[Preparation and characteristics of the human single-chain Fv antibodies to NS5B].

AIM: To prepare and identify the human single-chain Fv (scFv) antibody to hepatitis C virus(HCV) NS5B. METHODS: After the peripheral blood mononuclear cells were collected from HCV patients, the single-chain antibody Fv library was constructed by amplifying the whole variable region of heavy chain(V(H);) and variable region of light chain(V(L);). Targeting NS5B, the scFv gene was cloned into pET16b, and then transfected into BL21 cells. The scFv was induced to express in the cells. Sequencing and analysis of the scFv were performed. RESULTS: The single-chain antibody Fv library was successfully constructed; two scFvs with higher binding activity were expressed; the sequencing result showed that the scFv was consistent with the expectation. CONCLUSION: Preparation of the human scFv antibody to HCV NS5B provides a basis for immunotherapy to HCV.

Mechanisms underlying rhythmic locomotion: dynamics of muscle activation.

We have studied the dynamical properties of tension development in leech longitudinal muscle during swimming. A new method is proposed for modeling muscle properties under functionally relevant conditions where the muscle is subjected to both periodic activation and rhythmic length changes. The 'dual-sinusoid' experiments were conducted on preparations of leech nerve cord and body wall. The longitudinal muscle was activated periodically by injection of sinusoidal currents into an identified motoneuron. Simultaneously, sinusoidal length changes were imposed on the body wall with prescribed phase differences (12 values equally spaced over 2π radians) with respect to the current injection. Through the singular value decomposition of appropriately constructed tension data matrices, the leech muscle was found to have a multiplicative structure in which the tension was expressed as the product of activation and length factors. The time courses of activation and length factors were determined from the tension data and were used to develop component models. The proposed modeling method is a general one and is applicable to contractile elements for which the effects of series elasticity are negligible.

Analysis of impulse adaptation in motoneurons.

Animal locomotion results from muscle contraction and relaxation cycles that are generated within the central nervous system and then are relayed to the periphery by motoneurons. Thus, motoneuron function is an essential element for understanding control of animal locomotion. This paper presents motoneuron input-output relationships, including impulse adaptation, in the medicinal leech. We found that although frequency-current graphs generated by passing 1-s current pulses in neuron somata were non-linear, peak and steady-state graphs of frequency against membrane potential were linear, with slopes of 5.2 and 2.9 Hz/mV, respectively. Systems analysis of impulse frequency adaptation revealed a static threshold nonlinearity at -43 mV (impulse threshold) and a single time constant (tau = 88 ms). This simple model accurately predicts motoneuron impulse frequency when tested by intracellular injection of sinusoidal current. We investigated electrical coupling within motoneurons by modeling these as three-compartment structures. This model, combined with the membrane potential-impulse frequency relationship, accurately predicted motoneuron impulse frequency from intracellular records of soma potentials obtained during fictive swimming. A corollary result was that the product of soma-to-neurite and neurite-to-soma coupling coefficients in leech motoneurons is large, 0.85, implying that the soma and neurite are electrically compact.

Muscle function in animal movement: passive mechanical properties of leech muscle.

We investigated passive properties of leech body wall as part of a larger project to understand better mechanisms that control locomotion and to establish mathematical models that predict such dynamical behavior. In tests of length-tension relationships in 2-segment-long preparations of body wall through step-stretch manipulations (step size = 1 mm), we discovered that these relationships are nonlinear, with significant hysteresis, even for the relatively small changes in length that occur during swimming. We developed a mathematical model comprising three nonlinear springs, two in series with nonlinear dashpots that describe well the tension statics and dynamics for step-stretch experiments. This model suggested that body wall dynamics are slow enough to be neglected when predicting the tension generated by imposed sinusoidal length changes (about +/-10% of nominal) at 1-3 Hz, mimicking swimming. We derived a static model, comprising one nonlinear spring, which predicts sinusoidal data accurately, even when preparations were exposed to serotonin (0.1-10 microM). Preparations bathed in saline-serotonin had significantly reduced steady-state and peak tensions, without alterations in tension dynamics. Anesthetizing preparations (8% ethanol) reduced body wall tension by 77%, indicating that passive tension in the obliquely striated longitudinal muscles of leeches results primarily from a resting tonus.