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V Lombardi

Publications and source records attributed to V Lombardi.

At least 37 records · Page 2Linked to original sources
PubMed1998

On the working stroke elicited by steps in length and temperature.

Muscle contraction is generally thought to involve tilting of the light chain region of the myosin head. This could account for 5-10 nm of axial displacement as it moves from nearly perpendicular to the filament axis (the state at the beginning of the working stroke) to the rigor conformation (at the end of the working stroke). According to the kinetic model of Huxley and Simmons, the extent that a cross-bridge progresses through the working stroke depends on the mechanical conditions. A large tilting occurs only when the fibre is allowed to shorten. Evidence for such tilting was provided by the changes in intensity of the third myosin meridional reflection (M3) following a step release. However, there is little change in the M3 intensity when a force increase is elicited by a 10 degrees C temperature jump, and these results were interpreted to indicate that tilting is not the structural transition responsible for force generation. Here we present a simulation of the changes in the intensity of the M3 reflection elicited by step changes in either length or temperature, based on the atomic model of the actin-myosin head complex. The results show that the same set of assumptions for the motions associated with the working stroke can predict the response to both kinds of perturbation. The main difference is due to the larger extent of the working stroke elicited by the length step.

Animals↗
PubMed1998

Myosin head movements during isometric contraction studied by X-ray diffraction of single frog muscle fibres.

Time resolved X-ray diffraction experiments in single muscle fibres of the frog at 2.15 microns sarcomere length and 4 degrees C were performed at ID2 (SAXS), ESRF, Grenoble (France) to investigate the structural aspects of cross-bridge action during the development of the isometric tetanic tension (T0). Changes in the low angle myosin-based reflections were measured with 5 ms time resolution by signal averaging data collected with a 10 m camera length and a 2D gas-filled detector. Upon activation the intensity of the first order myosin layer line reflection, I(M1), and the intensity of the second order meridional reflection, I(M2), reduced practically to zero with a half-time which leads the tension rise by 15-20 ms. The complex changes of the intensity of the third order myosin meridional reflection, I(M3), and the increase of its axial spacing from 14.34 nm (at rest) to 14.57 nm (at T0) could be analysed by assuming that they were the result of the combination of the time dependent modulation in intensity of two closely spaced periodicities, one at 14.34 nm, characteristics of the myosin molecule at rest and the other at 14.57 nm, assumed by the myosin as a consequence of the activation and force production. I(14.34) drops monotonically in advance to isometric tension development with a half-time similar to that of I(M1) and I(M2), while I(14.57) rises from zero to a maximum in parallel with tension.

Animals↗
PubMed1998

The stiffness of skeletal muscle in isometric contraction and rigor: the fraction of myosin heads bound to actin.

Step changes in length (between -3 and +5 nm per half-sarcomere) were imposed on isolated muscle fibers at the plateau of an isometric tetanus (tension T0) and on the same fibers in rigor after permeabilization of the sarcolemma, to determine stiffness of the half-sarcomere in the two conditions. To identify the contribution of actin filaments to the total half-sarcomere compliance (C), measurements were made at sarcomere lengths between 2.00 and 2.15 microm, where the number of myosin cross-bridges in the region of overlap between the myosin filament and the actin filament remains constant, and only the length of the nonoverlapped region of the actin filament changes with sarcomere length. At 2.1 microm sarcomere length, C was 3.9 nm T0(-1) in active isometric contraction and 2.6 nm T0(-1) in rigor. The actin filament compliance, estimated from the slope of the relation between C and sarcomere length, was 2.3 nm microm(-1) T0(-1). Recent x-ray diffraction experiments suggest that the myosin filament compliance is 1.3 nm microm(-1) T0(-1). With these values for filament compliance, the difference in half-sarcomere compliance between isometric contraction and rigor indicates that the fraction of myosin cross-bridges attached to actin in isometric contraction is not larger than 0.43, assuming that cross-bridge elasticity is the same in isometric contraction and rigor.

Actins↗
PubMed1998

Free radicals in rabbit spinal cord ischemia: electron spin resonance spectroscopy and correlation with SOD activity.

