. The sensitivity of dot-ELISA was compared with that of plate ELISA, the indirect hemagglutination test (IHA), and the indirect fluorescent antibody test (IFA) for the diagnosis of amebiasis. Of 37 serum samples from patients with documented amebiasis, 36 (97.3%) were positive by dot-ELISA. There was consistency among the results of dot-ELISA, plate ELISA, and IFA, although the positive rate of IHA was lower than that of the others (78.4%; 29 of 37 cases were positive). The specificities of dot-ELISA and plate ELISA were assessed using a total of 68 sera, collected from 38 patients infected with seven different parasites other than Entamoeba histolytica, 10 patients showing diarrhea or liver abscess without parasitic infection, and 20 healthy individuals. The two assays showed no false-positive results. There were no differences in sensitivity and specificity between dot-ELISA and plate ELISA. However, the dot-ELISA technique seems to be more feasible for clinical application than plate ELISA techniques, because the assay does not require any specific equipment. Received: July 8, 2002 / Accepted: December 7, 2002 RID="*" ID="*" A summary of this paper was presented at the 74th General Meeting of the Japanese Association for Infectious Diseases (Fukuoka, April 2000). Acknowledgments The authors are indebted to Professor Tsutomu Takeuchi and Dr. Seiki Kobayashi, Department of Tropical Medicine and Parasitology, Keio University School of Medicine, for supplying E. histolytica antigen, and to Dr. Hiroshi Yamasaki, Department of Parasitology, Juntendo University School of Medicine, for supplying recombinant Toxocara canis antigen for this study.     

Identification and quantitation of platelet-associated fibronectin antigen.

Platelet-associated fibronectin antigen has been identified by radioimmunoassay and immunofluorescent techniques. In radioimmunoassay, platelet fibronectin was immunochemically indistinguishable from plasma fibronectin. Platelet and plasma fibronectin were bound and eluted from gelatin-sepharose under similar conditions. The level of platelet fibronectin in detergent extracts of washed platelets from 12 healthy adults was 2.85 +/- 1.24 microgram/10(9) platelets. Immunofluorescence with F(ab')2 fragments of immunochemically purified antifibronectin showed that all platelets stained with a discrete punctate pattern. The identification of platelet fibronectin antigen raises the possibility that this protein may participate in platelet-platelet or platelet-surface interactions.     

Lebenslange Nachbehandlung und Sekund?rprophylaxe nach Herzinfarkt

Ein 60-jähriger Patient mit langjährigem Hypertonus, diätetisch schlecht eingestelltem Diabetes mellitus mit HbA_1c-Werten von 10%, Adipositas (Körpergröße 165 cm, Gewicht 85 kg) sowie Hyperlipoproteinämie kommt nach einem 2-wöchigen stationären Krankenhausaufenthalt wegen eines erlittenen Herzinfarkts in die ärztliche Praxis. Während des stationären Aufenthalts war im Rahmen des akuten Vorderwandinfarkts eine notfallmäßige Herzkatheteruntersuchung durchgeführt worden, die eine koronare Eingefäßerkrankung mit Verschluss der LAD erbracht hatte. Das Gefäß war mittels perkutaner transluminaler Koronarangioplastie (PTCA) rekanalisiert und mit einem Stent versorgt worden. Der Patient wurde aus dem Krankenhaus mit einer Medikation, bestehend aus Acetylsalicylsäure (ASS) 100 mg, Clopidogrel 75 mg, einem Angiotensinkonversionsenzym-(ACE-)Hemmer, einem -Blocker, einem Statin und einem Diuretikum, entlassen, nachdem er im Rahmen des Myokardinfarkts eine Linksherzinsuffizienz mit Lungenstauung sowie Beinödeme entwickelt hatte. Zurzeit ist der Patient klinisch beschwerdefrei und berichtet, dass er vor dem Akutereignis keine starke Angina pectoris verspürt habe. Beim Treppensteigen seien ihm aber schon eine gewisse Kurzatmigkeit und ein leichtes Druckgefühl in der Brust aufgefallen, denen er jedoch keine wesentliche Bedeutung beigemessen habe.Beruflich ist der Patient vornehmlich als freier Außendienstmitarbeiter mit dem Auto unterwegs oder am heimischen Schreibtisch tätig. Sportliche Aktivitäten werden verneint, dafür werden ca. zehn Zigaretten am Tag geraucht.     

Accumulation of methotrexate polyglutamates in lymphoblasts is a determinant of antileukemic effects in vivo. A rationale for high-dose methotrexate.

