In vitro assessment of factors affecting the apparent diffusion coefficient of Jurkat cells using bio-phantoms.

It is well known that many tumor tissues show lower apparent diffusion coefficient (ADC) values, and that several factors are involved in the reduction of ADC values. The aim of this study was to clarify how much each factor contributes to decreases in ADC values. We investigate the roles of cell density, extracellular space, intracellular factors, apoptosis and necrosis in ADC values using bio-phantoms. The ADC values of bio-phantoms, in which Jurkat cells were encapsulated by gellan gum, were measured by a 1.5-Tesla magnetic resonance imaging device with constant diffusion time of 30sec. Heating at 42℃ was used to induce apoptosis while heating at 48℃ was used to induce necrosis. Cell death after heating was evaluated by flow cytometric analysis and electron microscopy. The ADC values of bio-phantoms including non-heated cells decreased linearly with increases in cell density, and showed a steep decline when the distance between cells became less than 3µm. The analysis of ADC values of cells after destruction of cellular structures by sonication suggested that approximately two-thirds of the ADC values of cells originate from their cellular structures. The ADC values of bio-phantoms including necrotic cells increased while those including apoptotic cells decreased. This study quantitatively clarified the role of the cellular factors and the extracellular space in determining the ADC values produced by tumor cells. The intermediate diffusion time of 30msec might be optimal to distinguish between apoptosis and necrosis.

Extracellular sheets and tunnels modulate glutamate diffusion in hippocampal neuropil.

Although the extracellular space in the neuropil of the brain is an important channel for volume communication between cells and has other important functions, its morphology on the micron scale has not been analyzed quantitatively owing to experimental limitations. We used manual and computational techniques to reconstruct the 3D geometry of 180 µm(3) of rat CA1 hippocampal neuropil from serial electron microscopy and corrected for tissue shrinkage to reflect the in vivo state. The reconstruction revealed an interconnected network of 40-80 nm diameter tunnels, formed at the junction of three or more cellular processes, spanned by sheets between pairs of cell surfaces with 10-40 nm width. The tunnels tended to occur around synapses and axons, and the sheets were enriched around astrocytes. Monte Carlo simulations of diffusion within the reconstructed neuropil demonstrate that the rate of diffusion of neurotransmitter and other small molecules was slower in sheets than in tunnels. Thus, the non-uniformity found in the extracellular space may have specialized functions for signaling (sheets) and volume transmission (tunnels).

Knee bioelectric impedance assessment in healthy/with osteoarthritis subjects.

The present study analyzes parameters estimated by bioelectric impedance spectroscopy (BIS) in subjects with healthy and with osteoarthritis (OA) knees. Thirty-two male volunteers, members of the Parachute Military Infantry Brigade of Rio de Janeiro, Brazil, participated in the study (62 knee joints). Clinical specialists used the Dejour scale for OA classification and divided the subjects into a control (without OA) and a pathological group (with different degrees of OA). BIS data were obtained in a standing position using a BIS technique based on the current response to a step voltage excitation. Differences between groups were measured by means of a Wilcoxon-Mann-Whitney test. Results indicate that raw bioimpedance parameters seem to be sensitive to the physiological changes associated with OA. Thus, data indicate that extra-cellular resistance (Re) and reactance of the equivalent capacitance (Xcx) increase according to the disease intensity (p < 0.001). In conclusion, the BIS technique seems to be able to provide the objective and non-invasive basis for helping the diagnosis of knee OA.

Bioimpedance spectroscopy for clinical assessment of fluid distribution and body cell mass.

Body composition assessment has been used to evaluate clinical interventions in research trials, and has the potential to improve patient care in the clinical setting. Body cell mass (BCM) is an important indicator of nutrition status; however, its measurement in the clinic has been limited. BCM can be estimated by the measurement of intracellular water (ICW). The assessment of extracellular water (ECW) is also important because many clinical populations undergo alterations in fluid distribution, particularly individuals with wasting, those receiving dialysis, and obese individuals. Bioimpedance spectroscopy (BIS) is a unique bioimpedance approach that differs in underlying basis from the more readily recognized single-frequency bioelectrical impedance analysis (SF-BIA) in that it does not require the use of statistically derived, population-specific prediction equations. It has the potential advantage of not only measuring total body water (TBW), as does SF-BIA, but also offering the unique capacity to differentiate between ECW and ICW and, thus, to provide an estimate of BCM. This literature review was conducted to compare available BIS devices to multiple dilution for measuring fluid compartments or BCM in a number of populations. Variable results regarding the ability of BIS to measure absolute volumes, as well as the observation of wide limits of variation, make BIS problematic for individual assessment in the clinic, particularly in populations with abnormal fluid distribution or body geometry. BIS has been found to be more accurate for measuring changes in fluid volumes or BCM, particularly in post-surgical and human immunodeficiency virus (HIV)-infected individuals. It is certainly possible that population-specific adjustments may improve the accuracy of BIS for assessing individuals in the clinical setting; however, additional research and development is needed before the method can be accepted for routine clinical use.