1. In nonanesthetized rabbits temporal occlusion of the abdominal aorta was used to induce oxidative stress in the lower part of the body including distal segments of the spinal cord. 2. Spinal cord samples were taken from the animals exposed to 25-min aortic occlusion (AO) or to occlusion followed by 1- or 2-hr reperfusion (AO/R1 or AO/R2, respectively) or from sham-operated animals (C). The presence of free radicals (FR) in the spinal cord samples frozen in liquid N2 was assessed by ESR spectroscopy without spin trapping. Moreover, superoxide dismutase (SOD) activity and conjugated diene (CD) levels were measured in the samples. 3. In the AO group FR were detected in the spinal cord regions close to the occlusion (lower thoracic and distal segments) along with a decrease in SOD activity. The calculated g value (g = 2.0291) indicated that the paramagnetic signal recorded might be attributed to superoxide radicals. FR were absent in the AO/R1 group. Concurrently, the SOD activity revealed a significant tendency to return to the control level. FR appeared again in the AO/R2 group, mostly in the upper and middle lumbar regions, along with a decrease in SOD activity. No sample from the C group revealed FR. A significant increase in CD levels was observed in the thoracolumbar region only in the AO/R2 group. The temporary absence of FR in the AO/R1 group suggests activation of defense antioxidant mechanisms (e.g., specific enzymatic systems such as SOD), which might have been exhausted later. 4. Changes in SOD activity similar to those observed in the thoracolumbar region, though less noticeable, occurred in the obviously noncompromised tissue (upper cervical region). This points to a kind of generalized response of the animal to aortic occlusion. 5. Direct ESR spectroscopy revealed the presence of FR as well as their time course in the spinal cord during the early phase of ischemia/reperfusion injury and the inverse relationship between FR and SOD activity.

Animals↗
PubMed1998

[Diverticulosis of the gallbladder. A case report and review of the literature].

We report a case of diffuse diverticulosis of the gallbladder in a 32 year old patient, who presents epigastric pains and decayed general conditions. With the suspicion of a primitive tumor of the gallbladder, we make a cholecystectomy and the intraoperative histological examination shows a "diffuse diverticulosis of the gallbladder wall"; it displays micro-stones inside diverticulums, too. The patient, after the surgical treatment, has no problem and he gets back to his normal weight. The literature shows the rarity of this disease; its etiopathogenesis is connected with congenital factors. In our opinion, the many terms used (cholecystitis glandularis proliferans, hypertrophic, adenomyomatosis) to define those conditions are not appropriate for our case but rather for the gallbladder's chronic inflammation characterized by the Rokitansky-Ashoff sinuses.

Adult↗
PubMed1997

Cross-bridge detachment and attachment following a step stretch imposed on active single frog muscle fibres.

1. The time course of cross-bridge detachment-attachment following a step stretch was determined in single frog muscle fibres (at 4 degrees (1 and 2.1 microns sarcomere length) by imposing, under sarcomere length control by a striation follower, test step releases of various amplitudes (2-13 nm per half-sarcomere) at successive times (4-55 ms) after a conditioning stretch of approximately 4 nm per half-sarcomere. 2. The comparison with the control tension transients, elicited by releases not preceded by the conditioning stretch, shows that, early after the conditioning stretch, the quick tension recovery following small releases is depressed and the quick tension recovery following large releases is potentiated. Both effects are expected as a consequence of the strain produced in the cross-bridges by the conditioning stretch. 3. These effects disappear and the tension transient is reprimed, indicating substitution of freshly attached cross-bridges for strained cross-bridges, with a time constant of approximately 10 ms. 4. A novel multiple-exponential equation, based on the hypothesis of complete substitution of freshly attached cross-bridges for the cross-bridges that underwent the stretch, has been used to fit the whole tension transient following step stretches of different sizes (2-6 nm per half-sarcomere). For a stretch of 4 nm, the time constant of the exponential process responsible for cross-bridge detachment (tau d, 9.3 ms) almost coincides with the time constant of repriming as measured by the double-step experiments. The time constant of the exponential process representing the cumulative effects of attachment and force generation (tau 3) is 13.6 ms. 5. For stretches of different sizes the amount of quick tension recovery attributable to the reversal of the working stroke elicited by the stretches is estimated by subtracting, from the original tension transient, the contribution to tension recovery due to detachment-attachment of cross-bridges as estimated by the multiple-exponential analysis. Following this calculation, the structural change in the myosin heads responsible for the reversal of the working stroke can be 2 nm at maximum, suggesting that the elastic component in the cross-bridges is at least twice as rigid as previously thought.