Methotrexate (MTX) is one of the most widely used drugs for the treatment of childhood acute lymphoblastic leukemia (ALL) and is commonly given in high doses. However, the rationale for high-dose MTX (HDMTX) has been challenged recently. To determine whether higher MTX polyglutamate (MTXPG) concentrations in ALL blasts translate into greater antileukemic effects, 150 children with newly diagnosed ALL were randomized to initial treatment with either HDMTX (1,000 mg/m2 intravenously over 24 h) or lower-dose MTX (30 mg/m2 by mouth every 6 h x 6). ALL blasts accumulated higher concentrations of MTXPG and long-chain MTXPG (MTXPGLC) after HDMTX (P < 0.00001). Of 101 patients evaluable for peripheral blast cytoreduction, MTXPG concentrations were higher in patients whose blast count decreased within 24 h (P = 0.005) and in those who had no detectable circulating blasts within 4 days (P = 0.004). The extent of inhibition of de novo purine synthesis in ALL blasts was significantly related to the blast concentration of MTXPGLC (IC95% = 483 pmol/10(9) blasts). The percentage of patients with 44-h MTXPGLC exceeding the IC95% was greater after HDMTX (81%) than LDMTX (46%, P < 0.0001). These data indicate that higher blast concentrations of MTXPG are associated with greater antileukemic effects, establishing a strong rationale for HD-MTX in the treatment of childhood ALL.     

A knowledge based interpretation system for EMG abnormalities

The conventional method of diagnosis in electromyography is complex and time consuming, not only due to the large number of parameters, to be considered for diagnosis, but also because of the usual procedure of evaluating the different parameters of EMG signal by visual scanning of the plotted signal. So there is a clear need to make use of computer aided decision support system. In the present work an attempt has been made in the direction of integration into one automated system, the qualitative knowledge of the physician, with possibly sophisticated signal analysis tools which must replace the visual scanning. A software program (in Turbo-C) on a PC-AT has been developed to evaluate the different parameters of MUAP's (motor unit action potential) in a EMG signal. Then an Expert system (in Turbo-Prolog) has been implemented for diagnostic purposes of different muscular abnormalities by making a knowledge base from the different parameters involved in the decision making procedure of clinical electromyography. A hybrid model of rule and frame based Expert system is implemented. An attempt has been made for making a complete system, i.e., for recording, analysis and decision making for diagnosis.     

中度低温体外循环对大鼠海马c-fos,bcl-2及bax mRNA表达的影响

目的:观察中度低温体外循环(CPB)对大鼠海马即刻基因c-fos、凋亡相关基因bcl-2和baxmRNA表达的影响。方法:雄性SD大鼠随机分为CPB组(n=6)及假手术对照组(Sham组,n=6)。所有动物在咪唑安定、芬太尼麻醉后经口插管控制呼吸,置入颈静脉流出管和尾动脉输入管,肝素抗凝(500U/kg)。CPB组采用中度低温CPB(26℃—28℃),经尾动脉灌注、颈静脉右心房-腔静脉引流,灌流量160mL·kg-1·min-1,总转流时间2h,Sham组除不经历CPB外,其余操作同CPB组相同。实验中进行动脉压、ECG及动脉血气监测。术后1h处死动物,立即断头取脑分离海马组织,放入-70℃液氮罐保存。基因mRNA检测采用逆转录多聚酶联反应(RT-PCR)方法。结果:海马c-fos、bcl-2和baxmRNA表达CPB组显著高于Sham组;bax与bcl-2mRNA表达比值CPB组显著高于Sham组。结论:中度低温体外循环可引起海马即刻基因c-fos、凋亡相关基因bcl-2和baxmRNA的表达增加。     

Influence of Olfactory Epithelium on Mitral/Tufted Cell Dendritic Outgrowth

Stereotypical connections between olfactory sensory neuron axons and mitral cell dendrites in the olfactory bulb establish the first synaptic relay for olfactory perception. While mechanisms of olfactory sensory axon targeting are reported, molecular regulation of mitral cell dendritic growth and refinement are unclear. During embryonic development, mitral cell dendritic distribution overlaps with olfactory sensory axon terminals in the olfactory bulb. In this study, we investigate whether olfactory sensory neurons in the olfactory epithelium influence mitral cell dendritic outgrowth in vitro. We report a soluble trophic activity in the olfactory epithelium conditioned medium which promotes mitral/tufted cell neurite outgrowth. While the trophic activity is present in both embryonic and postnatal olfactory epithelia, only embryonic but not postnatal mitral/tufted cells respond to this activity. We show that BMP2, 5 and 7 promote mitral/tufted cells neurite outgrowth. However, the BMP antagonist, Noggin, fails to neutralize the olfactory epithelium derived neurite growth promoting activity. We provide evidence that olfactory epithelium derived activity is a protein factor with molecular weight between 50–100 kD. We also observed that Follistatin can effectively neutralize the olfactory epithelium derived activity, suggesting that TGF-beta family proteins are involved to promote mitral/tufted dendritic elaboration.