Use of bioelectrical impedance in hydration status assessment: reliability of a new tool in psychophysiology research.

Adequate hydration is crucial in maintaining optimal physical and mental functioning and the need for a fast and reliable hydration status assessment in behavioral medicine research has become increasingly important. The goal of this study was to determine the reliability of bioelectrical impedance assessment (BIA) in assessing total body water (TBW), extracellular water (ECW) and intracellular water (ICW) and to assess whether individuals can be reliably classified as being hypohydrated or hyperhydrated using lower and upper quartiles, respectively. TBW, ECW and ICW were assessed via BIA (Bodystat, Isle of Man, UK) in 52 male and 48 female college students on 2 separate days within 1 week. Results revealed strong test-retest reliability for TBW (r=0.983), ECW (r=0.972) and ICW (r=0.988) (all P's<0.001). Following the initial and follow-up assessments, participants were then classified as being either hypohydrated or hyperhydrated based on the percentage of body weight accounted for by TBW. Test-retest reliability of hydration status within classifications was then assessed by gender. Test-retest reliability was found for TBW, ECW and ICW among hypohydrated (r=0.985, r=0.972 and r=0.99, respectively) and hyperhydrated (r=0.994, r=0.989 and r=0.994, respectively) males (all P's<0.001). Significant test-retest correlations were also found for females classified as being hypohydrated (r=0.97, r=0.956 and r=0.976, respectively) and hyperhydrated (r=0.973, r=0.976 and r=0.976, respectively) (all P's<0.001). These findings suggest that hydration status, as indexed by bioelectrical impedance technique, is reliable across time and is also reliable within individuals who are chronically hyperhydrated or hypohydrated.

Assessment of skeletal muscle triglyceride content by (1)H nuclear magnetic resonance spectroscopy in lean and obese adolescents: relationships to insulin sensitivity, total body fat, and central adiposity.

The metabolism and composition of skeletal muscle tissue is of special interest because it is a primary site of insulin action and plays a key role in the pathogenesis of insulin resistance. Intramyocellular (IMCL) triglyceride stores are an accessible form of energy that may decrease skeletal muscle glucose utilization, thereby contributing to impaired glucose metabolism. Because of the invasive nature of muscle biopsies, there is limited, if any, information about intramuscular lipid stores in children. The development of (1)H nuclear magnetic resonance (NMR) spectroscopy provides a unique noninvasive alternative method that differentiates intracellular fat from intercellular fat in muscle tissue. The present study was performed to determine whether IMCL and extramyocellular (EMCL) lipid contents are increased early in the development of juvenile obesity and to explore the relationships between IMCL and EMCL to in vivo insulin sensitivity, independently of total body fat and central adiposity in obese and nonobese adolescents. Eight nonobese (BMI 21 kg/m(2), age 11-16 years) and 14 obese (BMI 35 +/- 1.5 kg/m(2), age 11-15 years) adolescents underwent 1) (1)H-NMR spectroscopy to noninvasively quantify IMCL and EMCL triglyceride content of the soleus muscle, 2) a 2-h euglycemic-hyperinsulinemic clamp (40 mU.m(-2).min(-1)) to assess insulin sensitivity, 3) a dual-energy X-ray absorptiometry scan to measure total percent body fat, and 4) magnetic resonance imaging to measure abdominal fat distribution. Both the IMCL and EMCL content of the soleus muscle were significantly greater in the obese adolescents than in the lean control subjects. A strong inverse correlation was found between IMCL and insulin sensitivity, which persisted and became even stronger after controlling for percent total body fat and abdominal subcutaneous fat mass (partial correlation r = -0.73, P < 0.01) but not when adjusting for visceral fat (r = - 0.54, P < 0.08). In obese adolescents, increase in total body fat and central adiposity were accompanied by higher IMCL and EMCL lipid stores. The striking relationships between both IMCL and EMCL with insulin sensitivity in childhood suggest that these findings are not a consequence of aging but occur early in the natural course of obesity.