Animals↗
PubMed1997

Cross-bridge kinetics studied with staircase shortening in single fibres from frog skeletal muscle.

The kinetics of actin-myosin interaction has been studied in single active muscle fibres by repetitively eliciting tension transients with staircase shortening, consisting in a sequence of step releases of identical size (1-5 nm per half-sarcomere) imposed at regular time intervals (3-11 ms). Under sarcomere length-clamp conditions, the quick phase of tension recovery following each step in the staircase is the manifestation of the working stroke by synchronized cross-bridges. Different average shortening velocities are obtained by varying both the size of the step and the time interval between steps. Ti, the tension just before each step in the sequence, T2, the tension attained at the end of the quick phase of tension recovery, decrease with the number of steps, reaching a steady state value, which is lower the larger the shortening velocity. In agreement with previous results on tension response to steady shortening, the overall shortening necessary to approach the steady state values of Ti and T2 is about 15 nm. The normalized amplitude of quick tension recovery (T2r), which is measured by the ratio of the amount of tension recovered at the end of the quick phase (T2-T1) over the tension drop simultaneous with the step (Ti-T1), has been used to measure the extent of the working stroke elicited by each step in the staircase. The steady state value of T2r decreases progressively with the increase of shortening velocity. At velocities higher than 0.5 microns s-1 per half-sarcomere the steady state value of T2r is attained after a transitory depression, which reaches a maximum for an amount of overall shortening increasing from about 8 nm up to about 13 nm with increase in shortening velocity from 0.5 to 1.4 microns s-1 per half-sarcomere. The velocity-dependent transitory depression of T2r can be explained with the mechanical-kinetic model described previously. In the model cross-bridges cycle through two pathway distinct for the kinetics of the detachment/reattachment process. Shortening promotes a redistribution of cross-bridges interacting in the isometric conditions among the various states of the force-generating process. Shortening at high speed, preventing most of cross-bridges from undergoing the relatively fast (100 s-1) detachment/reattachment process, uncovers a rate limiting step in the cycle at the end of the 12 nm working stroke. Under these conditions, the finding that the fraction of the working stroke elicited by each step is transitory depressed with respect to the steady state value reveals that in the original isometric state a large fraction of interacting cross-bridges was accumulated near the beginning of the working stroke.

Animals↗
PubMed1997

1H NMR ganglioside ceramide resonance region on the differential diagnosis of low and high malignancy of brain gliomas.

1. The high-resolution 1H NMR (MRS) spectra of human brain tumor homogenates revealed a broad resonance at 5.3-5.4 ppm in glioblastoma multiforme (N = 16) and brain metastases (N = 3). The broad resonance was identified as ceramide, a sphingosine-fatty acid combination portion of ganglioside, indicating an elevated abundance of monounsaturated fatty acids. GLC analysis of gangliosides in the highly malignant glioblastoma multiforme revealed that the elevated monounsaturated fatty acid is oleic acid (C18:1). The resonance at 5.3-5.4 ppm region was not detectable in normal human brain (N = 2), in meningiomas (N = 2), or in low-grade astrocytomas (N = 12). In normal human brain the abundance of monounsaturated fatty acid is minimal. 2. This investigation was made possible because the method of producing homogenate resulted in (i) no loss of lipids during the process and (ii) a well-homogenised sample, with (iii) no loss in chemical integrity. 3. The properties of tumor gangliosides include antigenic specificity and immunosuppressive activity and the ceramide, a sphingosine-fatty acid combination, noticeably influences the ganglioside immunosuppressive activity. 4. The observation of 1H NMR ceramide resonance in high-malignant brain tumors emphasizes the dramatic role of aberant gangliosides and ceramide precursors on the grade of malignancy and invasiveness. 5. Further insight into the specific nature of the ceramide portion of gangliosides in grading the malignancy of brain tumors should be investigated further.

Astrocytoma↗
PubMed1997

[Insulinoma. Clinical and surgical considerations concerning a case].

Insulinoma is the most common endocrine tumor of the pancreas. It arises from the beta-islet cells of Langerhans. Insulinomas synthesize and secrete insulin autonomously in the presence of low blood glucose levels, causing spontaneous hypoglycemia and characteristic clinical symptoms. The authors examined data the from the most important international research projects on this topic during the past 20 years. Insulinomas are rare, with an annual incidence of 0.5 per million population. Up to 90% patients have benign solitary pancreatic insulinomas. People of all ages can be affected with this neoplasm. The authors reported a case of a large insulinoma of the body and tail of the pancreas, with atypical psychic symptoms. A distal pancreatectomy with splenectomy was performed. No surgical complications occurred in the postoperative course. The psychic symptoms were emphasized with refusal of food. The patient underwent Parenteral Nutrition and was discharged 24 days after the operation. The surgical removal of the tumor permitted the patient to recover completely, with glucose and insulin blood levels in normal range.

Diagnosis, Differential↗
PubMed1996

Simulation of the rapid regeneration of the actin-myosin working stroke with a tight coupling model of muscle contraction.

A. F. Huxley's suggestion in Nature (1992) that a structural modification in the myosin head driven by phosphate release can explain the rapid regeneration of the working stroke, which follows the quick recovery elicited by a step release of moderate size (3-6 nm per half-sarcomere), has been tested with a theoretical model. It is assumed that, in the shortening muscle, cross-bridges can undergo their work producing interaction in two ways distinct for the biochemical state and for the amount of filament sliding allowed. During shortening at low speed, as well as after a shortening step of moderate size, phosphate release from the cross-bridge in the AM-ADP-P state promotes a 100 s-1 structural change which resets the myosin head in a configuration that allows for a new complete working stroke in the AM-ADP state. In this case the total sliding distance for interaction is about 15 nm. With the increase in shortening velocity a progressively larger fraction of interacting cross-bridges remains in the AM-ADP-P state throughout the working stroke and the sliding distance for interaction is about 11 nm. Reattachment of detached cross-bridges occurs at moderate rate whichever is the pathway from which they originate. The model predicts satisfactorily the time course of the rapid regeneration of the working stroke in double step experiments, but fails to simulate the transition to the steady state response in staircase experiments, the maximum power output during steady shortening and the decrease in rate of energy liberation at high shortening velocities. These results strengthen the conclusion of our previous modelling work where we demonstrated that the condition necessary to fit the mechanical and energetic properties of shortening muscle is to assume two pathways for cross-bridge cycling distinct for the kinetics of detachment and reattachment.

Actins↗
PubMed1996

A technique for the fast sampling of biological tissues for electron paramagnetic resonance spectroscopy.

A simple technique for the fast sampling of biological tissues for electron paramagnetic resonance (EPR) spectroscopy is described. By using tube-like stainless steel cutter, a cylindrical piece of tissue is dissected free; while holding the sample and cutter on a silicone rubber cutting base, the sample is transferred directly into the EPR test tube by pushing the tube into the instrument. The main features of the technique are its simplicity, speed, elimination of the necessity to manipulate frozen tissue (e.g., breaking, chopping, etc.), precise information on the sample volume, the ability to adjust sample size according to need, and avoidance of the use any foreign compounds (such as spin traps). The technique has been successfully applied to the EPR detection of free radicals in rabbit spinal cord exposed to ischemia. It can be easily modified for manipulation with samples in an inert atmosphere.

Animals↗
PubMed1995

Elastic distortion of myosin heads and repriming of the working stroke in muscle.