Assessment of fluid status in peritoneal dialysis patients.

OBJECTIVES: To assess the influence of abnormalities in fluid status and body composition on agreement between multifrequency bioimpedance analysis (MF-BIA), segmental BIA (sigmaBIA), the Watson formula, and tracer dilution techniques. DESIGN: Cross-sectional. SETTING: Multicenter. PATIENTS: 40 patients (29 males, 11 females) on peritoneal dialysis (PD). MAIN OUTCOME MEASURES: Agreement between the various techniques used to assess total body water (TBW) [MF-BIA, deuterium oxide (D2O), and the Watson formula] and extracellular water (ECW) [MF-BIA, bromide dilution (NaBr), and sigmaBIA], also in relation to the relative magnitude of the body water compartments [ECW (NaBr):body weight (BW) and TBW (D2O):BW] and body composition (DEXA). Second, the relation between body water compartments with echocardiographic parameters. RESULTS: Wide limits of agreement were observed between tracer dilution techniques and MF-BIA [TBW (D2O - MF-BIA) 2.0 +/- 3.9 L; ECW (NaBr - MF-BIA) -2.8 +/- 3.9 L], which were related to the relative magnitude of the body water compartments: r = 0.70 for ECW and r = 0.40 for TBW. sigmaBIA did not improve the agreement [ECW (NaBr-sigmaBIA): 3.7 +/- 2.9 L]. Also, wide limits of agreement were observed between D2O and the Watson formula (-2.3 +/- 3.3 L). The difference between D2O and Watson was related to hydration state and to percentage of fat mass (r = 0.70 and r = -0.53, p < 0.05). Both ECW and TBW as assessed by BIA and tracer dilution were related to echocardiographic parameters. CONCLUSION: Wide limits of agreement were found between MF-BIA and sigmaBIA with dilution methods in PD patients, which were related to hydration state itself. The disagreement between the Watson formula and dilution methods was related to both hydration state and body composition.

Role of bioimpedance spectroscopy in assessment of body water compartments in hemodialysis patients.

Bioimpedance spectroscopy (BIS) has been advocated as a tool to assess fluid status in hemodialysis (HD) patients. However, uncertainty remains about the reliability of BIS in patients with abnormalities in fluid status. Aims of the study are to assess the agreement between total-body water (TBW) and extracellular volume (ECW) measured by BIS and tracer dilution (deuterium oxide [D(2)O] and sodium bromide [NaBr]), the influence of the relative magnitude of water compartments (expressed as TBW(D(2)O) and ECW(NaBr):body weight) on the agreement between BIS and tracer dilution, and the ability of BIS to predict acute changes in fluid status. BIS and tracer dilution techniques were performed in 17 HD patients before a dialysis session. Moreover, the relation between BIS and gravimetric weight changes was assessed during both isolated ultrafiltration and HD. Correlation coefficients between TBW and ECW measured by BIS and tracer dilution were r = 0.71 and r = 0.71, respectively. Mean differences (tracer-BIS) were 6.9 L (limits of agreement, -1.5 to 21.6 L) for TBW and 2.3 L (limits of agreement, -1.7 to 9.7 L) for ECW. There was a significant relationship between the relative magnitude of TBW and ECW compartments and disagreement between BIS and tracer dilution (r = 0.65 and r = 0.77; P < 0.05). During both isolated ultrafiltration and HD, there was a significant relation between gravimetric changes and change in ECW(BIS) (r = 0.83 and r = 0.76; P < 0.05), but not with change in TBW(BIS). In conclusion, agreement between BIS and tracer dilution techniques in the assessment of TBW and ECW in HD patients is unsatisfactory. The discrepancy between BIS and dilution techniques is related to the relative magnitude of body water compartments. Nevertheless, BIS adequately predicted acute changes in ECW during isolated ultrafiltration and HD, in contrast to changes in TBW.

Non-invasive assessment of fluid volume status in the interstitium after haemodialysis.