Muscle contraction is driven by a cyclical interaction between the globular head domain of myosin and the actin filaments. We used quick stretches of 5 nm per half sarcomere to synchronize the movements of myosin heads in active single muscle fibres. The intensity of the 14.5 nm X-ray reflection decreased during the stretch, showing that the instantaneous elasticity of muscle involves distortion of myosin heads. Head movement continued at about 1,500 s-1 after the stretch, accompanied by partial force recovery. This indicates a reversal of the force-generating 'working stroke' in the myosin heads that is smaller and faster than assumed previously. By 50 ms after the stretch, myosin heads have regained both their original conformation and the ability to execute a normal working stroke. This 'repriming' process is slower than that following shortening but much faster than the ATP turnover rate per myosin head.

Animals↗
PubMed1995

Angiopathy affects circulating endothelin-1 levels in type 2 diabetic patients.

To evaluate the role of endothelin-1 (ET-1), a vasoconstrictor and mitogenic peptide synthesized by endothelial cells, on the endothelial dysfunction in non-insulin-dependent diabetic (type 2) patients, we have measured the circulating ET-1 levels in 25 patients with and without clinically evident vascular complications and in a control group. Circulating ET-1 levels were significantly higher in diabetic patients with angiopathy than in diabetics without angiopathy and in controls (7.02 +/- 2.9 pg/ml vs 4.4 +/- 1.1 pg/ml and 3.08 +/- 0.7 pg/ml, respectively; P < 0.001). No difference was demonstrated between diabetic patients without angiopathy and controls. These findings suggest that ET-1 may be a marker for arterial vascular disease only in patients with overt angiopathy. It is unclear whether it participates in the endothelial injury process or it is merely released from damaged endothelial cells.

Biomarkers↗
PubMed1995

A cross-bridge model that is able to explain mechanical and energetic properties of shortening muscle.

The responses of muscle to steady and stepwise shortening are simulated with a model in which actin-myosin cross-bridges cycle through two pathways distinct for the attachment-detachment kinetics and for the proportion of energy converted into work. Small step releases and steady shortening at low velocity (high load) favor the cycle implying approximately 5 nm sliding per cross-bridge interaction and approximately 100/s detachment-reattachment process; large step releases and steady shortening at high velocity (low load) favor the cycle implying approximately 10 nm sliding per cross-bridge interaction and approximately 20/s detachment-reattachment process. The model satisfactorily predicts specific mechanical properties of frog skeletal muscle, such as the rate of regeneration of the working stroke as measured by double-step release experiments and the transition to steady state during multiple step releases (staircase shortening). The rate of energy liberation under different mechanical conditions is correctly reproduced by the model. During steady shortening, the relation of energy liberation rate versus shortening speed attains a maximum (approximately 6 times the isometric rate) for shortening velocities lower than half the maximum velocity of shortening and declines for higher velocities. In addition, the model provides a clue for explaining how, in different muscle types, the higher the isometric maintenance heat, the higher the power output during steady shortening.

Actins↗
PubMed1995

Changes in the x-ray diffraction pattern from single, intact muscle fibers produced by rapid shortening and stretch.

Changes in the x-ray diffraction patterns produced by 100-microseconds-length steps imposed during tetanic stimulation were recorded from single intact fibers of frog tibialis anterior muscle. For shortening steps, a staircase length change was applied, with a 20-ms interval between steps. For stretches, each 20-ms cycle started with a stretch, followed after 4 ms by shortening to the original length. Each shortening step in the staircase and each stretch in the stretch/shortening protocol produced a response similar to that of a single step from the isometric steady state. The intensity of the 14.5-nm x-ray reflection arising from the axial repeat of the myosin heads along their filaments decreased after both shortening and stretch; this decrease was not accompanied by broadening along or across the meridian. The relationship between the intensity after the length step and step amplitude was approximately linear for both stretches and shortening steps, extrapolating to zero intensity for 11-nm stretches and 13-nm shortening steps, but there was no significant intensity change for the first approximately 2 nm of shortening. These results are broadly consistent with conventional models of muscle contraction in which myosin heads move through about 10 nm during the working stroke in the shortening direction, but an additional distortion of the myosin heads may be produced by a stretch.

Animals↗
PubMed1994

The effect of hypertonicity on force generation in tetanized single fibres from frog skeletal muscle.

1. We compared the tension transient that follows a step change in sarcomere length in normal Ringer solution with that in Ringer solution made hypertonic by the addition of 98 mM sucrose. Steps were applied on tetanized single muscle fibres during either the isometric plateau or the steady force response to lengthening at low speed. Sarcomere length was controlled on selected fibre segments by a striation follower. Analysis is limited to phase 1 (the tension change simultaneous with the length step, mainly due to cross-bridge elasticity) and phase 2 (the quick phase of tension recovery, a manifestation of the cross-bridge elementary force-generating process). 2. At the isometric tetanus plateau the steady force is reduced by 19% in hypertonic solution, and the stiffness is slightly increased. During slow lengthening both steady force and stiffness are similar in normal solution and in hypertonic solution. In hypertonic solution the tension-to-stiffness ratio, a measure of the mean cross-bridge extension before the step, is markedly reduced in isometric conditions (-23%), but not during lengthening (-2%). 3. The plots of instantaneous tension versus the length change during the step show that in hypertonic medium the elasticity of the fibre is almost undamped. Thus the increase in stiffness cannot be attributed to an increase in viscosity. 4. In isometric conditions (T2-T1)/(Ti-T1), the proportion of the initial tension drop recovered at the end of phase 2, is not affected by hypertonicity for releases of moderate and large size (> 2 nm) and is reduced for small releases (< 2 nm) and for stretches. The abscissa intercept of the relation (T2-T1)/(Ti-T1) versus step amplitude is the same in both media. During lengthening, for releases of small and moderate size, (T2-T1)/(Ti-T1) is 20% lower in hypertonic solution. For large releases the slope of the relation is lower so that the abscissa intercept is not changed. 5. The speed of quick tension recovery following a step length change imposed in isometric conditions is slightly depressed in hypertonic solution. The relation between speed of recovery and step amplitude maintains its shape and is shifted downwards. During lengthening, the speed of quick tension recovery in hypertonic solution is less dependent on step amplitude than in normal solution, as if a more linear viscoelasticity is responsible for a large fraction of residual recovery.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1993

Kinetics of regeneration of cross-bridge power stroke in shortening muscle.

The force developed by a muscle during steady shortening is due to cyclic interactions between the cross-bridges extending from the thick myosin filament to the thin actin filament. Each interaction consists of a power stroke of the myosin molecule that accounts for a limited amount of sliding between the two sets of filaments (about 12 nm according to quick release experiments), and is widely believed to be coupled to the hydrolysis of one ATP molecule. On the other hand both energetics studies in muscle and in vitro motility assays, indicating that shortening per ATP split is much larger than 12 nm, postulate that during shortening cross-bridges interact at a rate much faster than the ATP splitting rate. In the experiments reported here, made on intact fibres from frog skeletal muscle, the rate of regeneration of the power stroke was determined. Tension transients were elicited by imposing test step releases at different times (2-20 ms) after a conditioning release of about 5 nm. When the test step was imposed at 2 ms after the conditioning step, the tension attained at the end of the quick phase of recovery (T2, due to the force generating stroke of the attached cross-bridges) was depressed and the T2 curve (the plot of T2 tension versus size of the test step) intercepted the length axis to the right, with respect to the intercept of the control T2 curve, by an amount similar to the size of the conditioning step. By increasing the interval between conditioning and test step the T2 tension increased progressively and the T2 curve intercept approached the intercept of the control curve with a time constant of 6-7 ms. These results indicate that the force generating stroke elicited by a shortening step is followed by a relatively rapid process of detachment and reattachment by most of the cross-bridges, allowing for the generation of another power stroke. The rate of this process, 150/s, is one order of magnitude higher than that expected from the ATPase rate, suggesting that several actomyosin interactions occur in shortening muscle by the time one ATP is split. The results are stimulated with a mechanical kinetic model of contraction, in which, for a critical amount of shortening, cross-bridges can detach, rapidly reattach and generate force before the completion of the "normal" isometric cycle.

Animals↗