During dialysis excess fluid is removed from uraemic patients. The excess fluid is mainly located in the skin and subcutaneous tissues. In this study we wished, with two noninvasive techniques, the IM (impression method) and BIA (bioimpedance analysis), to study what mechanical (IM) and electrical cellular membrane (BIA) effects the fluid withdrawal has on these tissues. The IM measures the resistive force of the tissues when mechanically compressed. From the force curve two parameters are calculated, the F(0), indicative of interstitial tissue pressure and the FT corresponding to the translocation of tissue fluid (interstitial movable water). The BIA phase angle shift (phi), i.e. geometrical angular transformation of the ratio between reactance and resistance, which has been associated with cellular membrane function, was used as a measurement of electrical cellular membrane effects. Twenty patients were studied before and after haemodialysis measuring the F(0), FT and phi. Theresults showed that the patients lost a median of 3.7 kg during the haemodialysis. F(0) increased until after dialysis, but did not reach significant values, whereas FT increased significantly after dialysis, p < 0.001, as compared with before. After a peak at one hour postdialysis the FT value returned to predialysis values at four hours after termination of dialysis. Also phi increased from before to after dialysis, p < 0.001, but already after one hour it returned to predialysis values. It is common knowledge that dialysis alters the dynamics of fluid in the interstitium of the skin and subcutis. We conclude that the impression method is sensitive enough to detect and chronicle these changes. Furthermore, with the BIA, (phase angle) signs of changes in the electrical properties of the tissues, possibly reflecting cellular membrane function, could be detected.

Analytic assessment of the various bioimpedance methods used to estimate body water.

Knowledge of patient fluid distribution would be useful clinically. Both single-frequency (SF) and impedance modeling approaches are proposed. The high intercorrelation between body water compartments makes determining the best approach difficult. This study was conducted to evaluate the merits of an SF approach. Mathematical simulation was performed to determine the effect of tissue change on resistance and reactance. Dilution results were reanalyzed, and resistance and parallel reactance were used to predict the intracellular water for two groups. Results indicated that the amount of intracellular and extracellular water conduction at any SF can vary with tissue change, and reactance at any SF is affected by all tissue parameters. Modeling provided a good prediction of dilution intracellular and extracellular water, but an SF method did not. Intracellular, extracellular, and total body water were equally predicted at all frequencies by SF resistance and parallel reactance. Extracellular and intracellular water are best measured through modeling, because only at the zero and infinite frequencies are the results sensitive only to extracellular and intracellular water. At all other frequencies there are other effects.

Patient assessment and examples of a method of treatment. Use of the circulator boot in peripheral vascular disease.

Effective peripheral blood flow is positively related to cardiac output and gravity (part dependent) and inversely related to gravity (part elevated), venous pressure, interstitial fluid pressure, degree of peripheral neuropathy, arteriosclerotic and thrombotic arterial obstructions, and infection. These factors are considered in the operation of the end-diastolic pneumatic boot in the treatment of illustrative patients with lymphedema, venous stasis disease, peripheral arteriosclerosis obliterans, peripheral neuropathy, cellulitis, and osteomyelitis and the failing heart. A method of treatment that includes the use of the boot and the injection of local antibiotics is described.

Comparison of bio-impedance spectroscopy and multi-frequency bio-impedance analysis for the assessment of extracellular and total body water in surgical patients.

1. Measurements of extracellular and total body water provide useful information on the nutritional status of surgical patients and may be estimated from whole-body bio-impedance measurements at different frequencies. 2. Resistance and reactance were measured at 50 frequencies from 5kHz to 1MHz in 29 surgical patients (17 males, 12 females) with a wide range of extracellular to total body water ratios. 3. A fit to the spectrum of reactance versus resistance data gave predicted resistances at frequencies zero and infinity. Values of extracellular and total body water determined by this bio-impedance spectroscopy technique were regressed against values obtained from radioisotope dilution. The standard errors of the estimate were 1.8931 and 3.2591 respectively. 4. Resistance indices (height2/resistance) at selected frequencies gave the highest correlations with extracellular and total body water at 5kHz and 200kHz respectively, and prediction equations derived from multiple stepwise regressions also showed these to be the optimum frequencies. The standard errors of the estimate for this multi-frequency bio-impedance analysis method were 1.9371 and 2.6061 for extracellular and total body water respectively. 5. To assess the ability of the two methods to measure changes in extracellular and total body water, reproducibility was assessed from repeat measurements 10 min apart in a subgroup of 15 patients. Bio-impedance spectroscopy gave mean coefficients of variation for extracellular and total body water of 0.9% and 3.0% respectively. For multi-frequency bio-impedance analysis the corresponding coefficients of variation were 0.9% and 0.6%. 6. It is concluded that a simple impedance analyser operating at only two frequencies compares favourably with the more complex spectroscopy technique for the determination of extracellular and total body water in surgical patients.

Impression technique for the assessment of oedema: comparison with a new tactile sensor that measures physical properties of tissue.

To measure tissue oedema, the impression technique and a new tactile sensor technique are compared and evaluated in a silicone rubber model and in an in vivo rat testis model. The principles of the two techniques differ in that the impression technique evaluates interstitial fluid flow FT and peak force F(O) when tissue is compressed, whereas the tactile sensor evaluates the hardness/softness or change in resonance frequency delta f when a vibrating rod is attached to tissue. Both techniques can detect changes in silicone hardness/softness or in hormone-induced changes of testis interstitial fluid. Although both F(O) and FT are significantly correlated to delta f in the experiments, it is concluded that F(O) is the most promising impression parameter to give valuable information about the hardness of living tissue as compared with delta f. The comparison indicates that the impression technique is the most easy to interpret, non-invasive tool to assess tissue oedema so far developed.

Assessment of refill and hypovolaemia by continuous surveillance of blood volume and extracellular fluid volume.

During renal replacement therapy hypovolaemia due to ultrafiltration (UF) may, when not sufficiently counteracted by refill from the interstitium, result in hypotension. Combining two recently developed methods the haemodynamic process of refill was studied in order to find characteristics featuring hypotension. Relative blood volume (BV) and extracellular fluid volume (EFV) were measured continuously in 40 stable haemodialysis patients by means of an optical and a conductivity technique respectively. Regarding their postdialytic (PD) EFV the patients were divided into three groups: normohydrated (N, n = 20), dehydrated (D, n = 11) and overhydrated (O, n = 9). Significant differences between the groups were assessed in BV decrease (after 2 h until the end of treatment P < 0.05 and after 3 h P < 0.01), EFV decrease (after 3 h P < 0.05) and occurrence of hypotensive episodes (N,5; D,7; O,none; P < 0.01). During the entire session the speed of BV decrease was significantly higher in hypotensive patients (H) than in non-hypotensive patients (non-H). At the moment of hypotension (after 141 +/- 49 min) residual BV was less (P < 0.0005) in H (87.7 +/- 5.2%) than at the corresponding moment in non-H patients (96.5 +/- 4.0%). PD BV and PD EFV, both expressed as a percentage of the starting value, correlated significantly (r = 0.63, p < 0.005) and UF-volume (differences between the groups were not significant) correlated to EFV decrease (r = 0.45, P < 0.005). In conclusion, the combination of both non-invasive methods elucidates the pathophysiology of UF-induced hypotension and provides a means of reducing dialysis morbidity. The influence of tissue hydration state on these variables has been shown.

The assessment of colonic water retention prior to double-contrast enema.

The detection of colonic water retention prior to double-contrast enema (DCE) was studied in 41 outpatients. Ultrasound examination of the cecum (USC) correctly showed 4 of the 5 patients with water retention as well as all 14 cases with dry colon. Changes in patient's weight or in the number of cleansing enemas were not associated with water retention. The USC seems to offer a rapid and accurate method for the detection of colonic water retention prior to DCE. Aspects of the handling of colonic water retention are also discussed.

The role of Ca influx in cardiac muscle excitation-contraction coupling. Assessment by extracellular Ca microelectrodes.

Tension, dT/dt, and extracellular free [Ca] [( Ca]0) were continuously measured in isolated rabbit right papillary muscles at 28 degrees C. Double-barreled Ca-selective microelectrodes incorporating Simon's neutral Ca exchange resin (ETH-1001) were used to monitor [Ca]0. Decreases of [Ca]0 were seen during the course of single twitches, before the development of significant tension. This depletion of Ca0 probably represents Ca influx into the cells from the extracellular space. The magnitude of this Ca0 depletion is decreased by Co, verapamil, long rest intervals, and reduction of [Ca]0. The magnitude is increased by catecholamines, reduction of [Na]0, caffeine, continued pacing, and elevation of [Ca]0. After 3-min rest, stimulation (0.5-1 Hz) produces a biphasic tension response (larger first beat, small second beat, and monotonic rise to control). Caffeine (5 mM) changes the pattern after rest to a monotonic increase. Ca influx increases monotonically in both cases. Addition of 20 mM Co during the rest reduces tension of all beats by similar amounts in the presence of caffeine. In the absence of caffeine, Co has a much weaker effect on the first beat than on subsequent beats. The results suggest that caffeine inhibits an intracellular component of activator Ca that is more important after a rest interval, but that Ca influx becomes increasingly more important during continued pacing under the conditions used here.